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Some effects of supplemental carbon dioxide on the physiology of plant growth and development Hicklenton, Peter R. 1978

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SOME EFFECTS OF SUPPLEMENTAL CARBON DIOXIDE ON THE PHYSIOLOGY OF PLANT GROWTH AND DEVELOPMENT  by  PETER ROBERT HICKLENTON B.Sc, University of Wales (Swansea) 1973 M.Sc, McGill University 1975  A THESIS SUBMITTED IN PARTIAL FULFILMENT OF THE REQUIREMENTS FOR THE DEGREE OF DOCTOR OF PHILOSOPHY in  THE FACULTY OF GRADUATE STUDIES (Department of Plant Science)  We accept this thesis as conforming to the required standard  THE UNIVERSITY OF BRITISH COLUMBIA October 1978  ©  Peter Robert Hicklenton  In p r e s e n t i n g t h i s  thesis  an a d v a n c e d  degree at  the L i b r a r y  s h a l l make  I  f u r t h e r agree  for  the U n i v e r s i t y it  freely  f u l f i l m e n t o f the r e q u i r e m e n t s of B r i t i s h  available  s c h o l a r l y p u r p o s e s may be g r a n t e d by  this  written  thesis  It  permission.  Department  of  The U n i v e r s i t y  ? w f t t » . T  ^ c x E A t t t  o f B r i t i s h Columbia  2075 Wesbrook Place Vancouver, Canada V6T 1W5  Date  gain shall  Occet\te»-  i?. v^iP  I agree  for  that  and s t u d y .  copying of t h i s  thesis  t h e Head o f my D e p a r t m e n t  is understood that  for financial  Columbia,  for reference  that permission for extensive  by h i s r e p r e s e n t a t i v e s . of  in p a r t i a l  or  copying or p u b l i c a t i o n  n o t be a l l o w e d w i t h o u t my  ABSTRACT  This research was concerned with some physiological effects of supra-normal CG^ concentrations on cucumber, tomato and Japanese Morning Glory (Pharbitis  nil),  and with measurement of CO^ levels i n a  commercial greenhouse. Measurements of CG^ concentrations in a cucumber greenhouse showed that, i n the early stages of crop development, early morning CC^ levels reached 0.18% as a result of straw decomposition i n the plant beds. Later i n crop development, daytime  levels were much lower and  required gas combustion to restore high concentrations.  Stomatal  resistances i n cucumber leaves were relatively insensitive to high greenhouse  concentrations.  Variation i n stomatal resistance  through the crop canopy was, however, detected.  Generally, the two  most recently developed leaves showed higher resistances than those of a s l i g h t l y greater physiological age.  Differences i n leaf irradiance  could not f u l l y explain this effect, which may be related to the stage of leaf development. Subsequent experiments on greenhouse tomato crops showed that C^-enriched  (0.09% CT^) plants flowered earlier and produced 30% more  f r u i t than those grown in normal a i r .  Photosynthetic rates were  inherently higher i n apical and basal leaves developed under CO^ enrichment -1 -2.  at irradiances above 50  m. s .Behavioral indicies of photosynthetic  efficiency indicated an enhanced capacity to u t i l i z e C O 2 in enriched plants.  Measurements of CO^ exchange i n leaves of plants grown i n chambers at 3 CG^ concentrations (0.03, 0.1 and 0.5%) confirmed the enhancement of inherent photosynthetic rates i n young leaves of 0.1% grown plants. Reduced rates df photorespiration, total 0^ inhibition of photosynthesis, glycolate oxidase (GaO) a c t i v i t y , and an increased rate of ribulose-biphosphate-carboxylase (RuBP-case) a c t i v i t y , contributed to this enhancement. Maximum photosynthetic rates i n young leaves developed at 0.5% CO^ were similar to those developed in 0.03% CO^.  Growth rates of the 0.1% C02~grown plants were higher than the  similar rates of plants from the 0.03 and 0.5% regimes.  Apparently  maximum benefit from CO2 enrichment i s achieved by maintaining atmospheric CO2 concentrations close to 0.1%.  At a later stage of development,  however, GaO and RuBP-case a c t i v i t i e s were similar i n the 0.03 and 0.1% C02-grown plants and photosynthetic rates did not d i f f e r between growth regimes. Observations on the effects of 0.03, 0.1, 1.0 and 5.0% C0 on development i n the Short-Day Plant  Pharbitis nil  2  revealed that 1.0  and 5.0% CO2 modified normal flowering. These concentrations induced a weak flowering response in Long-Days and Short-Days and promoted stem elongation and leaf production under both photoperiods.  These  modifications were apparently unrelated to patterns of CO2 exchange which showed a relatively small increase above 0.5% CO2. These results are discussed i n relation to possible mechanisms for the effects of supra-normal CO^- concentrations on development. The diversity of physiological effects mediated by CX^, and their relationship to one another are discussed.  iv  TABLE  OF  CONTENTS Page  GENERAL INTRODUCTION PART I  1  Studies i n a cucumber greenhouse: Atmospheric CO^ concentration and leaf stomatal resistance.  INTRODUCTION  8  MATERIALS AND METHODS i) ii)  Conductimetric CO^ Analyser  12  Greenhouse Description and Crop Management Practices  iii) iv)  17  Measurements of Greenhouse CO^ Concentration . . 19 Measurements of Stomatal Resistance  21  RESULTS AND DISCUSSION i) ii)  Greenhouse CO^ Concentrations (Experiment 1) . . 23 In situ  Measurements of Stomatal Resistance  (Experiment 1)  33  PART I I The effect of atmoshperic CO^-enrichment on the growth, yield and gas-exchange physiology of the tomato plant.  .......  INTRODUCTION  47  PART I I , SECTION 1 Experiments on Greenhouse Crops MATERIALS AND METHODS i)  Selection of Suitable Greenhouses for a Comparative Study (Experiment 2)  52  V  Page  ii)  CC^ Enrichment Experiments (Experiments 3 -  and 4)  55  PART I I , SECTION 1 RESULTS AND DISCUSSION i)  Comparative Study Concerning Growth of Tomato Crops i n Unenriched Greenhouses (Experiment 2).  ii)  C0  2  64  Concentrations i n CO^-Enriched and Control  (Normalair) Greenhouses  71  iii)  Plant Growth and Y i e l d (Experiment  iv)  Leaf Photosynthesis (Experiment 4)  78  Resistances to C0 Assimilation  86  v) vi)  3)  73  2  Behavioral Indicies of Photosynthetic Responses to C0 Enrichment  90  2  PART I I , SECTION 2  Experiments on seedlings grown under controlled environment  conditions  MATERIALS AND METHODS i) ii) iii)  Growth Chambers and Control Systems  96  Gas Exchange Measurement System  102  Experimental Design  105  PART I I , SECTION 2 RESULTS AND DISCUSSION i)  Analysis of Plant Growth i n Relation to C0 Concentration (Experiment 5)  2  113  Page ii)  Effects of Growth CO^ Concentration on Photosynthesis, Photorespiration and Enzyme A c t i v i t y i n Leaf 3 at LPI^^.O  (Experiments  6 and 8) iii)  122  Comparison of the Effects of Growth CO^ Concentration on Photosynthesis, Photorespiration and Enzyme A c t i v i t y i n Leaf 3 at LPI^:7.5 and LPI :2.0 (Experiments 3  iv)  6 and 8)  Replacement Experiment (Experiment  145 7)  162  '  169  PART I I DISCUSSION  PART I I I The influence .of supra-normal CO^ concentrations on photosynthesis, and vegetative and reproductive development of Pharbitis  n i l plants i n Long and  Short Days INTRODUCTION  178  MATERIALS AND METHODS i) ii)  Growth System  183  Experiments a)  Effects of Growth CO^ Concentration and Photoperiod on Development (Experiment  b)  9) • 187  E f f e c t s of Growth CO2 Concentration and Photoperiod on Photosynthesis Rate and Stomatal Resistances (Experiment  10). . . . 190  vii  Page RESULTS i)  E f f e c t s of CO^ Concentration on Flowering and Vegetative Growth i n LD and SD (Experiment 9) .  ii)  192  Effects of 0.03 and 1.0% C0 During Growth i n 2  LD and SD on Subsequent Rates of Net PhotosynC • thesis and Stomatal Resistances (Experiment ". 10)  208  DISCUSSION  214  GENERAL DISCUSSION AND CONCLUSIONS  228  LITERATURE CITED  238  APPENDIX 1  261'  APPENDIX 2 .  263  APPENDIX 3  265  viii  LIST  OF TABLES  .  Page II-l  Functions of plant dry weight and leaf area with time derived from multiple regression equations for tomato plants grown i n experimental greenhouses A and B, without CC^ enrichment  II-2  65  Mean f r u i t y i e l d from tomato plants grown i n experimental greenhouses A and B, from March 16 to May 18, 1978, without CC^ enrichment  II-3  70  Time course of leaf development i n tomato plants grown with or without CO^ enrichment  II-4  74  compensation points f o r leaves of tomato plants grown with b r without CO 2 enrichment  84  N  II-5  Behavioral indices: for photosynthetic  exchange:  u t i l i z a t i o n ef f iciency (E^), radiant energy u t i l i z a t i o n e f f i c i e n c y (E^.)', and maximum photosynthetic C0 II-6  2  flux (F.^)  9  1  Comparison of observed rates of net photosynthesis with rates predicted by the model of Van Bavel, for a test C0 concentration of 340 y l l "  1  2  II-7  Experiment  94  6 - CO2 exchange of leaf 3 of tomato  plants grown at 3 CO2 concentrations i n chambers, at 2 stages of development: design  Summary of experimental 110  ix  Page II-8  E f f e c t s of CC^ c o n c e n t r a t i o n on n e t a s s i m i l a t i o n r a t e (NAR), r e l a t i v e growth r a t e (RGR) and l e a f ratio  area  (LAR) i n tomato p l a n t s grown a t 3 CC^  c o n c e n t r a t i o n s , i n chambers II-9  11-10  120  compensation p o i n t s of l e a f 3 of tomato p l a n t s grown a t 3 CO2  c o n c e n t r a t i o n s , measured a t 2 stages  o f development  (Leaf P l a s t o c h r o n index, LPI)  132  Components of oxygen i n h i b i t i o n of p h o t o s y n t h e s i s i n l e a f 3 o f tomato p l a n t s grown a t 3  CO2  c o n c e n t r a t i o n s , a t L e a f P l a s t o c h r o n Index:  2  . (LPI :2)  138  3  11-11  Components of oxygen, i n h i b i t i o n of p h o t o s y n t h e s i s i n l e a f 3 o f tomato p l a n t s grown a t 3 CO2 a t Leaf P l a s t o c h r o n Index:  11-12  (LPI :7.5) 3  t r a n s f e r ( r ) f o r l e a f 3 of tomato p l a n t s grown m  a t 0.03  or 0.1%  a common 0.03%  CO^ b e f o r e and a f t e r replacement i n CO2  atmosphere*  164  R e l a t i v e growth r a t e s (RGR) o f Pharbitis l e a v e s exposed to p h o t o p e r i o d  p l a n t s and  and CO2 treatments  d u r i n g a 14 day p e r i o d III-2  154  Rates of p h o t o r e s p i r a t i o n (R^) and m e s o p h y l l r e s i s t a n c e to CO2  III-l  7.5  concentrations  193  S i g n i f i c a n c e l e v e l s of terms i n A n a l y s i s o f V a r i a n c e on v a r i o u s parameters of growth and development i n Phavbitis and C0  o  p l a n t s subjected to d i f f e r e n t treatments  photoperiod 194  Page  III-3  Mean Plastochron Index of Pharbitis  seedlings at  end of 14 days i n photoperiod and CC^ treatments . . . 195 III-4  Mean change i n stem height (mm) of Pharbitis  plants  exposed to photoperiod and CG^ treatments during a 14 day period III-5  197  Mean number of f l o r a l buds formed on Pharbitis  plants  after 14 days i n photoperiod and CC^ treatments, and 7 days after the end of treatments III-6  198  Mean node of f i r s t f l o r a l bud formed on  Pharbitis  plants after 14 days i n photoperiod and CC^ treatments . . .....,....«...,.., III-7  ..  ««....<....  . ... 203  Combined leaf and a i r (boundary layer) resistances to CC^ transfer i n the second leaf of Pharbitis  plants grown  under LD or SD photoperiods and 0.03 or 1.0% C0  2  . . . 211  xi  LIST  OF  FIGURES  Page  1-1  Flow diagram of conductimetric CG^ analyzer used i n Experiment 1  13  1-2  Calibration curve for conductimetric CC^ analyzer. . . . 15  1-3  CG^ concentrations measured at 3 locations inside and outside the greenhouse on January 24, 1977  1-4  24  CO^ concentration measured inside the greenhouse on February 7, 1977  1-5  26  CC^ concentration measured inside the greenhouse on March 7, 1977.  1-6  30  Stomatal diffusion resistance of 4 leaves i n the cucumber canopy, and greenhouse CG^ concentration on February 23, 1977.  Figures i n parentheses indicate  irradiance at leaf level during resistance measurement -2 -1 (yE m s . 400 - 700 nm) 34 1-7  Stomatal diffusion resistance of 4 leaves i n the cucumber canopy, and greenhouse CO2 concentration on March 7, 1977.  Figures i n parentheses indicate  irradiance at leaf level during resistance measurement -2 -1 (yE m s , 400 - 700 nm) .- , 36 1-8  Stomatal diffusion resistance and irradiance at leaf level of successive leaves i n the cucumber canopy, at 4 times during the day on February 23, 1977.  Times  xii  Page indicate s t a r t of measurement sequence .  .  ......_ 1-9  stomatal d i f f u s i o n resistance quantum f l u x density  39  Stomatal d i f f u s i o n resistance and irradiance at leaf l e v e l of successive leaves i n the cucumber canopy, at 4 times during the day on March 7, 1977., Times indicate s t a r t of measurement .  II-l  sequence.  .  stomatal d i f f u s i o n resistance  .  quantum flux density  40  Spectral energy d i s t r i b u t i o n of 100 W incandescent lamp and 300 W cool beam lamp used to illuminate assimilation chamber i n Experiment 4  II-2  61  Time course of RGR derived from f i t t e d curves, f o r tomato crops' grown i n experimental greenhouses A and B without CO^ enrichment. .  I1-3  66  Time course of NAR derived from f i t t e d curves, for tomato crops grown in. experimental greenhouses A and B without CO^ enrichment  II-4  Time course of CGR derived from values of NAR  67 and LAI,  for tomato crops grown i n experimental greenhouses A and B without CO^ enrichment II-5  68  Atmospheric CO^ concentrations i n C02~enriched and normal a i r (unenriched) greenhouses  72  xiii  Page II-6  Truss development. regime.  Open bars:  Cross-hatched bars:  regime-(cross-hatching A:  CO^-enriched growth normal a i r growth  superimposed).  Number of trusses developed with f r u i t set on at least one flowerhead.  B:  Number of trusses developed with open flowerheads.  C:  Number of trusses developed with closed flowerheads.  NOTE:  Data are means for 17 r e p l i c a t e tomato plants per treatment.  At each date, A + B + C are  s i g n i f i c a n t l y d i f f e r e n t between treatments according to Witney-Mann test f o r nonparametric data (P < 0.001) II-7  7 5  Cumulative f r u i t y i e l d for plants grown i n CO2 enriched and normal a i r growth regimes. NOTE:  Data are means of 17 tomato plants per treatment. Differences at any date after May 24 are s i g n i f i c a n t according to t-test (P < 0.05) . . 77  II-8  Rates of apparent photosynthesis i n basal leaf 5 of plants grown i n normal a i r ( . _ =  (, NOTE:  ..) and  ,) and C02~enriched growth regimes. Data are means of ,2 tomato plants per "treatment.-  Points marked with the -same l e t t e r  xiv  Page  are not significantly different (P > 0.05) according to SNK multiple range test II-9  79  Rates of apparent photosynthesis i n f i r s t unrolled apical leaf of plants grown i n normal a i r (. and C0 -enriched 9  (,  ,)  ,) growth regimes.  NOTE: Data are means of 2 tomato plants per treatment. Points marked with the same l e t t e r are not significantly different (P > 0.05) according to SNK multiple range test 11-10  80  Differences i n apparent photosynthesis rates beweeen tomato plants grown i n CO^-enriched and normal a i r growth regimes (APs), i n relation to (A) C0 concentration and 2  .(B). quantum flux density.  Points represent mean APs  for a fixed level of one factor over a l l tested levels of the other. A  11-11  Apical leaves:  A  A; Basal leaves: 81  k  Leaf resistances to C0 transfer. 2  NOTE: Data for rs (stomatal resistance) are means of 2 determinations per treatment.  r represents m  mesophyll resistance calculated as described i n the text.  C0 represents test C0 concentration. 2  2  Bars show one standard deviation either side of the mean for test C0 concentration = 340 y l 1  1  2  (standard deviations for other r means were of s similar magnitude)  87  X  Page  11-12  Diagram of 8-chamber growth system used for Experiments 5, 6, 7, and 8  11-13  97  Diagram.of one chamber of the system shown i n Figure 11-12 ( f o i l facing removed to show internal detail). . 98  11-14  Spectral energy distribution of mixed fluorescent and incandescent lamps used i n the system shown i n Figure 11-12.  11-15  100  Diagram of assimilation chamber used i n Experiments 6 and 7  11-16  .104  Plastochron development of tomato plants grown at 3 C0„ concentrations as a function of time. NOTE: Points are means of measurements on a total of 6 plants, i n two chambers shown ± one standard deviation  11-17  114  Growth i n length of successive leaves of tomato plants grown at 0.03% C0 as a function of LPI . • < • . . - . . 2  NOTE: Points are means of measurments on a total of six plants i n two chambers 11-18  115  Growth i n length of successive leaves of tomato plants grown at 0.1% CO2 as a function of LPI. NOTE: Points as for Figure 11-17  11-19  116  Growth i n length of successive leaves of tomato plants grown at 0.5% CO2 as a function of LPINOTE: Points as for Figure 11-17  117  V  xvi  Page 11-20  Apparent rates of photosynthesis  i n r e l a t i o n to  quantum flux density i n leaf 3 of tomato plants grown at 3  concentrations.  Test C0 concentration: 2  Leaf temperature: LPI: NOTE:  330 y l l "  1  30 ± 2°C  2 ± 0.5 Points bearing the same l e t t e r at any irradiance are not s i g n i f i c a n t l y different according to SNK multiple range test (P > 0.05)  11-21  1  2  3  Stomatal resistance to CO2 transfer i n r e l a t i o n to quantum -flux density i n leaf 3 of tomato plants grown at 3 CO2 concentrations. Test C0  2  concentration:  Leaf temperature: LPI: NOTE:  330 y l 1  _ 1  30 ± 2°C  2 ± 0.5 Mean resistances at each irradiance are not s i g n i f i c a n t l y d i f f e r e n t (P > 0.05) according to SNK multiple range test. ...... .  11-22  Apparent rates of photosynthesis  127  i n r e l a t i o n to  i n t e r c e l l u l a r CX^ concentration i n leaf 3 of tomato plants grown at 3 CO2 concentrations. -2 Radiant flux density: Leaf temperature LP.I: 2 ± 0.5 . .  520 yE m  -1 s  , 400-700 nm  30 ± 2°C 129  xvii  Page  11-23  Rates nf phntorespir^tion (R^), and mesophyll resistance (rm ) for leaf 3 of tomato plants grown at 3 C0 concentrations and measured at LPI: ?  2 ± 0.5 NOTE: Values of R or r represented by bars L m T  bearing the same l e t t e r are not significantly different  (P > 0.05) according to SNK multiple  range test.  Rates of R^ and r ^ derived from  regressions relating apparent photosynthesis to intercellular CO^ concentration, as described i n the text 11-24  130  Effect of CO2 concentration on the percentage t o t a l oxygen inhibition of photosynthesis i n leaf 3 of tomato plants grown at 3 CO 2 concentrations. -2 -1 quantum flux density: Leaf temperature: LPI:  11-25  520 yE m  s  , 400-700 nm  30 ± 2°C  2 ± 0.5  136  A c t i v i t i e s of RuBP carboxylase and glycolate oxidase extracted from leaves of tomato plants grown at 3 CO2 concentrations.  PI: 5 ± 0.5  NOTE: Values are means of 3 replicate samples per CO2 regime shown ± one standard deviation. . . 142  xviii  Page  11-26  Apparent rates of photosynthesis i n relation to quantum flux density i n leaf 3 of tomato plants grown at 3 CO^ concentrations. Test C0 concentration: 2  Leaf temperature: LPI:  330 y l 1  _ 1  30 ± 2°C  7.5 ± 0.5  NOTE: Points at any irradiance are not significantly different from one another (P > 0.05) according to SNK multiple range test 11-27  146  Apparent rates of photosynthesis in relation to i n t e r c e l l u l a r CO^ concentration i n leaf 3 of tomato plants grown at 3 CO^ concentrations. -2 -1 quantum flux density: Leaf temperature: LPI:  11-28  520 yE m  s , 400 - 700 nm  30 ± 2°C  7.5 ± 0.5  149  Rates of photorespiration (R^) and mesophyll resistance ( r ) for leaf 3 of tomato plants grown at 3 CO m  concentrations and measured at LPI: 7.5 ± 0.5. NOTE: Values of R^ or  represented by bars  bearing the same l e t t e r are not significantly different (P > 0.05) according to SNK multiple range test . .Rates.of R_.and.r. „ „ L m  xix  Page derived from regressions r e l a t i n g apparent photosynthesis  to i n t e r c e l l u l a r CC^  concentration, as described i n the text. . . . 11-29  Stomatal resistance to CO^  151  transfer i n r e l a t i o n to  quantum flux density i n leaf 3 of tomato plants grown at 3 CO^  concentrations.  Test CC> concentration: 2  Leaf temperature: LPI: NOTE:  7.5 ±  330 y l  1  _ 1  30 ± 2°C  0.5  Mean resistances at each irradiance are not s i g n i f i c a n t l y different  (P > 0.05)  according  to SNK multiple range test •  11-30  E f f e c t of C0  2  concentration on the t o t a l oxygen  i n h i b i t i o n of photosynthesis plants grown at 3 G0  2  i n leaf 3 of tomato  concentrations. -2  quantum f l u x density: Leaf temperature: LPI: 11-31  152  520 yE m  -1 s  , 400-700 nm  30 ± 2°C  7.5 ± 0.5  155  A c t i v i t i e s of RuBP carboxylase and glycolate oxidase extracted from leaves of tomato plants grown at 3 CO NOTE:  concentrations.  Plastochron Index:  Values are means of  10.5±0.5.  3 r e p l i c a t e samples per  treatment shown ±-one standard deviation . . . 157  XX  Page  11-32  Apparent rates of photosynthesis i n relation to i n t e r c e l l u l a r CO^ concentration i n leaf 3 of tomato plants grown at 0.03% or 0.1% CO^ before placement i n common 0.03% atmosphere. -2 -1 quantum flux density:  520 liE m  s , 400-700 nm  Leaf temperature: 30 ± 2°C LEI: 11-33  2 ± 0.5  163  Apparent rates of photosynthesis i n relation to i n t e r c e l l u l a r GO^ concentration i n leaf 3 of tomato plants grown at 0.03% or 0.1%/CO , 14 -days after placement i n common 0.03% CO^ atmosphere. -2 -1 quantum flux density:  520 pE m  s  Leaf temperature: 30 ± 2°C LPI: III-l  12 ± 0.5  166  Diagram of gas flow system used i n Experiments 9 and 10 . .  III-2  plants A and B. A exposed to 14 SD cycles  Pharbitis  and 0.1% C0 ; B exposed to 14 SD cycles and 1% C0 . 2  2  Plants photographed 7 days after return to uniform LD conditions (0.03% CO^ III-3  Pharbitis  199  plants C and D. C exposed to 14 SD cycles  and 0.03% C0 ; D exposed to 14 LD cycles and 5% 2  C0 . 2  Note the flower formed.on Plant C. Plants  photographed 7 days after return to uniform LD conditions (0.03% C0 ) 2  200  xx i  Page  III-4  F i r s t f l o r a l bud a t node 6 on plant D (see Figure III-3) exposed to 14 LD cycles and 5% CCy Photograph at same time as Figure I I I - 3 . . .  III-5  201  D e t a i l of developing f l o r a l buds and one flower on plant C (see Figure III-3) exposed to 14 SD c y c l e s and 0.03% CO.  Photograph at same time as  Figure I I I - 3 III-6  204  Third f l o r a l bud at node 6 on plant A (see Figure III-2) exposed to 14 SD cycles and 0.1% C0 . 2  Photograph  at same time as Figure III-2 III-7  205  D e t a i l of plant A showing terminal f l o r a l bud (see also Figure I I I - 2 ) .  Plant exposed to 14 SD cycles and  0.1% (X> . Photograph at same time as Figure III-2 . . 2  III-8  206  D e t a i l of f i r s t f l o r a l bud at node 7 on plant B (see Figure III-2) exposed to 14 SD cycles and 1% C0 . 2  Photograph at same time as Figure III-2 III-9  Apparent rates of photosynthesis i n r e l a t i o n to C0 concentration i n Phavbitis or 1.0% C0  111-10  207  2  seedlings grown i n 0.03%  i n Long Days  209  Apparent rates of photosynthesis i n r e l a t i o n to C0 concentration i n Phavbitis or 1.0% C0  2  2  i n Short Days  2  seedlings grown i n 0.03% 210  xxii  Page  A3-1  RuBP-case a c t i v i t y i n relation to substrate concentration at pH 7.8, 30°C  A3-2  RuBP-case a c t i v i t y i n relation to pH at saturating substrate concentration  A3-3  2 6 7  268  GaO a c t i v i t y i n relation to pH at saturating substrate concentration, 27°C  269  xxiii  ACKNOWLEDGEMENTS  I wish to take t h i s o p p o r t u n i t y to thank the many people who  have a s s i s t e d me  throughout  e s p e c i a l l y l i k e to express my a d v i c e of my  Joyce  Ilmars D e r i c s , and  conduct  to  Mr.  the  I would  friendly technical  the s k i l l and p a t i e n c e of  my  Hollands.  Thanks are a l s o due to  a p p r e c i a t i o n of the h e l p and  r e s e a r c h s u p e r v i s o r , Dr. P e t e r J o l l i f f e ,  a s s i s t a n c e of Mr. t y p i s t Ms.  the course of t h i s r e s e a r c h .  to Mr.  A l b e r t Van Marrewyk f o r p e r m i s s i o n  experiments i n h i s greenhouse i n P i t t Meadows, B.C.,  and  R. D. B u l l i v a n t f o r the b a s i c d e s i g n of the a s s i m i l a t i o n chamber  used i n P a r t I I , S e c t i o n 2 and P a r t I I I . Finally, l o v e and of  to my w i f e Joyce, my  support which have h e l p e d me  t h i s work.  deepest  g r a t i t u d e f o r the  i n so many ways d u r i n g the  course  ABBREVIATIONS USED IN THE TEXT Abbreviations  Meaning  A  Leaf area  ANOVA  Analysis of variance  CGR  Crop growth rate(s)  C.  Intercellular (inside leaf)  1  CO2 concentration EDTA E  c  Ethylenediaminetetracetic acid Efficiency of CO^ u t i l i z a t i o n (see Appendix 2)  E  I  Efficiency of radiant energy u t i l i z a t i o n (see Appendix 2)  F  Flux density of CC^  IM  Maximum possible CC^ flux (see  F  Appendix 2) GaO  Glycolate oxidase  GSH  Glutathione  I  Quantum flux density (400 - 700  P  (see Appendix 2)  IRGA  Infra-Red Gas Analyzer  LAR  Leaf-area ratio  LD  Long-Day  LDP  Long-Day Plant(s)  XXV  Abbreviations LPI.:: j 1  J  Meaning Leaf plastochron  index f o r  leaf i = j NAR  Net  PhAR  Photosynthetically  a s s i m i l a t i o n rate(s)  radiation PI  Plastochron  % I  tot  active  (400 - 700 nm) Index  T o t a l percentage i n h i b i t i o n of apparent p h o t o s y n t h e s i s  by 21%  oxygen Rate of p h o t o s y n t h e s i s  i n 0 - 1%  oxygen Ps  21  Rate o f apparent  photosynthesis  i n 21% oxygen •ab  Stomatal r e s i s t a n c e measured on abaxial leaf  L  ad  Stomatal r e s i s t a n c e measured on adaxial leaf  m  surface  surface  M e s o p h y l l r e s i s t a n c e to CO^ t r a n s f e r T o t a l stomatal r e s i s t a n c e t o CO^ transfer  RGR  R e l a t i v e growth rate(s)  \  Rate o f p h o t o r e s p i r a t i o n  RuBP  D-Ribulose-l,5-biphophate  xxvi  Abbreviations  RuBP-case  Meaning  Ribulose-1,5-biphosphate carboxylase  SD  Short-Day  SDP  Short-Day Plant(s)  SNK  Student-Newman-Keuls multiple range test  t  Time  W  Plant dry weight  APs  Difference i n photosynthesis rate between tomato plants grown in CCv, enriched and normal a i r greenhouses (Part I I , Section 1)  T  CC^ compensation point  yE  Micro-Einstein  GENERAL INTRODUCTION The role of carbon dioxide as a primary reactant i n the process of photosynthesis i s fundamental to l i f e on earth since i t represents the sole inorganic carbon source available to green plants for the production of carbohydrate.  The concentration of CO^ i n  the atmosphere i s maintained by the respiration of plants, animals, and micro-organisms, geothermal a c t i v i t y and, i n recent times by the extensive combustion of f o s s i l fuels.  Due to the low, natural  concentrations of the gas (approximately0.033% by volume), extremely large quantities of a i r are required by plants to produce r e l a t i v e l y small amounts of dry matter.  For example, Norman (219) has calculated  that the production of 5500 pounds (2500 kg) of organic carbon i n a corn crop required 20,000 pounds (9091 kg) of CX^ drawn from some 21000 tons (19091 metric tons) of a i r . Current rates of plant photosynthesis are sustained by a complex series of reactions (the carbon cycle) which result i n e f f i c i e n t CO,, replenishment on a global scale.  Probably the largest  single contribution to the atmospheric pool of CO^ i s the microbial decay of organic matter i n the s o i l , oceans, lakes and r i v e r s .  The  large water masses of the earth represent an important buffering system i n the regulation of atmospheric CO2 concentration but recent work has suggested that the world's forests are an even more important reservoir i n the carbon cycle (300).  C0„ content of the atmosphere  2  changes significantly with season, with highest concentrations occurring i n late winter and the lowest i n late summer (33) .  The  most striking global variation i n CC^ concentration, however, i s the steady year to year increase (300).  U n t i l very recently this  increase (which has raised the atmospheric C0^ level from approximately 0.029% i n 1850 to 0.033% i n 1977) was attributed to the: increased combustion of f o s s i l fuels.  I t now seems l i k e l y that  an even more important cause i s the clearing of forested land which accelerates the breakdown of tree biomass and s o i l humus and removes a potentially large carbon reservoir from the biosphere (300). Despite the recent increases i n  concentration, the  atmosphere contains considerably lower CO^ levels today than were present i n primordial times.  The most l i k e l y estimates of the  content of the early atmosphere on earth range between 0.3 and 3% (229).  In most present day land plants using the  system of CO2  assimilation, rates of photosynthesis:.increase very l i t t l e above 0.15 - 0.2% CO2 under optimal conditions of irradiance (60, 103, 134, 252).  The a b i l i t y of plants to u t i l i z e  concentrations i n excess  of the ambient atmospheric level of 0.03% may be a remnant, of evolutionary development during a period when the atmosphere contained much greater quantities of the gas. selection may gradually reduce the  The pressures of natural concentration required for  photosynthetic saturation since i t seems l i k e l y that, i n previous ages, plants could u t i l i z e even higher levels of CO2 than are shown today.  3  The promotive effect of increasing CO^ concentration on photosynthetic fate was f i r s t noted i n the late nineteenth  century  (166) and was confirmed several years later by the work of Blackman and Smith (27) on the aquatic plants Elodea and Fontinalis...  These  early studies provided the background for further research into the effects of supra-normal CO^ concentrations on plant growth and development. At f i r s t , studies concentrated  on plant responses to  increased CC^ concentrations which seemed to be d i r e c t l y correlated with enhanced photosynthesis.  Demoussy (74) showed that dry weight  increases of 158% could be obtained i n plants grown with additional CO^,  and i n Europe and North America, experiments were soon undertaken  to determine the potential benefits of CC^ enrichment for greenhouse -  crops (69, 70).  The commercial application of greenhouse CO^ enrichment -  in North America was not f u l l y substantiated, however, u n t i l the results of a series of experiments on several crops were collated i n 1964  (298). At about that time, Krotkov  and co-workers (279) presented  the results of their studies on l i g h t stimulated respiration (photorespiration) i n tobacco.  Further work established that rates of  photorespiration increased with oxygen concentration (280).  The  inhibition of apparent photosynthesis by oxygen i n plants had f i r s t been demonstrated by Warburg i n 1920( 287), but i t was not u n t i l  1966  that the physiological components of this i n h i b i t i o n were f u l l y recognized (96, 280).  Those studies established, not only that  photorespiration was increased by high oxygen concentration, but that  4  oxygen also had a direct Inhibitory effect on photosythesis.  A  few years later further work showed that high levels of CG^ could markedly reduce the oxygen inhibition of apparent photosynthesis (150).  This finding put into perspective the results of Mortimer  (207) who showed that the production of serine and glycine (two intermediates of photorespiration) was reduced at CO^ of 2.5%.  concentrations  I t became evident that supra-normal levels of CO^ exerted  their effects on plant growth, not merely through increased true photosynthetic rates but also via a reduction i n photorespiration. This was further supported by subsequent studies which showed that the.synthesis of glycolate, the probable substrate for photorespiration, i s inhibited at high C0  2  concentrations (35, 83, 172,, 239, 258).  While much research into the physiological role of C0  2  in  higher plants has been concerned with photosynthesis and photorespiratory metabolism, a large amount of information has also accumulated on the effects of C0 (174) f i r s t noted that C0  2  2  on stomatal movement.  Linsbauer  free a i r causes maximum stomatal opening  in both l i g h t and darkness.  This early observation was confirmed by  a number of subsequent studies (119, 120, 122, 157). • I t . i s presently unclear exactly how CI^ concentration affects stomatal action although a number of detailed hypotheses have been put forward (173;, 235) . In addition to i t s effects on stomatal movement and photosynthetic physiology, CO  appears to affect a number of other  5  processes involved in plant growth and development. The growth of root tissue (58, 260) and coleoptile sections (39, 80, 90) are influenced by increasing CO^ concentrations over a range of 0 to 0.5%.  Other studies have shown that CO2 i s required for the  effective inhibition of flowering i n Xanthium pennsylvanicum  during  a single red light break in the middle of an inductive dark period, and for seed germination following red illumination i n lettuce (13). At higher concentrations (1% and above) C0  2  affects budset in  Douglas f i r (247) , and flowering in a number of SD* and LD plants (51, 234).  CO^ may also affect plant survival under adverse  environmental conditions as exemplified by the increased salt tolerance of tomatoes grown at 0.8% as compared with those grown at 0.035% C0  2  (85). The wide variety of the effects of CO^ on the physiology  of plants provided the background for the present studies. While considerable attention has previously been paid to the effects of C0  2  enrichment on yield in greenhouse crops, there remains a lack of  information on two important aspects of this practice. In the f i r s t case, many growers and horticulturists are acutely aware of the need to assess ambient C0  2  concentrations within greenhouses under  enriched and non-enriched conditions.  Secondly, very few studies  have been concerned with the physiological adjustments which may occur when plants are grown at supra-normal C0  2  concentrations.  *Abbreviations used here and throughout the thesis are explained and l i s t e d on p.xxiv  6  The f i r s t part of the research i n this thesis (Part I) consisted of experiments designed to provide information on CO^ levels within a commercial greenhouse at various stages i n the development of a cucumber crop.  In addition, measurements were conducted during  the course of the work to assess the influence of varying ambient CO^ concentrations on stomatal resistance i n this crop.  The scope of these  studies were limited, however, since i t was impractical to conduct a more detailed investigation of other physiological responses of these plants to  enrichment.  The studies were extended by further  experiments designed to investigate growth and photosynthetic metabolism of tomato plants grown under CG^-enriched conditions i n a greenhouse (Part I I , Section 1). The results of these experiments prompted further investigation into the physiological mechanisms underlying observed changes i n photosynthetic capacity i n the greenhouse-grown plants. Subsequent studies on photosynthesis, photorespiration, and enzyme a c t i v i t i e s associated with these processes were, therefore, conducted on plants grown at 3 levels of CC^ i n controlled environment chambers (Part I I , Section 2). Previous studies on plant response to CO^ have suggested that the gas may mediate physiological changes i n systems other than those directly involved i n photosynthetic metabolism.  To investigate such  effects, the f i n a l part of this research (Part III) examined the relationship between CO^ enrichment during growth and vegetative and reproductive development i n Pharbitis  nil  seedlings.  The particular  interest i n these responses originated i n the results of earlier studies which had demonstrated marked effects of high CO^ levels on flowering in Pharbitis  and other photoperiodically sensitive plants (51, 234).  PART I  Studies  in a cucumber greenhouse:  concentration  and  leaf  stomatal  Atmospheric resistance.  8  INTRODUCTION  Over the past 15 years, CO^-enrichment has become widely adopted by commercial growers engaged i n greenhouse vegetable crop production throughout Europe and North America.  The introduction of  the procedure i n the early part of the 1960's and i t s rapid acceptance by the greenhouse industry represented an outstanding example of cooperation between research workers and the growers to improve crop production techniques. supplemental  The beneficial effects of  during crop growth, i n terms of increased earliness,  yield and quality have been recognized since the early part of this century (32, 69, 74, 255).  In more recent years i t has been  demonstrated that the yields of a wide range of greenhouse crops show an increase with CO^-enrichment to a concentration of approximately 1000 y l 1  (0.1.%), although the most extensive literature refers to  responses of lettuce (85, 295, 298), tomato (47, 49, 50, 163, 165, 189, 273, 298), and cucumber (72, 85, 135, 245, 298).  Of these three  crops, cucumbers have received the least attention with regard to the effects of C^-enrichment.  This seems to be due, primarily, to the  fact that for many years a r t i f i c i a l means of supplementing  supply  in cucumber greenhouses (by combustion or injection of pure gas) were thought to be unnecessary.  CO2 concentrations i n these situations  attained naturally high levels as a result of decomposition of organic matter (straw and manure) traditionally used i n preparation of  the cropping beds (245, 162).  Recently, the highly encouraging  results obtained by tomato growers using CO^-enrichment i n their greenhouses throughout the growth of the crop, have prompted many B r i t i s h Columbia  cucumber growers to i n s t a l l CO^-enrichment equipment to  supplement production from the soil-straw growing medium (W. Gates, personal communication).  From the lower Fraser Valley region of  B r i t i s h Columbia (where this research project was conducted) there are, as yet, few data to suggest whether the new CO^-enrichment practices have proved beneficial i n terms of increased f r u i t yields. The introduction of a new cultural practice i n crop production always raises important questions, not only with regard to effectiveness i n yield improvement but also concerning possible detrimental side effects.  Some of the questions may be answered i n  experiments conducted under controlled environmental conditions i n growth chambers. The answers to other questions must rely on results from experiments carried out under f i e l d conditions. This i s especially true of C02~-enrichment, since the constant concentration conditions of most growth chamber experiments do not reflect the situation i n greenhouses where co2 levels often fluctuate widely and rapidly.  In early experiments on CO^-enrichment, Brown and Escombe (41)  found that C0 concentrations of 1100 ]il 1  1  2  inhibited flowering  and caused bud abortion i n a number of plants grown i n closed containers.  A few years l a t e r , Demoussy (74) pointed out that their  results were almost certainly due to toxic impurities i n the gas  10  supplied to the plants, and since that time considerable care has been taken to purify gas used i n laboratory experiments.  The  potential problems of toxicity are, however, often present i n greenhouse situations.  Sulphur dioxide (114) and nitrogen oxides  (54, 55) at concentrations sufficient to inhibit tomato growth are sometimes present i n greenhouses using hydrocarbon burners for enrichment.  Incomplete combustion of natural gas may give rise to  detrimental concentrations of ethylene i n greenhouse atmospheres (17).  Whenever CO2 enrichment i s introduced as a new procedure, and  particularly when i t i s practiced on a new crop, careful attention should be paid to the method of enrichment and the composition of the resulting gas.  Even i n the absence of toxic impurities some crops  respond unfavourably to very high concentrations of CO2.  Cucumbers  seem particularly susceptible i n this respect. Two studies have indicated that atmospheric enrichment to 2000 or 3000 y l 1  1  can result  i n severe leaf necrosis, whereas a constant concentration of 1000 u l 1 produces beneficial effects on growth and f r u i t yield (72, 295). The research which i s the subject of Part I of the thesis was prompted by the decision of a cucumber grower i n the lower Fraser Valley to i n s t a l l natural gas burning, CO^ enrichment equipment i n his greenhouse.  Prior to the i n s t a l l a t i o n , supplementary CO2 had been  provided solely from decomposition of straw bales covered with s o i l i n the cropping beds.  No information was available on CO^ levels  attained i n the house atmosphere using these cultural procedures.  In  view of the considerations mentioned above, there was a clear need to determine ambient concentrations achieved with arid without supplementary CC^ and, also, the composition of the gas produced by the burners.  The present study was concerned, primarily, with  the f i r s t of these objectives. The generally high cost, inaccuracy or inconvenience of most C0 measuring 2  equipment currently available for use i n  greenhouses (3) prompted the development of a new system for this study. I t was, also, of interest to measure the stomatal resistance of leaves within the developing cucumber crop under naturally fluctuating levels of C0 and irradiance. 2  Although C0 concentrations i n excess of those 2  normally present i n the atmosphere have often been shown to result in stomatal closure and a reduced potential for photosynthesis (123, 152) i n a number of plants, no data were available for cucumbers grown as a crop.„ The following studies were, therefore, carried out to establish the levels of C0 present i n one cucumber greenhouse at 2  various stages of crop growth and with different methods of C0 supply. 2  Some measurements were also made to determine the variation of stomatal resistance i n the crop canopy under different environmental and developmental conditions. The applicability of the present findings to other greenhouses and other crops awaits further investigation. I t i s hoped, however, that the information provided here, and the design of the CC> measuring 2  equipment developed for these studies, w i l l  prove useful i n other studies on greenhouse CO - enrichment.  12  MATERIALS AND METHODS i)  Conductime trio  CO„ Analyzer  The requirement for a portable analyzer to monitor ambient concentrations of CC^ was met with a conductimetriee measurement system based on a design principle f i r s t described by James (145). The system i s shown diagramatically i n Figure 1-1. Deionized water was induced to flow through the system by means of a small electric pump. Water passed v e r t i c a l l y through a glass column (length: 40cm, internal diameter: 2 cm) incorporating a lower side arm to which a one-way valve was f i t t e d . entry for the sample a i r stream.  This formed the point of  At the top of the bubble column  the glass was extruded into an atrium with an upper port for exhaust of the a i r stream and a side arm from where the water flowed, through plastic tubing, into the conductivity c e l l .  From the  conductivity c e l l water returned directly to the pump and was thus driven through a c y l i n d r i c a l deionizing column (length: 15 cm, internal diameter: 6 cm) packed with approximately 450 g of Amberlite MB-3 ion exchange resin (Mallinckrodt Inc.).  During  operation, water flow rate was maintained at a constant 100 ml min and a sample a i r stream was introduced into the bubble column at a rate of 90 ml min . 1  The a i r supply was maintained by a small  diaphragm pump (Universal Electric Co., Owosso, MI) and i t passed through a rotameter type flowmeter f i t t e d with a precision needle  1  12a  FIGURE 1-1:  Flow diagram of c o n d u c t i m e t r i c CC^ used i n Experiment  1.  analyzer  13  air  /N  outlet J, conductivity transmitter  upper atrium wateTTeveT r  thermi stor (for temperature compensation) bubble  column  water pump  air inlet  deionizing column  L E G E N D  -» direction of water flow :-- electrical connections  valve (Matheson Ltd; tube number 610) before entering the column. As the a i r bubbles rose with the water, some of the CT^ contained i n the sample became dissolved resulting i n a change i n conductivity (according to the equilibrium between CT^ i n solution, and carbonic acid i n i t s associated and dissociated forms).  The  conductivity of water i n the system was sensed by a flow-through c e l l with wall-embedded electrodes (Electronic Switchgear, London Ltd.:  Model EFA/01/NPT). The c e l l was used i n conjunction with a  model TX2/2P conductivity transmitter, and conductivity could be measured on any one of three ranges (0-1, 0-10 and 0-100 micromhos/cm Changes i n temperature have a marked effect on conductivity. In this system the effects of temperature changes over a wide range (between 0 and 100°C) were automatically corrected by means of a temperature sensor connected to the conductivity transmitter. Temperature changes also affect water flow rate through a change i n density, but this effect i s small (16) and was insignificant i n the operation of the system.  considered  Calibration of the  instrument was achieved by bubbling gases of known CO^ concentration into the water column and recording relative conductivity readings. A zero point was set by bubbling a sample of nitrogen gas through the system. A calibration curve for the analyzer i s shown i n Figure 1-2.  Conductivity over the range 0 :to 1650 y l 1  1  changed  quadratically although the deviation from a linear relationship was not great.  A linear change was assumed i n subsequent calculation  FIGURE 1-2:  C a l i b r a t i o n curve f o r c o n d u c t i m e t r i c C0„  analyzer.  180Ch  1600H  1  2  3 RECORDER READING  4  5  6  16  of a i r sample CO^  concentration,  since only r e l a t i v e l y  assessments of ambient greenhouse l e v e l s were r e q u i r e d .  gross  17  ii)  Greenhouse Description  and  Crop Management  Practices  The study was concerned with a greenhouse located i n P i t t Meadows, B r i t i s h Columbia.' I t was typical of many modern cucumber-producing  houses i n the lower Fraser Valley, with a 2  total area under glass of approximately 2250 m for a crop of 2800 plants. sativus  L. cv. Farbio was  giving enough space  In early January 1977 seed of Cuaumis sowm i n 15 cm square pots (1 seed  per pot) containing a mica peat mix supplemented with Magamp and dolomitic limestone. Germination was evident i n 95% of the pots after two days at a temperature of 24°C. This temperature was 2 maintained i n a section of the greenhouse (approximately 200 m ) throughout the propagation period of three and a half weeks. During this time, preparation of the plant beds was carried out by the greenhouse staff.  230 straw bales were l a i d i n  shallow trenches and thoroughly moistened before the addition of a potassium nitrate/lime mix at a rate of approximately 1 kg per 50 kg straw.  The straw was allowed to decompose for several days  before i t was covered with 15-20 cm of loam. The cucumber seedlings were transplanted into the beds i n the f i r s t week of February.  Each plant had formed at least two true leaves by this  date. A seven-day period had elapsed since i n i t i a l moistening and f e r t i l i z e r addition to the straw, thus ensuring adequate decomposition of this substrate and dissipation of gaseous ammonia. During transplanting, the seedlings were planted i n double rows  18  at a spacing sufficient to allow 0.5 m  of ground area per plant.  Subsequently standard management practices were used on the crop, and growth was vigorous throughout the course of this study (January to March, 1977). Daily i r r i g a t i o n and sprinkling of plants were carried out automatically.  Superphosphate was supplied as a s o i l dressing  and applications were repeated as necessary during growth of the crop.  Other nutrients, including ammonium, calcium and potassium  nitrates, were supplied via the i r r i g a t i o n system.  Rates of  nutrient application varied somewhat throughout the study period but broadly conformed to that recommended i n the 1976-77 B.C. Greenhouse Tomato and Cucumber Production Guide (4). Approximately three weeks after the seedlings were transplanted, two natural gas-burning CO^ generators (Priva Ltd., Netherlands) were installed in the house. Each burner incorporated a fan to disperse the CO2 and, according to the manufacturers specificiations, was capable 2 of enriching a covered area of 12000 to 16000 m . Unfortunately no details of the rate of CO^ production were;provided.by the manufacturer.  During the f i r s t four weeks after transplanting  the burners were not available to provide CO2- enrichment within the greenhouse.  Enrichment was commenced on a daily basis at  the beginning of March. The i n s t a l l a t i o n and operation of the CO2 enrichment equipment and a l l crop management procedures were undertaken by the greenhouse staff.  19  iii)  Measurement of Greenhouse CO  Concentrations  CC>2 concentrations inside the greenhouse were measured throughout the daylight period on four occasions (January 24, February 7, 23 and March 7, 1977) during the early stages of crop growth and development. At the f i r s t sampling date the seedlings had not been transplanted and were enclosed i n the propagation area which was closed off from the main greenhouse. • Within the remainder of the house the beds had been p a r t i a l l y prepared for transplanting (straw bales had been l a i d and moistened but were not covered with s o i l ) .  Sampling of the atmosphere was  commenced i n the early morning and consisted of half-hourly periods during which air was drawn into the analyzer alternately from the seedling enclosure and a randomly selected sampling site i n the main part of the greenhouse. Upon changing from one sampling location to another, spot checks were made on ambient external CC^ levels by drawing an air sample from the atmosphere above the greenhouse roof.  A l l measurements within the greenhouse were based on a i r  samples drawn from a height of 2 m above the ground surface.  Within  the seedling enclosure, the open end of the sample tube was positioned directly above the cucumber seedlings which were arranged i n rows on slatted benches. In the main greenhouse the open end of the sample tube was arranged to face upwards above the cropping beds. Measurements of the greenhouse atmosphere on the other dates (February 7 and 23, and March 7) were carried out on a i r  samples from a similar height.  On these occasions the a i r  samples were drawn continuously into the analyzer from randomly selected sites within the greenhouse. The output from the conductivity transmitter was recorded on a strip chart recorder (Yellow Spring Instrument Co. Model 80A) and the CO^ concentrations were integrated over successive fifteen minute periods throughout the day.  21  iv)  Measurements of Stomatal  Resistance  Stomatal resistance was determined on leaves of one plant i n the crop on February 23, and March 7, 1977. On each date, plant selection was at random but care was taken to exclude any plant growing on the end of a row where i t might be subjected to different light and temperature conditions, and other edge effects.  At each date stomatal resistance measurements were  conducted on successive leaves i n the canopy on four occasions during the daylight period (measurements starting at 0820, 1030, 1300 and 1630 h).  A diffusion resistance porometer (Lambda  Instrument. Co. Inc., Lincoln, Nebraska; Model LI-60) was used for the measurements. On February 23 the f u l l complement of 13 leaves was used for stomatal resistance determinations.  On March 7,  when each plant had i n excess of 20 leaves, only the f i r s t thirteen counting from the newly expanded leaf at the top of the canopy, were used.  Throughout a l l measurements a specific sequence  of operations was followed for each leaf, whereby the porometer was clasped to the abaxial leaf surface, leaf temperature was recorded (by means of a thermistor incorporated i n the porometer sensor), and a resistance measurement was taken. was repeated for the adaxial surface.  The sequence  PhAR incident upon each leaf  was measured concurrently with the resistance measurements using a Lambda Instrument Co. Model LI 190S quantum sensor with a LI 185 meter.,  22  Calibration of the porometer was carried out i n the laboratory<as recommended by the manufacturer (Lambda Instrument Model LI 60 operation manual).  Total leaf resistance was  calculated as:  2 / r  s  =  1 / r  ad  +  1 / r  ab  where rs = total leaf resistance .r ad. •= resistance of adaxial leaf surface r ab , = resistance of abaxial leaf surface (156) Measurements on a l l thirteen leaves occupied approximately one hour so that there was considerable temporal as well as spatial v a r i a b i l i t y i n the radiation and, possibly, the stomatal resistance data.  For descriptive convenience, i n Figures 1-8 and 1-9 and  throughout the text, measurement times are referred to by the time at which the series of measurements commenced. In Figures 1-6,and 1-7. resistance values are plotted at the actual time that they were taken.  23  RESULTS AND DISCUSSION i)  Greenhouse CO  Concentrations.  (Experiment 1) *  Measurements of CC^ levels i n the greenhouse atmosphere indicated that considerable increases i n ambient CC^ concentrations can be achieved simply as a result of straw decomposition within the plant beds. Figure 1-3 i l l u s t r a t e s the diurnal pattern of CC^ concentration within the greenhouse during seedling propagation. Spot checks on external atmosphere concentrations showed l i t t l e variation with concentrations remaining between 320 and 330 y l 1 throughout the daylight period.  1  In the bulk atmosphere within  the greenhouse, CO^ concentrations were also relatively stable and were some 130 y l 1 higher than external a i r values.  Within  1  the propagation area, CO^ concentrations i n the early morning (0830 - 0845 h) were approximately 420 y l 1 , but by the next _1  sampling period at 0930 h levels had declined to 385 y l 1 and 1  thereafter remained quite steady u n t i l measurements were terminated at 1600 h. Evidently, the elevated greenhouse were due to  concentrations inside the  production within the beds where straw  bales had been allowed to settle and undergo some decomposition for a week prior to the January 24. measurements. In the early morning,  levels i n the enclosed propagation area were similar  to those i n the bulk greenhouse atmosphere, but by mid-morning concentrations had been depleted to a value intermediate between ^Experiments are numbered sequentially throughout the thesis and are l i s t e d in Appendix 1.  FIGURE 1-3:  CC^ concentrations measured at 3 locations inside and outside the greenhouse on January 24, 1977.  500  bulk greenhouse atmosphere  inside seedling enclosure  400-  external atmosphere 3001  200 0800  1  1  1  1  i  0900  1000  1100  1200  1300  TIME OF DAY  i  U 0 0  n  1500  r  1600  25  the greenhouse and external a i r l e v e l s .  V e n t i l a t o r s were not  opened i n the propagation area on this day and the CO^ depletion was  apparently due to photosynthetic uptake by the young plants. The most complete information on ambient CC^ levels i n  a cucumber house i n which beds were prepared with a sub-soil straw layer i s provided by Klougart of approximately  4500 y l 1  (162).  He reported  concentrations  within a 20-metre greenhouse during  1  the Spring pre-planting period suggesting that decomposition already proceeding at a fast rate. that CO^  was  The present r e s u l t s show  levels i n the house used i n this study were an order of  magnitude lower than Klougart's  (162) data.  It i s l i k e l y that straw  decomposition had not reached maximum rate during the week preceding the January 24 measurements and the young seedlings were probably CO^-limited during this early stage of growth. high CO^  The  concentrations often described i n cucumber houses well  supplied with organic matter (162, 298) have been assumed to be more than adequate for seedling propagation. however, indicate that CO^  data,  levels could be u s e f u l l y raised (by  alternate methods of CO^ production) preparation i n operations  The present  for at least a week after bed  where young plants are r a i s e d i n the  main greenhouse. Figure 1-4 shows the diurnal trend of greenhouse CX^ concentration on February 7.  Plants had developed 5 leaves by  this date and had already been transplanted into the beds.  CO  25a  FIGURE 1-4:  CO^ concentration measured inside the greenhouse on February 7, 1977.  27  levels i n the early morning were over 5 times higher than normal atmospheric concentrations, and i t was apparent that straw decomposition was contributing significantly to CO^ production within the house.  I t i s noticeable, however, that concentrations  as high as 1800 y l 1  1  (measured at 0800 h) were quickly  depleted during the morning and by 1030 h had f a l l e n to a constant -1 daytime level of 600 y l 1  . Similar decrements i n house CO^  concentration during the f i r s t few hours after dawn have been reported i n previous studies (162), and may be due to a number of factors.  Even i n late winter some ventilation i s frequently  necessary to control greenhouse temperature and this occurred at intervals during the sunny morning of February 7.  The opening of  roof vents quickly reduces internal CX^ concentration (298) and this effect i s enhanced by strong wind (17, 253).  In addition,  CO^ consumption by the developing crop undoubtedly contributed to the drop i n ambient concentration during the early morning on this day.  Despite the rapid decline i n CO^ levels between 0800 and  1030 h, greenhouse concentrations remained close to 600 y l 1  1  as a result of the continued production of CO^ from the soil-straw substrate.  Considerable benefit i n terms of increased photosynthesis  and growth might be obtained at this stage of crop development by the injection of additional CO^ into the house atmosphere. I t i s evident, however, that even without CO^ generating equipment the replenishment of  lost by crop consumption and ventilation i s  28  quite e f f i c i e n t i n cucumber houses prepared with a sub-soil straw layer.  Morris et at.  (206) calculated that 10 a i r changes per  hour were necessary to produce a non-diminished growth rate of tomato plants i n a conventional greenhouse where no supplementary CO^ source was provided. This value i s clearly dependent upon season, stage of crop development and cultural conditions (45, 206). The present results suggest that ventilation of many cucumber houses i s unnecessary for CO^ replenishment, at least during the f i r s t 2-3 weeks of crop development.  Frequent ventilation might,  i n fact, be expected to reduce internal concentration close to that of the external atmosphere. In recent years, various management d i f f i c u l t i e s commonly associated with growing greenhouse crops i n s o i l (e.g. control of pests and levels of essential nutrients) have prompted some growers in the Lower Fraser Valley to change to a soilless-culture for growing cucumbers (W. Gates, personal communication).  One of the  disadvantages of this method of production w i l l be a lack of supplementary CO^ produced from the traditional manure/straw and s o i l cropping beds.  For some growers this has been an important  reason fo^- i n s t a l l i n g equipment to provide an alternate means of CO2~ enrichment. available  Since l i t t l e accurate information has been  on CO2 levels i n commercial greenhouses, previous  recommendations have tended to underestimate the importance of e f f i c i e n t f a c i l i t i e s for CO - enrichment i n cucumber houses (245). It i s clear from the profiles of diurnal CO2 concentrations presented above that the addition of C0 from a r t i f i c i a l sources 9  may  be of b e n e f i t even i n houses where t r a d i t i o n a l  cropping  systems are used. The  r e s u l t s o b t a i n e d from measurements of i n t e r n a l  c o n c e n t r a t i o n s on March 7 are shown i n F i g u r e 1-5. date  the n a t u r a l gas burners  on. .February  i n the greenhouse  A s i m i l a r pattern of C ^ — d e p l e t i o n to that 7 was  were turned on, reflected  although  to house CO  lower c o n c e n t r a t i o n s at 0800 h c o n t r i b u t i o n of o r g a n i c  f a l l e n to approximately  were turned on and  the c o n c e n t r a t i o n was  1530  h.  without and  lil 1  1  w i t h i n 2 hours.  550  probably  Ul 1  When the 1  the  enrichment procedures  r e s t o r e d to between  Enrichment was  temperatures were low and  from the c o n d u c t i m e t r i c  minimal.  t h e r e was  1100  d i s c o n t i n u e d at staff analyzer,  were s i m i l a r to those c a r r i e d out  day when house v e n t i l a t i o n was  internal  burners  A l l o p e r a t i o n s were performed by the greenhouse r e f e r e n c e to the output  burners  substrate  l e v e l s i n e a r l y March.  c o n c e n t r a t i o n had  1200  observed  e v i d e n t i n the e a r l y morning b e f o r e the  the d i m i n i s h e d  decomposition  and  this  were used from mid-morning u n t i l  l a t e a f t e r n o o n to r e p l e n i s h CO^ supply atmosphere.  On  CO^  :  each  On March 7 o u t s i d e  l i t t l e need f o r v e n t i l a t i o n .  Burners would have been turned o f f f o r p e r i o d s on o t h e r days when r i s i n g house temperatures n e c e s s i t a t e d vent  opening.  The  enrichment  p e r i o d on March 7 appeared to be q u i t e e f f i c i e n t i n m a i n t a i n i n g house l e v e l s near optimum c o n c e n t r a t i o n s p l a n t had  f o r crop growth.  formed at l e a s t 20 l e a v e s by t h i s time and had  Each set several  FIGURE 1-5:  CC^ concentration measured inside the greenhouse on March 7, 1977.  T I M E  OF  DAY  31  fruit.  I t i s at this stage of development that serious CO^  limitations may occur inside closed greenhouses.  These c l e a r l y  cannot be remedied by v e n t i l a t i o n or by natural CC^ production processes even i n the presence of abundant organic matter i n the cropping beds.'  Nevertheless, the use of burners i n the early  morning was evidently unnecessary  i n a house of this type since  CC^ concentrations were high, presumably as a result of night r e s p i r a t i o n by the extensive crop biomass and residual production, from the plant beds. For most growers who lack the instruments necessary to monitor CO^ levels i n their greenhouses on a daily basis, the operation of enrichment equipment i s dictated by the recommendations of a g r i c u l t u r a l b u l l e t i n s and manuals, although few recommendations r e l a t i n g to cucumber culture are available.  The usual recommendation  i s that CO^ addition should be commenced soon after dawn and continued u n t i l 1 1/2 hours before sunset, when l i g h t levels become severely l i m i t i n g to photosynthesis (e.g. 3, 257). Based upon the present r e s u l t s , i t appears that these recommendations may be legitimately applied to Spring-grown cucumber crops, although some economic benefit might be derived by s l i g h t l y delaying the onset of enrichment i n the morning.  In this study, however, i t i s evident  that enrichment could have been usefully commenced at least an hour e a r l i e r on March 7, and i t i s doubtful that the f i n a n c i a l saving i n f u e l costs by this delay i s very significant'. Calvert and Slack (50) , i n their studies  on tomatoes,  found t h a t any r e d u c t i o n  i n the d a i l y  enrichment  p e r i o d , e x t e n d i n g from s u n r i s e to sunset,had a marked e f f e c t on f i n a l y i e l d s which was not o f f s e t by s a v i n g s c o s t of enrichment.  Their findings advise  morning C0 -enrichment 0  i n cucumber houses.  detrimental i n the  caution i n delaying  33  ii)  In situ •Measurements of Stomatal Resistance  (Experiment 1)  Stomatal closure during CC^ enrichment has been the subject of several studies, which have disagreed on the importance of this effect i n l i m i t i n g photosynthesis and growth (138, 152, 225). It now seems probable that the magnitude of stomatal response to increasing CO^ concentration i s species-specific, with tomatoes apparently showing a rather low sensitivity (138, 225).  There  are, apparently, no previous reports dealing with the response of stomates .in cucumber leaves to elevated CO^ concentrations, despite the fact that CO^-enrichment by organic matter fermentation or gas combustion i s frequently practiced i n cucumber greenhouses. Figure 1-6 shows the stomatal resistance of four leaves from the top, middle and bottom of the crop canopy throughout the daylight period on February 23.  Also shown on this figure i s the  pattern of internal C0 concentration on this day, during which no 2  additional CO2 was supplied to the atmosphere from the gas burners. Stomatal resistances i n leaves 4, 8, and 13 tended to increase throughout the day, reaching a maximum i n the late afternoon between 1630 and 1700 h.  Radiation incident on individual leaves  (figures i n parentheses) was variable on this ^predominantly sunny day, with only leaf 13 (at the bottom of the canopy) receiving relatively low irradiance at each measurement time. The progression of stomatal resistance from low to high values i n leaves 4, 8, and 13 during the day was not obviously correlated  33a  FIGURE 1-6:  Stomatal d i f f u s i o n r e s i s t a n c e of 4 l e a v e s i n the cucumber  canopy, and greenhouse CC^ c o n c e n t r a t i o n  on February 23, 1977.  F i g u r e s i n parentheses  indicate irradiance at leaf l e v e l  during  r e s i s t a n c e measurement (yE m  » 400 - 700  s  nm).  35  with decreasing levels of radiation or increasing CO^ concentration.  In the early morning resistances i n a l l leaves  were at their lowest daily value, when atmospheric  CC^  concentration was highest. Leaf 2 exhibited considerably higher stomatal resistance than the other leaves except i n the late afternoon (1630 h) and did not show the same gradual increase i n this parameter. A similar trend of increasing resistance through the day was shown by leaves.4, 8, and 13 on March 7 (Figure 1-7), although resistance values were generally higher than on February 23. In contrast to that day, March 7 was predominantly overcast, as indicated by the r e l a t i v e l y low values of leaf irradiance, which may have accounted for :the observed differences.  Resistances were  higher during the CO^r-enrichment period (from 1030 to 1530 h) than i n the early morning, but there was no indication of a decrease during the f i n a l measurements, roughly an hour after the CO^ burners were turned off. The similar trends i n resistance throughout the daylight period i n leaves 4, 8, and 13 on March 7 (when (X^-enrichment was practiced) and February 23 (without enrichment) do not suggest that this parameter was strongly affected by ambient CO^ concentration. The pattern of gradually increasing resistance on both days may be related to a diurnal rhythm of stomatal opening potential which has been shown to result i n maximum aperture i n mid-morning followed by gradual closure through the afternoon and early evening (155, 197, 261).  35a  FIGURE 1-7:  Stomatal d i f f u s i o n resistance of 4 leaves i n the cucumber canopy, and greenhouse CC^ concentration on March 7, 1977.  Figures i n parentheses indicate  irradiance at leaf l e v e l during resistance —2 measurement (yE m  —1 s ,  400 - 700 nm) .  T I M E  O F  DAY  37  It i s worthwhile to mention at this point, some shortcomings of these experiments.  F i r s t l y , on both dates  that stomatal measurements were taken no adequate controls were available against which to compare the responses of the selected leaves to the greenhouse environment.  In each case the results  are purely descriptive and conclusions must, therefore, remain tentative.  Secondly, measurements of radiation incident on the  leaves were instantaneous values and may not have provided a true assessment of general irradiance conditions at that time of the day.  This problem was probably greatest on February 23, since  ambient light levels below the canopy tended to fluctuate rapidly, during alternate periods of bright sun, and cloud.  On March 7 the  sub-canopy radiation environment was much more constant.  A third  problem was the confounding of the effects of varying CO^ concentration and irradiance on stomatal response, which was unavoidable i n the greenhouse. The apparent lack of an effect of C0 concentrations 2  around 1000 y l 1  1  (present i n the greenhouse atmosphere during  the enrichment period on March 7 and at the 0830 h measurement time on February 23) i s not consistent with the results..of Heath and Russell (123).  They showed that the diffusive resistance of  illuminated stomata increased markedly between 170 and 1840 y l 1 i n wheat leaves.  The results of the present study are, however,  similar to those of Hurd (138) i n studies on a greenhouse tomato  i  38  crop.  He,itoo, discovered a p a t t e r n o f v a r i a t i o n i n stomatal  r e s i s t a n c e o f mid-canopy l e a v e s throughout  the d a y l i g h t p e r i o d  which was u n a f f e c t e d by CO^ enrichment t o 1000 y l 1 . -  X  High  l e v e l s o f CO^ i n greenhouse atmospheres may, t h e r e f o r e , be o f s m a l l consequence i n d e t e r m i n i n g cucumber and tomato c r o p s .  the use e f f i c i e n c y o f the gas i n C o n f i r m a t i o n of t h i s s u g g e s t i o n r e q u i r e s  f u r t h e r work p a r t i c u l a r l y w i t h cucumber p l a n t s , t e s t e d under c o n t r o l l e d c o n d i t i o n s o f CO^ c o n c e n t r a t i o n and i r r a d i a n c e . A c l e a r e r assessment o f t h e p a t t e r n o f d i u r n a l and s p a t i a l v a r i a t i o n i n s t o m a t a l r e s i s t a n c e w i t h i n t h e crop canopy can be made from the d a t a p r e s e n t e d i n F i g u r e s 1-8 and 1-9. show s t o m a t a l r e s i s t a n c e s o f the f i r s t  These  13 l e a v e s o f cucumber  p l a n t s measured on 4 o c c a s i o n s d u r i n g the day on February 23 and March 7.  At most measurement times, minimal  r e s i s t a n c e was found  to occur i n l e a v e s 3 and 4, s i t u a t e d j u s t below the top o f t h e canopy.  I n v e r y few cases were these low v a l u e s found to  c o r r e s p o n d w i t h the h i g h e s t i r r a d i a n c e i n the canopy.  Sometimes  r a d i a t i o n i n c i d e n t on these l e a v e s a t t h e time o f measurement, was lower  than t h a t on l e a v e s o f h i g h e r and/or lower  (e.g. February corresponded tomato crop  23 and March 7, 1030 h ) .  These o b s e r v a t i o n s  r e a s o n a b l y w e l l w i t h o t h e r in situ  (138).  l e a v e s 8 and 9.  I n t h a t case, minimal  insertion  measurements on a  r e s i s t a n c e s were found i n  I n tomato, however, i t was not p o s s i b l e to  determine whether l i g h t p l a y e d a r o l e i n these e f f e c t s , s i n c e  38a  FIGURE 1-8: Stomatal diffusion resistance and irradiance at leaf level of successive leaves i n the cucumber canopy, at 4 times during the day on February 23, 1977. Times indicate start of measurement sequence. .  . stomatal diffusion resistance  ••  •* quantum flux density  or UJ CO  top of  0830h  1 • 2 3i U  1030h  canopy  \\ 78H 9-  101H 12-  13H  b o t t o m of 1 c a n o p y 1630h  1300h  rg r 0  200  400  t o p of  canopy  (s c m " 1 )  600  800  QUANTUM FLUX  1000  1200  DENSITY  200  400  600  800  1000  1200  (uE m" s"UoO-700nm) 2  ID  39a  FIGURE 1-9:  Stomatal d i f f u s i o n resistance and irradiance  at leaf  l e v e l of successive leaves i n the cucumber canopy, at 4 times during the day on March 7, 1977.• Times indicate s t a r t of measurement sequence. •  •  stomatal d i f f u s i o n resistance  •  -  quantum flux density  LEAF  NUMBER  LEAF  NUMBER  41  irradiance within the canopy was not measured. The present results are also i n agreement with the work of Fischer (93) who demonstrated that stomata i n epidermal strips from leaf 5 to 7 of tobacco showed greater opening a b i l i t y than those from younger (numbers 1 to 4) and older (> 5) leaves. Thus, maximum aperture under test conditions was found i n the newest f u l l y expanded leaves (93). The results obtained i n the present study seem to indicate a similar effect of leaf age on stomatal resistance to that shown by Fischer's work.  In view of the uncertainty  that instantaneous measurements of irradiance on individual leaves were truly representative of effective, longer term radiation conditions, the p o s s i b i l i t y exists that minimal stomatal resistancesin leaves 3 and 4 were mediated by higher irradiances than those experienced by leaves at other levels. a  This seems to be  somewhat unlikely explanation, however, since the shading  effects of the upper leaves and neighbouring plants were considerable at most times during the day and the measurements showed no consistent trend of higher irradiance just below the top of the canopy. On most occasions during the present study, the highest stomatal resistances were shown by the oldest leaves at low levels i n the crop  canopy (Figures 1-8 and 1-9: leaves 11-13).  These high values were, for the most part, correlated with the lowest irradiances within the canopy. The results may be  42  consistent with those of Turner (785) who showed that increases i n stomatal resistance at progressively lower levels i n canopies of corn, poplar and dogwood could be explained entirely i n terms of reduced irradiance, u n t i l senescence induced leaf chlorosis. Davis e t al.  (73) have pointed out  that the effect of irradiance on stomatal resistance has two components. These consist of a conditioning effect whereby leaves adapt physiologically and structurally to ambient radiation conditions, and an instantaneous effect by which stomates are influenced by short term fluctuations i n radiant flux density. The physiological age of individual leaves was found to exert an influence on stomatal resistance which was exclusive of both components. Results obtained i n the present experiment at 1630 h on February 23 and March 7 (Figures 1-8 and 1-9) suggested that low irradiance during measurement had a more pronounced effect on stomatal resistance i n bottom leaves than i n those higher i n the canopy.  In addition, the increase i n stomatal resistance below  leaf 7 at 1300 h on March 7 (Figure 1-9) did not correlate with irradiance level which remained r e l a t i v e l y constant with leaf position.  These results may be due to the f i r s t component of the  irradiance effect mentioned previously, or to the greater age of leaves at progressively lower insertions. Unfortunately, the two factors were confounded i n these experiments.  Some support for the  involvement of leaf age as a determinant of stomatal resistance i n this  43  crop comes from the work of Hurd (138) who showed that leaf resistances in exposed tomato plants (at the edge of greenhouse rows) were higher i n old leaves from the low canopy than i n uppermost, young leaves. It appears from the data presented i n Figures 1-8 and 1-9 that high stomatal resistance i n leaves 1 and 2 i s a r e l a t i v e l y constant feature of these greenhouse cucumber plants. On several occasions on February 23 and March 7 resistances i n these two leaves were higher than i n those at the bottom of the canopy (e.g. 0830 and 1030 h, Figure 1-8), and at practically every measurement time they exceeded values for leaves 3 and 4. These observations are again i n accord with those of Hurd (138) who found that leaves i n the uppermost part of a greenhouse tomato canopy sometimes showed the highest resistances of a l l leaves on the plant.  This effect may have been due to dehydration of the  upper leaves which were often out of the range of the mid-canopy sprinkling system.  In this case i t i s , however, d i f f i c u l t to  explain the much lower resistances generally shown by leaves 3 and 4, which were also situated well above the sprinkling system. Another p o s s i b i l i t y i s that the higher resistances of leaves 1 and 2 are due to a closing response induced by high levels of irradiance, sometimes experienced at the top of the canopy, although such responses are more prevalent i n old than i n young leaves (182). The appeal of this explanation i s diminished by the observation that r e l a t i v e l y high resistances occurred i n these leaves i n the late afternoon (1630 h; Figure 1-8) when almost uniform irradiance conditions  prevailed at successive leaf insertions.  I t seems more l i k e l y  that the effects were related to the developmental stage of the leaves and possibly, a reduced stomatal density i n comparison with leaves of a s l i g h t l y greater physiological age (265). In summary, the p r o f i l e of stomatal resistance through the cucumber canopy displays a d i s t i n c t pattern quite similar to that of a greenhouse tomato crop (138).  As i n that case,  stomatal resistance of leaves from a l l levels i n the canopy did not show a clear response to CCX, enrichment. Resistances varied -  considerably according to the position of the leaf and level of irradiance, and i t was apparent that the lowest stomatal resistances occurred i n leaves just below the top of the canopy. Measurements of CC^ concentration using a conductimetric analyzer showed that greenhouse CC^ concentrations were normally maintained at a f a i r l y high level throughout the day i n the early stages of crop growth.  This effect probably resulted from CC^  production by decomposition of straw and organic material within the cropping beds.  I t was evident, however, that by the time the  crop canopy had developed appreciably, daytime CO^ levels became depleted below those necessary to maintain maximum rates of photosynthesis under optimal irradiance.  CO^ production from  natural gas burners was effective i n restoring CO^ concentrations to optimal daytime levels.  In recommending suitable enrichment  procedures for cucumber crops, i t i s worth bearing i n mind that  supplementary CG^ may is  (from combustion o r o t h e r a r t i f i c i a l  sources)  be of b e n e f i t f o r c e r t a i n p e r i o d s immediately a f t e r the crop transplanted.  As w i l l be shown i n P a r t I I of t h i s t h e s i s , the  b e n e f i c i a l e f f e c t s of CC^-enrichment to a p p r o x i m a t e l y 1000 i n another crop (tomato) may  yl 1 ^  be enhanced by commencing an  e f f e c t i v e enrichment program as e a r l y i n the growing p e r i o d as possible.  In cucumbers  proportion of  t h i s may mean g r a d u a l l y i n c r e a s i n g the  s u p p l i e d by a r t i f i c i a l  means, as the r a t e o f  d e c o m p o s i t i o n i n t h e s t r a w - s o i l c r o p p i n g bed slows. P r e v i o u s to t h i s study t h e r e has been l i t t l e i n f o r m a t i o n on the e f f e c t i v e l e v e l s of C0 houses u t i l i z i n g  ?  i n commercial  t r a d i t i o n a l c r o p p i n g methods.  c o n s i d e r a b l e u n c e r t a i n t y amongst  available cucumber  There was  growers as to whether  additional  CO2~ enrichment equipment, s i m i l a r to t h a t used f o r o t h e r greenhouse c r o p s , would be b e n e f i c i a l t o t h e i r o p e r a t i o n s .  Using a r e l a t i v e l y  simple apparatus, i t has been p o s s i b l e to demonstrate t h a t c o n c e n t r a t i o n s i n one house may  be i n s u f f i c i e n t  ambient  to promote  maximum crop growth w i t h o u t the i n j e c t i o n of a d d i t i o n a l CO2 the  atmosphere.  I t i s hoped t h a t t h i s system can be used  elsewhere i n cucumber  greenhouses to e s t a b l i s h ambient  concentrations i n different to  into  situations.  CO2  Such surveys s h o u l d  serve  p r o v i d e b a s i c i n f o r m a t i o n on the a d v i s a b i l i t y o f i n s t a l l i n g  enrichment  equipment.  CO2-  46  PART  The growth,  effect yield  of  atmospheric and gas  II  CO^ enrichment  exchange  tomato  plant.  -physiology  on  the  of  the  PART II  - INTRODUCTION  Tomato crops have been the subject of a large number of studies concerned with plant growth and yield responses to CC^ enrichment.  -  This considerable interest seems to have arisen for  several reasons.  Probably the most important of these i s the crop's  very great economic importance i n the greenhouse industry. In an early study, Bolas and Melville (32) showed that yield could be increased between 14 and 24% i n tomato crops grown i n a i r enriched with  to approximately 850 y l 1 .  Later work has established  X  that yield increases of 40% over normal air-grown tomato plants are possible with CX^-enrichment of crops grown i n the Spring (298). plants also respond to enhanced C0  2  The  concentrations by the earlier  production of ripe f r u i t and a reduction of the harvest period (115). As a result of these findings, CO^-enrichment i n tomato greenhouses has been enthusiastically adopted throughout Europe and North America (295) . The result's of early studies on tomato response to CO2"" enrichment also provided a strong impetus for similar research in a number of other greenhouse crops.  Yet, despite numerous studies  on the direct effects of supplemental CO2 on yield and growth under various conditions of "light, temperature and nutrient status (47, 52) very l i t t l e research has been concerned with the physiological basis for these effects (190).  Yield increases due to CO^-enrichment of  greenhouse atmospheres may be due to increased rates of photosynthesis  as a result of a steepened CG^ diffusion gradient from the bulk atmosphere to the sites of f i x a t i o n at the chloroplasts (103, 104). It i s , however, doubtful that this represents the only important factor.  Reduction i n l i g h t compensation point  of glycolate" production and photorespiration  (121), (310),  suppression of oxygen inhibition of photosynthesis  lower rates and the  (149, 150,  167)  may also result from high  concentrations and contribute to higher  net rates of photosynthesis.  I t i s also worthwhile to note that the  growth of plants and plant parts i s the result of interactions between many factors, and the l i n k between yield and photosynthetic rate i s often complex. The information available concerning the effects of  C^  -  enrichment during plant growth on subsequent rates of photosynthesis i s very limited.  Three studies have indicated that some persistent  changes may occur i n the photosynthetic response of enriched plants (21, 102, 185).  In each case, the physiological basis for these change  was not investigated. Recently, however, studies have been carried out which may help to determine the basis for the persistent changes i n photosynthetic metabolism observed i n other work. CO^- enrichment has been found to increase the a c t i v i t y of a number of enzymes associated with CO2 fixation i n a variety of plants (59, 92, 112), and to decrease a c t i v i t i e s of others important i n photorespiration (92).  Thes  results suggest that some changes i n photosynthetic a c t i v i t y associated with C0„-enrichment may be due to a modification of enzyme systems i n  the leaves of enriched plants.  Other studies have indicated  significant increases i n carbohydrate levels i n plants grown under COy-enriched conditions (184, 186).  In several cases, the high leaf  starch content resulted i n chloroplast deformation (186) which seemed to be correlated with a decrease i n apparent photosynthesis (143, 185,  190). The present study consists of 2 sections, the f i r s t dealing  with the effects of supplementary CO^ on greenhouse-grown tomato plants, and the second with the effects of 3 different levels of CO ~enrichment on tomato seedlings raised i n growth chambers. The f i r s t section of the study was prompted by the lack of information on the relative photosynthetic responses of greenhouse crop plants grown under C02~enriched and normal a i r conditions.  The  primary objective was to compare the growth and gas exchange physiology of two greenhouse tomato crops grown with and without CO2-enrichment. Two greenhouses were available for this part of the project, so preliminary experiments were required to determine the f e a s i b i l i t y of conducting a comparative study where supplementary CO^ would be introduced into only one house.  Once this was established, a further  series of experiments to elucidate the effects of CO — enrichment on growth, development, and CO^ exchange were begun. The approach taken was f i r s t l y to measure rates of leaf and flower formation, and y i e l d i n each crop.  Secondly rates of CO^ and water vapour exchange were  monitored to assess photosynthetic potential and diffusion resistances  in leaves developed under CO^—enriched and normal a i r atmospheres. Of particular interest at the outset of this research was the p o s s i b i l i t y that supplementary CO^ during growth might induce persistent changes i n the photosynthetic system. Accordingly, due attention was paid to this p o s s i b i l i t y and to whether such changes might be influenced by leaf age. The greenhouse experiments established that, i n fact, distinct differences i n development and changes i n photosynthetic physiology were evident i n plants grown under CO^-enriched conditions. Upon completion of this part of the study the physiological basis for these effects was s t i l l not f u l l y clear.  I t was decided, therefore, to  investigate this problem i n greater depth. Conventional greenhouse environments were unsuitable for this second study since they did not permit maintenance of constant CO2 concentration.  Therefore, a series  of laboratory experiments was undertaken using small growth chambers within which ambient CO^ levels could be closely controlled and monitored. The greenhouse study clearly demonstrated an increase i n inherent photosynthetic capacity of leaves developed at a CO2 concentration of approximately 0.09%y  I t was not evident whether  similar effects would be shown at higher growth concentrations which may be encountered i n greenhouses lacking control systems on CO2enrichment equipment.  The scope of the laboratory study was expanded  to investigate the growth and physiology of tomato plants at 0.5%  i n addition to 0.1% and 0.03%''.CO . An attempt was.made to determine whether the observed differences i n photosynthetic response between CO2—enriched and non-enriched plants would persist after return to a common, normal a i r environment.  The results of these  studies provided considerable additional information on the response of tomato plants to supplemental C0 during growth, i n terms of 2  observable physiological changes and the underlying mechanisms for these changes.  \  52  PART ii, SECTION:.! EXPERIMENTS ON GREENHOUSE CROPS MATERIALS AND METHODS i)  Selection  of Suitable  Greenhouses for  a Comparative Study (Experiment 2)  During the Spring of 1976, an experiment was conducted to compare the growth and f r u i t yield of two tomato crops grown under similar temperature and s o i l conditions i n adjacent greenhouses of glass construction. Each house had a floor area of 58 m2 and a i r volume of 182 m3. They w i l l be referred to as House A and B. Seeds of tomato {Lycopersi con esculentum L.) cv. Vendor, (from Stokes Seeds Ltd.,  St. Catharines, Ont.) were germinated i n  moist s o i l i n f l a t s during February 1976.  At the beginning of  March, 160 seedlings were transplanted into buckets containing 9 l i t r e s of s t e r i l i z e d s o i l with an admixture of 3 g;l  1  Osmocote  14-14-14 slow release f e r t i l i z e r (Sierra Chemical Co., Milpitos, Ca.).  Eighty seedlings were placed i n rows i n each house with 2  approximately 0.3 m of bench space provided per plant. Hand watering was carried out daily and the plants were subject to occasional  pest control procedures (monthly fumigation with  Plant Fume 103, Plant Products Co., Bramalea, Ont.).  Daytime  temperatures i n both houses were maintained at 24°C i n the early season, and roof ventilators were set to open at this temperature. Night temperatures varied between 15 and 18°C.  Three weeks after transplanting, four plants were harvested  from each house to determine t o t a l leaf area and plant  dry weight.  Three s i m i l a r harvests took place at 21 day i n t e r v a l s  thereafter.  Relative Growth Rate (RGR) and Net Assimilation Rate  (NAR) were calculated by means of functions derived from fittedcurves of ^he primary values against time (t) (286) .  (leaf area (A) and plant dry weight (W))  A multiple regression program was used  to derive suitable curves using the U.B.C. IBM 370 d i g i t a l computer. Input to the program consisted of the dependent v a r i a b l e (W or A) and  two independent variables (t and the second degree  polynomial,  2 t ).  Parabolic functions of W and A against time were calculated  for the two crops.  Analysis of Variance  of each regression showed  a l l the relationships to be highly s i g n i f i c a n t (P < 0.01). 1 dW 1 dW Using accepted d e f i n i t i o n s of RGR = — and NAR = — — the following revised formulae were established i n terms of multiple regression functions X(t) = W and Y(t) = A.  RGR  N A R N  A  K  dt  =  X(t)  dX£tl . _1_ dt Y(t)  Mean Leaf Area Index (LAI) was estimated  at several dates a f t e r  transplanting from the regression of A against t, and the bench  54  area occupied by each plant i n the respective houses.  Crop Growth  Rate (CGR) was derived from the product of LAI and NAR at selected dates (169,.). Plants i n both houses began to set f r u i t during the second week i n A p r i l .  Fruit y i e l d was determined by picking and weighing  marketable f r u i t from plants as i t became ripe. u n t i l May 18, 1976.  Picking continued  55  ii)  CO,, Enrichment Experiments  (Experiments  Cl  3 and 4)  During January 1977, tomato seeds of the same cultivar as used i n the preliminary experiments were germinated i n moist micapeat.  Upon emergence i n late January, seedlings were planted  i n mica-peat i n 40 small pots, each containing one plant. The pots were randomly divided into two groups of 20 and placed into the two greenhouses used i n the preliminary study.  When the f i r s t  two true leaves were developed, i n mid-February, 1977, the seedlings were transplanted into separate plastic bags which contained c_a. 9 1 of Douglas F i r (Pseudotsuga menziesii  (Mirb.)  Franco) sawdust. The bulk sawdust contained an admixture of 2.55 g 1  1  of 19.5% superphosphate, 4.18 g 1 of dolomitic limestone 1  and 1.0 ml 1 of minor element solution (183). 1  During growth,  nutrient solution containing 5.8 g 1 KNO^ plus 0.15 or 0.27 1  or 0.40 g 1 NH^NO^ (different concentrations associated with 1  different stages of development as specified by Maas and Adamson (183)) was supplied via separate plastic feeding tubes to each bag. The rate of supply of the nutrient solution to each bag was increased i n stages from 450 ml day mid-April.  1  i n February to 2500 ml day- i n 1  This nutrient regime conformed to that recommended by  Maas and Adamson (183) except that iron was supplied i n the minor element solution as FeEDTA (ferrous ethylenediaminetetraacetic acid containing 5 g 1 available iron) at the rate of 0.51 ml per 1  1 of sawdust.  56  Temperature regimes i n each greenhouse were similar to those used i n the 1976 study although daytime temperatures i n both houses rose to 27°C or above during many warm sunny days i n A p r i l and May. In general, CC^—enrichment procedures were similar to those i n use i n the local greenhouse industry.  The house i n  which CO-enrichment was applied i n 1977 was that which had produced the quantitatively lower yield i n the 1976 study.  C0 -enrichment 2  was begun at the time of seedling emergence and continued u n t i l April.  Each day CO^ was added between 30 min before sunrise and  30 min after sunset. The source was a propane burner (No. 1332, Johnson Gas Appliance Co., Cedar Rapids, Iowa) which produced C0 at a constant daytime rate. 2  The burner was thermostatically  controlled to switch off i f the house temperature was above 24°C i n order to prevent the addition of C0 at times when the roof 2  vents were open.  After late A p r i l , continued C0 —enrichment was not 2  feasible because the roof ventilators remained open v i r t u a l l y a l l day. At two week intervals during the experiment, C0 concentrations 2  i n each house were checked over a 24 h period using the conductimetric analysis system described i n Part I of this thesis. During early growth, leaf development of plants i n the enriched and normal a i r regimes (as the greenhouse treatments w i l l be referred to throughout this text) :was assessed using the plastochron index. (87). A reference length of 30 mm was chosen and leaf lengths were measured  57  on 3 replicate plants i n each regime using a d i a l micrometer. These measurements were discontinued after March..18 because the i n i t i a t i o n of flowering i n some plants could have produced non-linearity i n the relationship between plastochron index and time (87).  After March 18, plant development was assessed i n  terms of flowering and truss development. At approximately 3day intervals each plant was assigned a grade on the basis of the number of trusses developed with closed flowers, the number with open flowers, and the number with one f r u i t set on at least one flowerhead.  These data were collected u n t i l the study ended on  July 9. Fruit yield was determined as described previously with harvests taken at irregular intervals ranging from 3 to 7 days apart.  Due to the death from Fusarium w i l t  of 3 plants i n the  normal a i r growth regime i n late May, the experimental design became unbalanced.  This problem was remedied by randomly discarding  f r u i t yield values for 3 plants from the enriched regime.  Cumulative  yields are therefore expressed as means of 17 replicate plants per growth regime from the beginning of the harvest period. Analysis of flowering, leaf development and yield i n the two crops constituted experiment 3 of the thesis research. Experiment 4 was concerned with simultaneous  determinations  of CC>2 and water vapour exchange by attached leaves carried out during a 3 week period from mid-April to early May. Measurements  58  were conducted on true leaf 5 ("basal" l e a f ) , counting from the f i r s t above the cotyledons, and on the most recently unrolled apical leaf on the main axis of the same plant.  Gas exchange  determinations were made in situ on two replicate plants per CO^ regime by enclosing the terminal leaflet portion of a compound leaf i n a 43 ml, trap type Plexiglas chamber. The leaf chamber was connected i n an open gas flow system, and the i n l e t and outlet ports of the chamber were arranged to create turbulence i n the gas stream passing by the enclosed leaf.  Gas of known CC^ concentration  was supplied to the chamber by mixing CO^-free compressed a i r with a i r containing 1.0% Ingoing  using a Matheson 7321 gas proportioner.  concentrations were quite stable and were checked  frequently with a Beckman IR 215 CO2 analyzer (IRGA).  During  measurements, the gas supply to the leaf chamber was maintained at 825 ml min . x  The IRGA was used i n i t s d i f f e r e n t i a l mode to  determine the change i n CO2 concentration as the gas passed through the leaf chamber. The CO2 concentration d i f f e r e n t i a l across the chamber was never greater than 40 u l 1  _ 1  and was- usually less than 20  y l 1 .. The ingoing a i r was humidified to a relative humidity i  between 65 and 75%, and an electric hygrometer (Model 880, EG & G Ltd., Waltham, Mass.) was used to measure the dew point of the gas before and after the leaf chamber. The returning a i r stream was dried by passing through a calcium sulphate drying column before entering the IRGA.  59  Rates of gas exchange were calculated from the gas flow rate and the changes i n CO^ or water vapour concentration as the gas stream traversed the chamber. After a l e a f l e t was used for a series of measurements i t s outline was traced onto coarse f i l t e r paper.  The outlined leaf model was cut from the paper, moistened,  and used to determine a i r boundary layer resistance to gas transfer. Stomatal resistances were calculated from rates of transpiration, boundary layer resistances and water vapour pressure within the leaf chamber. The logarithmic relationship discussed i n yiarvis and S a t s k y ' (147) was used to estimate mean CG^ and water vapour levels i n the chamber atmosphere.  Intercellular space CG^  concentrations were calculated as indicated by Moss and Rawlins (210). Me'sophyll resistances  (  r  m  )  t o GO^  uptake w e r e calculated from t h e  reciprocal of the slope of the regression relating intercellular space  concentrations (Ci) minus CO^ compensation concentration  (17) to the flux of C0 i n net photosynthesis 2  (F) (146, 177, 282).  Mesophyll resistances were not calcuated for the apical leaves of plants grown i n the normal a i r regime since the relationship between CO^ flux and internal CO^ concentration was not significantly linear i n that case. Rates of gas exchange were measured under a l l combinations of 4 levels of photosynthetically active radiation (PhAR; 50, 150, -2 250, and 750 yE m C0  -1 s  , 400 to 700 nm) and 4 concentrations of  (240, 340, 440 and 540 y l 1 ) . _ 1  0  The enclosed leaf was allowed  60  to e q u i l i b r a t e f o r 20 to30 minutes a f t e r each change of t e s t conditions.  Leaf and a i r temperatures were monitored continuously  using 0.2 mm diameter copper-constantan thermocouples pressed to the a b a x i a l surface of the l e a f and positioned i n the a i r stream adjacent to the gas i n l e t ports.  L i g h t was provided by  e i t h e r a 100 W incandescent lamp s i t u a t e d d i r e c t l y above the chamber or by a 300 W cool beam lamp (General E l e c t r i c Ltd.) 1 m -2 above the chamber ( f o r 750 yE m  -1 s  only).  Figure I I - l shows  the s p e c t r a l energy d i s t r i b u t i o n of r a d i a t i o n from each lamp as measured by an ISCO model SR spectroradiometer.  PhAR was determined  by a Lambda Instruments Inc. ( L i n c o l n , Nebraska) L I 190 S Quantum Sensor w i t h a L I 185 meter.  Several layers of cheesecloth  interposed between the lamps and the chamber were used to adjust the PhAR l e v e l s .  Leaf temperature i n the chamber was maintained at  25 ± 2°C by c i r c u l a t i n g water through a 3 cm deep jacket attached to the top of the chamber. Rates of dark r e s p i r a t i o n and CO^ compensation points were determined f o r a p i c a l and basal leaves on three randomly selected plants from each growth regime.  Compensation points f o r  i r r a d i a n c e were c a l c u l a t e d by i n t e r p o l a t i o n between the CO2 f l u x of the lowest l e v e l of PhAR and the relevant mean dark r e s p i r a t i o n rate.  These measurements involved a simple closed gas a n a l y s i s  system incorporating an a i r pump, the l e a f chamber and the CO2 analyzer.  Leaf areas were determined using a p h o t o e l e c t r i c  planimeter (Hayashi Denko Co. L t d . , Japan).  66a  FIGURE I I - l :  Spectral energy d i s t r i b u t i o n of 100 W incandescent lamp and 300 W cool beam lamp used to illuminate assimilation chamber i n Experiment 4.  62  Certain behavioral indices of photosynthesis were calculated for apical and basal leaves grown under CG^-enrichment, and for basal leaves grown i n normal a i r . The indices were derived from a model for leaf CO^ assimilation described by Van Bavel (14). The model allows the flux of CC^ i n photosynthesis to be separated into two elements depending upon:  (a) the radiant  flux density and associated photochemical u t i l i z a t i o n efficiency of incident radiation, and (b) the internal CC^ concentration and associated CC^ u t i l i z a t i o n efficiency.  The model yields two  parameters, E and E respectively defined as the efficiencies of radiant energy and CO^ u t i l i z a t i o n .  The model can also be used  to estimate the theoretical maximum flux of C0„, F ,, which would 2 IM Tl  occur at f u l l y saturating levels of PhAR and CG^ concentration. The calculation of each parameter was carried out as described by Van Bavel (14), and i s reviewed i n Appendix 2'.... S t a t i s t i c a l analysis of the gas exchange results was carried out .by multiway analysis of variance (ANOVA) on an IBM 370 d i g i t a l computer.  In order to elucidate differences i n photosynthesis  rates between growth regimes, separate ANOVA models were used for basal leaves and apical leaves.  This approach provided an  alternative to pooling the data for different ages which preliminary tests had shown to be significantly (P < 0.05) different. stage multiple range test (Student-Newman-Keuls,  A two  a = 0.05) was  included i n the analysis to provide a method of mean separation.  63  Statistical  tests applied  to o t h e r d a t a i n the study are  i d e n t i f i e d , wherever a p p r o p r i a t e , i n the t e x t and i n f i g u r e table  headings.  and  64  PART II,  SECTION 1  RESULTS AND DISCUSSION i)  Comparative Study Concerning Growth of Tomato Crops in Unenriched Greenhouses (Experiment 2) The functions derived from multiple regression analysis of the primary growth parameters A and W against t are l i s t e d i n Table II—1.  These functions were subsequently used to calculate  values f o r RGR, NAR and CGR by d i f f e r e n t i a t i o n at various dates after transplanting and to produce the curves shown i n Figures II-2 to II-4.  The forms of the expressions  used to estimate the  respective growth parameters are shown i n the corner of each f i g u r e . Mean RGR and NAR as well as CGR were considerably higher i n house A as compared with house B.  S p e c i f i c reasons f o r the  discrepancies i n vegetative growth between the two crops were not investigated.  Temperature regimes and c u l t u r a l conditions were  unlikely sources of the v a r i a t i o n between the two houses.  It i s  more probable that the differences were due to higher incident l i g h t levels throughout the growing period i n house A, and this unfortunately represented an important uncontrolled variable throughout these and subsequent experiments.  The importance of  irradiance i n determining growth rates and NAR f o r plants and crops i s well established.  In general, RGR, CGR, and NAR decline  proportionally with a reduction i n incident radiation (28, 289).  TABLE I I - l :  Functions of plant dry weight and leaf area with-time derived from m u l t i p l e regression equations f o r tomato plants grown i n experimental greenhouses A and B, without CO^ enrichment.  HOUSE B  HOUSE A Coefficient: Leaf Area (A)  Coefficient: Dry Weight (W)  Coefficient: Leaf Area (A)  Coefficient: Dry Weight (W)  Const, 5215.8 A  Const, 16.6 W  Const, 2065.2 A  Const, 10.0 w  1.4  119.4  169.2  W c-, W  -2.1  -0.004  Function f o r W = Consty + b ( t ) + c y ( t ) with time w  Function f o r A = Const + t> (t) + c ( t ) with time A  A  A  Where t = time i n days from transplanting  0.1  W c W  1.6 -0.002  65a  FIGURE I I - 2 :  Time course of RGR  d e r i v e d from f i t t e d  f o r tomato crops grown i n e x p e r i m e n t a l A and B w i t h o u t  CO  -enrichment.  curves, greenhouses  D A Y S  F R O M  T R A N S P L A N T I N G  66a  FIGURE I I - 3 :  Time course of NAR  d e r i v e d from f i t t e d c u r v e s , f o r  tomato crops grown i n e x p e r i m e n t a l greenhouses A and B w i t h o u t C0„  enrichment.  DAYS FROM  TRANSPLANTING  67a  FIGURE I I - 4 :  Time course of CGR d e r i v e d from v a l u e s o f NAR and LAI, f o r tomato crops grown i n e x p e r i m e n t a l greenhouses A and B w i t h o u t  C0  o  enrichment.  DAYS FROM TRANSPLANTING  The and  c l e a r c u t d i f f e r e n c e s i n r a t e s of v e g e t a t i v e growth  dry matter a s s i m i l a t i o n c o n t r a s t e d s h a r p l y w i t h s i m i l a r mean  f r u i t y i e l d s on a per p l a n t b a s i s o b t a i n e d (Table I I - 2 ) .  i n each greenhouse  E v i d e n t l y , the environmental f a c t o r or f a c t o r s which  caused the growth d i s c r e p a n c i e s d i d not a f f e c t f r u i t p r o d u c t i v i t y on the p l a n t s i n the two  houses.  c l o s e s i m i l a r i t y , i t was  decided  experiments i n these l o c a t i o n s . greenhouse and  treatment  L a r g e l y on the b a s i s of  this  to proceed w i t h  the CO - enrichment  The  confounding of  unavoidable  (C02~enrichment v e r s u s normal a i r )  a m e l i o r a t e d by s u p p l y i n g a d d i t i o n a l CO2  was  i n the house which produced  lower growth and net a s s i m i l a t i o n r a t e s and q u a n t i t a t i v e l y lower fruit yield reasonable  ( i . e . house B), i n the p r e l i m i n a r y study. to assume t h a t any  I t seemed  demonstration of i n c r e a s e d  fruit  y i e l d , r a t e of v e g e t a t i v e or r e p r o d u c t i v e development by a crop grown i n t h i s house would c o n s t i t u t e s t r o n g evidence e f f e c t i v e n e s s of the enrichment regime.  f o r the  TABLE I I - 2 :  Mean f r u i t y i e l d from tomato plants grown i n experimental greenhouses A and B, from March 16 to May 18, 1976C without CO -enrichment.  House A  House B  Mean f r u i t fresh weight (g per  606.7*  plant)  * d i f f e r e n c e i n y i e l d between houses not s i g n i f i c a n t (P > 0.05) by t - t e s t .  598.8  71  ii)  CO ^ Concentrations  in  CO -Enriched  and  Control  (Normal-Air)  Greenhouses  The results i n Figure II-5 are representative of the house CO^ concentration measurements taken during early and late stages of the second part of the greenhouse experiment. roof vents opened infrequently and daytime remained between 800 arid 1000 y l 1 .  On March 12,  concentrations  On warmer days-(as exemplified  X  by data for A p r i l 23), vent opening was more frequent and CO2 levels i n the enriched house ranged between 550 and 850 y l 1 . x  Daytime maximum and minimum CO2 levels recorded i n the unenriched house were often well below 300 y l 1 . x  The lower concentrations  on A p r i l 24 compared to March 13 occurred despite more frequent vent opening and may be correlated with an increase i n f o l i a g e within the house, between these two dates.  71a  FIGURE I I - 5 : Atmospheric CO^ c o n c e n t r a t i o n s  in C O 2 -  e n r i c h e d and normal a i r (unenriched) greenhouses.  \C0  2  enriched house  A March 12  C 0 enriched house 2  0700 0900  1100  1300 1500  1700  1900 2100 TIME  0700 0900 1100 OF  1300  / \ April 23  1500 1700 1900 2100  DAY  K)  73  iii)  Plant Growth and Held  (Experiment 3)  Early leaf production by plants grown i n the CO^-enriched house was quantitatively higher than production by plants grown i n normal a i r (Table II-3) but this difference was not s t a t i s t i c a l l y significant.  This observation corresponds closely with those of  previous studies where CG^-enrichment to 1000 y l 1 has been 1  shown to have negligible or very small effects upon the rate of leaf development (137, 215).  In this respect the effects of CX^-  enrichment d i f f e r from those of other environmental parameters known to enhance tomato growth such as high light intensities and temperatures.  In several cases these parameters have been shown  to increase leaf production (46).  If should be noted, however,  that since leaf production i n the present study was recorded only i n terms of the plastochron index, i t i s possible that unobserved aspects of leaf production, such as area or dry weight accumulation, might have been influenced by  regime during growth.  Wittwer  and Robb (29,8), for example, noted an increase of ca. 25% i n total dry weight of lettuce leaves grown under 1000 y l 1 compared to 1  leaves produced under 400 y l 1-CO . The impact on vegetative growth 1  of similar levels of CO^—enrichment has also been demonstrated i n terms of increased relative growth rates and net assimilation rates (273). In the present study, CO — enrichment caused a d i s t i n c t advancement of truss development (Figure II-6).  Flowers opened ca. 3  TABLE I I - 3 :  Time course of l e a f development i n tomato p l a n t s grown w i t h o r w i t h o u t CO enrichment.  P l a s t o c h r o n Index Date (1976)  A i r Regime  CO - E n r i c h e d Regime  Feb. 28  5.30±0.89*  5.99±0.37  Mar.  2  5.91±0.62  6.69±0.44  Mar.  8  8.73±1.31  9.16±0.47  Mar. 12  9.72±1.32  10.9'3+0.53  Mar. 15  11.11±1.17  12.31+0.67  Mar. 18  11.58±1.60  13.43±0.58  *Values shown a r e means ± 1 s t a n d a r d d e v i a t i o n from 3 r e p l i c a t e p l a n t s p e r regime. D i f f e r e n c e s between regimes a t any date were not s i g n i f i c a n t (P > 0.05) by t - t e s t .  74a  FIGURE I I - 6 :  Truss development. growth regime.  Open b a r s :  C02~enriched  Cross-hatched b a r s :  normal  air  growth regime  ( c r o s s - h a t c h i n g superimposed):  A:  Number o f t r u s s e s developed w i t h f r u i t s e t on a t l e a s t one flowerhead.  B:  Number of t r u s s e s developed w i t h open flowerheads.  C:  Number of t r u s s e s developed w i t h c l o s e d flowerheads.  NOTE:  Data a r e means f o r 17 r e p l i c a t e p l a n t s per treatment.  tomato  A t each date,  A + B + C are s i g n i f i c a n t l y  different  between treatments a c c o r d i n g to WhitneyMann t e s t  f o r non-parametric  d a t a (P < 0.001).  7.0  FRUIT SET  60 504 030 2-010-  TRUSSES WITH OPEN FLOWERS  TRUSSES WITH CLOSED  2 5 - 3 28-3 2 - 4  5 - 4 1 3 - 4 18-4 2 6 - 4 2 9 - 4  3-5  FLOWERS  9-5 1 3 - 5 1 6 - 5 1 9 - 5 2 4 - 5 3 0 - 5  DATE (day-month)  2-6  6-6  76  days e a r l i e r i n the enriched plants, beginning at about 6 weeks a f t e r seedling emergence, and the number of trusses bearing f r u i t after A p r i l 13 was always s i g n i f i c a n t l y higher under the enriched treatment.  These results are i n agreement with most other studies  i n which flowering and f r u i t set i n a number of d i f f e r e n t tomato c u l t i v a r s has been shown to be increased by 3 to 5 days as a result of C0 -enrichment to 1000 y l 1 2  _ 1  (49, 116, 298).  They do not  correspond with the results of Knecht and O'Leary (163) who found no s i g n i f i c a n t difference between numbers of open flowers sixty-four days after seeding on tomato plants grown at 400, 800, and 1200 y l 1  X  Those plants, however, were cultivated i n  growth chambers under a r t i f i c i a l l i g h t i n contrast to the other, greenhouse experiments, which may explain some of the differences between them. Plants exposed to the higher CO^ levels also showed greater f r u i t y i e l d s throughout the harvest period (Figure II-7) and differences i n cumulative f r u i t y i e l d were accentuated end of the experiment.  toward the  Y i e l d increases i n response to CO^-enrichment  i n tomatoes have been widely reported (165, 189, 297., 298), the magnitude of the increase over normal a i r grown plants depending on other c u l t u r a l conditions, notably l i g h t and temperature .regimes (116).  The percentage increase (ca. 30%) i n marketable f r u i t  y i e l d as a .result of CO-enrichment i n the present experiments was consistent with that obtained i n many commercial and experimental greenhouses for Spring-grown tomato crops (165).  J  76a  FIGURE II-7: Cumulative f r u i t y i e l d f o r plants grown i n CO2enriched and normal a i r growth regimes. NOTE:  Data are means of 17 tomato plants per treatment.  Differences at any date after  May 24 are s i g n i f i c a n t according to t-test (P < 0.05).  77  78  iv)  Leaf Photosynthesis  (Experiment 4)  It i s often assumed that growth and y i e l d increases resulting from CO^-enrichment are brought about by increased rates of photosynthesis.  There i s , however, very l i t t l e information -  relating to the photosynthetic responses of plants which have been grown at supra-normal CO^ concentrations (185).  In order to  elucidate the basis for the differences i n yield and plant development between CO^-enriched and normal a i r grown plants i t was decided to evaluate some components of photosynthetic CO^ exchange i n plants from each regime. Under comparable test conditions of PhAR and CO^ concentration, the net rates of photosynthesis of basal (Fig. II-8) or apical (Fig. II-9) leaves were generally greater i n leaves developed under CO - enrichment.  The overall pattern of the  responses to variations i n PhAR and test CO2 concentration, however, was similar for leaves grown i n the enriched and normal a i r regimes. S t a t i s t i c a l analysis of these results showed significant (P <0.05) interactions between growth regime, PhAR, and CO^ concentration. A more direct impression of the effect of growth regime on leaf CO^ exchange i s given by Fig. 11-10 which shows the difference in net photosynthesis rate (APs) between leaves grown i n the two regimes.  In both basal and apical leaves, APs tended to increase  with increasing C0 concentration (Fig. II-10A). 9  Increasing PhAR  78a  FIGURE: II-8: Rates of apparent photosynthesis i n basal leaf 5 of plants grown i n normal a i r (-  --•• ) and  CC^-enriched (•———•) growth regimes. NOTE: Data are means of 2 tomato plants per treatment.  Points marked with the same  l e t t e r are not s i g n i f i c a n t l y different (P > 0.05) according to SNK multiple range test.  79a  FIGURE I I - 9 :  Rates of apparent photosynthesis i n f i r s t unrolled apical leaf of plants grown i n normal a i r (and C02~enriched (• NOTE:  • ) growth regimes.  Data are means of 2 tomato plants per treatment.  Points marked with the same  letter are not significantly different (P > 0.05) according to SNK multiple range test.  •*)  APPARENT  >  oo o  80a  FIGURE  11-10:  Differences i n apparent photosynthesis rates between tomato plants grown i n C02~enriched and normal a i r growth regimes (APs), i n relation to (A) concentration and (B) quantum flux density. Points represent mean APs for a fixed level of one factor over a l l tested levels of the other. Apical leaves: A  A;. Basal leaves: A  A.  apical leaves  g  basal leaves  1  200  QUANTUM  1  400  1  600  1  800  F L U X DENSITY (JJE m ' s" . 2  1  400-700nm)  82  also increased APs i n basal leaves, but no consistent trend was evident with apical leaves (Fig. II-10B).  The data suggest that  APs was more responsive to changes i n test CC^ concentrations than changes'in PhAR. Thus, i n basal leaves, a 2.25-fold increase i n test CC^ concentration caused a 1.8-fold increase i n APs, while a 15-fold increase i n PhAR resulted i n a 4.5-fold increase i n APs.  I t i s worth pointing out, however, that over the range of  test levels of PhAR used i n these experiments, leaf photosynthesis undoubtedly varies between a state of irradiance limitation to near irradiance saturation. At the highest test CG^ concentration, on the other hand,, photosynthesis i s probably proceeding at less than half i t s maximum rate (103).  These differences probably hold  important consequences for the relative effects of varying PhAR and CO2 concentration on APs. The tendency of APs to increase at progressively higher levels of PhAR and CO2 concentration i s i n contrast to the results of:some other studies. Bishop and Whittingham (21) found persistent increases i n photosynthesis rate of tomato seedlings grown i n 1000 y l 1 CO2 following transfer to normal a i r at low l i g h t intensity, but not at high intensity.  More recently, i t was found (102) that the  transfer of cucumber seedlings from a growth concentration of 1500 y l 1 ' _ . CO2 to normal a i r resulted i n lower photosynthesis rates -2 at an irradiance of 98 W m air.  than i n seedlings cultivated i n normal  At approximately half that irradiance, there was no difference  83  between the two regimes.  I t has been suggested (21) that the  modification of photosynthesis rates following CC^-enrichment may be related to changes i n pool sizes of photosynthetic intermediates.  There are many- other possible explanations and some recent  work has suggested that after growth of plants:in CO^-enriched conditions changes i n a c t i v i t i e s of enzymes involved i n CC^ exchange may be important (92). that  The results shown in Table II-4 indicate  compensation points were reduced following growth under  CO^- enrichment.  The magnitude of the CO^ compensation point often  reflects the rate of photorespiration (7$, 196 , 278) . The present ;  results, therefore, raised the p o s s i b i l i t y that changes i n photorespiratory a c t i v i t y may contribute to the alterations i n net photosynthetic rates.  The CC^ compensation point differences  prompted further study of photorespiration and the a c t i v i t y of enzymes involved i n CC^ exchange i n CG^ enriched plants. -  The results  of these studies w i l l be reviewed i n Part I I , Section 2 of this thesis. It i s well known that the photosynthetic capacity of leaves changes with age (6, 97, 148, 161, 223, 248, 256). rates of photosynthesis  rise to a maximum shortly after leaf  emergence and thereafter decline. photosynthesis  Generally  In accord with this, net  rates i n apical leaves tended to be about double  the rates of older basal leaves (Figures II-8 and II-9). The results also indicate a different response to growth under C0„-  TABLE II-4:  compensation points f o r leaves of tomato plants grown with or without C0„-enrichment.  CO2 Compensation Point ( y l 1 ^) A i r Regime  C0 -Enriched 2  Apical leaves  55 * b  46 a  Basal leaves (5th)  55 b  44 a  *Values are means of 3 r e p l i c a t e determinations. Values designated with the same l e t t e r are not s i g n i f i c a n t l y different (P > 0.05) by • AN OVA and SNK test.  Regime  enrichment between the apical and basal leaves.  In apical leaves,  values of APs tended to be much more sensitive to changes i n test CC>2 concentrations (Fig. II-10A), and possibly less sensitive to changes i n PhAR (Fig. II-10B), than i n basal leaves.  CO^  compensation points, however, were not significantly affected by leaf age (Table  II-4).  The differences i n effects of CO^-  enrichment between apical and basal leaves could be due to a number of factors. Apical leaves clearly emerged much later than basal leaves and were therefore exposed to the a e r i a l environment for a shorter time period.  I t i s possible that different times of  exposure to CO^-enriched conditions or d i f f e r e n t i a l effects of elevated CO^ concentration on the photosynthetic system of young and older leaves were responsible for the observed differences.  86  v)  Resistances  to C0„  Assimilation  Stomatal behavior was also influenced by growth regime and by conditions during the gas exchange measurements (Fig. 11-11). In plants grown i n normal a i r , stomatal resistances under comparable test conditions were lower i n apical leaves than i n basal leaves.  Similar differences were not obvious with leaves  from the CG^-enriched plants, and this may indicate a need to consider apsects of leaf development and growing conditions when undertaking studies on stomatal physiology.  Stomatal resistances  tended to decrease with increasing PhAR irrespective of growth regime, although this tendency was not strong i n apical leaves from the normal a i r regime which consistently had low resistances. Overall, the effect of test  concentration on stomatal  resistance was not s t a t i s t i c a l l y significant (P < 0.05). Other investigations have often observed increased stomatal resistances at higher than normal CO^ concentrations i n various plant species (99, 123, 152). • The relative stomatal insensitivity-to test ' C0  2  concentration found i n the present study, however, coincides  with the observations of Pallas (225) who examined tomato leaves over a wide range of CO2 concentrations. Mesophyll resistance was always considerably greater than stomatal resistance at equivalent levels of PhAR (Fig. 11-11), and i t was therefore the dominant resistance to CO 2 assimilation under the conditions tested i n this study.  Because of the procedure  used for i t s calculation, mesophyll resistance i s i n turn composed of elements related to l i q u i d phase transport of CO2 and to carboxylation at the photosynthetic sites within leaf c e l l s (146, 171)-;  Since  FIGURE 11-11: Leaf resistances to C 0 transfer. 2  NOTE: Data for r (stomatal resistance) are means s of 2 determinations per treatment, r m represents mesophyll resistance calculated as described i n the text. test C 0 concentration. 2  C 0 represents 2  Bars show one  standard deviation either side of the mean for test C 0 concentration = 340 y l 1  1  2  (standard deviations for other r means were s of similar magnitude).  87  —i  200  1  1  I  400  600  800  QUANTUM  '  FLUX  1  1  1  1  200  400  600  800  D E N S I T Y (uE m"  2  s\400-700nm)  88  carboxylation i s dependent on photochemical events, mesophyll resistance can be strongly affected by PhAR (e.g. 19, 291).  The contribution  of carboxylation resistance to mesophyll resistance i s very large at low PhAR levels and i s minimum at light saturation.  Also,  Jarvis (146) and Whiteman and Roller(292)have noted that mesophyll resistance calculated as described i n this study i s overestimated  unless the gas exchange determinations are made using  conditions of light saturation and CO^ limitation.  Clearly  (Figs. II-8 and II-9), many of the conditions used i n the present study do not satisfy those c r i t e r i a . The values for mesophyll resistances found i n these experiments (Fig. 11-11) are considerably larger than those reported for some other plant species. resistances between  Gaastra (103) reported mesophyll  2 and 10 s cm  i n turnip leaves, and  1  Bierhuizen and Slatyer (19) found similar values for cotton leaves. The values calculated from the present data are, however, of similar magnitude to those reported by Ku et al. leaves.  (167) for potato  Mesophyll resistances tended to be higher i n basal  leaves (Fig. 11a) than i n apical leaves (Fig. l i b ) on CO^-enriched plants.  These differences were particularly evident at low  (50 yE m~  2  s" ) and high(/50 yE m~ s ) levels of PhAR. In 1  2  _ 1  comparing basal leaves from the two growth regimes, similar mesophyll resistances were obtained at the higher PhAR levels -2 -1 used, but at low levels of PhAR (50 and 150 yE m s ) these  89  resistances were much lower i n leaves from the CO^-enriched regime.  As discussed above, i n d i v i d u a l contributions of the  components of mesophyll resistance to these differences could not be assessed, and i t i s possible that they were due either to induced changes i n l i q u i d phase resistance to CC^ transfer i n the mesophyll, or to changes i n carboxylation  efficiency  dependent upon enzyme a c t i v i t y (RuBP carboxylase) or upon photochemical reactions.  More than one of these factors may,  of  course, be implicated, but the d i f f e r e n t i a l e f f e c t s of low and high irradiance strongly suggest the involvement of changes i n photochemical-dependent  f i x a t i o n reactions.  90  vi)  Behavioral  Indiaies  of Photosynthetic  Responses to CO ^-Enrichment  The confounding of components within mesophyll resistance described above has made i t desirable to model photosynthesis without reference to internal inseparable resistances. The behavioral model of Van Bavel (14) which describes leaf assimilation i n terms of radiant flux density, internal C0  2  concentration and associated u t i l i z a t i o n efficiencies (Ej and E^), i s particularly relevant to the present study. Values of E„, C EI, and FIM calculated from the present T  T>;r  data are summarized i n Table II-5.  E^, was s i g n i f i c a n t l y higher i n  basal leaves of CO^-enriched plants than i n comparable leaves of those grown i n normal a i r . between those treatments.  However, values of E^ did not d i f f e r In the CO -enriched treatment E was 2.  t»  somewhat lower i n apical than in basal leaves but E^. was almost twice as great.  Clearly, the relatively high photosynthesis  rates and F ^ value for these apical leaves was associated with a higher efficiency of PhAR u t i l i z a t i o n .  This i s consistent with the  results of McCree (181) who found that the highest quantum y i e l d (rate of photosynthesis per unit rate of quanta absorption) was obtained for sections of the youngest leaves of corn plants.  In the terms of the  Van Bavel model, E^. i s the maximum possible quantum yield although i n these calculations, this parameter i s based on incident rather than  TABLE I I - 5 :  B e h a v i o r a l i n d i c e s f o r p h o t o s y n t h e t i c C O 2 exchange: C 0 u t i l i z a t i o n e f f i c i e n c y ( E ) , r a d i a n t energy u t i l i z a t i o n e f f i c i e n c y ( E T ) , and maximum p h o t o s y n t h e t i c CO flux ( F ) . 2  c  I M  Growth Regime  Leaf Basal  Air C0 ~Enriched 2  Apical  Air CO - E n r i c h e d  E  r  I _ i _ (g y E i n s t e i n x 10 ) E  5  2.8±0.5* 10.9+0.6  —  7.5±0.5  *Values shown a r e means ± 1 s t a n d a r d d e v i a t i o n .  g m 2^ s  x  > x 10 -  0.27±0.07  0.85±0.20  0.27±0.02  1.00±0.20  —  0.50±0.03  —  1.50±0.21  -  5  92  absorbed quanta.  In view of the low reflectance and trans-  mittance relative to the absorbance of leaves over the waveband 400-700 nm, i t i s doubtful that the difference between incident and absorbed PhAR results i n more than a small, constant error i n values of E^.. The value of E  for basal leaves from the normal a i r  regime i s of similar magnitude to that reported previously for the 12th leaf of a sunflower plant (14). The values obtained for the other leaves i n these experiments are considerably higher than this, suggesting that E^ may be very sensitive to supplemental during growth and leaf age.  Values for E^. and F  are about  an order of magnitude lower than those reported for the sunflower leaf, and this may account for the r e l a t i v e l y low rates of net photosynthesis i n tomato compared with those reported for sunflower (14). Although i t i s clear that E^, and E^ are influenced by growth regime and/or leaf age, the physiological changes underlying these responses are obscure.  Modifications i n leaf structure,  chloroplast photophysiology, and the a c t i v i t i e s of photosynthetic enzymes may be involved, but the relative importance of such modifications i s not evident from the present data.  In studies  of the aging of bean leaves, Fraser and Bidwell (97) observed a repetitive pattern of changes i n photosynthetic a c t i v i t y i n . successive leaves. They suggested that this pattern may be controlled by i n t r i n s i c factors.  The data i n Table 5 seem to  93  indicate that aging i s accompanied by changes i n efficiencies of both radiation and possibly carbon dioxide u t i l i z a t i o n . The v a l i d i t y of the behavioral model (14) used i n these calculations i s supported by the accuracy of prediction of net CC^ assimilation rates from values of E^, E^., and  (Table II-6).  Calculated rates (only those for a test CC^ concentration of 340 pi. .1 have been shown here) were closely correlated with observed rates over the range of CO^ concentrations and light intensities used i n these experiments.  The present study thus demonstrated the  a p p l i c a b i l i t y of the model to an additional plant species grown under various environmental conditions. This research has, therefore, indicated that the increases in growth and yield which result from C02 enrichment may not _  simply be due to an increased concentration gradient driving the diffusion of CO2 from the greenhouse a i r to the chloroplasts. Physiological modifications occurred i n plants grown at high CO2 levels which increased inherjentt rates of C0  2  assimilation and  decreased CO2 compensation points relative to those plants grown i n normal a i r .  Also,the responses to CO2 enrichment during -  growth differed i n leaves of different physiological ages.  It i s  apparent that increased tomato yields following CO2-enrichment are at least p a r t i a l l y due to these changes. The experiments so far performed were designed to provide general descriptive information on the physiology of tomato plants grown under C0 -enriched and normal a i r conditions. No special ?  TABLE II-6:  Comparison of observed rates of net photosynthesis with rates predicted by the model of Van Bavel, for a test CO. concentration of 340 y l 1 . ±  Leaf Basal  Apical  Incident Radiation yE m~2 ~ (400-700 nm)  Observed  Predicted  Air  50 150 250 750  0.42 1.13 1.74 2.04  0.29 1.29 1.91 3.05  co2Enriched  50 150 250 750  0.61 1.70 2.42 3.02  0.62 2.33 3.09 4.88  co2Enriched  50 150 250 750  1.20 2.14 3.45 6.07  1.30 3.20 3.90 6.90  Growth Regime  x  s  Rate of Net Photosynthesis (g/m""2 s - x l 0 ) ]  4  emphasis was p l a c e d on determining observed  u n d e r l y i n g mechanisms f o r the  e f f e c t s of CO^-enrichment on the p h y s i o l o g y o f CO^  exchange.  The f o l l o w i n g s e c t i o n w i l l d e s c r i b e a s e r i e s of  experiments which were d e v i s e d to focus more a t t e n t i o n on mechanisms.  these  96  PART II,  SECTION 2  EXPERIMENTS ON SEEDLINGS GROWN UNDER CONTROLLED ENVIRONMENT CONDITIONS MATERIALS AND METHODS i)  Growth Chambers and Control Systems  An integrated system of 8 plant chambers (Fig. 11-12) was designed to satisfy the specific needs of the laboratory experiments.  The skeleton of the system was as previously described  (234) but the chambers and control systems they used were considerably modified for use i n the present study.  Each chamber  was a 30 x 30 x 46 cm plexiglas box surmounted by a 6 cm deep water bath (Fig. 11-13). a i r and CO^ supply  Separate inlet ports were provided for  to each chamber, and the internally mounted  fan f a c i l i t a t e d mixing of the atmosphere. Different concentrations of CO^ were maintained i n different chambers by supplying compressed a i r from a central source and pure, compressed CO^ at different flow rates. The flow rates needed to achieve a given atmospheric  concentration  varied only s l i g h t l y from chamber to chamber. Where normal a i r concentrations (0.03%) were required the compressed a i r flow rate was maintained at 4.5 1 min ^ and no additional CO2 was added. Gas flow to each chamber was controlled by a pair of rotameter flowmeters incorporating needle values (Matheson tube 600 or 610  96a  FIGURE 11-12:  Diagram of 8-chamber growth system used for Experiments 5, 6, 7, and 8.  FIGURE 11-13:  Diagram of one chamber of the system shown F i g u r e 11-12 ( f o i l internal  detail).  f a c i n g removed to show  98  99  for CC^j and tube 603 for a i r ) .  Each pair was connected to one  chamber by means of 0.6 cm internal diameter ygon plastic tubing. T  Gas outlet ports, near the top of each chamber, were connected to three-way solenoid values which normally exhausted the chamber a i r .  Monitoring of the outlet gas stream from a single  chamber atmosphere was possible by activating the appropriate solenoid valve, thus directing gas flow from the chamber through a manifold, a calcium sulphate drying column,and into the sample c e l l of an IRGA (Beckman Instruments Ltd., Model 864). Illumination of the chambers was by two banks of mixed i  i  fluorescent and incandescent l i g h t s . Each bank (illuminating four chambers) consisted of 20 high output fluorescent tubes (135 W) and 15 incandescent bulbs (40 W).  The lamps were positioned at a  height above the chambers sufficient to provide an irradiance of -2 -1 150 UE m level.  s  (400 - 700 nm) (approximately 12,000 lux) at plant  The radiation was f i l t e r e d through 6 cm of cooled water  i n the chamber bath before i t was incident on the plants.  The  radiant spectral energy distribution did not vary between the chambers, and a representative plot based on data obtained with an ISCO spectroradiometer on several occasions during the four month course of the experiments, i s shown i n Figure 11-14. In these experiments, photoperiod was kept constant at 12 hours light and 12 hours darkness; the lamps were set to switch on and off automatically at 0700 h and 1900 h each day.  The sides  99a  FIGURE 11-14: Spectral energy distribution of mixed fluorescent and incandescent lamps used i n the system shown i n Figure 11-12.  100  o  w u  *x  *x  (N  cn  "x ^  UD S^OM ri A1ISN3Q  * in  x  ro  " I V H I O B d S  x *-  101  and bottom of the chambers were shielded by facing with aluminum f o i l , and, during the dark period, a screen of black cloth moved automatically over the chamber tops to exclude extraneous room light.  A i r temperature was not controlled but was continously  monitored by means of a shielded copper-constantan thermocouple placed i n each chamber. Temperatures were quite consistent between chambers during the light and dark periods, a d i f f e r e n t i a l of 4°C being the maximum observed under any conditions.  Excessive heat  was removed by means of the flowing water bath located on each chamber top (Figures 11-12, 11-13).  Chamber a i r temperatures varied  between 30 and 35°C during the light period whereas during darkness they were consistent at approximately 25°C. Chamber humidity varied between 80 and 90% relative humidity throughout the experiments. Throughout the experiments plants within each chamber werf. watered two times per week. On each occassion the nutrient solution of Epstein (86) was used except that a 1 M stock solution of potassium dihydrogen phosphate was used i n place of ammonium dihydrogen phosphate, and no potassium nitrate was added.  102  ii)  Gas Exchange Measurement System  The system used to measure rates of CO^ and water vapour exchange i n these laboratory studies was similar to the portable system used i n the greenhouse study.  The fixed  location of the apparatus, however, allowed certain improvements to be made to some of the components, which w i l l be described here.  The system was, once again,of an open flow design.  The IRGA  (Beckman Instruments Inc. Model 1^215) was used to establish the appropriate  concentration i n the a i r stream supplied to the  chamber and to measure the CO2 d i f f e r e n t i a l of ingoing and outgoing streams. Throughout measurements of gas exchange at normal (21%) oxygen concentrations, C02~free a i r and 1% CO2 i n a i r were mixed to give a required CO2 concentration as described previously (Part I I , Section 1: Materials and Methods). For measurements at low oxygen concentration, nitrogen and 3% CO2 i n nitrogen were similarly mixed, and the composition of the resulting gas was checked by means of the IRGA and a Servo.mex model OA. 184 oxygen analyzer.  Chamber i n l e t a i r was humidified to between 65  and 75% relative humidity as measured by the EG'&G Ltd. Model 880 e l e c t r i c hygrometer, and flow rate was monitored by means of a Datametrics model 800 VTP mass flow meter and sensor.  Throughout  a l l gas exchange measurements, gas flow was maintained at a s u f f i c i e n t l y high velocity to l i m i t the C0 d i f f e r e n t i a l across o  the chamber to < 40 y l 1 . x  For most measurements, this was  achieved by maintaining gas flow at 800 ml min . x  Details of the assimilation chamber used i n these studies are shown i n Figure 11-15. The fan mounted i n the bottom of the chamber maintained conditions of turbulence i n the a i r about the leaf.  A i r .and leaf temperatures were measured using  0.2 mm diameter copper-constantan thermocouples placed near the a i r i n l e t ports and against the abaxial leaf surface, respectively. Light was provided to the chamber by two 300 W cool beam lamps (General Electric Ltd.) vand was f i l t e r e d through 5 cm of water before i t was incident on the chamber. Irradiance was varied by means of cheesecloth f i l t e r s and by varying the number of lamps used.  PhAR was measured as described previously (Part I I , Section 1  Materials and Methods).  The spectral energy distribution of  the lamps was similar to that shown for the 300 W lamp i n Figure I I During a l l experiments leaf temperature was maintained between 30 and 33°C by circulating water through the 2 cm deep bath mounted on the top plate of the chamber.  FIGURE 11-15:  Diagram of assimilation chamber used i n Experiments 6 and 7.  104  SIDE  VIEW  105  iii)  Experimental  Design  i i  The laboratory study consisted of four experiments. The f i r s t to be described here (Experiment 5), occupied approximately 6 weeks commencing i n early A p r i l 1978.  I t was  designed to provide information on vegetative growth and development of plants exposed to different CO2 concentrations during growth. Seeds of Lycopersicon  esculentum L. (cv. Vendor) were  germinated as described i n Part I I , Section 1: Materials and Methods^at three day intervals and allowed to develop under a 12 hour photoperiod (14000 lux;180 yE m~ s 2  _ 1  (400 - 700 nm)) for  14 days from sowing on a greenhouse bench. Daytime and nightime temperatures were maintained at 25°C and 18°C respectively. At the end of the 14 day period, s i x uniformly developed seedlings were transplanted into 10 cm square pots (one seedling per pot) which were transferred to one of the plexiglas growth chambers. The CO^ concentration within the chamber was maintained at 0.03% as described previously. At three day intervals oth°r seedlings were similarly transplanted and transferred to other chambers supplied with a i r containing 0.1 and 0.5% C0 , respectively. Other 2  growth conditions were maintained as described above. A total of s i x chambers were used i n this study, providing two replicate locations for each CO treatment (0.03, 0.1 and 0.5%). Before placement i n the chambers, three of the six plants were randomly selected for subsequent harvest.  The others were  106  l a b e l l e d for i d e n t i f i c a t i o n , and the lengths of a l l unrolled leaves on these plants were measured.  The leaf measurements were  then repeated at three day intervals for the next three weeks. The mean developmental stage of plants at each measurement date i n the three growth regimes, was calculated in terms of Plastochrons from the leaf length data (Part I I , Section 1: Materials and Methods).  At the end of the f i r s t week i n the growth  regimes (three weeks from germination) the previously selected plants were harvested to determine leaf area and t o t a l plant and leaf dry weight.  Three weeks l a t e r , the rest of the plants were  s i m i l a r l y harvested.  The data obtained were used to calculate  NAR and Leaf Area Ratio.(LAR) of plants i n each chamber.  RGR,  The growth  analysis formulae given by Kvet et al. (169) were used i n each case.  Single factor ANOVA was used i n conjunction with a  two-stage SNK test (a = 0.05)  to distinguish between the mean values  f o r growth analysis parameters i n each growth regime. Experiment 6 occupied a period of three months from January to March 1978 and was designed to study effects of CO^ concentration during growth on subsequent CO2 exchange response to irradiance and.CO2 concentration i n atmospheres containing normal (21%) and low (0 - 1%) oxygen.  Tomato plants were raised from  seed as described for Experiment 5. chamber at three day i n t e r v a l s . were the same as i n Experiment 5.  Six plants were placed in each  CO^ concentrations and growth conditions The course of plant development  was  107  followed by monitoring Plastochron Index (PI). When a majority of plants within any one chamber had attained a PI value of 5, one of the majority was randomly selected for use i n measurements of leaf gas exchange. Upon completion of the measurements the plant was returned to the same chamber to permit continued development under the same growth conditions as before.  A second  series of measurements were carried out on the same leaf when the plant had attained a PI value of 10.5.. These gas exchange measurements were carried out on leaf 3 (counting from the f i r s t true leaf above the cotyledons).  Each  measurement series was begun by measuring photosynthesis rate at 500 -2 - l s or520 yE m s (400 — 700 nm, irradiance under normal atmospheric conditions, and this was followed by measurements at -2 -1 4 lower irradiances (^00, 200, 130, 32 yE m s ) and complete darkness. Leaf irradiance was then readjusted to a level -2 -1 sufficient to saturate photosynthesis  (520 yE m  s  i n each case),  prior to a series of measurements at 5 different CO^ concentrations (540, 440, 340, 240, 140 u l 1 ) . _ 1  After each change of  irradiance and/or CO 2 concentration, the leaf was allowed to adjust to the new chamber conditions for at least 30 minutes. Measurements under conditions of saturating irradiance and the same five CO2 concentrations were subsequently repeated i n atmospheres containing 0 - 1% 0^.  The CO 2 compensation point for the leaf  was then determined using a simple closed gas exchange system as  108  described i n Part I I , Section 1: the; leaf was Area was  Materials and Methods.  Finally  removed from the chamber and c a r e f u l l y photocopied.  determined from the leaf outline using a photoelectric  planimeter  (Hayashi Denko Co. Ltd., Japan). In these and the subsequent replacement experiments,  rates of gas exchange were again calculated from gas flow rates and measured CO2  and water vapour concentration differences between  the leaf chamber ingoing and outgoing gas streams. space CO2  concentrations,  Intercellular  and boundary layer and stomatal  resistances were also calculated as described i n the greenhouse study.  Mesophyll resistance to C0^ uptake was  calculated from  the r e c i p r o c a l of the regression slope r e l a t i n g the i n t e r c e l l u l a r space C0  2  concentration  (C^) plus CO2  (P) to rate of net photosynthesis  compensation  concentration  (F), so that estimates of  photorespiration rate (R ) could be obtained by extrapolating JLi  each regression to zero F (246). F to test CO2  From the data obtained  concentration at 21% and 0 - 1% O2,  i n h i b i t i o n of photosynthesis the following formula (22, (Ps = tot where PSQ represents Ps„, represents  (% I  ) • In a i r was  relating  the percentage calculated from  283) - Ps  ^  ) —  " 100  PSQ  the rate of net photosynthesis  the rate i n 21% 0 .  i n 0 - 1% O2  and  Experiment 6 had a randomized block design i n which a complete series of gas exchange measurements on plants from each growth regime constituted a single block.  Each series was  replicated three times to provide three complete blocks. The experimental design i s summarized i n Table II-7. analysis of the calculated values of  Statistical  and r ^ was performed by  means of covariance analysis i n combination with a Student-NewmanKeuls multiple range test (a = 0.05) (303).  Tests were applied  to the regression relationships relating C\ to F. Experiment 7 (Replacement Experiment) was designed to investigate whether permanent changes occurred i n the gas exchange physiology of leaf 3 after a period of growth under supra-normal CO^ concentrations.  Plants were grown, as described above, for  approximately two weeks under 0.03% or 0.1% CO,,. At PI = 5, two plants were removed from each growth regime. Measurements df CT^ and watp.r vapour exchange at 5 Ct^ concentrations (540, 440, 340, 240 and 140 y l 1 ) at saturating irradiance were conducted on x  leaf 3. The plants were removed to a greenhouse bench and l e f t for a two week period under conditions of light and temperature similar to those used for early propagation of the seedlings. Then, f i n a l l y , the plants were returned to the laboratory and the same series of measurements was repeated on leaf 3. Experiment 8 .'investigated a c t i v i t i e s of the enzymes RuBP-case and GaO i n leaves 2, 3, 4, and 5 of plants grown at  TABLE I I - 7 : Experiment 6 - C O 2 exchange of leaf 3 of tomato plants grown at 3 C O 2 concentrations i n chambers, at 2 stages of development: Summary of experimental design.  Number of Levels  Factor  Definition  Blocks (B)  3  Replicate measurements: 1 block representing 1 complete set of the other factors, randomized i n space (chambers) and staggered i n time  Growth CC^ concent r a t i o n (C0 )  3  0.03,  B x C0  9  Error term  Plastochron index (PI)  2  PI = 5; PI = 10.5  PI x C0  6  Interaction of leaf age (defined by PI), and growth C0„ concentration  0.1, 0.5% C0  2  2  2  o  Residual error  Error term  Ill  0.03, 0.1 and 0.5% C^.  500 mg of leaf material was cut into  pieces to exclude the petiole and major veins.  Homogenization  was carried out using a mortar and pestle at 4°C i n 5 ml of a medium containing 0.5 mM potassium phosphate buffer (pH 7.3), 0.2 yM EDTA and 1 mg solid sodium glycolate.  The homogenate was  decanted and centrifuged at 12,000 g for 15 minutes. GaO a c t i v i t y was determined spectrophotometrically i n the unpurified extract by the rate of phenylhydrazine oxidation to phenylhydra'zone, using the reaction mixture and procedures described by Baker and Tolber.t (10). A Gilford 2400S spectrophotometer was used i n a l l determinations. The assay procedure for RuBP-case was similar to that of Wishnick and Lane (294). Enzyme a c t i v i t y was determined from 14 the amount of  C radioactivity fixed i n 0.1 ml aliquots of the  unpurified leaf extract reaction mixture incubated for 10 minutes i n a water bath at 39°C. Each 0.1 ml aliquot was dried i n a forced a i r draft i n a fume hood for 60 minutes and then resolubilized i n 10 ml of aquasol (Nuclear Chicago Ltd.) before counting on a Nuclear Chicago Mark 1 l i q u i d s c i n t i l l a t i o n counter. The reaction mixtures for each enzyme assay are summarized i n Appendix 3" at the end of this thesis. Before commencing Experiment 8,trials were conducted with each assay procedure to determine the pH optimum and saturating substrate concentration for GaO and RuBP-case.  The  112  details of these preliminary experiments are also presented i n Appendix 3. Final pH of reaction mixtures i n experiment 8 were adjusted to 8.3 and 7.8 for GaO and RuBP-case, respectively. A l l pH adjustments were carried out by dropwise addition of 1 M Tris-HCl buffer. A c t i v i t y assays were replicated three times using plants grown i n different chambers. With the exception of the f i n a l replicate, plants for assay were selected from those which were not used for gas exchange determinations, two plants being harvested at PI = 5 and another pair at PI = 10. Thus the enzyme assays were conducted, for the most part, at the same time as the gas exchange, determinations on plants from each growth regime. f i n a l replication was done without concurrent measurements of gas exchange but otherwise growth conditions were identical to those.described above.  The  113  PART II,  SECTION 2  RESULTS AND DISCUSSION i)  Analysis  of Plant Growth jv> Relation  to CO„ Concentration  (Experiment  The effects of greenhouse CO ^- enrichment on tomato leaf development i n terms of plastochron index have been reviewed i n Part I I , Section 1.  Analysis of the results of the present  experiment confirmed the earlier finding that supplemental CO^ had l i t t l e effect on this aspect of vegetative development:. A close similarity between the i n i t i a l slopes of the lines i n Figure 11-16 indicated that plastochron index increased linearly with time u n t i l a value of between 8 and 9 was attained. After this point there was some indication of a decreasing rate of development i n plants grown i n 0.1 and 0.5%, but not i n 0.03% C0 . 9  Rogan and  Smith (240) noted a transition to a lower rate of leaf i n i t i a t i o n and emergence between the appearance of leaves 6 and 7 i n Agropyron: repens, which seems to correspond reasonably well with the present data.  However, i n these experiments the trend at any  CO2 concentration was not marked and may be attributable to measurement v a r i a b i l i t y . Figures T.I-17 to 11-19 show the development of leaf length i n the f i r s t five leaves of tomato seedlings exposed t<~> 0.03, 0.1 and 0.5% C0 , i n relation to Leaf Plastochron Index (LPI). ?  At each CO2 concentration, the rate of length increase i n the early exponential growth phase appeared to be similar irrespective of leaf number, although measurements were not available for leaves 1 and 2 at negative values of LPI.  This confirms the results of  5)  FIGURE 11-16: Plastochron development of tomato plants grown at 3 CG^ concentrations as a function of time. NOTE: Points are means of measurements on a t o t a l of 6 plants, i n two chambers shown f one standard deviation.  10H  10  11  -1  1  12  13  1  1  15  16  T  U  DAYS FROM I N I T I A L M E A S U R E M E N T tdayO is arbitrary for each growth regime)  f 17  114a  FIGURE 11-17:  Growth i n length of successive leaves of tomato plants grown at 0.03% CO^ as a function of LEI. NOTE:  Points are means of measurements on a t o t a l of s i x plants i n two chambers.  LEAF  PLASTOCHRON  INDEX  115a  FIGURE 11-18:  Growth i n l e n g t h of s u c c e s s i v e l e a v e s of tomato p l a n t s grown at 0.1% C 0 NOTE:  2  as a f u n c t i o n of L P I .  P o i n t s as f o r F i g u r e  11-17.  LEAF  PLASTOCHRON  INDEX  FIGURE 11-19:  Growth i n l e n g t h o f s u c c e s s i v e l e a v e s of tomato p l a n t s grown a t 0.5% NOTE:  as a f u n c t i o n of L P I .  P o i n t s as f o r F i g u r e  11-17.  117  250-1  230H  LEAF  PLASTOCHRON  INDEX  118  Coleman and Greyson (64) who noted, i n addition, that the LPI at which a single tomato leaf l e f t the exponential growth phase depended on i t s s e r i a l number on the main shoot of the plant.  This  observation also corresponded with the present data where leaves 1 and 2 had attained maximum length at an LPI of between 3 and 4 in contrast to higher leaves which showed exponential growth u n t i l they had attained an LPI of between 4 and 6. These transitions from phase 1 (exponential) growth to phase 2 (mature) growth (64) were apparently independent of CO^ concentration although the mean lengths of individual leaves at the transition point were CO^-dependent.  In general, leaves which developed at a growth  concentration of 0-5% were shorter at the growth curve asymptote than those developed at 0.03%. This difference was p a r t i c u l a r l y marked above leaf 2. At the start of phase 2, the lengths of leaves 1 and 2 which were developed i n 0.1% CO^ were greater than at the other two concentrations.  This difference was not evident  between the 0.03 and 0.1% CO^~grown plants at higher leaf numbers. The lack of effect of either 0.1% or 0.5% CT^ during growth on leaf production i s consistent with results from the previous greenhouse study (Part I I , Section 1).  I t i s , nevertheless,  interesting to note that lengthening of leaves 1 and 2 was respectively inhibited and promoted i n plants grown at 0.5% and 0.1% CX^. To my knowledge there are no previous reports on the effect of atmospheric CO,, concentrations on the development of leaf length i n tomato or other plants.  119  An  i n t e r e s t i n g comparison may  o b s e r v a t i o n s and  the v a l u e s of LAR  t h r e e growth CO^ c o n c e n t r a t i o n s decreased  by 0.1  and  0.5%  C0  (Table I I - 8 ) .  LAR  was  r e l a t i v e to the 0.03%  2  these  f o r p l a n t s exposed to  i n accordance w i t h s e v e r a l p r e v i o u s The  be made between  the  significantly  treatment  s t u d i e s (95, 136,  140,  273).  d i f f e r e n c e s i n l e a f l e n g t h at the s t a r t of phase 2 growth  were not r e f l e c t e d i n d i f f e r e n t v a l u e s f o r LAR grown i n 0.1  and  0.5%  CO^.  T h i s may  suggest  between p l a n t s  t h a t h i g h growth  c o n c e n t r a t i o n s e x e r t morphogenetic i n f l u e n c e s on r e d u c t i o n a t 0.5%  C0  2  and  i n c r e a s e at 0-1%)  the l e a v e s  which a r e  C0  2  (length  compensated  by d i f f e r e n t i a l p r o d u c t i o n and/or p a r t i t i o n i n g of dry matter w i t h i n the p l a n t .  In a l a t e r s e c t i o n ( P a r t I I I ) d a t a w i l l be  which i n d i c a t e t h a t h i g h C 0 o v e r a l l morphological  2  concentrations g r e a t l y i n f l u e n c e  development i n  Compared w i t h 0.03% grown a t 0.1%  C0  Photosynthetic 0.1% was  and  2 >  but at 0.5%  C0 ,  RGR  2  C0  Pharbitis  RGR  2 >  at 0.03%  C0 . 2  NAR  sufficient  LAR  RGR ambient C0  i n the 0.5%  to i n c r e a s e RGR  the reduced  o  was  not  was  values.  2  II-8).  highest at  I t may  be  COp-grown p l a n t s was  over p l a n t s grown i n 0.03%  a t the h i g h e r C 0  and NAR  changed ( T a b l e  of p l a n t s grown under 0.5%  i n t e r m e d i a t e between the othe*" two  that the i n c r e a s e i n NAR  nil.  were i n c r e a s e d i n p l a n t s  e f f i c i e n c y , as i n d i c a t e d by NAR,  lowest  reviewed  C0 , 2  C0  2  noted not due  to  concentration.  have p r e v i o u s l y been shown to i n c r e a s e w i t h  c o n c e n t r a t i o n i n a number nf s p e c i e s (25, 139,  140,  273).  120  TABLE I I - 8 :  E f f e c t s of CO2 concentration on net a s s i m i l a t i o n rate (NAR), r e l a t i v e growth rate (RGR) and l e a f area r a t i o (LAR) i n tomato plants grown at 3 C0^ concentrations, i n chambers.  Growth CO2 Concentration (%)  NAR (mg cm day"-*-) -2  -  RGR (mg g ~ l day ^) -  LAR* cm mg~ 2  0.03  0.40a**  229.40a  0.55a  0.10  0.65b  234.90b  0.35b  0.50  0.59c  215.60a  0.36b  *LAR determined 22 days a f t e r placing plants i n growth regimes. *Mean values i n any column designated with the same l e t t e r are not s i g n i f i c a n t l y d i f f e r e n t (P > 0.05) according to SNK m u l t i p l e range t e s t .  121  Basing their conclusions on detailed mathematical  analysis,  Thornley and Hurd (271) have suggested that the main e f f e c t of CO^" enrichment RGR  i n tomatoes i s upon NAR,  with other changes i n  and LAR r e s u l t i n g d i r e c t l y from these e f f e c t s .  The present  data are not e n t i r e l y inconsistent with this view although they are more e a s i l y explained by direct effects of CO^ on NAR which thus determine RGR.  and  LAR  Support for this explanation i s provided  by the data of Monsi et al. (202) concerning the e f f e c t of another environmental factor, l i g h t intensity, upon these three growth parameters.  Monsi at al. (202) showed that, i n mung bean, a  reduction i n l i g h t intensity decreased NAR but concurrently increased LAR so that RGR  remained constant.  It i s important to emphasize that the increase i n NAR  i n the 0.5% CO2 grown plants was not s u f f i c i e n t to promote the  rate of dry matter production. gain by the plant.  NAR  represents net^photosynthetic  Differences in NAR between plants may be a  r e s u l t of differences i n rates of r e s p i r a t i o n or photosynthesis (290) .  The contribution of changes i n either of these processes  to the reduction i n NAR  i n tomato plants grown at 0.5%  CO2  r e l a t i v e to those grown at 0.1% CO2 i s unclear from the present data.  The results described later i n this Section w i l l serve  partly to c l a r i f y the relationships between NAR concentration.  and growth CO2  122  Effects  of  Growth  Photorespiration  C0„ and  Concentration Enzyme  on  Activity  in  Photosynthesis„ Leaf  3 at  LPI  : 2.0  (Experiment  In Part I I , Section 1 of this thesis i t has been shown that tomato plants grown at ambient CO^ concentrations of approximately  900 y l 1  x  (0.09%) exhibit higher rates of net  photosynthesis and higher e f f i c i e n c i e s of CO^ u t i l i z a t i o n than those grown i n normal a i r . In order to determine i f these effects were evident at higher concentrations experiments were conducted to investigate the effects of 0.5,  0.1,  and 0.03%  during  growth on subsequent rates of photosynthesis i n r e l a t i o n to irradiance and  concentration.  At LPI :2 net rates of photosynthesis were highest i n 3  -2 plants grown at 0.1%  CO2  at irradiances of 130 yE m  above, when test CO^ concentration was  -1 s  and  330 y l 1 ^ (Figure 11-20) .  This confirmed e a r l i e r observations and indicated a s i m i l a r response of young leaves to growth CO^ concentration under greenhouse conditions and i n small controlled environment chambers. growth CO2,  however, a different response pattern was -2 -1  At irradiances above 200 yE m  s  , and a test  At  0.5%  observed.  concentration  of 330 y l 1 , net photosynthetic rates were similar to those of x  leaf 3 on plants grown at 0.03% rates i n both 0.03  and 0.1%  CC^'  CO^ grown plants were s i g n i f i c a n t l y  higher than i n those from the 0.5% the  At lower irradiances, leaf  CO2  a p p l i c a t i o n of supplementary C0  9  regime.  It was  clear that  during growth at concentrations  6&8)  122a  FIGURE 11-20: Apparent rates of photosynthesis i n relation to quantum flux density in leaf 3 of tomato plants grown at 3 Test  concentrations.  concentration:  Leaf temperature: LPI:  330 y l 1  x  30 ± 2°C  2 ± 0.5  NOTE: Points bearing the same l e t t e r at any irradiance are not significantly different according to SNK multiple range test (P > 0.05).  124  5 times the normal commercial enrichment level of 1000 y l 1 (0.1%) resulted i n depressed leaf photosynthetic rates at low irradiances when plants were returned to a normal (0.03%) CO^ atmosphere. This effect partly resembles that described for cucumber seedlings returned to 320 y l 1 enriched conditions (102).  CO^ after growth under CO^-  In that case, the photosynthetic  rates i n three newly emerged leaves were lower when plants grown at 1500 y l 1 ^ CO^ were measured at 320 y l 1 ^ at an irradiance -2 of 98 W m  . Yet, there are many differences between the present  results and those obtained i n that study.  Notably, the  discrepancies i n photosynthetic rates i n cucumber plants were not evident at low irradiances, and the growth CO^ concentration  of  1500 y l 1 ^ was more similar to the 0.1% treatment used i n the present study.  I t i s not possible to resolve these differences  at present, but the lack of uniformity between the two studies, i n terms of growth conditions and plant type,may be important. The results showing net photosynthetic rate differences at low irradiances (Figure 11-20) may be due to differences i n rates of dark respiration.  In plants grown at 0.5% CO^, dark respiratory  rates were higher than i n those grown at either 0.1 or 0.03% CO^. Furthermore, compensation point for irradiance (estimated from the intercept of the curve relating CO^ exchange to irradiance, with the abscissa) was much higher i n plants from the 0.5% C0 regime ?  125  than i n those grown at 0.3 or 0.1%  CO^-  The reasons for the  higher rates of dark respiration i n the 0.5% are not clear from the present  C0 -grown plants 2  data.  Madsen (184) has noted that the starch content of plants grown at CO^ concentrations of 0.22  to 0.5%  may  be as high as  13% of t o t a l plant dry matter as compared with 2 to 6% i n plants grown i n normal a i r or 0.1%  CO^.  It seems possible that such large  accumulations of starch may  result i n an increase i n mitochondrial  r e s p i r a t i o n through increased a v a i l a b i l i t y of respiratory substrates (266).  Similar increases i n dark r e s p i r a t i o n rates of  young tomato leaves grown i n C0 - enriched a i r have been described 2  by Ito (143).  I t has often been observed that dark r e s p i r a t i o n  i n plant leaves continues under conditions of low irradiance (126, 132) although  the rate of  production i s i n h i b i t e d  apparently as the result of d i r e c t i n h i b i t i o n of breakdown (132).  (193),  carbohydrate  Inhibition of dark r e s p i r a t i o n increases with  increasing l i g h t intensity, and, i n plants u t i l i z i n g only the pathway of photosynthesis,is replaced by peroxisomal photorespiration -2 (144).  At irradiances below 200 yE m  -1 s  i n the present  experiments, i t seems l i k e l y that net photosynthetic rate leaves of plants grown at 0.5% grown at 0.03  or 0.1%  C0  2  i s reduced r e l a t i v e to those  because of greater production of  by mitochondrial r e s p i r a t i o n . for the observed e f f e c t s .  CO^  in  C0  2  There are a l t e r n a t i v e explanations  It might be suggested, for example, that  126  higher quantum yields i n the 0.03 and 0.1% CO^- grown plants could cause the r e l a t i v e l y greater photosynthetic rates i n those cases.  However, the close similarity between the i n i t i a l  slopes of the irradiance response curves i n Figure 11-20  tends  to refute this hypothesis. Another possible source of the differences i n photosynthetic rates i s that stomatal resistances to CO^ transfer were inherently higher at low irradiances i n the 0.5% CO^-grown plants, but this i s not borne out by the data shown i n Figure 11-21. V a r i a b i l i t y i n stomatal resistance within plants from any one growth regime was high at each test irradiance. Resistance values for leaf 3 of plants grown at 0.5% CO^ were quantitatively lower than for those from the 0.03% regime at low irradiance, but analysis revealed no s t a t i s t i c a l differences between growth regimes. I t may be concluded that the particular CO^- enrichment regime under which tomato plants were grown did not influence stomatal resistance. The lack of stomatal acclimatization to increased CX^  concentration  during growth has been well demonstrated for lettuce and Xanthium (152).  I t has also been shown with two lettuce varieties,  that instantaneous rates of photosynthesis may be depressed at CO^ concentrations of 1700 y l 1 \ stomatal closure (238).  an effect which i s unrelated to  The present data suggest that variations  in photosynthetic rates i n plants grown under different CO concentrations are not related to differences in stomatal resistance.  126a  FIGURE 11-21:  Stomatal resistance to CO^ transfer i n relation to quantum flux density i n leaf 3 of tomato plants grown at 3 CO^ concentrations. Test C0 concentration: 2  Leaf temperature: LPI:  330 y l 1  _ 1  30 ± 2°C  2 ± 0.5  NOTE: Mean resistances at each irradiance are not significantly different (P > 0.05) according to SNK multiple range test.  20  n  18-  e  16  o  tu H o ?  12-  £  10-  tI/)  0 03% C 0  •0-5%CO, 8  _J  < o < r -  growth regime  0 1% CO,  LU  oc  2  Line:  y=10 52-1 58 ln(x)  [all  points]  6  CO  2  -r  100  200 QUANTUM  300 FLUX  ^00  D E N S I T Y (uE irf s"!400-700nm) 2  500  128  Some results discussed i n Part I I , Section 1 suggested that rates of photorespiration and magnitudes of mesophyll resistance at low irradiances were reduced i n leaves of tomato plants grown at approximately 900 y l 1 CO . In order to assess 1  the importance of these factors, two tests involving the effects of  and 0^ concentration on photosynthesis at saturating  irradiance were undertaken. Figure 11-22 i l l u s t r a t e s the relationship between CO^ concentration i n the intercellular spaces within the leaf and apparent rate of photosynthesis. at each value of  The enhancement of photosynthesis  i n leaf 3 of plants grown at 0.1% CX^ i s  clearly shown by these data.  The relationship also provides some  insight to the mechanisms responsible for this enhancement, and also the lack of effect of higher CO^ concentrations (0.5%) on inherent rates of net photosynthesis. The steep slope of the regression l i n e for the 0.1% CO^grown plants i s reflected i n a lower value for mesophyll resistance as compared to leaves of plants f rom the 0.03 or 0.5% CO^ growth regimes (Figure 11-23).  In addition, values of photorespiration  were s i g n i f i c a n t l y lower i n leaf 3 developed under 0.1% C0 . I t 2  was apparent that both factors played important roles i n determining the maximum photosynthetic rate i n the leaf at this developmental stage.  128a  FIGURE 11-22: Apparent rates of photosynthesis  i n relation to  i n t e r c e l l u l a r CO,, concentration i n leaf 3 of tomato plants grown at 3 CO quantum flux density: Leaf temperature: LPI:  concentrations. -2 -1  520 yE m  s  , 400 - 700 nm  30 ± 2°C  2 ± 0.5  NOTE: Extrapolation to zero CO^ concentration to estimate rate of photorespiration.  129a  FIGURE 11-23:  Rates o f p h o t o r e s p i r a t i o n ( R ^ , and m e s o p h y l l resistance  ( r ) f o r l e a f 3 o f tomato p l a n t s m  grown a t 3 CO^ c o n c e n t r a t i o n s  and measured a t  L P I : 2 ± 0.5. NOTE:  Values o f L o r r r e p r e s e n t e d L m bearing  by b a r s  the same l e t t e r a r e not s i g n i f i c a n t l y  different  (P > 0.05) a c c o r d i n g  m u l t i p l e range t e s t .  t o SNK  Rates o f R_ and r L m  d e r i v e d from r e g r e s s i o n s r e l a t i n g photosynthesis  apparent  to i n t e r c e l l u l a r CO2  c o n c e n t r a t i o n , as d e s c r i b e d i n t h e t e x t .  130  131  The rates of photorespiration i n leaf 3 of plants grown at 0.03, 0.1, and 0.5% CO^ correspond closely with the relative magnitudes of the observed CO^ compensation points (Table II-9) . Reduction i n photorespiratory a c t i v i t y can result i n substantial enhancement of photosynthesis i n plant species u t i l i z i n g only the photosynthetic system (62, 170).  The p o s s i b i l i t y of effectively  decreasing photorespiration by supplying additional CX^ to the atmosphere during plant growth has been discussed (110). 14 exposure to 0.1% C0  9  has been shown to reduce  Short-term  C incorporation  into the photorespiratory intermediates serine and glycine i n tomato leaves (172) but not i n leaves of Atriplex (258).  (224) or tobacco  U n t i l now the relationship between CO^-enrichment and  photorespiration has not, however, been well understood.  The  present results clearly indicate that after a period of growth at 0.1% CO^, photorespiration i s inhibited relative to that of normal a i r grown plants. Yet, an increase of growth CX^ concentration to 0.5% resulted i n similar photorespiratory rates to those observed i n leaves developed in normal a i r . There i s considerable evidence that the substrate for photorespiration i n  plants i s glycolate or a related metabolite  (79, 81, 144, 276, 308).  An increase i n ambient  concentration  from 0.03 to 0.1% has been shown to result i n a marked reduction in glycolate synthesis i n isolated chloroplasts (83, 239), tobacco leaf discs (310), and i n algae (35). A possible explanation for the  132  TABLE II-9: C0 compensation points of leaf 3 of tomato plants grown at 3 CO2 concentrations, measured at two stages of development (Leaf Plastochron Index, LPI). 2  Growth C0 Concentration (%) 2  CO2 Compensation Point at LPl3:2.0±0.5 (g m~ (ul l ) ) 3  - 1  CO2 Compensation Point at LPl3:7 ,5±0.5 (g m (ul I ) ) -3  - 1  0.03  0.095 (54)a*  0.069 (39)a  0.10  0.065 (37)b  0.076 (43)a  0.50  0.090 (51)a  0.083 (47)a  ^Values are means of 6 determinations per treatment. Values for each LPI designated with the same letter are not s i g n i f i c a n t l y different (P > 0.05) by ANOVA and SNK test.  133  results reported here i s that, i n plants grown under 0.1% the biochemical  CO^  capacity for glycolate production i s decreased  r e l a t i v e to those grown under 0.03% a v a i l a b i l i t y of photorespiratory to an i n h i b i t i o n of the o v e r a l l  .  In t h i s case the  substrate would be reduced leading process.  A r e l a t i v e l y large number of mechanisms have been proposed for the synthesis of glycolate i n green leaves (9, 37, 65, 276,  308).  these viz.  There i s currently f a i r l y wide support for two  the oxidation of the transketoluse-glycoaldehyde  of  complex  by hydrogen peroxide (produced by reoxidation of a photosynthetically produced reductant  - the Mehler reaction) (65, 249)  and the d i r e c t  oxidation of RuBP to phosphoglycolate i n a reaction catalyzed by RuBP carboxylase/oxygenase (37, 176,  276).  Both mechanisms may  operate i n some c e l l s as i s suggested by some recent work on the alga Hydrodiatyon of  serious  afvioanwn  (107, 108).  There are, however, a number  c r i t i c i s m s of the Mehler reaction pathway proposed by  Coombs and Whittingham (65) and i t seems that, i n most c e l l s , the r e l a t i v e a c t i v i t i e s of RuBP-carboxylase and oxygenase determine the rate of glycolate production and hence the rate of photorespiration.  Later i n this section i t w i l l be shown that,  i n plants grown i n 0.1%  CO2,  the carboxylation a c t i v i t y of t h i s  enzyme i s s i g n i f i c a n t l y increased r e l a t i v e to normal air-grown plants, thus supporting  the hypothesis of a reduced capacity f o r  glycolate production after growth at this  concentration.  134  I t i s p o s s i b l e that an i n c r e a s e of CO above a c e r t a i n o p t i m a l photorespiratory higher  l e v e l brings  concentration  about f u r t h e r changes i n  mechanisms, which may  ultimately result  in  r a t e s than e x h i b i f e d by p l a n t s grown i n normal a i r .  i s suggested from experiments on Hydrangea i n which i t observed t h a t an i n c r e a s e  i n ambient CO^  r e s u l t e d i n 25% more r e s p i r a t o r y CC^ compared to p l a n t s t e s t e d i n 0.03% important to p o i n t out  This  was to  1%  r e l e a s e i n the l i g h t  as  CO^  concentration  (216).  I t i s , however,  t h a t those experiments were concerned  e n t i r e l y w i t h i n s t a n t a n e o u s r a t e s of p h o t o r e s p i r a t i o n  and  w i t h the long  during  growth.  The  of l e a f CO^  term e f f e c t s of h i g h  concentrations  not plant  r e s u l t s p r e s e n t e d here have shown t h a t c h a r a c t e r i s t i c s exchange are determined, not  gaseous environment, but  a l s o by CO^  o n l y by  conditions  the  current  during  a  previous  p e r i o d of growth. That the growth c o n d i t i o n s a f f e c t e d o t h e r physiology  i s evident  from the lower v a l u e s  i n the young l e a f 3 o f p l a n t s grown at 0.1% I t i s worth r e c a l l i n g  two  of mesophyll r e s i s t a n c e (Figure  11-23).  s e p a r a t e components, one r e s u l t i n g  from the p h y s i c a l r e s i s t a n c e to CO2 through the m e s o p h y l l , and biochemical  leaf  t h a t mesophyll r e s i s t a n c e as c a l c u l a t e d  i n t h i s study i n c o r p o r a t e s  and  a s p e c t s of  t r a n s f e r i n the l i q u i d  another a t t r i b u t a b l e to the  r e s i s t a n c e s to CO2  I t i s not p o s s i b l e , a t p r e s e n t , t o  phase  photochemical  f i x a t i o n at the c h l o r o p l a s t s .  c o m p l e t e l y s e p a r a t e these  135  components since even under conditions of saturating irradiance, the slope of the line relating internal CO^ concentration to net photosynthesis w i l l depend on both physical transport processes across the mesophyll and the rate of dark carboxylation reactions (146).  Hesketh (125) has stated that inherent differences i n net  photosynthetic rates between species are p r i n c i p a l l y due to variations i n CO^ diffusion resistance across the mesophyll and the kinetics of the dark reactions of photosynthesis.  I t i s possible  that both factors play a role i n determining the magnitude of r ^ i n plants grown i n 0.03 and 0.1% CO^.  Other data to be presented  shortly, however, have provided strong support for the idea that differences i n the carboxylation efficiency of RuBP largely account for the observed differences i n r . m In leaf 3 at  L?Iy2,  there were certain close s i m i l a r i t i e s  between the effects of CO^~enrichment during growth on photorespiration and mesophyll resistance.  Just as photorespiration was suppressed  by growth at 0.1% but not at 0.5% CO2, mesophyll resistance was significantly decreased at 0.1% but, again, not at 0.5% (Figure 11-23).  CO2  The similar effects of supra-normal CO2  concentrations suggested a close correlation between these parameters which warranted careful examination. Figure 11-24 21% O2.  shows the percent i n h i b i t i o n of photosynthesis  Inhibition was highest at low test concentrations of CO^  (140 y l 1 ) i n plants from each growth regime, a finding which x  by  135a  FIGURE 11-24: Effect of CO^ concentration on the percentage t o t a l oxygen inhibition of photosynthesis i n leaf 3 of tomato plants grown at 3 CO^ concentrations. -2 quantum flux density: Leaf temperature: LPI:  2 ± 0.5  520 yE m  30 ± 2°C  -1 s  , 400 - 700 nm  136  137  corresponds with observations on oxygen i n h i b i t i o n of photosynthesis i n wheat (179).  (151), soybean (96, 170), and sunflower  Total oxygen inhibition of photosynthesis i n a i r among  plants appears to be of variable magnitude.  Experiments on  soybean and sunflower have indicated values between 40 and 45% (96, 179) whereas similar determinations for wheat have shown between 10 and 15% inhibition (149).  The present data for t o t a l  inhibition of photosynthesis i n leaves of tomato plants grown at 0.03 and 0.5% C0  2  f a l l between these values (Table 11-10).  The  10% inhibition shown by plants grown at 0.1% CO^ i s less than half the values for plants from the other regimes. The t o t a l inhibition of photosynthesis by 21% 0^ comprises two components, a stimulation of photorespiration and a direct 0^ inhibition of photosynthesis (96, 280).  Different  studies have produced differing estimates of the relative contributions of these components to overall i n h i b i t i o n . for example, Curtis et al.  In soybean,  (71) found that photorespiration  accounted for s l i g h t l y more than 50^ of t o t a l i n h i b i t i o n of photosynthesis whereas a later study by Laing et al. (170) showed that photorespiration was responsible for only 33%, the remaining 67% being contributed by direct oxygen inhibition.  Other studies  on sunflower (178, 179) support the findings of Laing et al.  (170).  An explanation for the differing results probably l i e s i n the methods used by different workers to determine photorespiratory "Total" refers to a l l components of oxygen inhibition of photosynthesis.  TABLE 11-10:  Growth C O 2 Concentration (%)  Components of oxygen i n h i b i t i o n o f p h o t o s y n t h e s i s i n l e a f 3 of tomato p l a n t s grown a t 3 CO c o n c e n t r a t i o n s , a t l e a f p l a s t o c h r o n i n d e x : 2 (LPI : 2 ) .  Mean I r r a d i a n c e S a t u r a t e d Rate of P h o t o s y n t h e s i s i n Normal A i r (20.8% 0 , 0.03% C 0 ) ,-1x l O ) (g m 9  2  4  Rate of Photorespiration (g m" s-ixlO ) 2  Inhibition of P h o t o s y n t h e s i s by Photorespiration in A i r  4  Direct Inhibition of P h o t o s y n t h e s i s by Oxygen in A i r %  T o t a l Oxygen I n h i b i t i o n of Photosynthesis in A i r  0.03  2.00  0.37  18.5  6.8  25.3  0.1  2.55  0.24  9.4  0.6  10.0  0.5  1.80  0.33  18.3  5.2  23.5  CO  139  Ludwig and Convin (179) and Laing et al.  rates.  (170) both  measured photorespiration directly by methods involving the d i f f e r e n t i a l uptake of  C0  2  and  CO^ (180), and the measurement  of CO^ efflux into CO^—free a i r , respectively. Curtis et  al.  (71), on the other hand, calculated soybean photorespiration from a simple relationship between net photosynthetic rate and the C0  2  compensation point.  The data obtained from graphically  determined rates of photorespiration i n the present study (246) , indicate that the major part of the total oxygen inhibition of  photosynthesis  was accounted for by photorespiratory C0  It i s  production.  2  noteworthy that i n plants grown i n 0.1% C0  2  both photorespiratory  and direct oxygen inhibition of photosynthesis were reduced relative to those from the 0.03 or 0.5% C0  2  regimes.  In the case  of 0.1% grown plants, some 94% of total inhibition was accounted for by photorespiration alone. These observations help to c l a r i f y the close relationship between reduced photorespiratory rate and mesophyll resistance i n leaves of 0.1% grown plants, discussed above. Direct oxygen inhibition of photosynthesis may be a result of depleted supplies of the C0  2  acceptor RuBP (65, 83, 230, 239, 249), a depressed  rate of pyridine nucleotide oxidation (284), or the competitive inhibition of RuBP-case by oxygen with respect to C0  2  (36, 221)  which reduces the a v a i l a b i l i t y of the carboxylating enzyme for photosynthetic C0  9  fixation.  I f the l a t t e r mechanism i s correct,  140  as i s strongly suggested by evidence implicating the RuBP carboxylase/oxygenase  enzyme system i n the production of  glycolate, then any increase i n carboxylase a c t i v i t y w i l l reduce biochemical carboxylation resistance. This i n turn, would lead to a reduction i n total mesophyll resistance as observed for plants grown at 0 . 1 % CG^ i n these experiments. A l l the available evidence thus points towards a closely integrated system of photosynthesis and photorespiration whereby an increase i n RuBP-case activity leads to a stimulation of photosynthesis and concomitant inhibition of photorespiration. This does not discount the p o s s i b i l i t y that other systems may be involved i n increasing the photosynthetic potential of leaves developed i n 0 . 1 % C^.  Increased carbonic anhydrase a c t i v i t y at this  C O 2 concentration has been demonstrated i n young cotton plants ( 5 9 ) and rate.  may be an important determinant of maximum photosynthetic The enhancement of photosynthesis i n leaf  3  at  L71^:2  of  tomato plants grown at 0 . 1 % CO^ may thus be a result of a number of changes i n leaf physiology. Experiment 8 , which was conducted to determine a c t i v i t i e s of two enzymes i n relation to growth CO^ concentration, provided further support for the hypothesis that ultimate control of potential net photosynthetic rate was through RuBP-case. In leaves 2 and 3 of tomato plants grown at 0 . 1 % C O 2 at a PI of 5 , a c t i v i t y of RuBP-case was well over twice that of the same leaves  141  developed i n normal a i r (Figure 11-25).  In leaves 4 and 5 the  differences were not as great, but a similar pattern was  evident.  In contrast to the effect of the 0.1% CX^ growth regime, the data obtained from leaves developed i n 0.5% CO^ showed lower a c t i v i t i e s i n a l l four tested leaves, as compared with those from the normal air-grown plants.  These data do not correspond  precisely with a previous study using barley, i n which growth of plants at supra-normal  concentrations resulted i n increased  RuBP-case a c t i v i t y in the f i r s t leaf on the main stem (92).  In  this case increased a c t i v i t i e s were noted after growth for two days in an atmosphere containing between 1 and 5% CO^.  I t i s possible  that after a longer growth period under C^ enriched conditions, -  reduced RuBP-case a c t i v i t i e s would have been evident i n the experiments with barley.  Treatment with high (1 - 5%)  concentrations for a short period may e l i c i t similar effects to the longer term 0.1% exposures used i n the present experiments, where plants were subjected to each growth regime for at least 14 days. Enhancement of RuBP-case a c t i v i t y i n leaf 3 of plants grown at 0.1% CO^ i s consistent with the concept that increased carboxylation results i n decreased photorespiration and oxygen inhibition of photosynthesis.  The similar rates of  photorespiration shown by the same leaf on plants grown i n normal air a"d 0.5% CO^ are not, however, reflected i n similar RuBP-case a c t i v i t i e s (Figure 11-25).  It i s apparent that a growth concentration  141a  FIGURE 11-25:  A c t i v i t e s of RuBP carboxylase and glycolate oxidase extracted from leaves of tomato plants grown at 3 CO^ concentrations. PI:  5 ± 0.5  NOTE: Values are means of 3 replicate samples per CO^ regime shown ± one standard deviation.  GLYCOLATE OXIDASE ACTIVITY (umoles glyoxylate formed gfw^mirJ) o  o  p  p  o  o  p  _  »  ro  _l_  I  m >  co  _ i  i _  __i  I—•—I  W-»-H  TI  c W  I  m  »  1  3J  • • •  o  en  o o  ho  o  o o  CO  cn  o o o KJ o m o KJ m -i  RuBP CARBOXYLASE ACTIVITY (umoles RuBP converted g f w~ min") 1  D  O  Co  o  O  O  O  i  I  I  <ln  - v j t]o  —'  —'  I  I  —»  co  —'  ai  L_  1  ro  N)  ^*  co  cn  '  '  I  M  N>  NJ  -sj io L_-_-l  CD  3  (0  m >  CO A  a;  m  cn  -•-I  143  of 0.5% CO2 i s inhibitory to carboxylation a c t i v i t y of RuBP-case which may explain reduced  assimilation rates and growth  i n some plants subjected to this and higher concentrations (11). Since the oxygenase a c t i v i t y of the enzyme was not measured i n these experiments, i t can only be speculated that the competitive advantage of the RuBP-oxygenase system, which in the absence of high rates of carboxylation would be expected to increase glycolate production and photorespiration, i s not realized due to an overall suppression of enzyme a c t i v i t y by high ambient (X^ concentrations during growth. The d i f f e r e n t i a l effects of growing plants at 0.1% as compared with 0.03% CO2 upon RuBP-case a c t i v i t y , are similar to those noted between "sun" and "shade" grown plants, where enzyme a c t i v i t i e s are higher i n plants developed under high l i g h t intensities (29, 67).  Such plants also show higher rates of net  photosynthesis when measured at high irradiances (30).  Clearly,  growth conditions which favour high instantaneous rates of photosynthesis seem to be closely correlated with higher RuBP-case a c t i v i t i e s and higher inherent rates of photosynthesis. The pathway of photorespiration i n higher plants and some algae after the synthesis of glycolate apparently proceeds through the enzymic oxidation of this substrate by glycolate oxidase (98, 274, 275).  A reduced a v a i l a b i l i t y of glycolate, which seems  to be a l i k e l y result of the 0.1% growth C0  9  treatment i n these  144  experiments, might be expected to result i n a reduced a c t i v i t y of glycolate oxidase relative to the 0.03% CO^-grown plants. This i s confirmed by the data shown i n Figure 11-25.  In leaf 3,  GaO activity was approximately halved i n the 0.1% CO^ grown plants. A similar reduction i n activity of a number of enzymes associated with photorespiratory metabolism including glycolate oxidase, resulted from the two day exposure of barley plants to 1-5% CO^ (92).  In the present experiments, growth at 0.5% CO^ once again  resulted i n suppression of enzyme a c t i v i t y relative to both the 0.03-and 0.1%- grown plants.  The poor correlation between glycolate  oxidase activites in leaf 3 of plants from the 0.03 and 0.5% CO^ growth regimes i s d i f f i c u l t to reconcile with their similar photorespiratory rates (Figure 11-23).  I t i s , however, important  to r e c a l l that the enzyme i s involved i n other aspects of carbon flow within the leaf including the oxidation of glyoxylate to oxalate, a reaction involved i n the proposed glycolate-glyoxylate shuttle (237, 274, 277). This oxidation does not contribute directly to the main stream photorespiratory pathway resulting i n the production of CO2 via the decarboxylation of glycine (7, 160). The lack of substrate s p e c i f i c i t y demonstrated by glycolate oxidase, and i t s consequent involvement i n other reactions within the peroxisome, may account for the poor correlation observed between i t s a c t i v i t y i n extracts and measured rates of photorespiration or C0 compensation points (91). ?  145  iii)  Comparison of the Effects Photosynthesis,  of Growth CO„ Concentration  Photorespiration  and Enzyme Activity  on in Leaf S  at LPI :7.5 and LPI : 2,0 (Experiments 6&8)  The effect of age on net assimilation rate i n plant leaves i s d i f f i c u l t to predict. Patterns of behavior tend to differ even }  within a single species (6) and are mediated by external factors such as light intensity (18) and the sequence of reproductive and vegetative development within the plant (97, 228). Figure 11-26 shows the results of a second series of measurements of net CC^ exchange i n relation to irradiance i n leaf 3 of tomato plants grown at 0.03, 0.1 and 0.5% CO^. At the time of this experiment, the leaves had attained a LPI of 7.5 and the plants had been exposed to the treatment regimes for approximately five weeks. Photosynthetic rates at saturating irradiance were reduced relative to rates at LPI^:2 i n leaves of plants from each growth regime (cf. Figure 11-20).  In the plants subjected to 0.03 -4  or 0.5% CO2 this reduction corresponded to approximately 0.9 x 10 -2 -1 g CO^ m s . In the case of the 0.1%-grown plants, however, -4 -2 -1 the reduction was greater, being 1.1 x 10  g CX^ m  resulting maximum rates of net photosynthesis  s . The  at this later stage  of leaf development were, therefore, similar irrespective of growth regime. A similar relationship was observed over the entire range of irradiances tested.  Lower rates of dark respiration i n  145a  FIGURE 11-26: Apparent rates of photosynthesis i n relation to quantum flux density in leaf 3 of tomato plants grown at 3 CO^ Test C0  2  concentrations.  concentration:  Leaf temperature: LPI:  330 y l l "  1  30 ± 2°C  7.5 ± 0.5  NOTE: Points at any irradiance are not s i g n i f i c a n t l y different from one another (P >  0.05)  according to SNK multiple range test.  147  leaf 3 or the 0.5% C^-grown plants at this stage apparently resulted i n higher rates of net assimilation at low irradiance -2 (30 yE m  -1 s  ) as compared with the response at LPI^:2  (Figure 11-20) . These results are not consistent with those obtained from the experiments on greenhouse grown plants (Part I I , Section 1), where differences between rates of net assimilation i n plants grown under enriched or non-enriched conditions were evident i n young and old leaves on the main stem. To my knowledge, there are no other reports dealing with the variation of net assimilation rate with age i n leaves developed under different CO 2 concentrations.  Some  data, however,are available on such variations i n plants grown under different irradiances. Osman and Milthorpe (223) found that leaves of plants grown at high irradiance show inherently higher maximum rates of photosynthesis  early i n development followed  by a rapid decline during aging to a rate equal to or lower than those grown at lower irradiance. The high irradiance leaves had a shorter l i f e span and senesced e a r l i e r .  Thus, conditions  resulting i n a high rate of net assimilation induced an early decline i n photosynthetic capacity,, an observation which may bear upon results of the present experiments. At LPI^:7.5, leaf 3 of plants grown at 0.1% CO^ could be at a more advanced stage of senescence than a leaf at the same developmental stage i n 0.03% CO . The data obtained i n this study do not allow further  148  interpretation of the situation, but i t should be borne i n mind that senescence i s a complex phas^ of development during which leaves pass through several changes of metabolism each of which i s susceptible to environmental modification (251, 301). Physiological differences between plants grown i n controlled environments and i n f i e l d situations present a constant source of d i f f i c u l t y i n the interpretation of experiments using a r t i f i c i a l growth regimes (88). The differences between results obtained i n the experiments discussed here and those involving greenhouse grown plants may be directly related to the more r e s t r i c t i v e growth conditions encountered i n the controlled environment chambers. An earlier onset of senescence i n the growth chambers would be l i k e l y to mask differences i n inherent photosynthetic rates discernable i n the older basal leaves i n the greenhouse experiment.  The particular susceptibility of the  tomato plant to continuous a r t i f i c i a l l i g h t (5, 128) may indicate a special d i f f i c u l t y i n growing these plants i n controlled conditions, although the 12-hour photoperiod used i n these experiments was selected to circumvent problems of early leaf senescence which can be induced by unusual light-dark cycles (2, 127). The close s i m i l a r i t i e s between photosynthetic rates i n leaf 3 (LPI^:7.5) among plants from each growth regime are also exhibited i n relation to leaf response to i n t e r c e l l u l a r CO^ concentration (Figure 11-27). Mesophyll resistance to CO^ transfer  148a  FIGURE 11-27: Apparent rates of photosynthesis  i n relation  to intercellular CO^ concentration i n leaf 3 of tomato plants grown at 3 CO^ concentrations. -2 -1 quantum flux density: Leaf temperature: LPI:  520 yE m  s  , 400 - 700 nm  30 ± 2°C  7.5 ± 0.5  NOTE: Extrapolation to zero CO^ concentration to estimate rate of photorespiration.  I — 1  150  and rate of photorespiration, as calculated from the regression relationship, were independent of growth regime (Figure 11-28). Mesophyll resistances showed a significant increase over the LPI^:2 stage of leaf development i n a l l plants, the average change being 3 s cm 1  x  i n the 0.03 and 0.5% CO^-grown plants, and 12 s cm  i n those from the 0.1% regime.  x  Increases i n internal resistance  with leaf age a"d nutrient deficiency have bee" shown to contribute significantly to an overall decline i n photosynthetic in a number of species (223, 242).  activity  In others this resistance  has been shown to remain quite constant with age (218).  The  decrease i n maximum rates of photosynthesis with age i n leaf 3 of tomato plants i n this study seemed to be closely connected to an increase i n mesophyll resistance. again, dominant i n the  This resistance was, once  transfer catena at a l l irradiances,  since values were always higher than stomatal resistances even when the l a t t e r were measured at low irradiance (Figure 11-29). Stomatal resistances were generally higher than at the LPI^:2 stage of development (cf. Figure 11-21).  The increase of this  resistance with age i s i n accord with a number of previous studies (73, 133, 254), but since i t was smaller than mesophyll resistance, the p o s s i b i l i t y that this represented a major determinant of photosynthetic capacity seems unlikely (24). In a l l plants except those grown under 0.1% 00^, rates of photorespiration were depressed at the LPI^:7.5 stage of development  150a  FIGURE 11-28: Rates of photorespiration (R^) and mesophyll resistance (rm ) for leaf 3 of tomato plants r  grown at 3 CO^ concentrations and measured at LPI: 7.5 ± 0.5. NOTE: Values of R or r represented by bars L m T  bearing the same l e t t e r are not significantly different (P > 0.05) according to SNK multiple range test.  Rates of R_ and r L m  derived from regressions relating apparent photosynthesis to intercellular CO^ concentration, as described i n the text.  151  0 Al  r  A0  E o CM  E  0-3 H  h30  i LU O  < hco  < cr CL  CO LU  •—i  LU  0-2H  h-20  OC  >x  o  r-  CL O CO LU  o  X Q_ U. O  cr  0-1-1  -10  LU  I— < OC  R  m 003  L  P  %C02  0-1  0 5  GROWTH REGIME  151a  FIGURE 11-29:  Stomatal resistance to CO^ transfer i n relation to quantum flux density i n leaf 3 of tomato plants grown at 3 CO^ concentrations. Test CO^ concentration: Leaf Temperature: LPI:  330 y.1 1 ^  30 ± 2°C  7.5 ± 0.5  NOTE: Mean resistances at each irradiance are not significantly different (P > 0.05) according to SNK multiple range test.  20-,  — I  100 QUANTUM  1  !  200  300  FLUX  '  A00  '  500  D E N S I T Y ( u E rrf s , 4 0 0 - 7 0 0 n m ) z  Ul  153  (Figure 11-28).  In many plants photorespiratory a c t i v i t y changes  with leaf age, although frequently the pattern of change i s dependent upon leaf type, developmental events taking place i n other parts of the plant (9 7) and on previous growth conditions (84).  Data obtained from the present experiments are inconsistent  with some previous work on leaves of tobacco and some citrus varieties, showing an increase i n photorespiration with leaf age (243).  ^ock and Krotkov (94), however, found an increase i n  photorespiratory a c t i v i t y i n young, expanding leaves of kidney bean, which paralleled an increase i n net photosynthesis. stage of leaf development both photosynthesis  At a later  and photorespiration  declined, a finding which i s comparable to the present results and to some previous observations on wheat (97). Clearly, the decline i n net photosynthetic rate with leaf age shown i n these experiments i s not attributable to increased photorespiration.  I t i s , however, interesting to note  that rates of photorespiration in leaf 3 of plants grown under 0.1% CO2 were unaffected by leaf age; but the advantage of those leaves in terms of net carbon gain at the LPI^:2 developmental stage was lost at LPI^:7.5.  This may be due, i n part, to a marked  increase i n the t o t a l oxygen i n h i b i t i o n of photosynthesis which, at LPI^:7.5 was of similar magnitude to t o t a l i n h i b i t i o n i n the 0.03 a™d 0.5% CO^-grown plants (Table 11-11, Figure 11-30). The increase i " the direct oxygen inhibition component largely accounts  TABLE 11-11:  Components of oxygen i n h i b i t i o n of photosynthesis i n leaf 3 of tomato plants grown at 3 CC> concentrations, at leaf plastochron index: 7.5 (LPI :7.5) . 2  Growth C0 Concentration 2  Mean Irradiance Saturated Rate of Photosynthesis i n Normal A i r (20.8% 0 , 0.03% C0 ) (g m-2 s x l 0 ) 2  2  - 1  4  3  Rate of Photorespiration (g m~ s-^xlO ) 2  I n h i b i t i o n of Photosynthesis by Photorespiration in A i r  4  Direct I n h i b i t i o n of Photosynthesis by Oxygen in A i r %  Total Oxygen I n h i b i t i o n of Photosynthesis in A i r  0.03  1.33  0.19  14.3  16.0  30.3  0.1  1.45  0.19  13.1  14.2  27.3  0.5  1.20  0.18  15.0  10.0  25.0  154a  FIGURE 11-30: Effect of CO,, concentration on the t o t a l oxygen inhibition of photosynthesis i n leaf 3 of tomato plants grown at 3 CO2 concentrations. -2 quantum flux density: Leaf temperature: LPI:  7.5 ± 0.5  520 yE m  30 ± 2°C  -1 s  , 400 - 700 nm  156  for the higher total inhibition of photosynthesis at LPI^:7.5 in the 0.1% CO^ grown plants (Table 11-11).  I t was notable  that, total i n h i b i t i o n i n leaves of plants grown at 0.03 and 0.5% C0  2  showed l i t t l e change from the LPI^:2.0 developmental  stage. A review of the present data yields a number of interesting points.  The decline i n net photosynthesis i n leaf 3 of plants  grown i n the 0.03 or 0.5% C0  2  regimes from LPI :2 to LPI :7.5, 3  3  appears to be directly related to an overall decline i n C0 activity.  2  exchange  Both photosynthesis and photorespiration are decreased  at the later developmental stage, while t o t a l oxygen inhibition of photosynthesis remains relatively unchanged.  In the 0.1% C0 -grown 2  plants, however, the lower photosynthetic rates exhibited at LPI^:7.5 are partly due to an enhanced t o t a l oxygen inhibition of photosynthesis, including a greatly increased direct oxygen inhibition component (Table 11-11).  The results provide further  evidence to suggest that leaves developed under 0.1% C0 show 2  a degree of physiological adjustment resulting i n different C0  2  exchange characteristics at both early and late stages of development, compared with leaves of 0.03 or 0.5% C0 ~grown plants. 2  Lower maximum rates of photosynthesis at LPI^:7.5 as compared to LPI^:2 i n plants from each growth regime are generally correlated with lower RuBP-case a c t i v i t i e s (Figure IT-31). In leaves of the 0.5% C0 -grown plants, a c t i v i t y was quite similar 0  156a  FIGURE 11-31: A c t i v i t i e s of RuBP carboxylase and glycolate oxidase extracted from leaves of tomato plants grown at 3 CO^ concentrations. Plastochron index:  10.5 ± 0.5  NOTE: Values are means of 3 replicate samples per treatment shown ± one standard deviation.  GLYCOLATE OXIDASE ACTIVITY (umoles gloxylate formed gfw" mirT ) 1  O  CO  O O o O O — * — * ~ * ~ *  | s J U ) * ~ - c n O ^ - v l O O * i D O — » N ) C 0 * ~ I I L- I 1 1 1 1 1 jl k ill »  M  m >  O  o  1  I I  A  •  1  (D  m 30  •  •  o  o  •.  cn  vP  CO  o o NJ  rO  m pi  r  r  -i  ' l  o  o  o  1  °  RuBP CARBOXYLASE ACTIVITY  m o m  (umoles RuBP converted g fw" min") 1  O  o  o  u>  o  ro I r->U  t~-  p  1  o  p  cn cn L  1  p  o  -si 6o 1  -  ib  ^  1 |»'•! i  1  -*  o  J  1  ro to I  1  1  r  O  eg  3  to  m >  0D  m  I  *- 4  cn  Z.ST  M — W — I  co 4  11  i  1  1  •  r  1  h  =4]  H  •i  1  i  — l  158  to that obtained at LPI^:2, and, as before, was considerably lower than values for equivalent leaves of plants from the 0.1 or 0.03% growth regimes.  The general decline i n RuBP-case a c t i v i t y  with leaf age i s consistent with data obtained with a number of other plants (77, 124, 195, 256, 288). Maximum carboxylation a c t i v i t y has been found to occur during the period of leaf expansion.  Thereafter, RuBP-case a c t i v i t y rapidly declines to a  level which remains relatively constant u n t i l long after expansion has ceased (262, 264). In the present experiments, leaf 3 of plants from each growth regime was f u l l y expanded at LPI^:7.5 (equivalent to PI:10.5, Figure 11-31), and the reduction i n RuBP-case a c t i v i t y at this later developmental stage therefore corresponded with those previous studies. The increases i n mesophyll resistance at LPI :7.5 i n 3  comparison with LPI^:2, may be correlated with a reduction i n RuBP-case a c t i v i t y resulting i n an increase i n carboxylation resistance.  Indirect support for this idea i s provided by a  recent study which showed that the  transfer component of  mesophyll resistance i n bean leaves actually decreased with the onset of leaf senescence, while the biochemical and/or photochemical components showed a marked increase (272). In several plants, variations i n in vitro  RuBP-case  a c t i v i t y p a r a l l e l changes i n maximum photosynthetic rate (23, 38, 124, 256, 263, 264, 281, 288). Results obtained  for leaf 3 of  159  plants from the 0.03 and 0.1% CO^ growth regimes seem to be consistent with those findings, but the close s i m i l a r i t i e s between a c t i v i t i e s at PI 5 and PI 10.5 i n the 0.5% C^-grown plants were unexpected.  In addition, similar rates of net photosynthesis i n  leaf 3 of plants grown under 0.5%  and leaves from the 0.03 or  0.1% CO2 growth regimes were not reflected i n similar RuBP-case activities.  Such results lend some support to the previously  discussed hypothesis that growth of tomato plants at very high ^'  (-0.-05% and above) ambient CO2 concentrations results in an overall suppression of RuBP carboxylase/oxygenase a c t i v i t y .  Although close  correlations can exist between photosynthetic rate and RuBP-case (214), inconsistencies have also been reported (38).  I t i s evident  from the present results that factors other than a c t i v i t y of the carboxylating enzyme are responsible for determining maximum photosynthetic rates i n plants grown under 0.5% CO2.  The light  intensity under which plants are grown, has been shown to play an important role i n determining a c t i v i t y and photosynthesis  the relationship between RuBP-case (38).  Perhaps a similar importance may  be ascribed to ambient CO2 concentrations during growth. GaO a c t i v i t i e s at PI:10.5 did not show a consistent trend from leaf to leaf between the three growth regimes.  The results  were quite variable at this developmental stage, but indicated that there was l i t t l e difference i n GaO a c t i v i t y among the leaves studied.  This i s consistent with the similar rates of photorespiration  160  observed i n leaf 3 from each growth regime (Figure 11-28), but represents a quite different situation to that described for the PI:5 stage of development. GaO a c t i v i t y i n leaves of plants grown under 0.5% CO2 no longer showed lower values than leaves grown at other CO^ concentrations.  In the 0.03 and 0.1% CO2- grown  plants GaO a c t i v i t i e s i n leaves 2-to 5 were reduced relative to the PI:5 stage of development, but not i n those developed under 0.5% CO2. A decrease i n GaO a c t i v i t y with leaf age has been described i n kidney bean (94).  Salin and Homann (243), on the other hand found  lowest GaO a c t i v i t i e s i n the youngest leaves of tobacco and some citrus plants.  In the latter study, variation i n GaO a c t i v i t y  corresponded closely with photorespiratory rate i n a l l plants tested, but i n kidney bean the decline i n a c t i v i t y was much greater than the accompanying decrease i n photorespiration.  The results of the  present study indicate that the relationship between GaO (and RuBPcase) a c t i v i t y , and leaf age may be altered by CO2 conditions during growth.  I t i s probable that many of the differences between  the other studies cited above are related to different growth conditions or possibly to differences i n vegetative or reproductive plant development (228). The results of experiments performed at the PI:10.5 stage of development (LPI^:7.5) have indicated physiological differences between plants grown under 0.03, 0.1 and 0.5% CO^. The patterns of response to the various measurement conditions were  quite different to those at the PI:5 (LPI :2) stage. Rates of 3  net photosynthesis and photorespiration were similar i n a l l of the plants at LPI ;7.5. 3  The relative changes i n these paramete  with respect to increasing leaf age, however, differed markedly between growth regimes.  162  iv)  Replacement Experiment',  (Experiment 7)  The growth of tomato plants at CG^ concentrations of 0.1% i n a i r results i n changes i n the photosynthetic and photorespiratory metabolism of young leaves.  The previous  sections have shown that these changes are affected by leaf age, and that the effects are more marked i n chamber-grown plants. None of the experiments described so far have indicated whether the physiological modifications induced by growth at 0.1%  are  retained after a subsequent period of growth i n a non-enriched (0.03% C^) atmosphere. The results shown i n Figures 11-32 and 11-33 provide some information on this question.  Figure 11-32  i l l u s t r a t e s the relationship between C:.. and apparent photosynthesis i n leaf 3 of tomato plants at LPI : 2 after a 14 day growth period i n 0.03 or 0.1% CO^. The measurements were analogous to those described previously, and the results corroborate those shown i n Figure 11-22. Mesophyll resistances and rates of photorespiration derived from the regression relationships shown i n Figure 11-32 are l i s t e d i n Table 11-12. Plants exposed to 0.1% CO^ during the early stages of growth showed lower rates of photorespiration and mesophyll resistance than those grown i n normal a i r (0.03% CO^) • Magnitudes of each of these parameters were similar to those determined i n previous experiments (Figure 11-23). After a further two week growth period, during which plants were placed i n a normal a i r atmosphere i n the greenhouse, the  162a  FIGURE 11-32:  Apparent rates of photosynthesis i n t e r c e l l u l a r CG^ concentration  i n r e l a t i o n to i n leaf 3 of  tomato plants grown at 0.03% or 0.1% CO^ before placement i n common 0.03% atmosphere. -2 quantum flux density: Leaf temperature: LPI: NOTE:  520 yE m  -1 s  , 400 - 700 nm  30 ± 2°C  2 ± 0.5 Extrapolation to zero CO^ concentration to estimate rate of photorespiration.  to  £9T  ,  TABLE 11-12: Rates of photorespiration (R^ and mesophyll resistance to CO2 transfer ( r ) for leaf 3 of tomato plants grown at 0.03 or 0.1% CO2 before and after replacement i n a common 0.03% CO2 atmosphere. m  Before Replacement R_ r (g m s xl0 ) (s cm  Growth C0 Concentration (%) 2  L  4  m  i  0.03  0.1  "  14 Days After Replacement _ R r _ (g m s ^xlO ) (s cm ) 2  L  4  0.30a*  20a  0.20c  22a  0.22b  15b  0.22cb .  20a  ^Values of R or r bearing the same letter are not significantly different (P > 0.05) to the SNK multiple range test. L  m  m  1  according  165  relationship between CL and apparent photosynthesis was similar in plants from each growth regime (Figure 11-33). Mesophyll resistance was increased relative to the previous measurements i n leaf 3 of plants grown i n i t i a l l y under 0.1% C0 , although i t 2  remained constant i n the 0.03% CO ~grown plants (Table 11-12). 2  Rates of photorespiration were decreased i n plants from the 0.03% C0 growth regime but not i n those grown i n i t i a l l y under 0.1% C0 2  2  These results are i n broad agreement with those obtained at the LPI :7.5 stage of leaf development for plants maintained i n growth 3  chambers throughout development (Figure 11-28). Interpretation of the results of these experiments i s complicated by the confounding of effects of leaf age and replacement to a normal a i r atmosphere.  Moreover, i t does not  seem possible to separate these effects by any other experimental design, and the conclusions that can be drawn from the present data are, therefore, tentative.  The results do suggest that the  relative changes i n mesophyll resistance and rates of photorespiration in plants grown under 0.03 and 0.1% C0 are determined by 2  increasing leaf age and not by prolonged exposure to the particular C0 concentration i n the growth chamber. Changes i n photosynthetic 2  or photorespiratory processes which occur i n the young leaf i n response to growth at 0.1% C0 are modified as the leaf ages. 2  Plant replacement to an atmosphere containing normal levels of C0 has no effect on this modification. o  FIGURE 11-33:  Apparent r a t e s of p h o t o s y n t h e s i s intercellular  i n r e l a t i o n to  concentration  i n l e a f 3 of  tomato p l a n t s grown a t 0.03% o r 0.1% C0 ,,14 days 2  a f t e r placement i n common 0.03% C 0  2  -2 quantum  flux density:  Leaf temperature: LPI: NOTE:  520 yE m  atmosphere. -1  s  , 400 - 700  30 ± 2°C  12 ± 0.5 Extrapolation  to zero  C0  2  concentration  e s t i m a t e r a t e of p h o t o r e s p i r a t i o n .  to  167  The results are again i n contrast to those obtained in the greenhouse study where mature basal leaves developed under CO^ enriched conditions showed inherently higher rates of -  photosynthesis than those developed i n normal a i r .  The i n i t i a l  growth environment i s clearly of great importance i n determining the effect of aging on plants grown at 0.1%  CO^.  It i s unfortunate that these experiments have provided no definite answer to the question of whether changes induced in young leaves i n response to CO^-enrichment to 0.1% are retained for an extended period upon replacement in normal a i r .  Bishop and  Wittingham (21) have suggested that inherently higher rates of photosynthesis in tomato plants grown in low light and 0.1%  CO^,  upon transfer to a 0.03% C^*-atmosphere, are related to a greater pool size of certain photosynthetic intermediates. As they are careful to point out, the v a l i d i t y of this assumption depends upon the demonstration that differences i n photosynthetic rates between 0.03 and 0.1%-grown plants are rapidly eliminated after replacement in a common, 0.03% CO  atmoshpere.  In general, studies on the effects  of other environmental variables (notably light intensity and temperature) have indicated a rapid and reversible adaptation of plants to a change in growth conditions. An increase in light intensity resulted i n a concomitant increase in maximum photosynthetic rate i n shade grown maize and Amaranthus, which was complete after 6 days (118).  An even faster adaptation (24 hours) was evident i n  Encelia  califovnica  and Polygonum bistortoides  grown at cool  temperatures and transferred to a 30°C growth chamber (204). The question of whether physiological adaptations:" induced by growth under 0.1% CO2 follow a similar pattern of rapid adjustment when plants are returned to a 0.03% CO2 atmosphere, remains to be answered. The results of experiments 4 and 6, which investigated CO2 exchange rates i n relation to irradiance and CO2 concentration over a period of hours, seem to indicate, however, that very short term changes i n physiology, which could be related to metabolic pool sizes, do not occur.  169  PART II  - DISCUSSION  The results of these experiments have focussed attention on the physiological adaptations of tomato plants to growth under supra-normal CG^ concentrations i n greenhouses and controlled environment chambers. Results of experiments performed on chamber-grown plants have provided a comprehensive description of the physiological modifications which can take place under CO^-enrichment.  They have  answered many of the questions relating to the underlying basis for the changes observed i n plants grown under supplementary CO^ i n the greenhouse (Part I I , Section 1).  I t i s clear, for example, that the  increased efficiency of CC^ u t i l i z a t i o n (E^,) (14) in CC^-enriched plants was due, at least partly, to changes i n photorespiration and the a c t i v i t i e s of two enzymes (GaO and RuBP-case).  The discovery of  these mechanisms extended the basic observations of the study that photosynthetic capacitywas improved as a result of CG^ enrichment to 0.1% during growth, and that this concentration was apparently optimal for maximizing inherent rates of photosynthesis. i n concentration (to 0.5%)  Further increases  exerted different effects on the  photosynthetic system i n C02~enriched plants which may have masked or counteracted the enhancement due to growth at 0.1% 00^.  These findings  corresponded with previous studies on tomato which showed that highest rates of photosynthesis occurred i n plants grown at 0.15% CO^ and that increasing growth concentrations above this level resulted in significant  170  rate depressions (185).  The present studies have demonstrated,  furthermore, that the enhancement of photosynthetic capacity apparently depends on the physiological age of the leaves since photosynthetic potential of older leaves of chamber-grown plants was independent of CO2  concentration. The results and observations of these studies may  considered on a number of levels.  be  F i r s t l y , i t has been demonstrated  that tomato plants can adapt photosynthetically to ambient CO2 concentrations.  The relationships between this adaptation process  and the more extensively investigated adaptations to irradiance and temperature are worth examining.  Secondly, the physiological changes  that take place are complex and require careful review to elucidate the response of plants to growth under CX^-enriched conditions. F i n a l l y , the relationship between growth CO2 concentration and photosynthetic capacity should be considered i n relation to the agricultural practice of greenhouse  enrichment.  I t may be asked,  for example, how production techniques may be modified to maximize the use efficiency of (X^-enrichment f a c i l i t i e s .  The present discussion  is concerned with each of these points, and, i n addition, serves to connect the growth analysis observations (Experiment 5) with the results of the other experiments performed on chamber-grown plants. The change i n photosynthetic capacity i n response to growth at 0.1% (X>2 shows many parallels with the response patterns of plants to growth under different light and temperature regimes.  The higher  171  maximum rates of photosynthesis shown by plants grown at 0.1% CO^ as compared with 0.03% CO^ in these experiments, correspond to similar differences between plants grown under high and low ambient light intensities (24, 38, 44, 67, 76, 111, 233, 299).  Similar  photosynthetic adaptation has "also been shown i n relation to growth temperature. Plants from various environments tend to show higher maximum photosynthetic rates after growth under warm conditions (204, 205) although rates of arctic and alpine plants may be reduced by this treatment (20) . Previously, plant adaptation to different light and temperature regimes has recieved much more attention than the physiological changes resulting from growth at supra-normal CO^ concentrations.  This emphasis i s probably due to the greater  v a r i a b i l i t y i n f i e l d light and temperature conditions as compared to atmospheric CO2 concentration (203).  The widespread manipulation  of CO^ levels i n greenhouses has recently provided a strong incentive to investigate physiological adaptation to high CO2 concentrations. The present results also allow some comparisons to be made between the underlying mechanisms involved in adaptive responses to different environmental variables. The importance of changes i n RuBP-case a c t i v i t y , photorespiration and the degree of oxygen inhibition i n the regulation of photosynthesis i n plants grown at 0.1% CO2 as compared with those grown at 0.03% CO,  has been emphasized i n the  preceding sections. As already discussed, variations i n RuBP-case a c t i v i t y also play an important role in the photosynthetic adaptation of plants to high irradiance. On the other hand, increases in maximum  172  rates of photosynthesis in response to increasing irradiance during growth cannot be attributed to changes i n photorespiration i n at least one plant species (Atriplex;  24).  The physiological basis for adaptation  to different temperature conditions has been the subject of some controversy (26, 57, 204), but recent studies have suggested that changes in RuBP-case a c t i v i t y , dark respiration rates and thermo-stability of photosystem 2 a c t i v i t y are important factors in Atriplex (226,' 227).  lentiformis  I t i s evident, therefore, that adaptation to a growth CT^  concentration of 0.1% involves some physiological changes which are common to other adaptation processes (changes i n RuBP-case activity) and others which are distinctive (changes i n photorespiration and degree of oxygen inhibition of photosynthesis). It has been shown in the present experiments that the photosynthetic adaptation to supra-normal on the level of CO^ enrichment. -  concentrations depends  Some of the mechanisms underlying the  enhancement of photosynthesis at a growth  concentration of 0.1%  have been characterized i n this study, but the reasons for the absence of a similar response when the concentration was raised to 0.5% requires further consideration. I t seems l i k e l y that an overall suppression of enzyme a c t i v i t y (particularly RuBP-case) may be partly responsible, although the reasons for such an inhibition are not clear. Some explanation may be provided by previous studies on the relationship between CO^ concentration, leaf starch content and photosynthesis i n tomato plants.  At a growth C0  o  concentration of 0.5% tomato leaves tend to  173  accumulate excess quantities of starch (184, 186) and become rolled and mishapen (188).  Madsen (190) postulated that these effects may result  in chloroplast deformation and could explain his previous results which showed a reduction in photosynthetic rates at growth concentrations above 0.15%  (185).  Another explanation may be that high leaf starch  contents cause a reduction i n photosynthesis possibly through a feedback control on carboxylating enzyme a c t i v i t y . photosynthesis  Reduction i n leaf  rate as a result of large accumulations of starch have  been demonstrated i n a number of species (75, 130, 143). however, photosynthesis  In others,  i s unaffected by starch content (63) and there  i s , as yet, no general support for a l i n k between leaf photosynthesis rate and level of accumulated assimilate (213).  The p o s s i b i l i t y  that other regulatory mechanisms contribute to the effects discussed above cannot be  discounted.  It would be unwise to conclude from the present data that the mechanisms involved i n the photosynthetic adaptation of tomato plants to growth at 0.1% CO^ have been f u l l y characterized.  Undoubtedly  increased RuBP-case a c t i v i t y plays an important role i n this adaptation. The relatively lower mesophyll resistances at low irradiance i n greenhouse C^-enriched plants (Figure 11-11) may not, however, be directly correlated with enhanced carboxylation by RuBP-case (a dark reaction).  Elevated  concentrations during growth may cause other  changes i n photochemical fixation reactions in the chloroplasts which \ relieve inhibition of fixation at low irradiances.  174  The finding that the photosynthetic capacity of tomato plants changes with growth CO^ concentration holds important implications for the use of supplemental CC^ i n the production of commercial greenhouse crops.  The data presented in Figure II-9a clearly indicate that  photosynthesis i n leaves of plants grown at approximately 0.1% (X^ i n the greenhouse are more responsive to subsequent increases i n ambient CO^ concentration, than leaves of unenriched plants. This i s confirmed by the relationship between i n t e r c e l l u l a r CO^ concentration and net photosynthesis for young leaves of chamber-grown plants (Figure 11-22). These observations suggest that maximum benefit i n terms of photosynthetic enhancement w i l l be obtained by exposing young tomato seedlings to supplemental  as early as possible. General  recommendations state that CO^-enrichment may be commenced at the time of "pricking out" the young plants (3, 48).  The present data indicate  that this practice results, not only i n immediately increased rates of photosynthesis, but also i n higher rates at a s l i g h t l y later stage of development than i f the onset of CO2-enrichment i s delayed.  The  growth chamber experiments indicated that the enhancement of photosynthesis as a result of growth at 0.1% CO2 was increased at higher irradiances and was maximal under conditions of saturating irradiance (Figure 11-20). This was not f u l l y supported by the results of the greenhouse experiments, although there were indications that increasing irradiance enhanced differences between enriched and non-enriched plants (Figure 9b).  If  subsequent work v e r i f i e s this effect, then the benefits obtained from  175  CO^ enrichment of greenhouses crops may be even greater than previously -  imagined. Not only are inherent rates of photosynthesis enhanced by growth at 0.1% CO^, but this enhancement may be increased by seasonally increasing light levels. The beneficial effects of supplementary CO^ in greenhouse atmospheres are well established and require no further experimental v e r i f i c a t i o n (297).  The results of the present studies have indicated,  however, that the control of C0 ~enrichment levels at around 0.1%  may  2  be more important than has been realized.  To i l l u s t r a t e this point  i t i s worthwhile to consider the results of growth analysis (Experiment 5) and i t s relationship to other experiments on chambergrown plants. The higher NAR and RGR of 0.1% C0  2  versus 0.03%  C0 ~ 2  grown plants (Table II-8) are readily explained in terms of enhanced leaf photosynthesis at 0.1% C0 . 2  This enhancement i s accentuated by an  upward s h i f t i n photosynthetic capacity.  The higher NAR in the 0.5%  C0 -grown plants as compared with those grown at 0.03% also resulted 2  from an increased photosynthetic rate in direct response to the increased C0  2  content of the a i r .  NAR of the 0.5% C0 -grown plants was, 2  nevertheless, lower than of those grown at 0.1% C0 2  This difference  undoubtedly reflects the higher inherent photosynthetic capcity of the latter plants. I t i s worth repeating that the mutually compensating effects of decreased LAR and increased NAR i n the 0.5% C0 resulted i n similar RGR to those grown at 0.03% C0  2>  2  plants  Plants grown at  0.1% C0 , on the other hand, showed sufficient increase in NAR to produce 2  a significant increase i n RGR as compared to the 0.03% C0 -grown 2  controls.  I t i s clear that increasing the CO  content of a tomato  176  greenhouse atmosphere to concentrations greatly i n excess of 0.1% i s unwise.  Enrichment to 0.5% i s not effective i n increasing NAR  values obtained at 0.1% CO^.  over  I t may, i n fact, result i n a similar  rate of vegetative development to that shown by plants grown without supplementary CO^.  These effects may be of special concern i n other  crops, particularly greenhouse cucumbers which are often subjected to very high levels of CO i n the early stages of growth (Part I; 162). It i s , perhaps, surprising i n view of the considerable research that has been undertaken on the subject of CO^-enrichment, that the phenomenon of photosynthetic adaptation to high CO^ concentrations has only recently been recognized (190).  The results  of these studies have determined the responses of tomato plants to different CO^ concentrations i n excess of normal atmospheric levels and, for the f i r s t time, have established physiological mechanisms for these responses.  The question of whether similar adaptations are  shown by other species opens a large and intriguing area for future research.  PART III The -influence of supra-normal CO^ concentrations photosynthesis,  and vegetative  '•productive development of Pharbitis  and  nil Ghoisy.  Long and Short Bays .  on  plants  178  INTRODUCTION  In the early work of Garner and Allard (105, 106), the phenomenon of photoperiodism was clearly established as the basic controlling mechanism for flowering i n at least some plant species. Since then some research has been concerned with characterizing other environmental variables, including light intensity, temperature, and carbon dioxide, which may substitute for, or modify the effect of the timing of light/dark periods. The importance of l i g h t intensity was established i n early work which found that for f l o r a l induction to occur i n obligate SDP, the long (longer than c r i t i c a l ) dark period must be followed by a suitable period of high intensity white light (34, 113, 191, 259). This light requirement may be replaced i n some plants by the a r t i f i c i a l application of sucrose and some tricarboxylic acid cycle intermediates to the leaves or stems of plants maintained i n continuous darkness (56, 159, 17/5, 19.8). Such studies strongly suggested that high intensity light preceding or following an inductive dark period was concerned with the photosynthetic production of respiratory substrates necessary for complete f l o r a l development.  This explanation for the  l i g h t intensity effects was supported by earlier work (34, 117) which established that atmospheric CO^ was required.during the inductive light period i n B i l o x i soybean and Kalanehoe'..  I t has since been  corroborated by studies which have demonstrated that flowering may be completely inhibited by the application of the photosynthetic inhibitor DCMU during a normally inductive photoperiod (15).  179  Temperature treatments have also been found to affect photoperiodic behavior, but the interpretation of these effects has not proven as straight forward i n the case of light intensity effects.  For example, De Zeeuw (304) showed that the SDP Xanthium  pennsylvanianm/was induced to flower under non-inductive LD conditions i f the plants were maintained at 4°C for part of the long light period.  Similar results were obtained with Xanthium pennsylvanioum  by Nitsch and Went (217) and with Pharbitis  nil by Ogawa (220). Some  indication of the complicated nature of these temperature effects was provided by the finding that i n Pharbitis,  the cold treatment must  be continuous throughout the l i g h t period (141), while i n Xanthium, temperatures of 4°C must follow or precede a 25°C treatment for flowering to occur. Temperature has also been shown to exert negative effects on flowering.  In Pharbitis,  the normally inductive effects  of a long dark period were n u l l i f i e d by low temperature treatment, the effect being most pronounced when low temperatures were applied near the middle of the dark period (212). • The mechanisms by which temperature affects flowering are not well understood. Nitsch and Went (217) explained their results with Xanthium by suggesting that low temperatures inhibited the light-mediated destruction of a l a b i l e flowering stimulus, formed i n the dark.  In LDP i t has been shown that  some gibberellins are effective i n promoting flowering i n SD (201) and are produced  under cold conditions.  Gibberellins do not,  however, substitute for SD i n SDP i n various temperature regimes (244).  180  It seems equally l i k e l y that temperature influences the flowering process by altering the rate of certain metabolic reactions (199), but the true mechanism probably involves effects on gibberellin production, reaction rates as well as other biochemical systems. Atmospheric carbon dioxide concentration has also been found to modify photoperiodic responses. concentrations of  In a recent, study, high  (1% and above) were shown to. i n h i b i t flower  formation i n two SDP {Pharbitis  and Xanthiim),  and to promote flowering  i n the LDP SiV&ne armeria when a l l were maintained under SD conditions (234).  In another study, 1% CO2 was found to inhibit  flowering under normally inductive conditions i n the SDP Lemna paucieostata  (perpusilla)  (232).  In each case i t was suggested that  the effect of the high (X^ concentration was due to increased accumulation of photosynthetic products which directly or indirectly prevented flowering.  Inhibitory effects of high CO2 concentrations are not  restricted to SDP. An earlier study with the LDP Lemna gibba (153), indicated that 3.5% GO2 suppressed flowering, and a subsequent review attributed this i n h i b i t i o n to an indirect effect of enhanced photosynthesis  (154).  Some research has also considered the effects of high CO2 concentrations on photoperiodically sensitive plants grown i n non-inductive photoperiods. Xanthiim  Studies on Sitene armeria (234) and  (51), however, have indicated that at least some long and  short day species may be induced to flower by the provision of  181  supplementary CO^ during a photoperiod normally ineffective for f l o r a l induction. The study of the effects of secondary factors such as l i g h t intensity, temperature, and CC^ on photoperiodism have, i n the past, yielded important information on the mechanisms involved i n f l o r a l induction.  I t has been shown that normal flowering can be  controlled and modified by changing environmental conditions exclusive of the length of the light/dark period.  Such modifications may hold  important implications for the control of flowering i n commercially important crops.  At this time the potential of CO^ enrichment for this  type of control i s unknown and, i n fact, the link between photoperiodic behaviour and CX^ concentration i s not firmly established. I t •seemed desirable therefore,,to carry out further investigations into the role of CO^ i n f l o r a l induction. In addition, the interpretation that very high CO2 concentrations affected flowering via enhancement of photosynthesis seemed inadequate.  the  Many plants attain  CO2 saturation of the photosynthetic system at concentrations between 0.1 and 0.2% (103), yet concentrations as low as this were ineffective in modifying the photoperiodic flowering response i n Xanthium (51). Furthermore, very l i t t l e information was available on the effects of high levels of CO2 on vegetative development i n photoperiodically sensitive plants. The general objective of the research described i n this part of the thesis was to assess the influence of low (0.1%) and high  182  (1.0 - 5.0%), supplementary physiology of Pharbitis  nil  concentrations on the developmental seedlings.  Investigations were carried  out to obtain....information on both vegetative and reproductive development under different conditions of photoperiod and concentration (Experiment 9). The relationship of the effect of growth concentration on development, and photosynthetic a c t i v i t y was studied by measuring the gas exchange characteristics of plants at C0  0  concentrations up to 2.5% (Experiment 10).  183  MATERIALS AND METHODS i)  Growth System  Due to the unsuitability of conventional growth chambers for work requiring the maintenance of constant supra-normal CC^ concentrations i t was necessary to construct a new growth system for these experiments.  The maintenance of appropriate conditions  of irradiance, temperature, photoperiod and CC^ concentration i n the plant environment were the general requirements of the experimental system, which was developed for use i n conjunction with four growth chambers. A flow plan i s shown i n Figure I I I - l . The central feature of the system was a group of four plywood cabinets lined with thin sheets of PVC.  Each cabinet was divided  i n two by another sheet of PVC to give a t o t a l of 8 individual treatment boxes of internal dimensions 55 x 60 x 25 cm.  Each of  these was completely self-contained and sealed from i t s neighbour and the external atmosphere with silicone rubber wherever PVC sheets abutted.  The top and bottom walls of each treatment  box were of clear Plexiglas and access was provided by a 15 x 35 cm opening, which i n normal operation was sealed with a Plexiglas plate tightly pressed to rubber gaskets i n the front of the treatment  box. During system construction, two treatment boxes (numbers  4 and 7, Figure I I I - l ) were randomly selected for subsequent use as normal a i r controls. These two were f i t t e d with bottom plates perforated with 2.5 cm diameter holes to ensure adequate  183a  FIGURE I I I - l :  Diagram of gas flow system used i n Experiments 9 and 10.  LEGEND F: MF:  Flow meter Variable connection to mass flow sensor (1 l i n e may be connected at one time)  C:  Flow control clamp  TB:  Treatment box  nc:  normally closed  no:  normally open  /:  clamps f o r optional closure of i n d i v i d u a l gas lines  air  pump  C1<>  \'  F2F3  C2C)C3<>C4c>C5<>C60  v  v  v  "  "  no/  185  v e n t i l a t i o n and exchange of a i r with the external atmosphere. Mixing of a i r within every treatment box was f a c i l i t a t e d by a small e l e c t r i c fan, mounted close to the top of the front wall. Each of the gas-tight boxes was provided with one outlet and two i n l e t ports for the input and through-flow of a i r and CO streams.  Tygon p l a s t i c tubing was used throughout  gas  for the gas  supply l i n e s . During the course of the experiments, a i r was supplied continuously to the treatment boxes from a single gas pump (Doerr Electronics Ltd.). 1.29  or 4.65  1 min"  respectively).  1  The a i r flow rate was maintained at  ( i n the 5.0, 1.0 and 0.1 % C0  2  0.25,  treatments,  Flow control was by means of screw-mounted tubing  clamps which allowed variable c o n s t r i c t i o n of the p l a s t i c tubing. Just a f t e r the flow control clamps, each supply l i n e was  divided  into two with one branch terminating i n an open-ended tube and the other passing d i r e c t l y to a treatment box.  A i r flow rates were  monitored by completely closing the l a t t e r branch and connecting a mass flow sensor (Datametrics Inc. - see Part I I , Section 2: Materials and Methods) to the open-ended tube.  Whenever this  was done an uninterrupted flow to the treatment box was  maintained  by completing the c i r c u i t , with the mass flow sensor i n place through another p l a s t i c supply l i n e , following the mass flow sensor.  In normal operation, these a u x i l l i a r y lines were completely  closed by appropriately placed tubing clamps (Figure I I I - l ) .  186  Supplementary  CG^ was supplied from a cylinder of pure,  compressed  CC^ and the flow of gas to each treatment box was metered through needle valves and rotameter flow meters (Matheson 610A). rates were varied between 10 and 40 ml min  Flow  depending on the  required CO^ concentration. The treatment box CO^ i n l e t ports were located d i r e c t l y behind the fans, and the i n l e t s for the a i r . stream, 10 cm to the l e f t .  This arrangement  mixing of the incoming gases.  allowed e f f i c i e n t  Concentrations of CO^ i n the  outgoing a i r streams were measured with a Beckman model 864 IRGA which had a range of 0 - 2.5% C0 . 2  In the case of the 5% C0  2  treatment, the outgoing a i r stream was mixed with nitrogen gas i n a 1:10 ratio by means of a gas mixing pump (H. Wosthoff,  BRD)  before entering the IRGA. For  the experiments, two treatment boxes were placed  in each of 4 separate growth chambers (Sherer model 4005).  The  growth chamber controls were programmed to provide a photoperiod of 16 hours l i g h t and 8 hours darkness (LD treatment) on four of the  treatment boxes, 8 hours l i g h t and 16 hours darkness (SD  treatment) on the other four boxes.  Light intensity at plant  l e v e l inside a l l treatment chambers was between 3000 and 4000 lux (approximately 50 - 70 liE m  s ., 400 - 700 nm), and a i r temperatures  throughout the experiments were maintained at 25° ± 1°C.  187  ii)  Experiments a.  Effects  of growth C0„ concentration  development (Experiment Two  and photoperiod  9)  r e p l i c a t e t r i a l s were conducted to investigate  the effects of CO^  concentration and photoperiod on vegetative  and reproductive growth of Pharbitis 9 was  on  nil  seedlings.  Experiment  designed i n a 2-way f a c t o r i a l arrangement with 4 levels  of CC>2 concentration (0.03, 0.1, 2 photoperiods objective was  1.0 and 5.0%  by volume) and  (LD and SD as specified previously).  The  to evaluate parameters of vegetative plant  development ( r e l a t i v e growth rates of whole plants and leaves, leaf plastochron index and change i n stem height over the treatment period), and flower development (number of flower buds formed and the s e r i a l number of the primary flower bearing node). Pharbitis  nil  seeds (cv. Imperialis Japanese from  Stokes Seeds Ltd., St. Catharines, Ont.) were s c a r i f i e d and sown i n moist mica peat.  Germination  took place i n three to  four days i n a growth chamber at a constant temperature of 25°C.  Immediately a f t e r the cotyledons emerged, seedlings  which were uniform i n height and development were i n d i v i d u a l l y transplanted to 10 cm square p l a s t i c pots containing micapeat which were then replaced i n the growth chamber. transplanting, 72 seedlings were maintained  After  for two weeks  188  under a photoperiod consisting of 16 hours l i g h t and 8 hours darkness.  The l i g h t intensity was 4000 lux (ca. 70 yE m"  2  s" ,400-700nm) 1  and consisted of a mixture of fluorescent and incandescent illumination.  Temperature was maintained at 25°C both day  and night. At the end of the 14 day preconditioning period, a l l plants had formed two to three true leaves and were quite similar i n terms of vegetative development. At this stage 6 seedlings were placed i n each of the eight treatment boxes, 4 of which were exposed to SD conditions and 4 to LD conditions under the range of CG^ concentrations previously described. Leaf and t o t a l plant dry weights were determined for each of the remaining twenty four seedlings (Harvest 1).  Also, at this  time, the stem height (distance between the peat surface and apical shoot tip) of 3 randomly selected plants i n each treatment box was measured. Before placing seedlings in the treatment boxes each pot was watered to f i e l d capacity with a nutrient solution (see Part I I , Section 2:  Materials and  Methods), and i n most cases further watering during the course of the experiment was unnecessary due to the high humidity maintained within the treatment boxes. Watering of plants i n those boxes open to the growth chamber atmosphere was required at approximately  4 day intervals.  In these cases the  treatment box front plate was removed and s o i l moisture was  189  readjusted  to f i e l d  capacity using  treatment p e r i o d of each t r i a l was gaseous c o n c e n t r a t i o n s  tap water. 14  The  days d u r i n g which  of t h i s p e r i o d t h r e e p l a n t s were h a r v e s t e d f o r the d e t e r m i n a t i o n  growth r a t e s f o r whole p l a n t s and data o b t a i n e d cited  the  w i t h i n the chambers were c o n s t a n t l y  monitored and m a i n t a i n e d at the d e s i r e d l e v e l s .  ( h a r v e s t 2)  CC^  at h a r v e s t s  1 and  ( P a r t I I , S e c t i o n 2:  At the  end  i n each treatment  of dry weights.  Relative  l e a v e s were c a l c u l a t e d from  2 using  the formulae p r e v i o u s l y  M a t e r i a l s and  Methods).  For the remaining t h r e e p l a n t s i n each treatment box,  the stem h e i g h t s were remeasured and  buds and  the number of  t h e i r n o d a l p o s i t i o n s were n o t e d .  assessment of v e g e t a t i v e  development was  flower  An a d d i t i o n a l  made by  determining  the p l a s t o c h r o n  index of each p l a n t based upon a l e a f  l e n g t h of 30 mm  ( P a r t I I , S e c t i o n 1:  M a t e r i a l s and  reference  Methods).  Upon c o m p l e t i o n of a l l measurements, these remaining p l a n t s were r e t u r n e d  to a growth chamber under i d e n t i c a l  to those used d u r i n g 0.03%  C^) .  p l a n t was  the p r e c o n d i t i o n i n g p e r i o d  Seven days l a t e r ,  was  25°C and  the number of f l o w e r buds per  i n accord with  u s i n g a n a l y s i s of v a r i a n c e  (P  m u l t i p l e range t e s t ,  the  = 0.05).  c a r r i e d out by the two-stage ..' •„  (SNK)  (LD,  reassessed. Data were a n a l y z e d  design  conditions  (a = 0.05).  factorial Mean s e p a r a t i o n  190  b.  Effects  of growth CO„ concentration  photosynthesis  and photoperiod  rate and stomatal resistances  on  (Experiment 10)  Sixteen plants for use in gas exchange t r i a l s were raised from seed and preconditioned as described above. After preconditioning, two plants were placed i n daily sequence in each of the eight treatment boxes.  The allocation of  plants to chambers was at random except that plants assigned to the same chamber were inserted on consecutive days so that interruption of the gas flush treatments was minimized. In four of the treatment boxes, CO^ concentration was controlled at 1.0% with two of the boxes under SD conditions and two under LD conditions. 0.03%  The control treatment consisted of  CO2 (maintained by the flushing of growth chamber a i r )  with two treatment boxes at each photoperiod.  3  Each plant was  subjected to fourteen consecutive 24 hour treatment cycles after which i t was removed for the gas exchange determinations. These determinations were conducted on the second true leaf and consisted of two complete sets of measurements performed daily between 0900 and 1500 h.  The f i r s t set u t i l i z e d a simple  closed gas exchange system for the determination of photosynthetic rates at concentrations between 2.5% and CO^.  0.1%  The design of the system was identical to that described  in Part I I of this thesis (Section 2:  Materials and Methods),  except for the use of a Beckman Instruments Ltd. Model 864  191  IRGA. An open flow gas system incorporating a water-bath immersed bubbler, flow,meter, dew-point hygrometer and leaf chamber (Part I I , Section 2:  Materials and Methods) was used  in the second measurement set to determine stomatal resistances i n relation to CO^ concentrations between 0.03% and 2.5%.  In this case . CG^ concentrations were established  by mixing CO^-free a i r and 3% CO^ i n nitrogen from compressed gas sources, and verified prior to passing through the leaf chamber, by means of the IRGA. The a i r stream was humidified to between 70 and 80% relative humidity and rates of transpiration of individual leaves were calculated from chamber inlet and outlet dew point temperatures, a i r temperature and flow rate.  From these data a combined estimate of stomatal and  boundary layer resistance was calculated (146). Upon completion of a l l measurements the leaf was excised and i t s area determined as described previously (Part I I , Section 1: Materials and Methods)..  1  192  RESULTS i)  Effects  of COr. Concentration  on Flowering  and Vegetative  Growth  in LB and SD (Experiment 9)  Mean relative growth rates of whole plants and leaves of Pharbitis  seedlings grown i n LD and SD and at 0.03, 0.1, 1.0, and  5.0% CO^, are shown i n Table I I I - l .  In LD treatments, rates  were increased at 0.1% CO2 relative to the 0.03% CO^ control, while 5% CO2 resulted i n suppression.  Overall, whole plant growth  rates were higher i n LD than i n SD at 0.03 and 0.1%, although at higher concentrations (1 and 5%) increasing daylength had no effect on this index of dry matter accumulation.  A similar pattern was  shown by relative leaf growth rates i n LD, with the exception that at 1% C0 , rates were s i g n i f i c a n t l y higher than at the same CO2 2  concentration i n SD. Further information was provided by consideration of a number of parameters of vegetative and reproductive development. In each case, the data indicated a significant interactive effect of CO2 concentration and photoperiod (Table III-2) . At the end of the 14 cycle, LD treatment, plants grown at 0.03% and 0.1% CO2 showed a s l i g h t l y more advanced stage of plastochron development than those grown under SD (Table III-3). At 1 and 5% C0 , however, the mean, f i n a l index 2  was similar for the two photoperiods and was s i g n i f i c a n t l y higher than i n those plants grown under the lower C0 concentrations. o  Relative growth rates (RGR) of Pharbitis plants and leaves exposed to photoperiod and CO treatments during a 14 day period.  TABLE I I I - l :  SD*  LD  5T0  0.03  0.165*  0.171*  0.179  0.188*  0.189*  0.195  oTl  lTo  RGR whole plant (g g day )  0.164 *** 0.159*  RGR leaves (g g day )  0.185*  0.03  - 1  - 1  *SD: LD:  3  1.0  0.1b  5.0v  0;.19.I  0.177  0.202°  0.194  C  ab  0.164*  -1  0.181*  b  b  0.185  -1  Short Day treatment (8 hours l i g h t , 16 hours dark). Long Day treatment (16 hours l i g h t , 8 hours dark).  **Percentages of CO2 i n treatments. ***Means i n either row of the table designated with the same l e t t e r are not s i g n i f i c a n t l y different according to SNK multiple range test (P > 0.05).  a  TABLE I I I - 2 :  Source of Variation  S i g n i f i c a n c e l e v e l s of terms i n a n a l y s i s o f v a r i a n c e on v a r i o u s parameters o f growth and development i n Pharbitis plants subjected to d i f f e r e n t photoperiod and C0„ treatments.  Number o f F l . Buds at End of 2 Week Treatment  Number of F l . buds 1 Week A f t e r End o f Treatments  Change i n P l a n t Height Over Treatment Period  Final Plastochron Index  Final Lf A r e a Per Plant  Final Lf Dry Wt Per P l a n t  Photoperiod  AA  AA  AA  A  AA  AA  CO  AA  • *  AA  AA  AA  AA  a. j.  AA  AA  AA  AA  AA  concentration  Photoperiod X C0„ concentration  * D i f f e r e n c e s i g n i f i c a n t a t 95% l e v e l . * * D i f f e r e n c e s i g n i f i c a n t a t 99% l e v e l .  TABLE III-3:  Mean plastochron index of Pharbitis.plants at end of 14 days i n photoperiod and CO treatments.  SD* 0.03**  0.1  5.63 ***  5.69  a  LD 1.0."  a  7.47°  5.0  0.03  0.1  7.81°  6.27  6.26  b  b  1.0"  5.0  7.44°  7.54°  *SD: Short Day treatment (8 hours l i g h t , 16 hours dark). ...LD: Long Day treatment (16 hours l i g h t , 8 hours dark). **Percentages of CO^ i n treatments. ***Means designated with the same l e t t e r are not signficantly different according to SNK multiple range test (P > 0.05).  196  Under SD conditions and either 0.03 or 0.1% C0 , plants 2  tended to remain small with l i t t l e elongation of the main stem. Once again, the response to 1 and 5% CO^ during growth was quite different.  Under those conditions the main axis of the plants  showed a marked tendency to elongate.  This effect i s clearly  indicated by the values for mean change i n stem length (Table III-4) and by the comparisons shown in Figure III-2.  Elongation of  the main stem in plants grown at the two highest CO^  concentrations  in SD was approximately twice that of the normal a i r controls. In Long Days a l l plants showed greater stem elongation but again, the effect was most marked in those grown under 1 and 5% C0  2  (Figure III-3, Table III-4).  It i s interesting to note  in the present context, that the stem elongation response exhibited at the two highest C0  2  concentrations was not accompanied  by an increase i n relative growth rate of the plants (Table I I I - l ) . The normal effects of photoperiod on flowering in Pharbitis also modified by 1 and 5% C0  2  during growth.  were  Table III-5 l i s t s  the number of flower buds produced at the end of the 14 day  C0  2  treatment period, and after an additional week, during which the plants were maintained under LD conditions. In normally non-inductive long photoperiods, flower bud formation occurred on some plants exposed to 1 and 5% C0  2  (Table III-5, Figure III-4).  This response was not always exhibited in this experiment.  In  the f i r s t t r i a l two plants, and i n the second only one out of three, showed flower bud development under LD and 1 or 5% C0„  TABLE III-4:  Mean change i n stem height (mm) of Pharbitis plants exposed to photoperiod and C02 treatments during a 14 day period.  SD* * 0.03**  0.16  339.5 ***  497.0  a  *SD: LD:  LD 1.0  a  5.0  700.0 ° 660.0 b  b  0.03  0.16  1.0  810.3°  717.5 ° 1214.0 1135.0 b  5.0  d  Short Day treatment (8 hours l i g h t , 16 hours dark). Long Day treatment (16 hours l i g h t , 8 hours dark)  **Percentages of CO^ i n treatments. ***Means designated with the same l e t t e r are not s i g n i f i c a n t l y different according to SNK multiple range test (P > 0.05).  TABLE III-5:  Mean number of f l o r a l buds formed on Pharbitis plants a f t e r 14 days i n photoperiod and C0„ treatments, and 7 days after the end of treatments.  SD*  LD  0.03**  0.1  1.0  End of treatments  cb 1.5***  2.75°  2.75  End + 1 week  4.75  3.50  4.5  b  b  b  C  5.0  1.0  0.1,  0.03  5.0  i.oo o.oo  a  o.oo  2.75°  2.0  o.oo  o.oo  3.25  4.00  ab  3.00  b  a  a  a  b  Cb  ( A l l plants replaced to LD conditions) *SD: LD:  Short Day treatment (8 hours l i g h t , 16 hours dark). Long Day treatment (L6 hours l i g h t , 8 hours dark).,  **Percentages of CO  i n treatments.  ***Means i n either row of the table designated with the same l e t t e r are not s i g n i f i c a n t l y different according to SNK multiple range test (P > 0.05).  b  198a  FIGURE III-2:  Pharbitis  plants A and B.  A exposed to 14 SD  cycles and 0.1% CO^; B exposed to 14 SD cycles and 1% CO^.  Plants photographed 7 days after  return to uniform LD conditions (0.03% CO ) .  1'99a  FIGURE I I I - 3 :  Pharbitis  plants  C and D.  C exposed to 14 SD  c y c l e s and 0.03% CO^; D exposed t o 14 LD c y c l e s and 5% CO^. Plants  Note the f l o w e r formed on p l a n t  photographed 7 days a f t e r r e t u r n  LD c o n d i t i o n s  (0.03% C 0 ) . o  C.  to u n i f o r m  200  200a  FIGURE III-4:  F i r s t f l o r a l bud at node 6 on plant D (see Figure III-3) exposed to 14 LD cycles and 5% CO .  Photograph at same time as Figure III-3.  201  202  conditions.  No flower buds were observed on plants maintained  under 0.03 or 0.1% C0 i n LD at the end of the C0 treatment 2  2  period, or 1 week later.  Higher than normal C0  concentrations  2  had no significant effect on the t o t a l number of flower buds formed i n SD.  However, growth at 1 and 5% C0 i n SD did have a 2  marked effect on the position of the f i r s t f l o r a l bud upon the main stem, and on the overall morphology of flowering i n these plants (Table III-6, Figures III-2 and III-5 to III-8).  Plants  grown at 0.03 or 0.1% C0 i n SD formed their f i r s t f l o r a l buds 2  in the axils of leaves 3 or 4 (Table III-6).  Development proceeded  with f l o r a l buds being formed at each subsequent node on the main stem axis (Figures III-5, III-6), and i n many cases a terminal flower bud was formed (Figure III-7).  On the other hand, at 1  and 5% C0 , the f i r s t flower buds were formed at nodes 6, 7, or 2  8.(Figure III-8, Table III-6) and the cessation of stem elongation by induction of a terminal flower bud was never observed. I t i s interesting to note that when flower buds were formed under LD conditions at high C0 concentrations, the f i r s t ones also 2  appeared at nodes 6, 7, or 8 (Figures III-4, Table III-6), with no f l o r a l bud development below this level on the stem. Visual observation of plants exposed to 1.0 and 5.0% C0 i n LD or SD 2  revealed a less vigorous flowering response i n comparison to those grown at 0.03 or 0.1% C0 i n SD (cf. Figures III-4 and III-5; 2  III-6 and III-8).  TABLE III-6:  Mean node of f i r s t f l o r a l bud formed on Pharbitis plants after 14 days i n photoperiod and CC^ treatments .  SD* 0.03**  0.1  3.5****  3.25  a  LD 1.0  5.0  7.0°  6.75  C  0.03  0.1.  nf  nf  b  b  1.0  5.0  7.25°  7.75  nf: No f l o r a l buds formed. *SD: "."Short Day treatment (8 hours l i g h t , 16 hours dark). **LD: Long Day treatment (16 hours l i g h t , .8 hours dark).. ***Means designated with the same l e t t e r are not s i g n i f i c a n t l y / different according to SNK multiple range test (P < 0.05).  C  203a  FIGURE III-5:  Detail of developing f l o r a l buds and one flower on plant C (see Figure III-3) exposed to 14 SD cycles and 0.03% C0 . Photograph at 2  same time as Figure III-3.  204a  FIGURE I I I - 6 :  T h i r d f l o r a l bud at node 6 on p l a n t A (see F i g u r e I I I - 2 ) exposed 0.1% CO^. Figure  to 14 SD c y c l e s and  Photograph a t same time as  III-2.  205  205a  FIGURE III-7:  D e t a i l of plant A showing terminal f l o r a l bud '(see also Figure III-2) . SD cycles and 0.1% CO^. as Figure III-2.  Plant exposed to 14 Photograph,at same time  206  206a  FIGURE I I I - 8 :  D e t a i l of f i r s t  f l o r a l bud a t node 7 on p l a n t  B (see F i g u r e I I I - 2 ) and 1% CO2. Figure  III-2.  exposed t o 14 SD c y c l e s  Photograph a t same time as  208  ii)  Effects  of 0.03 and 1.0% CO During Growth in LD and SD on  Subsequent Rates of Net Photosynthesis  and Stomatal  Resistances  (Experiment 10)  Figures III-9 and 111-10 depict the effects of C0 concentration on net photosynthesis Pharbitis  2  in the second leaf of  seedlings, grown under four combinations of C0 and 2  photoperiod.  Plants grown for 14 days at a continuous concentration  of 1% C0 showed lower inherent rates of net photosynthesis at 2  test concentrations above 0.5% as compared to those subjected to normal a i r (0.03% C0 ). These differences were independent of 2  photoperiod, except at a test concentration of 0.5% C0 where 2  rates were significantly different between plants grown i n LD and those grown i n SD.  An explanation for the rate reductions  in the high C0 -grown plants was sought i n the relative magnitudes 2  of leaf resistances at test C0 concentrations of 1.0% and 2.5% 2  (Table III-7).  I t was apparent that.increased stomatal resistance  was not a contributory factor, since, with values of 9.50 and 6.78 s cm , mean resistances for leaves of the 1%-grown plants were x  quantitatively, but not s i g n i f i c a n t l y , lower at 2.5% than for those of plants grown at 0.03%.. C0 . 2  Similarly, at a test concentration of  1.0% C0 no significant differences were found between plants subjected 2  to different growth C0 concentrations. 2  An increase i n test C0 concentration from 0.03 to ' 2  2.5% caused a marked increase i n stomatal resistance (Table III-7). The magnitude of the increase was somewhat variable depending  208a  FIGURE III-9:  Apparent rates of photosynthesis i n relation to CO^ concentration i n Pharbitis grown i n 0.03% or 1.0% CO  -seedlings  i n Long Days.  O  209a  FIGURE 111-10: Apparent rates of photosynthesis i n relation to CO^ concentration i n Pharbitis grown i n 0.03% or 1.0% CO  seedlings  i n Short Days.  to f—  1  o  TABLE III-7:  Combined leaf and a i r (boundary layer) resistances to CO-2 transfer i n the second leaf of Pharbitis plants grown under LD or SD photoperiods and 0.03 or 1.0% C0„.  SD (8" hours.'.light 16 hours dark) 0.03% C0 1.0% C0 2  2  LD (8 hours dark 16 hours light) 0.03% CO 1.0% C0  2  Test CO Concentration 0.03%  6.33*  6.87  6.09  5.35  0.1%  7.34  6.56  6.59  5.93  1.0%  8.13  9.21  8.91  6.45  2.5%  10.58  9.50  9.14  6.78  *Leaf + a i r (boundary layer) resistances i n s cm . Values i n any row of the table are not significantly different according to analysis of variance (P > 0.05).  212  upon the particular combination of growth photoperiod and CO^ concentration, but averaged 2.8 s cm ^ between the lowest and highest test concentration. With one exception (1.0% growth CO^ under SD photoperiod), resistance increments were also noted between 0.03 and 0.1, and 0.1 and 1.0% test C0 . 2  At the lower  concentrations these increments corresponded closely with those noted i n a number of previous studies (e.g. 123). Increased stomatal resistance at high CO^ concentrations apparently imposed l i t t l e r e s t r i c t i o n on photosynthetic a c t i v i t y . In plants grown at 0.03 or 1% C0 i n LD or SD, photosynthesis 2  approached saturation between 0.1 and 0.5% CO^ (Figures III-9 and 111-10) . The gradual l e v e l l i n g off of the C0 response curves above 2  0.5% C0 was probably due more to an i n a b i l i t y of the photosynthetic 2  apparatus to u t i l i z e additional CX^ than to the effects of stomatal closure.  This view i s supported by other studies which have shown  that increased CO  concentrations around the leaves of corn and  sunflower resulted i n increased intercellular concentrations and enhanced photosynthesis despite an increase i n stomatal closure (291). In plants grown at 1.0% CO^, rates of net photosynthesis tended to level off at somewhat lower test C0 concentrations, 2  compared to those grown i n 0.03% C^.  The response of plants  subjected to SD photoperiods and 1.0% CX^ to test concentrations above 0.5% was quite erratic, and rates at 1.0% and 2.5% (2.68 and  213  2.45  x 10  g CO2 m  s  r e s p e c t i v e l y ) were o f s i m i l a r magnitude  to those a t 0 . 1 % ( 2 . 6 6 x 1 0 ~ Plants  5  g C0  2  m~  s  2  _ 1  111-10).  grown a t 1 . 0 % CO^ i n LD showed a more c o n s i s t e n t  response ( F i g u r e I I I - 9 ) . photosynthetic  Nevertheless,  little  show t h a t net p h o t o s y n t h e t i c  at t e s t C 0  2  increase i n  r a t e was observed above 0 . 5 % C ^ .  In summary, t h e r e s u l t s p r e s e n t e d 111-10  ) (Figure  concentrations  i n Figures  I I I - 9 and  r a t e s a r e i n h e r e n t l y lower  above 1% i n p l a n t s grown a t 1 %  C0  2 >  as compared t o those grown i n normal a i r ( 0 . 0 3 % ) .  The lower r a t e s  cannot be e x p l a i n e d  resistance  but  appear t o depend on p h o t o s y n t h e t i c  leaves.  reactions i n s i d e the  i s e s s e n t i a l l y saturated  and 0 . 5 % C 0 2 i n a l l p l a n t s .  i n c r e a s e of photosynthesis was  stomatal  I t i s a l s o c l e a r from t h e s e r e s u l t s t h a t t h e p h o t o s y n t h e t i c  f i x a t i o n of C 0 2 0.1  i n terms o f i n c r e a s e d  evident  with  above 0 . 5 % CO..  at concentrations  Only a slow, and sometimes increasing test  C 0  2  between erratic,  concentration  214  DISCUSSION  The results obtained in these experiments indicate that very high CC^ concentrations (1 and 5%) have distinctive effects on the vegetative and reproductive growth of Pharbitis photoperiods.  nil  under different  The findings have provided more support for the  hypothesis, established by a number of previous workers (51, 234, 247), that these CO^ concentrations are effective i n modifying plant growth and development. ,  Further understanding of the developmental role of CG^  required close examination of the effects of very high CO^ concentration on vegetative and reproductive growth and how they relate to one another.  I t may be asked, for example, i f the effects share common  physiological mechanisms, and which ones may be involved?  The following  discussion i s concerned with these questions. The relationship between CO^ concentration during growth under LD or SD photoperiods and vegetative development i s well i l l u s t r a t e d by the data shown i n Tables I I I - l , III-3, and III-4.  The  results showing increased stem length and leaf production, but a similar RGR i n plants grown at 1 . 0 or 5.0% CO^ as compared with those grown at 0.03 or 0.1% C O 2 , suggested that very high CO^  concentrations  induced changes i n dry matter distribution within the plants. In a previous study Purohit and Tregunna (234) observed that  Pharbitis  seedlings grown i n SD and 1.5% C O 2 had fewer leaves per plant and were shorter than those grown at 0.1% C 0 9 . The discrepancies between  215  t h e i r results and those of the present study may have arisen as a r e s u l t of the d i f f e r e n t methods employed i n the control of gas concentrations, or because of differences i n irradiance supplied during growth.  The c o n f l i c t i n g results may also be due to an e r r a t i c  morphogenetic behaviour of plants to very high CO^ concentrations, corresponding with the photosynthetic response of some species to concentrations above 0.2% (207). It i s apparent that the increases i n stem length and leaf production shown by Pharbitis  seedlings i n the present study are not  simply, related to increased photosynthesis and growth caused by higher CO^ concentrations during development.  A number of studies on  the e f f e c t s of 00 - enrichment i n other plant species have shown that vegetative development i s more vigorous at C O 2 concentrations above 0.03%. In tomato plants, stem height increases with approximately  concentration up to  0.22% (188) and similar responses have been shown by roses  (250) Chrysanthemum (8) and Poinsettia  (109).  In each case, increasing  GO^ concentration from 0.03 to 0.1% also produced s i g n i f i c a n t effects on dry matter accumulation  and plant vigor, presumably through increased  photosynthetic production.  This i s c l e a r l y i n contrast to the present  r e s u l t s which indicated no s i g n i f i c a n t changes i n stem morphology by CO^-enrichment to 0.1%. On the other hand, a further increase i n growth C O 2 concentration to 1% produced certain changes i n morphological development which were apparently independent of r e l a t i v e growth rate.  The observations on f l o r a l bud formation (Tables III-5 and III-6) provide equally strong evidence for an effect of 1.0 or 5.0%  on flowering in Pharbitis  i n both LD and SD. As an aid  in recognizing these effects i t i s important to understand the normal course of development i n Pharbitis  under different photoperiods  It has been known for many years that Pharbitis  seedlings become  photoperiodically sensitive with the unfolding of the cotyledons two to three days after germiantion (168).  At t h i s , or a later, stage  of development, a dark period of 16 hours i n a single 24 hour cycle w i l l induce flowering (142).  Several repetitions of the cycle cause  the development of a terminal flower and the complete cessation of v e r t i c a l stem growth (269).  Under weaker stimulation, for example,  when plants are subjected to dark periods of only 13 or 14 hours i n 3 or 4, 24 hour cycles, the apex continues to grow vegetatively and flower Buds are only developed i n the axils of the leaves. The c r i t i c a l dark period i n most seedlings appears to be between 9 and 10 hours (142) and plants maintained under 24 hour cycles each incorporating a 16 hour light period show only vegetative growth for ah almost indefinite period (194).  Since Pharbitis  seedlings  w i l l i n i t i a t e flowering after a single SD, the 14 cycles each incorporating a 16 hour dark period given i n the present experiments constituted an extremely strong stimulus to flower.  This was shown by  217  the f l o r a l morphology of plants grown at 0.03% or 0.1% C0 i n 2  SD.  In these cases flower buds were formed i n the a x i l s of leaves  low on the stem (Table III-6) and frequently at the stem apex (Figure III-7).  The effect of 1.0 or 5.0% C0  2  was to i n h i b i t  flowering i n SD, since i n plants maintained under these conditions flower buds were formed at much higher nodes than i n the other C0  2  treatments  (Table III-6), and the stem apex remained vegetative i n  a l l cases.  This interpretation i s i n l i n e with the discussion by  Hillman (129) which established that, under two sets of treatment conditions, plants which have developed fewer leaves before the development of flower buds are considered to be at a more advanced flowering stage. Under non-inductive LD conditions, flowering was stimulated i n a number of cases by growth at 1 or 5% C0  2  (Table III-5).  It i s  important to note, however, that i n these cases morphology was similar to that shown by plants maintained at the same C0  2  concentrations  i n SD (cf. Figures III-2 and III-3). The results showing i n h i b i t i o n of flowering i n seedlings kept i n SD at 1 or 5% C0  2  Pharbitis  (Table III-6, c f . Figures III-6  and III-8) confirm the results of Purohit and Tregunna (234).  The  present study has also considerably extended the work of those authors by demonstrating  that these very high C0  2  concentrations can  promote flowering i n this species under normally non-inductive LD  218  conditions.  Generalizing, i t may be stated that very high CO^  concentrations tend to oppose the photoperiodic influences on flowering i n Pharbitis.  Similar effects have previously been shown  by studies on Xanthium pennsylvanicum  (51, 234) in which i t was  observed that CC^ concentrations i n excess of 1% inhibit flowering under inductive SD conditions and promote i t i n LD. It i s worthwhile at this point, to compare the effects of supra-normal CO^ concentrations on vegetative and reproductive growth in Pharbitis  i n order to determine i f a relationship could exist between  them. Under both LD and SD conditions an increase i n CO^ concentration from 0.03 to 0 . 1 % had apparently l i t t l e effect on vegetative growth or flowering. The modifying effects on both types of development occurred only when the CO^ concentration was raised to 1 or 5%. In addition, the vegetative and f l o r a l morphology were affected by these concentrations under both photoperiods.  These observations tend  to suggest that the effects of 1 or 5% C O 2 on vegetative and reproductive growth have a similar basis. I f , on the other hand, i t i s considered that flowering i s inhibited by 1 or 5% C O 2 i n SD but promoted by the same concentrations i n LD, the conclusions that are drawn from these results may be quite different.  In this case, i t  i s evident that conditions which inhibit flowering (LD and 0.03 or 0 . 1 % C O 2 ; SD and 1 or 5% C O 2 ) produce, respectively, the smallest and largest increases i n stem length (Table III-4).  This suggests,  conversely, a different basis for the C 0 ? effects on reproductive  219  and vegetative development. This suggestion may be considerably strengthened i f flower formation and vegetative growth in Pharbitis  can be shown to  be governed by separate physiological mechanisms, as i s the case i n certain rosette plants (306). The difference between the two viewpoints rests on the question of whether very high  concentrations influence  development which i s s t i l l primarily under photoperiodic control or whether they act by releasing development from this control.  Evidence  in favour of the latter mechanism would provide support for a common basis for the effect of C0^ on vegetative and reproductive growth.  This  question w i l l be addressed i n a later part of this discussion. The present results do not indicate the nature of the physiologi c a l mechanisms which may be important in the control of development i n Pharbitis  by 1 or 5% C O 2 .  They do, however, focus attention on the  interpretation of some previous results which have also demonstrated the regulatory effects of C0 on flowering i n a number of species (51, 153, 2  232, 234).  In 1971, Posner (232) noted that the inhibitory effects of  exogenously applied sucrose on flowering i n Lemna paucicostata were^slmilar to the effects of 1% (X^.  (perpusilla)  He postulated that the correlation  was due to an increase in photosynthesis, giving r i s e to greater sucrose production, i n the presence of high CO^ concentrations.  Similar observa-  tions and conclusions were made by Kandeler (153, 154), working with Lemna gibba.  In a more recent study, Purohit and Tregunna (234) suggested  that the inhibitory effects of 1% C0 on flowering in Pharbitis 2  might  be due to the increased production of a photosynthetic product which i s required i n minimum quantities for f l o r a l induction. In the  220  present study, however, i t was observed that, i n plants grown at -1%  CO^ i n SD, rates of net photosynthesis were quite erratic at  test  CO2  concentrations between  0.1  and 2.5% (Figure  111-10),  the  range over which large differences in development had been observed in Experiment 9 (e.g. Tables III-5 and III-6).  In contrast, when test  CO^ concentration was raised from 0.03 to 0 . 1 % in these plants, photosynthesis rates showed a clear increase from 1.81 to 2.66 x 10~* g -2 - 1 m  s  (Figure . 1 1 1 - 1 0 ) .  Yet, a similar increase i n growth  CO^  concentration was ineffective i n modifying development in the plants tested.  These results did not support the interpretation of previous  workers that the developmental effects of high C O 2 concentrations are mediated by increases i n photosynthetic rate. In fact, plants grown at 1% C O 2 i n SD tended to have lower inherent photosynthetic rates than those grown at  0.03%  C0  2  (Figure  In plants grown in LD at  111-10).  1%  t n e  response of net  photosynthesis was less erratic at high iGO^. concentrations (Figure III-9).  In this case, the gradual increase i n photosynthetic  rate between test concentrations of 0 . 1 and 1% C O 2 in both 0.03 and 1% C O 2 grown plants, might be connected with the differences i n development over this range.  As with plants grown i n SD, however,  i t i s d i f f i c u l t to explain the lack of developmental response to an increase i n test GO2 concentration from 0.03 to 0 . 1 % , which results i n a relatively large increase i n net photosynthesis. It i s important to distinguish between the apparent lack of correlation between photosynthetic rate and the very high C 0 0  221  concentration effects shown i n this study, and the involvement of photosynthesis i n flowering. Several studies, showing a requirement for high irradiance and atmospheric concentrations of CO^ during inductive light periods (15, 31, 34, 42, 100) have established a l i n k between photosynthesis and flowering i n both LDP and SDP. The precise role of photosynthesis i n the flowering process, however, i s not well understood.  Some workers have suggested that adequate rates  of photosynthesis alone are not sufficient for the induction of flowering.  Cumming (68) has presented evidence i n support of a dual  requirement for flower induction i n Chenopodium vubrum based on photoperiodic stimulation of the phytochrome system i n combination with sugar substrates produced by photosynthesis during the light period. On the other hand, induction of flowering took place i n dark-grown Pharbitis  seedlings following two short red irradiations i n a 24 hour  dark period, even though they showed no net photosynthesis (101). These studies clearly demonstrate the complexity of the relationship between photosynthesis and flowering. I t seems l i k e l y that the major changes in physiology which are associated with flowering may require substrates provided by net assimilation.  There  are, however, many other modifying factors which are apparently independent of photosynthesis.  Bassi et at.  (13) found that C0  2  removal during a short light break i n the middle of an inductive dark period i n Xanthium pennsylvanicum  reversed the inhibitory effects  of this treatment on flowering.  In a later study (12) i t was  222  demonstrated that CO exchange during similar light breaks was not correlated with flower induction.  The results of the present study  have demonstrated that flowering may also be modified by continuous treatment with very high CO^ concentrations, a factor which, once again, does not appear to be related to photosynthesis. The diversity of the effects of CO  on flowering suggests  that this gas may be involved through a number of physiological mechanisms.  The present data permit only a speculative approach to  the question of which ones may underlie the modification of flowering and/or vegetative development by 1 or 5% CO^.  The problem i s complex  since i t i s uncertain at present, whether similar mechanisms are involved i n the response of plants exposed to LD and SD conditions, or whether the mechanisms depend on photoperiod.  Each of these  p o s s i b i l i t i e s w i l l be considered i n the following discussion. The effects of CCv, concentration on respiration represent a possible control point for the action of 1 and 5% CO^.  Kidd  (158) noted that 1% CO^ could significantly suppress rates of dark respiration.  Such an effect might be expected to reduce energy  supply during an inductive dark period, which may be required for the synthesis of.a f l o r a l stimulus (306) and thereby i n h i b i t flowering under SD conditions.  Indirect support for this hypothesis i s provided  by the work of Nakaymama (212) who showed that flowering i n  Pharbitis  depends on respiration during a 16 hour dark period, since induction could be prevented by the application of various respiratory poisons. Some SDP show an increase i n respiratory rate after exposure to an  223  inductive photoperiod„(82) which may be essential for flowering. ?  In others, respiration follows a distinctive pattern with an increase i n rate during the last 7 hours of a long dark period which differs from the continual respiratory decline shown during a short, non-inductive night (211).  This explanation for the suppressive  effect of 1 or 5% CC^ on flowering of Pharbitis plausible.  i n SD seems, therefore,  I t i s d i f f i c u l t to imagine, however, how the same  mechanism relates to the flower inducing effects of these CO^ concentrations under LD conditions.  Campbell (51) postulated that  the inhibitory effects of 10% CO^ on flowering i n Xanthium pennsylvanieum in SD and the promotion of flowering by the same concentration i n LD, were mediated through separate mechanisms. Under LD conditions he suggested that CO^ may interact with some system to produce a f l o r a l stimulus or, alternatively, that CO^ acts i n some way to prevent the destruction of a l a b i l e stimulus.  I t i s unfortunate  that since the time that those hypotheses were advanced, no other studies have provided evidence to prove  or disprove them.  Another possible mode of action for the effects of 1 or 5% CO^ on development in Pharbitis of ethylene a c t i v i t y (1, 43).  i s via the competitive inhibition  Ethylene can promote flowering in a  number of plants (1). but-Siige (267) has shown that high -1 (100 y l 1 ) can completely inhibit flowering i n Pharbitis normally inductive conditions.  concentrations under  The antagonism between CO^ and ethylene  action represents a possible mechanism for the induction of flowering  224  in Pharbitis  i n LD.  This requires that ethylene i s produced under  LD conditions and i s normally effective i n inhibiting flowering, whereas under inductive photoperiods production i s eliminated or maintained at a low level.  Although this i s a somewhat complex  explanation, the role of ethylene i n flowering i s not well understood and its'.possibler.involvement with very high CC^ concentrations i s worthy of careful consideration. Clearly, i t i s d i f f i c u l t to propose a mechanism which adequately explains the effects of 1 and 5% CO^ on floweringiin Pharbitis  under both inductive and non-inductive photoperiods.  It  i s interesting to note, however, that there i s a close correlation between the responses shown in these experiments, and those caused by low temperature treatments.  In SD, low temperatures applied during  the inductive dark period inhibit flowering (212), whereas i n LD similar temperatures given during the light period cause flower induction (220, 268, 270). of C0  2  Campbell (51) showed that the effectiveness  flushes (10%) i n the control of flowering i n Xanthium  pennsytvanicum depended on whether the gas was supplied during the light or dark period i n either LD or SD.  Application times for  flower promoting or inhibitory effects were found to coincide precisely with those which were effective for low temperature treatments. similar finding for Pharbitis  A  would . largely substantiate the  correlation between these factors. Possibly the effects of very high CO^ concentration and low temperature share a common physiological  225  mechanism, or alternatively, they may operate by altering the rate of reactions involved i n flower induction. In an e a r l i e r part of this discussion, a hypothesis was introduced which stated that 1 and 5% CCXj may act by' releasing development i n Pharbitis  from photoperiodic control.  Such an effect  i s worth consideration since i t could provide a satisfactory explanation for the observed flowering responses under LD and SD conditions, and may also be related to patterns of vegetative growth. I t has been shown for a number of SDP and LDP that when seedlings are grown i n continuous darkness they are capable of producing flower primordia, providing that adequate reserve materials are available (222, 305).  I t i s , therefore, clearly possible to separate f l o r a l  induction from photoperiodism. There are, at present, no available data to suggest any mechanism through which very high could exert these effects.  The appeal  concentrations  of" this explanation i s ,  however, enhanced by the results which showed that flower development and vegetative morphology were very similar under 1 or 5% CO^ and both photoperiods.  As mentioned previously, i f this theory i s  proven correct i t w i l l provide evidence i n favour of a common basis for the CO^ effects on vegetative and reproductive growth.  It w i l l  also, however, raise other questions related to the interaction between these two aspects of development. The results obtained i n the present experiments suggest that flowering may not be easily controlled on a commercial scale  226  by atmospheric CO^-enrichment. At present, the development of many photoperiodically sensitive flower crops, notably  Chrysanthemum,  is regulated by black-out screening of supplementary illumination depending on the time of year and the type of development that i s required (231).  Due to the considerable cost of illuminating  plants during the winter months, there has been some interest i n developing new, cheaper methods of flowering control.  The p r a c t i c a l i t y  of gaseous treatments to maintain plants i n the vegetative condition has been demonstrated using low concentrations of ethylene (241), but .these treatments may also lead fco?a reduction i n growth. Although CC^-enrichment to 0 . 1 % has been used successfully to promote yield of many flower crops i n commercial greenhouses (131), this study has shown that plant development at very high C O 2 concentrations i n SD may normal CO^ concentrations.  '- -  not resemble that shown i n LD at  Further work i s necessary to determine  whether other flower crops show similar responses to those observed in the present study.  Ultimately, an assessment of the potential  of C^ enrichment as a control on flower development requires a -  thorough understanding of the effective concentrations, and the time and duration of their application. Although plant response to very high C O 2 concentrations may hold l i t t l e f l o r i c u l t u r a l significance, the present study has provided important insight into the physiological role of C O 2 i n plant development.  The results have confirmed the modifying effects  of 1 and 5 % C O 2 on flowering and vegetative development which have also  227  been shown to oppose the normal responses to photoperiod in Xanthium pennsylvanioum (51, 234),  and Pseudotsuga menziesii  (247).  For the f i r s t time since the early work of Campbell (51) on Xanthium, i t has been established that very high CO^ concentrations can affect flowering i n another SDP under both inductive and non-inductive photoperiods.  Clearly such effects are important  i n several species in which development i s under photoperiodic control. The pattern of plant response to 1 and 5% COv, established i n this and previous studies, has provided a strong basis for future research to discover the physiological mechanisms which are involved and to assess the significance of these C0 effects under natural conditions. o  GENERAL. DISCUSSION AND CONCLUSIONS  The experiments described i n this thesis have dealt  with a  wide range of CO^ effects on the physiology of plant growth and development.  Information has been provided on atmospheric CC^  concentrations within a commercial greenhouse, thus providing a factual basis for some decisions on the most effective use of natural and a r t i f i c i a l CO^-enrichment systems.  I t has been demonstrated that  stomates i n leaves of a cucumber crop are r e l a t i v e l y insensitive to greenhouse CO^ concentrations as high as 1600 y l 1 ^ (0.16%), but that stomatal resistance varies with leaf age and/or position i n the canopy. In tomato, the benefits of greenhouse CO^-enrichment to 0.09%  ca.  i n terms of reduced time to flowering, and f r u i t yield have been  confirmed.  I t i s clear from the results of these studies that the  supply of supplementary CO^ i s a worthwhile cultural practice for commercial tomato production in B r i t i s h Columbia. Investigations of CO^ exchange rates i n leaves of tomato plants grown at 0.09 - 0.1% C0 , and i n normal a i r (0.03%) have 2  established that plants grown with supplementary CO^ show higher inherent rates of net photosynthesis.  This finding has been confirmed  using both greenhouse and growth chamber-grown plants.  Laboratory  experiments have indicated that these differences i n photosynthetic physiology are at least partly due to changes i n photorespiration and a c t i v i t i e s of the enzymes GaO and RuBP-case.  I t has also been shown,  229  however, that different physiological changes occur when tomato plants are grown at higher CO^ concentrations (0.5%).  These changes  do not result i n higher rates of photosynthesis or lower rates of photorespiration as compared with normal air-grown plants. Moreover, RGR of plants grown i n 0.5% C O 2 correspond with those of plants grown in normal a i r , and contrast with the higher values shown by plants grown i n 0.1% CO^-  The relationship between inherent net photosynthetic  rate and growth C O 2 treatment has been shown to change with leaf age, i n chamber-grown plants. The effects of very high C O 2 concentrations (1 'and 5%) on vegetative and reproductive growth i n Pharbitis investigated.  nil were also  Seedlings grown i n LD or SD for 14 days i n continuous  1 or 5% C O 2 showed greater stem elongation and leaf development than those grown, i n 0.03 or 0.1% C O 2 under the same photoperiods.  Flowering  was induced by 1 and 5% C O 2 under normally non-inductive LD conditions, whereas i n SD the normal flowering response was less vigorous at these concentrations. It i s clear from the results obtained i n each of these studies that the effects of C O 2 concentration on the physiology of plant growth and development are diverse.  U n t i l quite recently, plant physiological  studies concerned with C O 2 have strongly emphasized i t s connection with photosynthetic metabolism. The great majority of previous investigations dealt only with changes i n photosynthetic or photorespiratory rate i n response to a short term change i n C O 2 concentration. The present studies have shown, however, that photosynthetic and  230  photorespiratory metabolism represent only part of the true physiological role of  CO^•  Data obtained from the experiments on tomato and have established two effects of  Pharbitis  concentration on growth and  development which have previously received l i t t l e attention. F i r s t l y , the results presented in Part I I have indicated that photosynthetic adaptation to growth at supra-normal C0^ concentrations can occur i n at least one plant species.  The generality of this response in the  plant kingdom requires further investigation, but the apparent ubiquity of the C 3 CO  assimilation system leads me to speculate that similar  adaptations may be shown by many other plants. Secondly, the experiments concerned with the effects of supplementary CO^ on the growth of Pharbitis  seedlings have demonstrated significant effects of the  gas at concentrations of 1 - 5 % , development.  on vegetative and reproductive  I t i s important to note that these effects do not appear  to be mediated by photosynthesis and are, therefore, quite separate to those discussed in Part I I . The mechanisms involved in the adaptation response of tomato plants to supplementary CO^ during growth have, to some extent, been characterized.  Future research w i l l undoubtedly reveal other  physiological modifications which occur during this adaptation, but the changes induced.! i n photorespiration and enzyme a c t i v i t i e s seem to be basic to the overall process.  On the other hand, insufficient  data areavailable at present to determine the.mechanisms which may be involved in the very high CO^ concentration effects on development.  The  231  apparent lack of correlation between CO^ effects on photosynthesis, and the developmental changes observed i n Pharbitis  should not be  surprising i n view of the number of non-photosynthetic effects of CG"2 on plant growth and development which have been reported i n the literature.  I t does, however, raise the question of whether  correlations exist between those effects and the response of  Pharbitis  seedlings to very high CO^ concentrations. Unlike the effects oh flowering and vegetative morphology i n Pharbitis,  many of the previously reported developmental responses  to CO^ have been shown to occur at r e l a t i v e l y low concentrations.  For  example, i t has been demonstrated that 0.03% CO 2 stimulates the elongation of Avena sativa  coleoptiles i n darkness (40) and root growth  i n tomatoes (260). Similar CO^ concentrations were found to be necessary to i n i t i a t e germination i n lettuce seeds exposed to red i i g h t , and for maximum effectiveness of a l i g h t break i n inhibiting flowering i n Xanthium pennsylvanioum  (13).  A number of mechanisms have been proposed to explain the action of  In these responses. Evans (89) postulated that the  elongation of dark grown coleoptile sections i n CO^-saturated water may be a response to lowered pH.  Some later work, however, clearly  separated the growth stimulating effects of CO^ and pH (236). More recently i t has been proposed that dark fixation reactions may play an important role i n the response of etiolated coleoptiles to (39, .40-; ,80)'. .  232  The competitive inhibition of ethylene action by CO^ (1, 43) has received some attention as a possible mechanism for several non-photosynthetic  effects of CO^ on plant development. For example,  Zeevart (306) has suggested that interactive effects of CO^ and ethylene may be responsible for the CO^ dependency of the l i g h t break phenomenon i n the inhibition of flowering i n Xanthium (13).  Ethylene  has also been shown to inhibit root growth ( 58) i n direct opposition to the stimulatory action of CX^ (260).  I t i s regrettable that no  investigation has been carried out, to date, to assess the importance of ethylene i n other CO^-mediated plant responses. Clearly, more than one of these mechanisms may be responsible for a single effect.  For example, both dark fixation reactions (260)  and the inhibition of ethylene action (58) could account for the stimulatory effect of atmospheric CC^ levels on root growth. The relationships between the known effects of C O 2 on plant development have received l i t t l e attention, however, and i t i s d i f f i c u l t to determine at present whether or not they share common mechanisms. From the data that are available, there appears to be l i t t l e support for a relationship between the effects of 1 and 5% CO^ on flowering and vegetative growth i n Pharbitis, the literature.  and other plant responses reported i n  Most non-photosynthetic  effects of CO^ develop at  atmospheric concentrations (0.03%) or lower. and vegetative development i n Pharbitis,  The effects on flowering  however, are shown at much  higher concentrations (1 - 5%) and normal development takes place under 0.03% C0„ i n LD and SD.  233  It i s tempting to suggest that there are at least 4 classes of CG^ effects viz:  (1) those associated with the requirement for C0  as a substrate for photosynthesis  2  (e.g. the instantaneous response  of photosynthetic rate to increased CO^ concentration);  (2) those  associated with adaptation of CO^ exchange systems to growth at supra-normal levels of CO^ (e.g.  the changes i n photosynthetic and  photorespiratory metabolism demonstrated i n Part I I ) ;  (3) those which  show a requirement for levels of CO^ approximately equal to atmospheric concentrations (ca. 0.01 to 0.03%)'(e.g. the stimulation of coleoptile elongation (40) , stomatal closure (122), and light-break mediated flowering inhibition i n Xanthium (13)); (4) those which result i n modification of growth and development when the CO^ concentration i s raised to approximately 1% (e.g. the effects on flowering i n Pharbitis, and Xanthium  (Part I I I , 51, 234)).  Siline,  Within each category there may be  responses involving different species and different aspects of physiology.  For example, i n addition to the very high C0  effects on flowering i n Pharbitis,  S'iiene, and Xanthium,  2  concentration  1% C0 has 2  been shown to influence budset and winter hardiness i n Douglas F i r seedlings (247).  The mechanisms involved i n the effects of classes (3)  and (4) are uncertain at present, and the p o s s i b i l i t y that some of these responses are mediated through similar physiological reactions cannot be discounted. C0  2  As discussed i n the preceding sections, the interaction of  and ethylene may underly the effects of both classes (2) and (3).  Furthermore, dark C0 fixation has previously been implicated only i n 2  low concentration responses (class 2), although i t has also been observed  234  that changes in protein and organic acid synthesis occur at much higher CG^ concentrations (260).  Such qualitative and quantitative  differences between the products of dark CO^ f i x a t i o n at low (0.03%) and high (1-5%) concentrations may relate to the effects on flowering and vegetative development in Pharbitis  (class 3).  Undoubtedly some  overlapping also occurs between class (1) and several of the other categories.  I t i s generally considered, for example, that the  relationship between stomatal opening and atmospheric CO^ concentration (class 3) depends partly upon the net assimilation response of the guard c e l l chloroplasts (235; class 1).  The fact that this i s not the only  factor involved i s demonstrated by the requirement for C O 2 to induce closure in stomata in complete darkness (123).  As indicated in previous  discussions, the interaction of CO^ and ethylene i s another possible mechanism underlying the effects of both classes 2 and 3. The categorization of CO2 effects creates arbitrary distinctions which may prove to be inaccurate as further studies are carried out.  This approach does, however, underline the varied role of  C O 2 in mediating the known physiological responses of plants.  In  view of t h i s , i t i s interesting to consider how the effects demonstrated in this research relate to current practices in crop production, and whether CX^-control has potential as a tool i n modifying plant growth. Reduction of photorespiration  and the inhibition of  photosynthesis in crop plants would probably result i n significant increases i n productivity (62, 192).  The results of the present studies  have shown that both these processes may be reduced by atmospheric CO —enrichment to 0.1%. The practical significance of these findings  235  rests f i r s t l y on the effectiveness of this method of controlling photorespiration relative to other approaches, and, secondly, on the f e a s i b i l i t y of C0 enrichment for crop plants. To date, attempts to _  2  reduce photorespiration have concentrated on chemical treatments (307, 309), and screening species of crop plants for mutant individuals which show low photorespiratory a c t i v i t y (200, 293). Although some success has been reported for the control of glycolate synthesis by application of glycidate (309), recent results have raised serious doubts about the effectiveness of this chemical in reducing photorespiration (61).  Screening programs have so far met with l i t t l e  success (53, 62, 209) despite the testing of numerous crop species and selections.  In view of these findings, i t seems that (^—enrichment  may be, at present, the most effective means of reducing photorespiration. This method not only results in increases in instantaneous rates of photosynthesis, but also persistent changes i n the photosynthetic system of enriched plants, which improve the efficiency of CO^ fixation. There i s l i t t l e doubt of the f e a s i b i l i t y of CO^-enrichment for greenhouse crops, but the question of whether these procedures are applicable i n the open f i e l d i s more contentious. Many major food crops respond to CO- enrichment by increased dry matter production and y i e l d (66, 164, 208, 296, 302, 135) and, as discussed above, i t seems l i k e l y that they may also exhibit the same physiological modifications shown by tomato, i n the present experiments.  The main  236  problem c l e a r l y l i e s i n the rapid atmospheric d i l u t i o n of supplementary when the gas i s applied i n a f i e l d s i t u a t i o n (208).  Some tests  have shown, however, that the procedure may be f e a s i b l e i n the v i c i n i t y of large natural CT^ sources (296). At present, i t seems that the p r a c t i c a l constraints on the use of CO^ enrichment i n the f i e l d may be too great for i t s -  adoption on a large scale.  These constraints are not encountered i n  crop growing enclosures, but the problems associated with CO^-enrichment in these situations nevertheless  require close attention.  The results  presented i n Part II suggest that, i n vegetable crops, c a r e f u l control of a 0.1% C O 2 l e v e l i s advisable. concentrations  The lack of information on CO^  i n commercial greenhouses i s regrettable, e s p e c i a l l y  since few growers are fortunate enough to have a feedback control system integrated with  enrichment equipment.  Knowledge of  ambient CT^ l e v e l s may be of importance e s p e c i a l l y where  concentrations  can achieve naturally high l e v e l s , as i n cucumber houses using t r a d i t i o n a l straw/soil mixtures i n the cropping beds.  Conductimetric  analysis systems s i m i l a r to that described i n Part I of this thesis, may f i l l present needs for C O 2 monitoring and control equipment i n greenhouses. It i s doubtful from the r e s u l t s of experiments concerned with the effects of CX^ on flowering i n Pharbitis,  whether adequate  control of blooming i n flower crops can be achieved by this method. It may be that other plants w i l l show complete suppression of  237  flowering i n inductive photoperiods at 1 - 5% CC^ (as already demonstrated with Xanthium. at a CC^ concentration of 10%; 51). The practical and economic f e a s i b i l i t y of this technique would, i n that case, rest on the exposure time necessary for effective flowering control. The potential uses and control of CO^ i n agriculture have been considered i n the light of present day crop production techniques and limitations on productivity.  As a f i n a l consideration on the  implication of this research i t i s interesting to note some recent predictions which have stated that the ambient atmospheric CO^ content could increase from the present 0.03%, to 0.06% by the year 2020 (300). One result of the increased C O 2 levels might be a considerable improvement i n crop productivity through an enhancement of photosynthesis. Unfortunately, such predictions ignore the other effects of the high CO^ concentrations on climate and biological systems, which would probably be detrimental to plant growth and the functioning of the entire biosphere. The potential for research into the effects of C O 2 on the physiology of plant growth and development i s enormous. The work described i n this thesis has provided some idea of the complex physiological role played by the gas, and yet has not considered many plant responses to C O 2 which have become evident through recent research.  I t i s my hope that this study w i l l provide a background  and incentive for future work into what i s a fascinating, yet relatively unknown area of plant physiology.  238  LITERATURE CITED 1.  ABELES, F.B. 1973. New York. 302  Ethylene i n Plant Biology. pp.  2.  ALLARD, H.A. and W.W. GARNER. 1941. Responses of some p l a n t s to equal and unequal r a t i o s of l i g h t and darkness i n c y c l e s r a n g i n g from 1 hour to 72 hours. J . A g r i c . Res. 63:305-330.  3.  ALLEN, P.G. 1973. Carbon d i o x i d e enrichment. In: Kingham H.G. (editor). The U.K. Tomato Manual. Grower Books, London, pp. 156-162.  4.  ANON. 1976. Greenhouse tomato and cucumber p r o d u c t i o n guide. B r i t i s h Columbia M i n i s t r y of A g r i c u l t u r e . 20 pp.  5.  ARTHUR, J.M., J.D. GUTHRIE and J.M. NEWELL. 1930. Some e f f e c t s of a r t i f i c i a l c l i m a t e s on the growth and c h e m i c a l c o m p o s i t i o n of p l a n t s . Amer. J . Bot. 17:416-482. .  6.  ASLAM, M., S.B. LOWE and L.A. HUNT. 1977. E f f e c t of l e a f age on p h o t o s y n t h e s i s and t r a n s p i r a t i o n of cassava (Manihot esculenta) Can. J . Bot. 55:2288-2295.  7.  ATKINS, C.A., D.T. CANVIN and H. FOCK. 1971. Intermediary metabolism of p h o t o s y n t h e s i s i n r e l a t i o n to carbon d i o x i d e e v o l u t i o n i n sunflower. In: Hatch, M.D., C B . Osmond and R.O. S l a t y e r ( e d i t o r s ) . P h o t o s y n t h e s i s and P h o t o r e s p i r a t i o n . W i l e y - I n t e r s c i e n c e , New York. pp. 497-505.  8.  AULENBACH, W. 1965. chrysanthemums.  9.  BAHR, J.T. and R.G. JENSEN. 1974. R i b u l o s e biphosphate oxygenase a c t i v i t y from f r e s h l y r u p t u r e d s p i n a c h c h l o r o p l a s t s . Arch. Biochem. Biophys. 164:408-413.  S o i l s , temperature, and C 0 Penn. Flower Growers B u l l .  Academic P r e s s ,  2  f o r cut 174.  10.  BAKER, A.L. and N.E. TOLBERT. 1966. G l y c o l a t e oxidase (Ferredoxinc o n t a i n i n g form). In: Wood, W.A. (editor). Methods i n Enzymology. IX. Carbohydrate Metabolism. Academic P r e s s , New York. pp. 338-342.  11.  BALLARD, L.A.T. 1941. The depressant e f f e c t of carbon d i o x i d e upon p h o t o s y n t h e s i s . New P h y t o l . 40:276-290.  12.  BASSI, P.K., E.B. TREGUNNA and P.A. JOLLIFFE. 1976. Carbon d i o x i d e exchange and phytochrome c o n t r o l of f l o w e r i n g i n Xanthium pennsylvanicum. Can. J . Bot. 54:2881-2887.  13.  BASSI, P.K. E.B. TREGUNNA and A.N. PUROHIT. 1975. Carbon d i o x i d e requirements f o r phytochrome a c t i o n i n p h o t o p e r i o d i s m and seed g e r m i n a t i o n . P l a n t P h y s i o l . 56:335-336.  239 14.  VAN BAVEL, C.H.M. 1975. A b e h a v i o r a l e q u a t i o n f o r l e a f d i o x i d e a s s i m i l a t i o n and a t e s t of i t s v a l i d i t y . P h o t o s y n t h e t i c a 9:165-176.  carbon  15.  BAVRINA, T.V., A.P. AKSENOVA and T.N. KONSTANTINOVA. 1969. The p a r t i c i p a t i o n of p h o t o s y n t h e s i s i n photoperiodism. Soviet P l a n t P h y s i o l . 16:315-327.  16.  BEGG, J . E . and J.V. LAKE. 1968. Carbon d i o x i d e measurement: A continuous c o n d u c t i m e t r i c method. Agr. M e t e o r o l . 5:283-290.  17.  VAN BERKEL, N. 1967. Some t e c h n i c a l a s p e c t s of CO Proc. XVII I n t . Hort. Cong. I I I . :333-341.  18.  BEURLEIN,,J.E. and J.W. PENDLETON. 1971. P h o t o s y n t h e t i c r a t e s and l i g h t s a t u r a t i o n curves of i n d i v i d u a l soybean l e a v e s under f i e l d c o n d i t i o n s . Crop S c i . 11:217-219.  19.  BIERHUIZEN, J . F . and R.O. SLATYER. 1964. P h o t o s y n t h e s i s of c o t t o n l e a v e s under a range of environmental c o n d i t i o n s i n r e l a t i o n to i n t e r n a l and e x t e r n a l d i f f u s i v e r e s i s t a n c e s . Aust. J . B i o l . S c i . 17:348-359.  20.  BILLINGS, W.D., P.J. GODFREY, B.F. CHABOT, and D.P. BOURQUE. 1971. M e t a b o l i c a c c l i m a t i o n to temperature i n a r c t i c and a l p i n e ecotypes of Oxyvia digyna. A r c t i c and A l p i n e Res. 3:277-289.  21.  BISHOP, P.M. and C P . WHITTINGHAM. 1968. The p h o t o s y n t h e s i s of tomato p l a n t s i n a carbon d i o x i d e e n r i c h e d atmosphere. P h o t o s y n t h e t i c a 2:31-38.  22.  BJORKMAN, 0. 1966. The e f f e c t of oxygen c o n c e n t r a t i o n on p h o t o s y n t h e s i s i n h i g h e r p l a n t s . P h y s i o l . Plantarum 19:618-633.  23.  BJORKMANN, 0. 1968. Carboxydismutase a c t i v i t y i n shade-adapted and sun-adapted s p e c i e s of h i g h e r p l a n t s . P h y s i o l . P l a n t 21:1-10.  24.  BJORKMAN, 0., N.K. B0ARDMAN, J.M. ANDERSON, S.W. TH0RNE, D.J. G00DCHILD and N.A. PYLI0TIS. 1972. E f f e c t of l i g h t i n t e n s i t y d u r i n g growth of Atviptex patula on the c a p a c i t y of p h o t o s y n t h e t i c r e a c t i o n s , c h l o r o p l a s t c o m p o s i t i o n and s t r u c t u r e . Carnegie I n s t . Yearbook. 71:115-135. Carnegie I n s t . Wash.  25.  BJORKMAN, 0., E. GAUHL, W.M. HIESEY, F. NICHOLSON and M.A. NOBS. 1969. Growth of Mimulus, Marahantia and Zea under d i f f e r e n t oxygen and carbon d i o x i d e l e v e l s . Carnegie I n s t . Yearbook. 67:477-479. Carnegie I n s t . Wash.  26.  BJORKMAN, 0. and R.W. PEARCY. 1971. E f f e c t s of growth temperature on the temperature dependence of p h o t o s y n t h e s i s in vivo and on CO^ f i x a t i o n by carboxydismutase in vivo i n C^ and C^ s p e c i e s . Carnegie I n s t . Yearbook. 70:511-520. Carnegie I n s t . Wash.  enrichment.  240 27.  BLACKMAN, F.F. and A.M. SMITH. 1911. E x p e r i m e n t a l r e s e a r c h e s on v e g e t a b l e a s s i m i l a t i o n and r e s p i r a t i o n . IX. On a s s i m i l a t i o n i n submerged w a t e r - p l a n t s , and i t s r e l a t i o n to the c o n c e n t r a t i o n o f carbon d i o x i d e and other f a c t o r s . P r o c . Roy. Soc. London. B83:389-412.  28.  BLACKMAN, G.E. and J.M. BLACK. 1959. P h y s i o l o g i c a l and e c o l o g i c a l s t u d i e s i n the a n a l y s i s o f p l a n t environment. X I I : The r o l e of the l i g h t f a c t o r i n ' l i m i t i n g growth. Ann. Bot. n.s. 23:131-145.  29.  BLENKINSOP, P.G. and J.E. DALE. 1974. The e f f e c t s o f shade treatment and l i g h t i n t e n s i t y on r i b u l o s e - 1 , 5 - d i p h o s p h a t e c a r b o x y l a s e a c t i v i t y and f r a c t i o n I p r o t e i n l e v e l i n the f i r s t l e a f of barley. J . Exp. Bot 25:899-912.  30.  BOARDMAN, N.K. 1977. Comparative p h o t o s y n t h e s i s o f sun plants. Ann. Rev. P l a n t P h y s i o l . 28:355-377.  31.  B0DS0N, M., R.W. KING, L.T. EVANS and G. BERNIER. 1977. The r o l e of p h o t o s y n t h e s i s i n f l o w e r i n g o f the long day p l a n t Sinapis alba. A u s t . J . P l a n t P h y s i o l . 4:467-478.  32.  BOLAS, B.D. and R. MELVILLE. 1935. The e f f e c t on the tomato p l a n t of carbon d i o x i d e produced by combustion. Ann. A p p l . B i o l . 22:1-15.  33.  BOLIN, B.  34.  BORTHWICK, H. and M.W. PARKER. 1938. P h o t o p e r i o d i c B i l o x i soybeans. Bot. Gaz. 100:374-387.  35.  BOWES, G. and J.A. BERRY. 1972. The e f f e c t o f oxygen on p h o t o s y n t h e s i s and g l y c o l a t e e x c r e t i o n i n Chlamydomonas reinhardtii. Carnegie I n s t . Yearbook. 71:148-158. Carnegie I n s t . Wash.  36.  BOWES, G. and W.L. OGREN. 1972. 0 i n h i b i t i o n and other p r o p e r t i e s of soybean r i b u l o s e 1,5-diphosphate c a r b o x y l a s e . Biochem. B i o p h y s . Res. Commun. 45:716-722.  37.  BOWES, G., W.L. OGREN and R.H. HAGEMAN. 1971. P h o s p h o g l y c o l a t e p r o d u c t i o n c a t a l y z e d by r i b u l o s e diphosphate c a r b o x y l a s e . Biochem. Biophys. Res. Comm. 45:716-722.  38.  BOWES, G., W.L. OGREN and R.H. HAGEMAN. 1972. L i g h t s a t u r a t i o n , p h o t o s y n t h e s i s r a t e , RudP c a r b o x y l a s e a c t i v i t y , and s p e c i f i c l e a f weight i n soybeans grown under d i f f e r e n t l i g h t i n t e n s i t i e s . Crop S c i . 12:77-79.  39.  BOWN, A.W. and T. AUNG. 1974. The i n f l u e n c e o f 0.03% d i o x i d e on p r o t e i n metabolism o f e t i o l a t e d Avena coleoptiles. P l a n t P h y s i o l . 54:19-22.  40.  BOWN, A.W., I . J . DYMOCK and T. AUNG. 1974. A s y n e r g i s t i c s t i m u l a t i o n of Avena sativa c o l e o p t i l e e l o n g a t i o n by i n d o l e a c e t i c a c i d and carbon d i o x i d e . P l a n t P h y s i o l . 54:15-18.  1970. The carbon c y c l e .  and shade  S c i . Amer. 223(3):125-158. perception i n  2  carbon  sativa  241 41.  BROWN, H.T. and F. ESCOMBE. 1902. The i n f l u e n c e o f v a r y i n g amounts of carbon d i o x i d e i n the a i r on the p h o t o s y n t h e t i c p r o c e s s o f l e a v e s and on the mode o f growth o f p l a n t s . P r o c . R o y a l Soc. 70: 347-413.  42.  BRULFERT, J . 1965. Etude e x p e r i m e n t a l e du developpement  et f l o r a l chez Anagallis  arvensis  vegetatif  L. spp. phoen-ioea Scop.  Formation de f l e u r s p r o l i f e r e s chez c e t t e meme espece. Rev. Gen. de Bot. 72:641-694. 43.  BURG, S.P. and E.A. BURG. 1965. E t h y l e n e a c t i o n and the r i p e n i n g of f r u i t s . S c i e n c e . 148:1190-1195.  44.  BURNSIDE, C.A. and R.H. BONNING. 1957. The e f f e c t o f prolonged shading on the l i g h t s a t u r a t i o n curves o f apparent p h o t o s y n t h e s i s i n sun p l a n t s . P l a n t P h y s i o l . 32:61-63.  45.  BUSSINGER, J.A. 1963. The g l a s s h o u s e c l i m a t e . I n : Van Wijk, W.R. (editor). The P h y s i c s o f P l a n t Environment, pp. 277-318.  46.  CALVERT, A. 1959. E f f e c t o f the e a r l y environment on the development of f l o w e r i n g i n tomato. I I . L i g h t and temperature i n t e r a c t i o n s . J . H o r t . S c i . 34:154-162.  47.  CALVERT, A. 1972. E f f e c t s o f day and n i g h t temperatures and carbon d i o x i d e enrichment on y i e l d o f g l a s s h o u s e tomatoes. J . Hort. S c i . 47:231-247.  48.  CALVERT, A. 1973. E n v i r o n m e n t a l r e s p o n s e s . I n : Kingham, H.G. (editor). The U.K. Tomato Manual. Grower Books, London, pp. 23-34.  49.  CALVERT, A. and G. SLACK. 1975. E f f e c t s o f carbon d i o x i d e enrichment on growth, development and y i e l d of g l a s s h o u s e tomatoes. I . Responses t o c o n t r o l l e d c o n c e n t r a t i o n s . J . Hort. S c i . 50:61-71.  50.  CALVERT, A. and G. SLACK. 1976. E f f e c t o f carbon d i o x i d e enrichment on growth, development and y i e l d o f g l a s s h o u s e tomatoes. I I . The d u r a t i o n o f d a i l y p e r i o d s o f enrichment. J . Hort. S c i . 51:401-409.  51.  CAMPBELL, CW. 1957. M o d i f i c a t i o n o f l i g h t c o n t r o l i n p l a n t development. Ph. D. T h e s i s . Purdue U n i v e r s i t y .  52.  CANHAM, A.E. 1974. Some e f f e c t s o f C 0 , a i r temperature and supplementary a r t i f i c i a l l i g h t on the growth o f young tomato p l a n t s . A c t a H o r t . 39:175-183.  53.  CANNELL, R.Q., W.A. BRUN and D.N. MOSS. 1969. A s e a r c h f o r h i g h net p h o t o s y n t h e t i c r a t e among soybean genotypes. Crop S c i . 9:840-841.  54.  CAPRON, T.M. and T.A. MANSFIELD. 1976. I n h i b i t i o n o f net p h o t o s y n t h e s i s i n tomato i n a i r p o l l u t e d w i t h n i t r i c o x i d e and n i t r o g e n d i o x i d e . J . Exp. Bot. 27:1181-1186.  2  242 55.  CAPRON, T.M. and T.A. MANSFIELD. 1977. I n h i b i t i o n of growth i n tomato by a i r p o l l u t e d w i t h n i t r o g e n o x i d e s . J . Exp. B o t . 28:112-116.  56.  CARR, D.J. 1957. On the n a t u r e o f p h o t o p e r i o d i c i n d u c t i o n . IV. P r e l i m i n a r y experiments on the e f f e c t of l i g h t f o l l o w i n g t h e i n d u c t i v e long dark p e r i o d i n Xanthium :pennsylvaniaum. Physiol. P l a n t . 10:249-265.  57.  CHABOT, B.F., J . F . CHABOT and W.D. BILLINGS. 1972. R i b u l o s e - 1 , 5-diphosphate c a r b o x y l a s e a c t i v i t y i n a r c t i c and a l p i n e p o p u l a t i o n s of Oxyria digyna. Photosynthetica 6:364-369.  58.  CHADWICK, A.V. and S.P. BURG. 1967. An e x p l a n a t i o n of t h e i n h i b i t i o n o f r o o t growth caused by i n d o l e a c e t i c a c i d . P h y s i o l . 42:415-420.  Plant  59.  CHANG, C.W. 1975. Carbon d i o x i d e and senescence i n c o t t o n P l a n t P h y s i o l . 55:515-519.  60.  CHAPMAN, H.W. and W.E. LOOMIS. 1953. P h o t o s y n t h e s i s i n the p o t a t o under f i e l d c o n d i t i o n s . P l a n t P h y s i o l . 28:703-716.  61.  plants.  14 CHOLLET, R. 1978. E v a l u a t i o n of l i g h t / d a r k C assay of Photorepiration. P l a n t P h y s i o l . 61:929-932.  62.  CHOLLET, R. and W.L. OGREN. 1975. R e g u l a t i o n of p h o t o r e s p i r a t i o n i n C and C s p e c i e s . B o t . Rev. 41:137-179. 3 4  63.  CLAUSSEN, W. and E. BILLER. 1977. D i e bedeutung der s a c c h a r o s e und s t a r k e g e h a l t e der b l a t t e r f u r d i e r e g u l i e r u n g der h e l t o - photosyntheseraten. Z. P f l a n z e n p h y s i o l . . Bd". 81: 189-198.  64.  COLEMAN, W.K. and R.I. GREYSON.  of the l e a f i n tomato  1976.  (Lycopersicon  The growth and development  esaulentum).  I . The  p l a s t o c h r o n index, a s u i t a b l e b a s i s f o r d e s c r i p t i o n . Bot. 54:2421-2428.  Can. J .  65.  COOMBS, J . and C P . WHITTINGHAM. 1966. The mechanism of i n h i b i t i o n of p h o t o s y n t h e s i s by h i g h p a r t i a l p r e s s u r e s of oxygen i n ChloreHa. P r o c . Roy. Soc. London. B 164:511-520.  66.  COOPER, R.L. and W.A. BRUN. carbon d i o x i d e - e n r i c h e d  67.  CR00KST0N, R.K. K . J . TREHARNE, P. LUDFORD and J . L . 0ZBUN. 1975. Response o f beans to shading. Crop S c i . 15:412-416.  68.  GUMMING, B.G. 1967. C i r c a d i a n rhythm responses i n Chenopodium rubvum: e f f e c t s of g l u c o s e and s u c r o s e . Can J . B o t . 45: 2173-2193.  69.  CUMMINGS, M.B. and C H . JONES. 1918. The a e r i a l f e r t i l i z a t i o n of p l a n t s w i t h carbon d i o x i d e . Vermont S t a t . B u l l . 211  1967. Response o f soybeans to a atmosphere. Crop S c i . 7:455-457.  243 70.  CUMMINGS, M.B. and C H . JONES. 1920. The a e r i a l f e r t i l i z a t i o n o f p l a n t s w i t h carbon d i o x i d e . J . Chem. Soc. 118:267.  71.  CURTIS, P.E., W.L. OGREN and R.H. HAGEMAN. 1969. V a r i e t a l e f f e c t s i n soybean p h o t o s y n t h e s i s and p h o t o r e s p i r a t i o n . Crop. S c i . 9:323-327.  72.  DAUNICHT, H.J. 1966. Methods and r e s u l t s o f experiments on the e f f e c t o f C0„ c o n c e n t r a t i o n on v e g e t a b l e c r o p s . Acta Hort. 4:116-125.  1  73.  DAVIS, S.D., C.H.M. VAN BAVEL and K.J. McCREE. 1977. E f f e c t o f l e a f aging upon s t o m a t a l r e s i s t a n c e i n bean p l a n t s . Crop. S c i . 17:640-644.  74.  DEMOUSSY, E. 1904. Sur l a v e g e t a t i o n dans des atmospheres r i c h e s en a c i d e carbonique. Comptes Rendus Acad. S c i . P a r i s 139: 883-885.  75.  D0R0KH0V, L.M. 1938. Dynamics o f c a r b o h y d r a t e a c c u m u l a t i o n i n p l a n t l e a v e s as i n f l u e n c e d by d i f f e r e n t c o n t e n t s o f CO i n a i r . C R . (Doklady) Acad. S c i . URSS 21 :72-76.  76.  DOWNES, R.W. 1971. A d a p t a t i o n o f sorghum p l a n t s t o l i g h t i n t e n s i t y : i t s e f f e c t on gas exchange i n response t o changes i n l i g h t , temperature and CO^- I n : Hatch, M.D., C.B. Osmond and R.O. S l a t y e r ( e d i t o r s ) . P h o t o s y n t h e s i s and p h o t o r e s p i r a t i o n . W i l e y - I n t e r s c i e n c e , New York. pp. 57-62.  77.  DOWNTON, J . and R.O. SLATYER. 1971. V a r i a t i o n s i n l e v e l s o f some l e a f enzymes. P l a n t a ( B e r l . ) 96:1-12.  78.  DOWNTON, W.J.S. and E.B. TREGUNNA. 1968. Carbon d i o x i d e compensation - i t ' s r e l a t i o n to p h o t o s y n t h e t i c c a r b o x y l a t i o n r e a c t i o n s , s y s t e m a t i c s o f the Gvaminae, and l e a f anatomy. Can. J . B o t . 46:207-215.  79.  DOWNTON, W.J.S. and E.B. TREGUNNA. 1968. P h o t o r e s p i r a t i o n and g l y c o l a t e metabolism: A r e - e x a m i n a t i o n and c o r r e l a t i o n o f some p r e v i o u s s t u d i e s . P l a n t P h y s i o l . 43:923-929.  80.  DYMOCK, I . J . , B. HILL and A.W. BOWN. 1977. An i n v e s t i g a t i o n i n t o the i n f l u e n c e o f IAA and malate on in vivo and in vitro r a t e s o f dark carbon d i o x i d e f i x a t i o n i n c o l e o p t i l e t i s s u e . Can. J . B o t . 55:1641-1645.  81.  EGLE, K. and H. FOCK. 1967. L i g h t r e s p i r a t i o n - c o r r e l a t i o n s between CO^ f i x a t i o n , 0^ p r e s s u r e and g l y c o l l a t e c o n c e n t r I n : Goodwin, T.W. ( e d i t o r ) : B i o c h e m i s t r y o f C h l o r o p l a s t s . V o l I I . Academic P r e s s , New York. pp. 79-87.  82.  ELLIOT, B.B. and A . C LEOPOLD. 1952. A r e l a t i o n s h i p between p h o t o p e r i o d i s m and r e s p i r a t i o n . P l a n t P h y s i o l . 27:787-793.  \  244 83.  ELLYARD, P.W. and M. GIBBS. 1969. by oxygen i n i s o l a t e d s p i n a c h 1115-1121.  I n h i b i t i o n of p h o t o s y n t h e s i s c h l o r o p l a s t s . P I . P h y s i o l . 44:  84.  EL-SHARKAWY, M.A., R.S. LOOMIS and W.A. WILLIAMS. 1968. P h o t o s y n t h e t i c and r e s p i r a t o r y exchange of carbon d i o x i d e by l e a v e s of the g r a i n Amaranth. J . A p p l . E c o l . 5:243-251.  85.  ENOCH, H.Z., N. ZIESLIN, Y. BIRAN, A.H. HALEVY, M. SCHWARZ, B. KESLER and D. SHIMSI. 1973. P r i n c i p l e s of C 0 nutrition research. A c t a H o r t . 32:97-118. 2  86.  EPSTEIN, E. 1972. 412 pp.  87.  ERICKSON, R.O. and Amer. J . Bot.  88.  EVANS, L.T. 1963. E x t r a p o l a t i o n from c o n t r o l l e d environments to the f i e l d . In: Evans, L.T. ( e d i t o r ) . Environmental C o n t r o l of P l a n t Growth. Academic P r e s s , New York. pp. 421-438.  89.  EVANS, M. 1967. K i n e t i c s t u d i e s of the c e l l e l o n g a t i o n phenomenon i n Avena c o l e o p t i l e segments. Ph. D. T h e s i s : U n i v e r s i t y of C a l i f o r n i a a t Santa Cruz, Santa Cruz.  90.  EVANS, M., P.M. RAY and L. REINHOLD. 1971. I n d u c t i o n of c o l e o p t i l e e l o n g a t i o n by carbon d i o x i d e . P l a n t P h y s i o l . 47:335-341.  91.  FAIR, P., J . TEW and C F . CRESSWELL. 1973. Enzyme a c t i v i t i e s a s s o c i a t e d w i t h carbon d i o x i d e exchange i n i l l u m i n a t e d l e a v e s of Hovdeum vulgave L. I . E f f e c t s of l i g h t p e r i o d , l e a f age and §§fXii?n ° ^ d i o x i d e compensation p o i n t (P) . Ann. Bot.;'37: n  c  a  M i n e r a l n u t r i t i o n of p l a n t s .  F . J . MICHELINI. 44:209-296.  r  o  1957.  The  W i l e y , New  plastochron  York.  index.  n  92.  FAIR, P., J . TEW and C F . CRESSWELL. 1973. Enzyme a c t i v i t i e s a s s o c i a t e d w i t h carbon d i o x i d e exchange i n i l l u m i n a t e d l e a v e s of Hovdeum vulgave L. I I . E f f e c t s of e x t e r n a l c o n c e n t r a t i o n s of carbon d i o x i d e and oxygen. Ann. Bot. 37: 1035-1039.  93.  FISCHER, R.A. 1967. Stomatal p h y s i o l o g y w i t h p a r t i c u l a r r e f e r e n c e to the a f t e r e f f e c t of water s t r e s s and to behavior i n epidermal s t r i p s . Ph. D. t h e s i s . U n i v . of C a l i f . , D a v i s .  94.  FOCK, H. and G. KROTKOV. 1969. R e l a t i o n s h i p between p h o t o r e s p i r a t i o n and g l y c o l a t e o x i d a s e a c t i v i t y in" sunflower and red kidney bean leaves. Can. J . Bot. 47:237-240.  95.  FORD, M.A. and G.W. THORNE. 1967. E f f e c t of C 0 c o n c e n t r a t i o n on growth of sugar-beet, b a r l e y , k a l e and maize. Ann. Bot. ns. 31: 629-644.  96.  FORRESTER, M.L., G. KROTKOV and C D . NELSON. 1966. E f f e c t of oxygen on p h o t o s y n t h e s i s and p h o t o r e s p i r a t i o n i n detached l e a v e s . I. Soybean. P l a n t P h y s i o l . 41:422-427.  2  245 97.  FRASER, D.E. and R.G.S. BIDWELL. 1974. P h o t o s y n t h e s i s and p h o t o r e s p i r a t i o n d u r i n g the ontogeny of the bean p l a n t . Can. J . Bot. 52:2561-2570.  98.  FREDERICK, S.E., P.J. GRUBER and N.E. TOLBERT. 1973. The o c c u r r e n c e of g l y c o l a t e dehydrogenase and g l y c o l a t e o x i d a s e i n green p l a n t s . An e v o l u t i o n a r y survey. P l a n t P h y s i o l . 52:318-323.  99.  FREUDENBERGER, H. 1940. D i e r e a k t i o n der S c h l i e s s z e l l e n auf Kohlensaure und s a u e r s t o f f entzug. Protoplasma. 35:15-54.  Brassioa  100.  FRIEND, D.J.C. 1968. P h o t o p e r i o d i c responses of cv. Ceres. P h y s i o l . P l a n t . 21:990-1002.  101.  FRIEND, D.J.C. 1975. L i g h t requirements f o r p h o t o p e r i o d i c s e n s i t i v i t y i n c o t y l e d o n s of dark-grown Pharbitis nil. P l a n t . 35:286-296.  oampestris  Physiol.  102.  FRYDRYCH, J . 1976. P h o t o s y n t h e t i c c h a r a c t e r i s t i c s of cucumber s e e d l i n g s grown under two l e v e l s of carbon d i o x i d e . P h o t o s y n t h e t i c a . 10:335-338.  103.  GAASTRA, P. 1959. P h o t o s y n t h e s i s of crop p l a n t s as i n f l u e n c e d by l i g h t , carbon d i o x i d e , temperature and s t o m a t a l d i f f u s i o n resistance. Meded. Landbou. Wageningen 59:1-68.  104.  GAASTRA, P. 1966. Some p h y s i o l o g i c a l a s p e c t s of CC^ a p p l i c a t i o n i n g l a s s h o u s e c u l t u r e . A c t a . H o r t . 4:111-116.  105.  GARNER, W.W. and H.A. ALLARD. 1920. E f f e c t of the r e l a t i v e l e n g t h of day and n i g h t and other f a c t o r s of the environment on growth and r e p r o d u c t i o n i n p l a n t s . J . A g r i c . Res. 18:553-606.  106.  GARNER, W.W. and H.A. ALLARD. 1923. F u r t h e r s t u d i e s on p h o t o p e r i o d i s m , the response of the p l a n t to r e l a t i v e of day and n i g h t . J . A g r i c . Res. 23:871-920.  length  107.  GLIDWELL, S.M. and J.A. RAVEN. 1975. Measurement of simultaneous oxygen e v o l u t i o n and uptake i n Hydrodiotyon afriaanum. J . Exp. Bot. 26:479-488.  108.  GLIDWELL, S.M. and J.A. RAVEN. 1976. P h o t o r e s p i r a t i o n : R i b u l o s e diphosphate oxygenase or hydrogen p e r o x i d e ? J . Exp. Bot. 27: 200-204.  109.  GOLDSBERRY, K.L. 1965. E f f e c t s of C 0 Growers Assn. B u l l . 187.  110.  GOLDSWORTHY, A. 501-502.  111.  GRAHL, H. and A. WILD. 1972. D i e v a r i a b i l i t a t der g r b s s e der photosyntheseeinheit b e i licht-und schattenpflanzen. Untersuchungen zur photosynthese von e x p e r i m e n t e l l i n d u z i e r t e n l i c h t - u n d s c h a t t e n t y p e n von Sinapis alba. Z. P f l a n z e n p h y s i o l . 67:443-453.  1969.  2  on p o i n s e t t i a .  R i d d l e of p h o t o r e s p i r a t i o n .  Colo.  Flower  Nature 224:  246  112.  HAALAND, E. 1.974. The effect of the growing conditions of the stock plants (Campanula isophylla) on the sucrose metabolism i n the cuttings during rooting. Root I n i t i a t i o n Symposium, Copenhagen. Sept. 2-4, 1974. (Cited by E. MADSEN, 1976 - see r e f . 190.)  113.  HAMNER, K.C. 1940. Interrelation of l i g h t and darkness i n photoperiodic induction. Bot. Gaz. 102:658-687.  114.  HAND, D.W. 1973. A i r p o l l u t i o n i n glasshouses and the effects of a e r i a l pollutants on crops. Scient. Hort. 24:142-157.  115.  HAND, D.W. and J.D. POSTLETHWAITE. 1971. The response to C0 enrichment of c a p i l l a r y watered single-truss tomatoes at d i f f e r e n t plant densities and seasons. J . Hort. S c i . 46: 461-470.  116.  HAND, D.W. and R.W. SOFFE. 1971. Light modulated temperature control and the response of greenhouse tomatoes to d i f f e r e n t CO regimes. J . Hort. S c i . 46:381-396.  117.  HARDER, R. 1941. Uber d i e bedeutung der kohlensaure und der photoperiodische belichtung fur die blutenbildung b e i Kalanahoe blossfeldiana. Naturwiss. 29:770-771.  118.  HATCH, M.D., CR. SLACK and T.A. BULL. 1969. Light-induced changes in the content of some enzymes of the C^-dicarboxylic acid pathway of photosynthesis and i t s effect on other c h a r a c t e r i s t i c s of photosynthesis. Phytochem. 8:697-706.  119.  HEATH, O.V.S. 1948. Control of stomatal movement by a reduction in the normal carbon dioxide content of the a i r . Nature 161: 179-181.  120.  HEATH, O.V.S. 1950. Studies i n stomatal behavior. V. The r o l e of carbon dioxide i n the l i g h t response of stomata, Part 1. Investigation of the cause of abnormally wide stomatal opening within porometer cups. J . Exp. Bot. 1:29-62.  121.  HEATH, O.V.S. and H. MEIDNER. 1967. Compensation points and " carbon dioxide enrichment for lettuce grown under glass i n winter. J . Exp. Bot. 18:746-751.  122.  HEATH, O.V.S. and F.L. MILTHORPE. 1950. Studies i n stomatal • behavior. V. The r o l e of carbon dioxide i n the l i g h t response of stomata. Part I I . Preliminary experiments on the i n t e r r e l a t i o n s of l i g h t intensity, carbon dioxide concentration, and rate of a i r flow i n controlling the movement of wheat stomata. J . Exp. Bot. 1:227-243.  123.  HEATH, O.V.S. and J . RUSSELL. 1954. Studies i n stomatal behavior. VI. An investigation of the l i g h t responses of wheat stomata with the attempted elimination of control by the mesophyll. Part 2. Interactions with external carbon dioxide, and general discussion. J . Exp. Bot. 5:269-292.  2  247 124.  HEDLEY, L.C.L. and D.M. HARVEY. 1975. The involvement of CC> uptake i n the f l o w e r i n g behavior of two v a r i e t i e s of Antirrhinum majus. In: M a r c e l l e , R. ( e d i t o r ) . Environmental and B i o l o g i c a l C o n t r o l of P h o t o s y n t h e s i s . Dr. W. Junk B.V., The Hague. 408 pp.  125.  HESKETH, J.D. 1963. L i m i t a t i o n s to p h o t o s y n t h e s i s r e s p o n s i b l e f o r d i f f e r e n c e s among s p e c i e s . Crop S c i . 3:493-496.  126.  HEW,  127.  HIGHKIN, H.R. and J.B. HANSON. 1954. P o s s i b l e i n t e r a c t i o n s between l i g h t - d a r k c y c l e s and endogenous d a i l y rhythms on the growth of tomato p l a n t s . P l a n t P h y s i o l . 29:301-302.  128.  HILLMAN, W.S. 1956. I n j u r y of tomato p l a n t s by continuous l i g h t and u n f a v o u r a b l e p h o t o p e r i o d i c c y c l e s . Amer. J . Bot. 43:89-96.  129.  HILLMAN, W.S. 1969. P h o t o p e r i o d i s m and v e r n a l i z a t i o n . In: W i l k i n s , M.B. (editor). The P h y s i o l o g y of P l a n t Growth and Development. McGraw-Hill, London, pp. 559-601.  130.  HOFSTRA, G. and J.D. HESKETH. 1975. The e f f e c t s of temperature and C0„ enrichment on p h o t o s y n t h e s i s i n soybean. I n : M a r c e l l e , R. (editor). Environmental and B i o l o g i c a l C o n t r o l of P h o t o s y n t h e s i s , Dr. W. Junk B.V., The Hague. 408 pp.  131.  H0LLEY, W.D. 1970. C0 enrichment f o r flower Amer. Soc. A g r i . Eng. 13:257-258.  132.  HOLMGREN, P. and P.G. JARVIS. 1967. Carbon d i o x i d e e f f l u x from l e a v e s i n l i g h t and darkness. P h y s i o l . P l a n t . 20:1045-1051.  133.  HOLMGREN, P., P.G. JAMES and M.S. JARVIS. 1965. R e s i s t a n c e to carbon d i o x i d e and water vapour t r a n s f e r i n l e a v e s of d i f f e r e n t plant species. P h y s i o l . P l a n t . 18:557-573.  134.  HOOVER, W.H., E.S. JOHNSON and F.S. BRACKETT. 1933. Carbon d i o x i d e a s s i m i l a t i o n i n a higher p l a n t . Smithsonian M i s c . P u b l . 87:1-19.  135.  HOPEN, H.J. and S.K. RIES. 1962. The m u t u a l l y compensating e f f e c t of carbon d i o x i d e c o n c e n t r a t i o n s and l i g h t i n t e n s i t i e s on the  2  C-S., G. KROTKOV and D.T. CANVIN. 1969. D e t e r m i n a t i o n of the r a t e of CO e v o l u t i o n by green l e a v e s i n l i g h t . Plant Physiol. 44:662-6707  2  growth of Cuoumis sativus  production.  Trans.  L. Proc. Amer. Soc. Hort. S c i . 81:  358-364. 136.  HUGHES, A.P. and K.E. COCKSHULL. 1971. The v a r i a t i o n i n response to l i g h t i n t e n s i t y and carbon d i o x i d e c o n c e n t r a t i o n shown by two c u l t i v a r s of Chrysanthemum morifotium grown i n c o n t r o l l e d environments at two times of year. Ann. Bot. ns. 35:933-945.  137.  HURD. R.G. 1968. E f f e c t s of C0 -enrichment on the growth of young tomato p l a n t s i n low l i g h t . Ann. Bot. ns. 32:531-542.  138.  HURD, R.G. 1969. Leaf r e s i s t a n c e i n a g l a s s h o u s e tomato crop i n r e l a t i o n to l e a f p o s i t i o n and s o l a r r a d i a t i o n . New P h y t o l . 68:265-273.  2  248 139.  HURD, R.G. 1972. E f f e c t s o f carbon d i o x i d e enrichment and humidity on tomato v e g e t a t i v e growth. Ann. Rep. Glasshouse Crops Res. I n s t . , L i t t l e h a m p t o n , U.K. 1972. pp. 44-45.  140.  HURD, R.G. and J.H.M. THORNLEY. 1974. An a n a l y s i s o f the growth of young tomato p l a n t s i n water c u l t u r e a t d i f f e r e n t l i g h t i n t e g r a l s and C0„ c o n c e n t r a t i o n s . I . P h y s i o l o g i c a l a s p e c t s . Ann. Bot. n.s. 38:375-388.  141.  IKEDA, K. and K. KIMURA. 1967. The temperature dependence o f floral initiation. In: Imamura, S. ( e d i t o r ) . Physiology of F l o w e r i n g i n Pharbitis nil. Jap. Soc. P l a n t P h y s i o l o g i s t s , Tokyo, pp. 95-105.  142.  IMAMURA, S. 1967. P h o t o p e r i o d i c i n d u c t i o n and the f l o r a l s t i m u l u s . I n : Imamura, S. ( e d i t o r ) . Physiology of Flowering i n Pharbitis nil. Jap. Soc. P l a n t P h y s i o l o g i s t s , Tokyo, pp. 15-28.  143.  ITO,  144.  JACKSON, W.A. and R.J. VOLK. 1970. P h o t o r e s p i r a t i o n . P l a n t P h y s i o l . 21:385-432.  145.  JAMES, D.B. 1964. L o c a t i n g earthquake v i c t i m s . p r o g r e s s . 3:4.  146.  JARVIS, P.G. 1971. The e s t i m a t i o n o f r e s i s t a n c e s t o carbon d i o x i d e t r a n s f e r . In: Sestak, Z., J . Catsky and P.G. J a r v i s (editors). Plant Photosynthetic Production. Manual o f Methods. Dr. W. Junk N.V., The Hague, pp. 566-631.  147.  JARVIS, P.G. and J . c'ATSKY'. 1971. General p r i n c i p l e s o f gasometric methods and the main a s p e c t s o f i n s t a l l a t i o n d e s i g n . I n : Sestak, Z., J . Catsky and P.G. J a r v i s ( e d i t o r s ) . Plant Photosynthetic Production. Manual o f Methods. Dr. W. Junk N.V. The Hague, pp. 49-110.  148.  JEWISS, O.R. and J . WOLEDGE. 1967. The e f f e c t o f age o n the r a t e of apparent p h o t o s y n t h e s i s i n l e a v e s o f t a l l f e s c u e (Festuaa arundinaoea Schreb.). Ann. Bot. n.s., 31:661-671.  149.  JOLLIFFE, P.A. 1970. P h o t o s y n t h e s i s , p h o t o r e s p i r a t i o n and r e l a t e d a s p e c t s o f CO^ exchange by wheat, corn and Amaranthus edulis. Ph. D. t h e s i s . Univ. o f B r i t i s h Columbia.  150.  JOLLIFFE, P.A. and E.B. TREGUNNA. 1968. E f f e c t o f temperature, C 0 c o n c e n t r a t i o n and l i g h t i n t e n s i t y on oxygen i n h i b i t i o n o f p h o t o s y n t h e s i s i n wheat l e a v e s . P l a n t P h y s i o l . 43:902-906.  151.  JOLLIFFE, P.A. and E.B. TREGUNNA. 1973. Environmental r e g u l a t i o n of the oxygen e f f e c t on apparent p h o t o s y n t h e s i s i n wheat. Can. J . Bot. 51:841-853.  T. 1973. P l a n t growth and p h y s i o l o g y o f v e g e t a b l e p l a n t s as i n f l u e n c e d by carbon d i o x i d e environment. Trans. Fac. Hort. Chiba Univ.,.Japan 7:1-34. Ann. Rev.  Ion exchange  2  JONES, R.J. and T.A. MANSFIELD. 1970. Increases i n the d i f f u s i o n resistances of leaves i n a carbon dioxide-enriched atmosphere. J . Exp. Bot. 21:951-958. KANDELER, R. 1964. Wirkungen des kohlendioxyds auf d i e blutenbildung von Lemna gibba. Naturwiss. 51:561-567. KANDELER, R., B. HUGEL and Th. ROTTENBURG. 1975. Relations between photosynthesis and flowering i n Lemnaceae. In. Marcelle, R. ( e d i t o r ) . Environmental and B i o l o g i c a l Control of Photosynthesis. Dr. W. Junk B.V., The Hague. 408 pp. KANEMASU, E.T. and C B . TANNER. 1969. Stomatal d i f f u s i o n r e s i s t a n c e of snap beans. I I . E f f e c t of l i g h t . Plant P h y s i o l . 44: 1542-1546. KANEMASU, E.T., G.W. THURTELL and C B . TANNER. 1969. Design, c a l i b r a t i o n and f i e l d use of a stomatal d i f f u s i o n porometer. Plant P h y s i o l . 44:881-885. KETELLAPER, H.J. 1963. Stomatal physiology. P h y s i o l . 14:249-270.  Ann. Rev. Plant  KIDD, F. 1917. The c o n t r o l l i n g influence of carbon dioxide. I I I . The retarding e f f e c t of carbon dioxide on r e s p i r a t i o n . Proc. Roy. Soc. B, 89:136-157. KIMURA, K. 1964. F l o r a l i n i t i a t i o n i n Pharbitis nil subjected to continuous i l l u m i n a t i o n at r e l a t i v e l y low temperatures. I I I . E f f e c t of i n t e n s i t y and q u a l i t y of l i g h t . Bot. Mag., Tokyo. 77:115-121. KISAKI, T.,,A. IMAI and N.E. TOLBERT. 1971. I n t r a c e l l u l a r l o c a l i z a t i o n of enzymes r e l a t e d to photorespiration i n green leaves. Plant C e l l P h y s i o l . 12:267-273. KISAKI, T., S. HIRABAYASHI and N. YANO. 1973. E f f e c t of the age of tobacco leaves on photosynthesis and photorespiration. Plant C e l l P h y s i o l . 14:505-514. KLOUGART, A. 1967. A look ahead based on research on CO^ and growth of h o r t i c u l t u r a l plants i n Europe. Proc. XVII. I n t . Hort. Cong. 111:323-332. KNECHT, G.N. and J.W. O'LEARY. 1973. Increased tomato f r u i t development by C0„ enrichment. J . Amer. Soc. Hort. S c i . 99: 214-216". KRENZER, E.G. and D.N. MOSS. 1975. Carbon dioxide enrichment e f f e c t s upon y i e l d and y i e l d components i n wheat. Crop S c i . 15:71-74. KRETCHMAN, D.W. and F.S. HOWLETT. 1970. C0 enrichment f o r vegetable production. Trans. Amer. Soc. A g r i . Eng. 13:252-256. 2  250 166.  KREUSLER, U. 1890. Beobachtunger uber a s s i m i l a t i o n und athmung der kohlensaureausscheidung seitens getodter exemplare; kohlensaureverbrauch, wenn ober-und u n t e r s e i t e der b l a t t e r dem l i c h t zugewendet. Landw. Jahrb. 19:649-668.  167.  KU, S-B., G.E. EDWARDS and C.B. TANNER. 1977. E f f e c t s of l i g h t , carbon dioxide and temperature on photosynthesis, oxygen i n h i b i t i o n of photosynthesis and t r a n s p i r a t i o n i n Solarium tuberosum. Plant P h y s i o l . 59:868-872.  168.  KUJIRAI, C. and S. IMAMURA. 1958. Uber d i e photoperiodische empfindlichkeit der kotyledonen von Pharbitis nil Chois. Bot. Mag., Tokyo. 71:408-416.  169.  KVET, J . , J.P. 0ND0K, J . NECAS and P.G. JARVIS. 1971. Methods of growth a n a l y s i s . In: s'estak, Z., J . Catsky and P.G. J a r v i s ( e d i t o r s ) . Plant Photosynthetic Production. Manual of Methods. Dr. W. Junk N.V., The Hague, pp. 343-411.  170.  LAING, W.A., W.L. OGREN and R.H. HAGEMAN. 1974. Regulation of soybean net photosynthetic CO^ f i x a t i o n by the i n t e r a c t i o n of C0„, 0„ and r i b u l o s e 1,5-dxphosphate carboxylase. Plant P h y s i o l . 54:678-685.  171.  LAKE, J.V. 1967. R e s p i r a t i o n of leaves during photosynthesis. I . Estimates from an e l e c t r i c a l analogue. Austr. J . B i o l . S c i . 20:487-493.  172.  LEE, R.B. and C.P. WHITTINGHAM. 1974. The i n f l u e n c e of p a r t i a l pressures of carbon dioxide upon carbon metabolism i n the tomato l e a f . J . Exp. Bot. 25:277-287.  173.  LEVITT, J . 1967. 74:101-118.  174.  LINSBAUER, K. 1916. Beitrage zur kenntnis der spaltoffnungsbewegungen. F l o r a . 109:100-143.  175.  LIVERMANN, J.L. and J . BONNER. 1953. Biochemistry of the photoperiodic process. The high l i g h t i n t e n s i t y r e a c t i o n . Bot. Gaz. 115:121-128.  176.  LORRIMER, G.H., T.J.. ANDREWS and N.E. T0LBERT. 1973. Ribulose diphosphate oxygenase. I I . Further proof of r e a c t i o n products and mechanism of a c t i o n . Biochemistry 12:18-23.  177.  LUDLOW, M.M. and P.G. JARVIS. 1971. Photosynthesis i n S i t k a Spruce (Bong.) Carr. I . General c h a r a c t e r i s t i c s . J . Appl. Ecol. 8: 925-953.  178.  LUDWIG, L.J. 1972. The r e l a t i o n s h i p between photosynthesis and r e s p i r a t i o n . In: Rees, A.R., K.E. Cockshull, D.W. Hand and R.G. Hurd ( e d i t o r s ) . Crop processes i n c o n t r o l l e d environments. Academic Press, New York. pp. 305-326.  The mechanism of stomatal a c t i o n .  Planta (Berl.)  LUDWIG, L . J . and D.T. CANVIN. 1971. The r a t e of photorespiration during photosynthesis and the r e l a t i o n s h i p of the substrate of l i g h t r e s p i r a t i o n to the products of photosynthesis i n sunflower leaves. Plant P h y s i o l . 48:712-719. LUDWIG, L . J . and D.T. CANVIN. 1971. An open gas-exchange system for the simultaneous measurement of the C0„ and ^ C0 fluxes from leaves. Can. J . Bot. 49:1299-1313. 4  McCREE, K.J. 1972. The a c t i o n spectrum, absorptance and quantum y i e l d of photosynthesis i n crop plants. A g r i c . Meteorol. 9: 191-216. MACDOWAL, F.D.H. 1963. Midday closure of stomata i n ageing tobacco leaves. Can. J . Bot. 41:1289-1300. MAAS, E.F. and R.M. ADAMSON. 1974. S o i l l e s s c u l t u r e of commercial greenhouse tomatoes. A g r i c u l t u r e Canada Publ. 1460. MADSEN, E. 1968. E f f e c t of C0 -concentration on accumulation of starch and sugar i n tomato leaves. P h y s i o l . Plant. 21:168-175. 2  MADSEN, E. 1971. The e f f e c t of carbon dioxide on the photosynthetic r a t e i n tomato leaves. Royal Vet. and A g r i c . Univ. Copenhagen, Denmark, Yearbook. 1971. pp. 195-200. MADSEN, E. 1971. C y t o l o g i c a l changes due to the e f f e c t of carbon dioxide concentration on the accumulation of starch i n chloroplasts of tomato leaves. Royal Vet. and A g r i c . Univ. Copenhagen, Denmark, Yearbook. 1971. pp. 191-194. MADSEN, E. 1973. The e f f e c t of C0 -concentration on development and dry matter production i n young tomato plants. Acta A g r i c . Scand. 23:235-240. 2  MADSEN, E. 1973. E f f e c t of C0 -concentration on the morphological, h i s t o l o g i c a l and c y t o l o g i c a l changes i n tomato plants. Acta A g r i c . Scand. 23:241-246. 2  MADSEN, E. 1974. E f f e c t of C0 -concentration on growth and f r u i t production of tomato plants. Acta A g r i c . Scand. 24:242-246. 2  MADSEN, E. 1976. E f f e c t of C0„-concentration on morphological, h i s t o l o g i c a l , c y t o l o g i c a l and p h y s i o l o g i c a l processes i n tomato plants. State Seed Testing S t a t i o n Press. Lyngby, Denmark. 75 pp. MANN, L.K. 1940. E f f e c t of some environmental f a c t o r s on f l o r a l i n i t i a t i o n i n Xanthium. Bot. Gaz. 102:339-356. MARX, J.L. 1973. Photorespiration: Key to increasing plant p r o d u c t i v i t y . Science 179:365-367.  252 193.  MARSH, H.V. J r . , J.M. GALMICHE and M. GIBBS. 1965. R e s p i r a t i o n during photosynthesis. In: Krogman, D.W. and W.H. Powers ( e d i t o r s ) . Biochemical Dimensions of Photosynthesis. Wayne State Univ. Press, D e t r o i t , pp. 95-107.  194.  MATHON, C.C. and M. STRAUN. 1962. Mise a f l e u r en jour continu et changement du type de r e a c t i o n photoperiodique en f o n c t i o n de l a q u a l i t e et du niveau de 1'eclairement chez l e Pharbitis nil. Choisy. Compt. Rend. Seance Soc. B i o l . 254:1478-1480.  195.  MEDINA, E. 1970. Relationships between nitrogen l e v e l , photosynthetic capacity and carboxydismutase a c t i v i t y i n Atriplex patula leaves. Carnegie I n s t . Washington Yearbook. 69:655-662.  196.  MEIDNER, H. 1967. Further observations on the minimum i n t e r c e l l u l a r space carbon-dioxide concentration (P) of maize leaves and the postulated r o l e s of 'photo-respiration' and g l y c o l l a t e metabolism. J . Exp. Bot. 18:177-185.  197.  MEIDNER, H. and T.A. MANSFIELD. 1968. Physiology of stomata. McGraw H i l l , New York. 178 pp.  198.  MELCHERS, G. and A. LANG. 1942. Auslosung von blutenbildung b e i Hyoscyamus niger durch i n f i l t r a t i o n der b l a t t e r mit zuckerlosungen. Naturwiss. 30:589-590.  199.  MELCHERS, G. and A. LANG. 1948. Die physiologie der blutenbildung. B i o l . Z e i t b l a t t . 67:105-174.  200.  MENZ, K.M., D.N. MOSS, R.Q. CANNELL and W.A. BRUN. 1969. Screening f o r photosynthetic e f f i c i e n c y . Crop. S c i . 9:692-694.  201.  MICHNIEWICZ, M. and A. LANG. 1962. E f f e c t of 9 d i f f e r e n t g i b b e r e l l i n s on stem elongation and flower formation i n c o l d r e q u i r i n g and photoperiodic plants grown under non-inductive conditions. Planta ( B e r l . ) . 58:549-563.  202.  MONSI, M.,H. IWAKI, S. KURAISHI, T. SAEKI and N. N0M0T0. 1962. P h y s i o l o g i c a l and e c o l o g i c a l analyses of shade tolerances of p l a n t s . 2: Growth of dark-treated greengram under varying l i g h t i n t e n s i t i e s . Bot. Mag., Tokyo. 75:185-194.  203.  MONTEITH, J.L. 1973. P r i n c i p l e s of Environmental Arnold, London. 241 pp.  204.  MOONEY, H.A. and F. SHROPSHIRE. 1967. Population v a r i a b i l i t y i n temperature r e l a t e d photosynthetic a c c l i m a t i o n . Oecolog. P l a n t . 2:1-13.  205.  MOONEY, H.A. and M. WEST. 1964. Photosynthetic a c c l i m a t i o n of plants of diverse o r i g i n . Amer J . Bot. 51:825-827.  206.  MORRIS, L.G., J.D. POSTLETHWAITE and R.I. EDWARDS. 1954. V e n t i l a t i o n and the supply of carbon dioxide to a glasshouse tomato crop. Rep. Nat. I n s t . A g r i c . Engng., S i l s o e , U.K. 1954. 14 pp.  ;  Physics.  Edward  253 207.  MORTIMER, D.C. 1959. Some short-term e f f e c t s of increased carbon dioxide concentration on photosynthetic a s s i m i l a t i o n i n leaves. Can. J . Bot. 37:1191-1201.  208.  MOSS, D.N. 1976. Studies on increasing photosynthesis i n crop p l a n t s . In: B u r r i s , R.H. and C.C. Black ( e d i t o r s ) . CO^ Metabolism and Plant P r o d u c t i v i t y . U n i v e r s i t y Park Press, Baltimore, pp. 31-42.  209.  MOSS, D.N., E.G. KRENZER and W.A. BRUN. 1969. Carbon dioxide compensation points i n r e l a t e d plant species. Science 164: 187-188.  210.  MOSS, D.N. and S.L. RAWLINS. 1963. Concentration of carbon dioxide i n s i d e leaves. Nature 197:1320-1321.  211.  MOUSSEAU, M. 1977. Night r e s p i r a t i o n i n r e l a t i o n to growth, photosynthesis and development of Chenopodium polyspermism i n long and short days. Plant Science L e t t e r s 9:339-346.  L  212.  NAKAYAMA, S. 1958. Studies on the dark process i n the photoperiodic response of Pharbitis seedlings.. The S c i . Rep. Tohoku Univ. Ser. IV B i o l . 24:137-183.  213.  NEALES, T.F. and L.D. INCOLL. 1968. The c o n t r o l of leaf photosynthesis r a t e by the l e v e l of a s s i m i l a t e concentration i n the l e a f : A review of the hypothesis. Bot. Rev. 34:107-125.  214.  NEALES, T.F., K.J. TREHARNE and P.F. WAREING. 1971. A r e l a t i o n s h i p between net photosynthesis, d i f f u s i v e r e s i s t a n c e and carboxylating enzyme a c t i v i t y i n bean leaves. In: Hatch, M.D., Osmond, C B . and S l a t y e r , R.O. ( e d i t o r s ) . Photosynthesis and Photorespiration. Wiley-Interscience, New York. pp. 497-505.  215.  NEWTON, P. 1966. The influence of increased C 0 concentration and supplementary i l l u m i n a t i o n on growth of tomato seedlings during the winter months. Ann. Appl. B i o l . 57:345-353.  216.  NISHIDA, K. 1962. Studies on the r e - a s s i m i l a t i o n of r e s p i r a t o r y C 0 i n i l l u m i n a t e d leaves. Plant and C e l l P h y s i o l . 3:111-124.  217.  NITSCH, J.P. and F.W. WENT. 1959. The i n d u c t i o n of flowering i n Xanthium pennsylvanicum under long days. In: Withrow, R.B. ( e d i t o r ) . Photoperiodism and r e l a t e d phenomena i n plants and animals. Amer. Assoc. Adv. S c i . , Wash. D.C. pp. 311-314.  218.  NOBEL, P.S., L . J . ZARAGOZA and W.K. SMITH. 1975. R e l a t i o n between mesophyll surface area, photosynthetic r a t e and i l l u m i n a t i o n l e v e l during development of leaves of Pleotranthus parviflorus Henckel. Plant P h y s i o l . 55:1067-1070.  219.  NORMAN, A.G. 1962. The uniqueness of p l a n t s .  220.  OGAWA, Y. 1960. Uber d i e auslosung der blutenbildung von Pharbitis nil durch niedere temperatur. Bot. Mag. Tokyo 73:334-335.  2  2  Amer. S c i . 50:436-449.  OGREN, W.L. and G. BOWES. 1971. Ribulose diphosphate carboxylase regulates soybean photorespiration. Nature New B i o l . 230: 159-160. OLTMANNS, 0. 1960. Uber den e i n f l u s s der temperatur auf die endogene tagesrhythmik und die bluhinduktion b e i der kurtztagpflanze Kalanchoe bZossfeldiana. Planta. 54:233-264. OSMAN, A.M. and F.L. MILTHORPE. 1971. Photosynthesis of wheat leaves i n r e l a t i o n to age, illuminance and n u t r i e n t supply. I I . Results. Photosynthetica 5:61-70. OSMOND, C.B. and 0. BJ0RKMAN. 1972. E f f e c t s of 0 on photosynthesis Simultaneous measurements of oxygen e f f e c t s on net photosynthesis and g l y c o l a t e metabolsim i n C^ and C^ species of AtvipZex. Carnegie I n s t . Yearbook. 71:141-148. Carnegie I n s t . Wash. 2  PALLAS, J.E. J r . 1965. T r a n s p i r a t i o n and stomatal opening with changes i n carbon dioxide content of the a i r . Science 147: 171-173. PEARCY, R.W. 1977. Acclimation of photosynthetic and r e s p i r a t o r y carbon dioxide exchange to growth temperature i n AtvipZex Zentifovmis (Torr.) Wats. Plant P h y s i o l . 59:795-799. PEARCY, R.W., J.A. BERRY and D.C. FORK. 1977. E f f e c t s of growth temperature on the thermal s t a b i l i t y of the photosynthetic apparatus of AtvipZex Zentifovmis (Torr.) Wats. Plant P h y s i o l . 59:873-878. PEET, M.M., A. BRAVO, D.H. WALLACE and J.L. 0ZBUN. 1977. Photosynthesis, stomatal r e s i s t a n c e and enzyme a c t i v i t i e s i n r e l a t i o n to y i e l d of field-grown dry bean v a r i e t i e s . Crop S c i . 17:287-293. PLASS, G.N. 41-45.  1959.  Carbon dioxide and climate.  S c i . Amer. 201(1):  PLAUT, Z. and M. GIBBS. 1970. Glycolate formation i n i n t a c t spinach c h l o r o p l a s t s . Plant P h y s i o l . 45:470-474. POESCH, G.H. 1935. Supplementary i l l u m i n a t i o n from mazda, mercury and neon lamps on some greenhouse p l a n t s . Proc. Amer. Soc. Hort. S c i . 33:637-638. P0SNER, H.B. 1971. I n h i b i t o r y e f f e c t of carbohydrate on flowering i n Lemna pevpusiZZa. I I I . E f f e c t s of r e s p i r a t o r y intermediates amino acids and C0„. Glucose-6-phosphate dehydrogenase a c t i v i t y . Plant P h y s i o l . 48:361-365. PRIOUL, J.L. A. REYSS and P. CHARTIER. 1975. Relationship between carbon dioxide t r a n s f e r resistances and some p h y s i o l o g i c a l and anatomical features. In: M a r c e l l e , R. ( e d i t o r ) . Environmental and b i o l o g i c a l c o n t r o l of photosynthesis. Dr. W. Junk N.V., The Hague. pp. 17-28  255 234.  PUROHIT, A.N. and E.B. TREGUNNA. 1974. E f f e c t s of carbon dioxide on Pharbitis, Xanthiwn and Silene i n short days. Can. J . Bot. 52:1283-1291.  235.  RASCHKE, K. 1975. 309-340.  236.  RAYLE, D.L. and R. CLELAND. 1970. Enhancement of w a l l loosening and elongation by a c i d s o l u t i o n s . Plant P h y s i o l . 46:250-253.  237.  RICHARDSON, K.E. and N.E. TOLBERT. 1961. Oxidation of g l y o x y l i c a c i d to o x a l i c acid by g l y c o l i c acid oxidase. J . B i o l . Chem. 236: 1280-1284.  238.  ROBELIN, M. et M. MARTIGNAC. 1975. Contribution a 1'etude de l a fumure carbonique sous serre. In: Chouard, P. and N. de B i l d e r l i n g ( e d i t o r s ) . Phytotronics i n A g r i c u l t u r a l and H o r t i c u l t u r a l Research. Bordas, Montreal, pp. 305-317.  239.  ROBINSON, J.M. and M. GIBBS. 1974. Photosynthetic intermediates, the Warburg e f f e c t and g l y c o l a t e synthesis i n i s o l a t e d . spinach c h l o r o p l a s t s . Plant P h y s i o l . 48:193-196.  240.  ROGAN, P.G. and D.L. SMITH. 1975. Rates of leaf i n i t i a t i o n and leaf growth i n Agropyron repens (L). Beauv. J . Exp. Bot. 26: 70-78.  241.  ROGERS, M.N. and B.O.S. TJIA. 1966. E f f e c t of ethylene i n the atmosphere on photoperiodic responses of chrysanthemums. Proc. XVII. I n t . Hort. Cong. Vol 1: Paper 471.  242.  RYLE, G.J.A. and J.D. HESKETH. 1969. Carbon dioxide uptake i n n i t r o g e n - d e f i c i e n t p l a n t s . Crop S c i . 9:451-454.  243.  SALIN, M.L. and P.H. HOMANN. 1971. Changes of photorespiratory a c t i v i t y with leaf age. Plant P h y s i o l . 48:193-196.  244.  SALISBURY, F.B. New York.  245.  SALTER, L.S. 1969. Production techniques i n d e t a i l . 11. Other c u l t u r a l treatments. In: S a l t e r , L.S. ( e d i t o r ) . Manual of Cucumber Production. H.M. Stationery O f f i c e , London, pp. 56-58.  246.  SAMISH, Y. and D. KOLLER. 1968. Estimation of photorespiration of green plants and of t h e i r mesophyll r e s i s t a n c e to C0„ uptake. Ann. Bot. n.s. 32:687-694.  247.  SCHEUPLEIN, C. 1978. E f f e c t s of C0 and daylength on growth, development and hardiness of Douglas F i r . Ph. D. t h e s i s , U n i v e r s i t y of B r i t i s h Columbia.  248.  SESTAK, Z. 1977. Photosynthetic c h a r a c t e r i s t i c s during ontogenesis of leaves: 2. Photosystems, components of e l e c t r o n transport chain, and photophosphorylations. Photosynthetica 11:449-474.  Stomatal a c t i o n . Ann. Rev. Plant P h y s i o l 26:  1963. The flowering process. 234 pp.  Pergamon Press,  2  256 249.  SHAIN, Y. and M. GIBBS. 1971. Formation of g l y c o l a t e by a r e c o n s t i t u t e d spinach c h l o r o p l a s t system. Plant P h y s i o l . 48: 325-330.  250.  SHAW, R.J. and M. RODGERS. 1964. Interactions on roses; temperature and C0 e f f e c t s . F l o r i s t s Rev. 135:21-22. 2  251.  SIMON, E.W. 1967. Types of l e a f senescence. In: Woolhouse, H.W. ( e d i t o r ) . Aspects of the Biology of Ageing. Academic Press, New York. pp. 215-230.  252.  SINGH, B.N. and K.N. LAL. 1935. L i m i t a t i o n s of Blackman's law of l i m i t i n g f a c t o r s and Harder's concept of r e l a t i v e minimum as applied to photosynthesis. Plant P h y s i o l . 10:245-268.  253.  SLACK, G. and A. CALVERT. 1972. Control of carbon dioxide concentration i n glasshouses by the use of conductimetric c o n t r o l l e r s . J . A g r i c . Engng. Res. 17:107-115.  254.  SLATYER, R.O. and J.F. BIERHUIZEN. 1964. The influence of several t r a n s p i r a t i o n suppressants on t r a n s p i r a t i o n , photosynthesis and water-use e f f i c i e n c y of cotton leaves. Aust. J . B i o l . S c i . 17:131-146.  255.  SMALL, T. and H.L. WHITE. 1930. Carbon dioxide i n r e l a t i o n to greenhouse crops. IV. The e f f e c t on tomatoes of an enriched atmosphere maintained by means of a stove. Ann. Appl. B i o l . 17:81-89.  256.  SMILLIE, R.M. 1962. Photosynthetic and r e s p i r a t o r y a c t i v i t i e s of growing pea leaves. Plant P h y s i o l . 27:716-721.  257.  SMITH, D.R. 1973. Plant propagation. In: Kingham, H.G. ( e d i t o r ) . The U.K. Tomato Manual. Grower Books, London, pp. 106-115.  258.  SNYDER, F.W. and N.E. TOLBERT. 1974. E f f e c t of C0 concentration on g l y c i n e and serine formation during photorespiration. Plant P h y s i o l . 53:514-515.  259.  SNYDER, W.E. 1840. E f f e c t of l i g h t and temperature on f l o r a l i n i t i a t i o n i n cocklebur and b i l o x i soybean. Bot Gaz. 102:302-322.  260.  SPLITTSTOESSER, W.E. 1966. Dark carbon dioxide f i x a t i o n and i t s r o l e i n the growth of plant t i s s u e . Plant P h y s i o l . 41:755-759.  261.  STALFELT, M.G. 1955. The stomata as a hydrophotic regulator of the water d e f i c i t of the plant. P h y s i o l . Plant. 8:572-593.  262.  STAMP, P. 1978. Der Chlorophyllgehalt, die PEP- und RuDPCarboxylase- A k t i v i t a t e n wahrend der Blattentwicklung einer ergrunenden Chlorophyllmutante und einer normalen l i n i e von Zea Mays L. Z. Pflanzenphysiol. Bd. 86:395-404.  2  257 263.  STEER, B.T. 1971. The dynamics of l e a f growth and photosynthetic capacity i n Capsicum frutescens L. Ann. Bot. n.s. 35:1003-1015.  264.  STEER, B.T. 1973. Control of ribulose-1,5-diphosphate carboxylase a c t i v i t y during expansion of leaves of Capsicum frutescens L. Ann. Bot. n.s. 37:823-829.  265.  STOCKER, 0. 1960. P h y s i o l o g i c a l and morphological changes i n plants due to water d e f i c i e n c y . In: Plant-Water Relationships i n A r i d and Semi-Arid Conditions. V o l . 15. Reviews of Research, UNESCO. pp. 63-94.  266.  STREET, H.E. and W. COCKBURN. 1972. Plant Metabolsim. Press, Toronto. 321 pp.  267.  SUGE, H. 1972. I n h i b i t i o n of photoperiodic f l o r a l induction i n Pharbitis nil by ethylene. Plant and C e l l P h y s i o l . 13:10311038.  268.  TAKIMOTO, A. 1960. E f f e c t of sucrose on flower i n i t i a t i o n of Pharbitis nil i n a s e p t i c c u l t u r e . Plant and c e l l P h y s i o l . 1:241-246.  269.  TAKIMOTO, A. 1967. General methods of experimentation with Pharbitis nil. I n : Imamura, S. ( e d i t o r ) . Physiology of Flowering i n Pharbitis nil. Jap. Soc. Plant P h y s i o l o g i s t s . Tokyo, pp. 1-5.  270.  TAKIMOTO, A., Y. TASHIMA and S. IMAMURA. 1960. E f f e c t of temperature on flower i n i t i a t i o n of Pharbitis nil cultured i n v i t r o . Bot. Mag. Tokyo 73:377.  271.  THORNLEY, J.H.M. and R.G. HURD. 1974. An a n a l y s i s of the growth of young tomato plants i n water c u l t u r e a t d i f f e r e n t l i g h t i n t e g r a l s and C0„ concentrations. I I . A mathematical model. Ann. Bot. n.s. 38:389-400.  272.  TICHA, I . and J . CATSKY. 1977. Ontogenetic changes i n the i n t e r n a l l i m i t a t i o n s to bean-leaf photosynthesis. 3. Leaf mesophyll s t r u c t u r e and i n t r a c e l l u l a r conductance f o r carbon dioxide t r a n s f e r . Photosynthetica 11:361-366.  273.  TOGNONI, F., A.H. HALEVY and S.H. WITTWER. 1967. Growth of bean and tomato plants as a f f e c t e d by root absorbed growth substances and atmospheric carbon dioxide. Planta 73:43-52.  274.  TOLBERT, N.E. 1971. Microbodies - Peroxisomes and Glyoxysomes. Ann. Rev. PI. P h y s i o l . 22:45-74.  275.  TOLBERT, N.E. 1973. Compartmentation and c o n t r o l i n microbodies. Symp. Soc. Exp. B i o l . 27:215-239.  276.  TOLBERT, N.E. 1973. Glycolate Biosynthesis. In: Horecker, B.L. and E.R. Stadtman ( e d i t o r s ) . Current Topics i n C e l l u l a r Regulation. V o l . 7. Academic Press, New York. pp. 21-50.  Pergamon  258 277.  TOLBERT, N.E. and R.K. YAMAZAKI. 1969. Leaf peroxisomes and t h e i r r e l a t i o n to photorespiration and photosynthesis. Ann. New York Acad. S c i . 168:325-341.  278.  TREGUNNA, B. 1966. F l a v i n mononucleotide c o n t r o l of g l y c o l i c acid oxidase and photorespiration i n corn leaves. Science 151:1239-1241.  279.  TREGUNNA, E.B., G. KROTKOV and C D . NELSON. 1961. Evolution of carbon dioxide by tobacco leaves during the dark period f o l l o w i n g i l l u m i n a t i o n with l i g h t of d i f f e r e n t i n t e n s i t i e s . Can. J . Bot. 39:1045-1056.  280.  TREGUNNA, E.B., KROTKOV and C D . NELSON. 1966. E f f e c t of oxygen on the rate of photorespiration i n detached tobacco leaves. P h y s i o l . Plant 19:723-733.  281.  TREHARNE, K.J. 1972. Biochemical l i m i t a t i o n s to photosynthetic r a t e s . In: Rees, A.R., K.E. Cockshull, D.W. Hand and R.G. Hurd ( e d i t o r s ) . Crop Processes i n Controlled Environments. Academic Press, New York. pp. 285-304.  282.  TR0UGHT0N, J.H. and R.O. SLATYER. 1969. Plant water status, leaf temperature and the c a l c u l a t e d mesophyll r e s i s t a n c e to carbon dioxide of cotton leaves. Aust. J . B i o l . S c i . 22:815-827.  283.  TURNER, J.S. and E.G. BRITTAIN. 1962. Oxygen as a f a c t o r i n photosynthesis. B i o l Rev. 37:130-170.  284.  TURNER, J.S., J.F. TURNER, K.D. SHORTMAN and J.E. KING. 1958. The i n h i b i t i o n of photosynthesis by oxygen. I I . The e f f e c t of oxygen on glyceraldehyde phosphate dehydrogenase from c h l o r o p l a s t s . Aust. J . B i o l . S c i . 11:336-342.  285.  TURNER, N.C 1969. Stomatal response to t r a n s p i r a t i o n i n three contrasting canopies. Crop S c i . 9:303-307.  286.  VERNON, A.J. and J.C.S. ALLISON. 1963. A method of c a l c u l a t i n g net a s s i m i l a t i o n r a t e . Nature 200:814.  287.  WARBURG, 0. 1920. Uber d i e geschwindigkeit der photochemischen kohlensaurezerset-zung i n lebenden z e l l e n . I I . Biochem. Z. 103:188-217.  288.  WAREING, P.F., M.M. KHALIFA and K.J. TREHARNE. 1968. Rate l i m i t i n g processes i n photosynthesis at saturating l i g h t i n t e n s i t i e s . Nature 220:453-457.  289.  WARREN-WILSON, J . 1966. E f f e c t of temperature on net a s s i m i l a t i o n r a t e . Ann. Bot. n.s. 30:753-761.  290.  WATSON, D.J. 1952. The p h y s i o l o g i c a l basis of v a r i a t i o n i n y i e l d . Adv. Agron. 4:101-145.  259 291.  WHITEMAN, P.C. and D. ROLLER. 1967. I n t e r a c t i o n s of carbon dioxide concentration, l i g h t i n t e n s i t y and temperature on plant resistances to water vapour and carbon dioxide d i f f u s i o n . New P h y t o l . 66:463-473.  292.  WHITEMAN, P.C. and D. ROLLER. 1968. Estimation of mesophyll r e s i s t a n c e to d i f f u s i o n of carbon dioxide and water vapour. In: Eckardt, F.E. ( e d i t o r ) . Functioning of T e r r e s t r i a l Ecosystems a t the Primary Production Level. Unesco, P a r i s , pp. 415-419.  293.  WIDHOLM, J.M. and W.L. OGREN. 1969. Photorespiratory-induced senescence of plants under conditons of low carbon dioxide. Proc. Nat. Acad. U.S.A. 63:668-675.  294.  WISHNICR, M. and M.D. LANE. 1971. Ribulose diphosphate carboxylase from spinach leaves. I n : San P i e t r o , A. ( e d i t o r ) . Methods i n Enzymology. V o l . XXIII Photosynthesis. Part A. Academic Press, New York. pp. 570-577.  295.  WITTWER, S.H. 1967. Carbon dioxide and i t s r o l e i n plant growth. Proc. XVII I n t . Hort. Cong. 111:311-322.  296.  WITTWER, S.H. 1974. Maximum production capacity of food crops. Bioscience 24:216-224.  297.  WITTWER, S.H. 1977. Carbon dioxide f e r t i l i z a t i o n of crop p l a n t s . In: Gupta V.S. ( e d i t o r ) . Crop Physiology. Oxford and IBM Publishing Co. Janpath, I n d i a , pp. 310-333.  298.  WITTWER, S.H. and W.M. ROBB. 1964. Carbon dioxide enrichment of greenhouse atmospheres f o r food crop production. Econ. Bot. 18:34-56.  299.  WOLEDGE, J . 1971. The e f f e c t of l i g h t i n t e n s i t y during growth on the subsequent r a t e of photosynthesis of leaves of t a l l Festuca (Festuaa avundinaaea Schreb.). Ann. Bot. n.s. 35: 311-322.  300.  WOODWELL, G.M. 1978. The carbon dioxide question. 238(1): 34-43.  301.  W00LH0USE, H.W. 1967. The nature of senescence i n p l a n t s . I n : Woolhouse, H.W. ( e d i t o r ) . Aspects of the Biology of Ageing. Academic Press, New York. pp. 179-214.  302.  YOSHIDA, S. 1972. P h y s i o l o g i c a l aspects of g r a i n y i e l d . Ann. Rev. P l a n t . P h y s i o l . 23:437-464.  303.  ZAR, J.H. 1974. B i o s t a t i s t i c a l A n a l y s i s . New Jersey. 620 pp.  304.  ZEEUW, D. DE 1957. flowering of Xanthivm under long-day conditions. Nature 180:558.  S c i . Amer.  Prentice H a l l ,  260 305.  ZEEVART, J.A.D. 723-731.  1962. Physiology of flowering.  Science 137:  306.  ZEEVART, J.A.D. 1976. Physiology of flower formation. Plant P h y s i o l . 27:321-348.  307.  ZELITCH, I . 1966. Increased rate of net photosynthetic carbon dioxide uptake cuased by the i n h i b i t i o n of g l y c o l a t e oxidase. Plant P h y s i o l . 41:1623-1631.  308.  ZELITCH, I . 1971. Photosynthesis, Photorespiration and Plant P r o d u c t i v i t y . Academic Press, New York. 347 pp.  309.  ZELITCH, I . 1974. The e f f e c t of g l y c i d a t e , an i n h i b i t o r of g l y c o l a t e sysnthesis, on photorespiration and net photosynthesis. Arch. Biochem. Biophys. 163:367-377.  310.  ZELITCH, I . and D.A. WALKER. 1964. The r o l e of g l y c o l i c acid metabolism i n opening of l e a f stomata. Plant P h y s i o l . 39: 856-862.  Ann. Rev.  APPENDIX 1 The f o l l o w i n g l i s t of experiments performed i n the course of t h i s thesis research, i s presented as an a i d to reference i n the text.  Experiment Number 1  Experiment T i t l e Measurement of stomatal resistances i n leaves of a greenhouse cucumber crop; concurrent measurement of greenhouse CO^ concentration.  2  Growth and y i e l d a n a l y s i s of tomato.crops, grown i n two greenhouses without C02~ enrichment (preliminary to Experiment 3).  3  Leaf development and y i e l d a n a l y s i s of greenhouse tomato crops grown with and without atmospheric C0^~enrichment.  4  Gas exchange measurements on greenhouse tomato crops grown with and without atmospheric CO^-enrichment.  5  Growth a n a l y s i s of tomato plants grown at 3 CO^ concentrations i n chambers.  6  CO^ exchange measurements on l e a f 3 of tomato p l a n t s grown at 3  concentrations i n  Chambers, at 2 stages of development.  262  Experiment Number 7  Experiment T i t l e Replacement Experiment: . CO^ exchange measurements on tomato plants grown a t 3 CC>2 concentrations  i n chambers, 14 days  a f t e r replacement i n a normal (0.03%) CO2 8  atmosphere.  Assays of RuBP-case and GaO a c t i v i t i e s i n leaves of tomato plants grown at 3 C O 2 concentrations  i n chambers, a t 2 stages of  development. 9  A n a l y s i s of flowering and vegetative development i n Pharbitis  nil  plants grown i n LD and  SD a t 4 C O 2 concentrations. 10  Gas exchange measurements on Pharbitis  nil  plants grown under 0.03% or 1.0% C O 2 i n LD and SD.  263  APPENDIX 2 The behavioral equation f o r net photosynthesis leaves"*"  of i n d i v i d u a l  used i n the a n a l y s i s of data i n Experiment 4, may be  summarized as follows:  F  IM I / E  +  ^p " V  1 / E  c  -2 Where:  F = Flux density of CO^ (g m I  +  (  C  I"  r )  -1 s )  = absorbed quantum f l u x density (400 - 700 nm) -2 (yEinstein m  -1 s  ) - i n t h i s study i n c i d e n t radiant f l u x  density was taken to equal t h i s parameter. IQ = the compensation point f o r i n c i d e n t p h o t o s y n t h e t i c a l l y -2 -1 a c t i v e .: r a d i a t i o n ( y E i n s t e i n m s ) C\ = CO^ concentration i n the i n t e r c e l l u l a r spaces w i t h i n the -3 l e a f (g m  ) '.- c a l c u l a t e d by the methods of Moss and  Rawlins^ -3 r = (X>2 compensation point (g m ) s a t u r a t i n g l e v e l s of atmospheric concentration and F^ = maximum p o s s i b l e CO^ f l u x from any one l e a f (obtained at M  Ej=  T  w  ~  2  I ) (g m s ) e f f i c i e n c y of u t i l i z a t i o n of radiant energy obtained at CO 2 s a t u r a t i o n , when C photosynthetic  places no l i m i t a t i o n on the  process (g y E i n s t e i n "*")  264  = e f f i c i e n c y of CC^ u t i l i z a t i o n obtained at s a t u r a t i n g 3 l e v e l s of I  (m  g  -1 )  In t h i s study, the terms I , F and T: were measured, 1^ p  was obtained by i n t e r p o l a t i o n between values of dark r e s p i r a t i o n , and net photosynthesis at an i r r a d i a n c e of 50 uE m E^. and E^, were c a l c u l a t e d .  -2  -1 s , and C., F , l IM T W  At each l e v e l of i r r a d i a n c e f o r i n d i v i d u a l  leaves, l i n e a r regressions were c a l c u l a t e d between r e c i p r o c a l values of ( C - T) and F.  From these p l o t s , values of CO^ f l u x at an  extrapolated value of s a t u r a t i n g C. (designated F ), and E X  X  L  were *  obtained from the i n t e r c e p t ^ and (slope ^ x i n t e r c e p t ) , r e s p e c t i v e l y . A mean value f o r E^, was obtained f o r each l e a f age i n each growth regime (except i n the case of a p i c a l leaves grown i n normal a i r where l a c k of a consistent l i n e a r r e l a t i o n s h i p between F precluded f u r t h e r a n a l y s i s of the data).  and (C - T) "*"  Other regressions were  c a l c u l a t e d between values of ( Ip - IQ) and F^. ^ to give F ^ and E^ -1 from the i n t e r c e p t  -1 and slope  . Again, average values were obtained  from a l l a v a i l a b l e regression data f o r each treatment. For a more complete d i s c u s s i o n and v a l i d a t i o n of the model, see reference"*". 2  Van Bavel, C. H. M. 1975. A behavioral equation f o r l e a f carbon dioxide a s s i m i l a t i o n and a test of i t s v a l i d i t y . Photosynthetica 9: 165-176. Moss, D. N. and S. L. Rawlins. 1963. Concentration of carbon dioxide i n s i d e leaves. Nature 197: 1320-1321.  APPENDIX 3 As a preliminary to the enzyme a c t i v i t y assays described i n M a t e r i a l s and Methods, Part I I , Section 2 of t h i s t h e s i s , pH optima were determined using the assay procedures of Wishnick and Lane  1  2 (RuBP-case) and Baker and Tolbert .(GaO).  Optimum substrate  concentration was also determined f o r RuBP-case, but not f o r GaO s i n c e , i n t h i s case, the amount of sodium g l y c o l a t e added i n the e x t r a c t i o n medium was s u f f i c i e n t to saturate a c t i v i t y .  The t r i a l s  were conducted on leaf m a t e r i a l of a uniform age from plants grown i n the greenhouse under the pretreatment  conditions mentioned i n the  t e x t (Materials and Methods, Part I I , Section 2 ) . 1.  RuBP-case The f o l l o w i n g r e a c t i o n mixture was used : x  1.0 M T r i s (Cl") Buffer 0.005 M  (100 ymoles)  D-ribulose-l,55blphosphate 14  0.5 M sodium bicarbonate,((  C)  (25 ymoles)  0.5 M magnesium c h l o r i d e  (5 ymoles)  0.006 M EDTA  (3 ymoles)  + enzyme e x t r a c t i n a t o t a l volume of 0.5 ml Wishnick, M. and M. D. Lane, 1972. Ribulose diphosphate carboxylase from spinach leaves. In: San P i e t r o , A. ( e d i t o r ) . Methods i n enzymology. V o l . XXIII Photosynthesis. Part A. Academic Press, New York. pp. 570-577. 2 Baker, A. L.„and N. E. Tolbert, 1966. Glycolate oxidase (Ferredoxin containing form). In: Wood, W. A. ( e d i t o r ) . Methods i n enzymology V o l . IX.Carbohydrate metabolism. Academic Press, New York, pp. 338-342.  Figure A3-1 depicts the r e l a t i o n s h i p between RuBP concentration and enzyme a c t i v i t y at a temperature of 30°C and pH 7.8. Saturating substrate concentration was found to be approximately 0.4 mM (equivalent to 0.2 ymoles per 0.5 ml of r e a c t i o n mixture. The r e l a t i o n s h i p between enzyme a c t i v i t y and pH a t a temperature of 30°C and s a t u r a t i n g substrate concentration i s shown i n Figure A.3-2. Optimum pH was approximately '7.8 which corresponds c l o s e l y w i t h that determined by Wishnick and Lane'*'. In Experiment 8 T r i s (Cl~) b u f f e r , pH 7.8 and 0.25 ymoles RuBP were used f o r a l l RuBP-case determinations. GaO 2 The f o l l o w i n g r e a c t i o n mixture was used : 0.04 M Sodium g l y c o l a t e  (20 ymoles)  0.1 M cysteine-HCl  (10 ymoles)  0.1 M phenylhydrazine-HCl  (10 ymoles)  0.1 M potassium phosphate b u f f e r  (200 ymoles)  + enzyme e x t r a c t i n a t o t a l volume of 2.9 ml The r e l a t i o n s h i p between a c t i v i t y and pH f o r t h i s enzyme i s shown i n Figure A3-3. pH optimum was estimated to l i e between 8.0 and 8.5. In Experiment 8 potassium phosphate b u f f e r , pH 8.3 was used f o r a l l GaO determinations.  FIGURE A3-1: RuBP-case a c t i v i t y i n r e l a t i o n to substrate concentration at pH 7.8, 30°C.  267  267a  FIGURE A3-2:  RuBP-case a c t i v i t y  i n r e l a t i o n to pH a t  s a t u r a t i n g s u b s t r a t e c o n c e n t r a t i o n , 30°C.  268a  FIGURE A3-3: GaO a c t i v i t y i n r e l a t i o n to pH at s a t u r a t i n g substrate concentration, 27°C.  ACTIVITY  1  1  (jjmoles g l y o x y l a t e formed g fw~' min o  V*  _i  o  1  o  I  o  I  I  o  o  i  o  i  o  i  O i  ) —-  i  '  

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