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Effects of carbon dioxide enrichment on growth, photosynthesis, and leaf senescence in bean (Phaseolus… Ehret, David Lloyd 1984

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EFFECTS OF CARBON DIOXIDE ENRICHMENT ON GROWTH, PHOTOSYNTHESIS, AND LEAF SENESCENCE IN BEAN ( PHASEOLUS VULGARIS L.) PLANTS by DAVID LLOYD EHRET B . S c , U n i v e r s i t y Of A l b e r t a , 1973, M . S c , U n i v e r s i t y Of I l l i n o i s , 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 P l an t Sc i ence We accept t h i s t h e s i s as conforming to the r e q u i r e d s tandard THE UNIVERSITY OF BRITISH COLUMBIA January 1984 © Dav id L l o y d E h r e t , 1984 In p r e s e n t i n g t h i s t h e s i s i n p a r t i a l f u l f i l m e n t o f the r equ i r ements f o r an advanced degree a t the U n i v e r s i t y o f B r i t i s h Co lumb i a , I agree t ha t the L i b r a r y s h a l l make i t f r e e l y a v a i l a b l e f o r r e f e r e n c e and s tudy . I f u r t h e r agree t h a t p e r m i s s i o n f o r e x t e n s i v e copy ing o f t h i s t h e s i s f o r s c h o l a r l y purposes may be g ran ted by the head o f my department o r by h i s o r her r e p r e s e n t a t i v e s . I t i s unde r s tood t h a t copy ing o r p u b l i c a t i o n o f t h i s t h e s i s f o r f i n a n c i a l g a i n s h a l l no t be a l l owed w i thou t my w r i t t e n p e r m i s s i o n . Department o f The U n i v e r s i t y o f B r i t i s h Co lumbia 1956 Main Mall Vancouve r , Canada V6T 1Y3 Date i i A b s t r a c t I n c r e m e n t a l i n c r e a s e s i n a b s o l u t e g r o w t h r a t e w i t h i n c r e a s i n g c a r b o n d i o x i d e c o n c e n t r a t i o n s r a n g i n g from 340 t o 3000- u l 1" 1 were o b s e r v e d i n c o n t i n u o u s l y c a r b o n d i o x i d e e n r i c h e d bean p l a n t s o v e r t h e c o u r s e of d e v e l o p m e n t . However, r e l a t i v e g r o w t h r a t e of e n r i c h e d p l a n t s was e n h a n c e d o n l y e a r l y i n " d e v e l o p m e n t . U n i t l e a f r a t e was a l s o i n c r e a s e d by e n r i c h m e n t p r i m a r i l y i n e a r l y d e v e l o p m e n t , e x p l a i n i n g , i n p a r t , t h e t r e n d s i n r e l a t i v e g r o w t h r a t e . C a r b o n d i o x i d e e n r i c h m e n t a l s o c a u s e d l a r g e i n c r e a s e s i n l e a f d r y w e i g h t , w h i c h c o n s t i t u t e d t h e major change i n c a r b o n p a r t i t i o n i n g among p l a n t p a r t s . L e a v e s o f e n r i c h e d p l a n t s showed a d e c r e a s e i n p h o t o s y n t h e t i c c a p a c i t y w h i c h was not a s s o c i a t e d w i t h c h a n g e s i n c h l o r o p h y l l c o n c e n t r a t i o n , or p h o t o r e s p i r a t i o n rate.. The d e c r e a s e . i n c a p a c i t y was l e s s e v i d e n t i n o l d e r l e a v e s , o r i n t h o s e m a i n t a i n e d a t low l i g h t i n t e n s i t y o r w i t h r e d u c e d c h l o r o p h y l l l e v e l s , s u g g e s t i n g t h a t t h e r e d u c e d p h o t o s y n t h e t i c c a p a c i t y was due t o t h e h i g h e r p h o t o s y n t h e t i c r a t e of t h o s e l e a v e s d u r i n g g r o w t h . R e s p i r a t i o n r a t e i n l e a v e s of e n r i c h e d p l a n t s was a l s o i n c r e a s e d , but o n l y u nder c o n d i t i o n s w h i c h c a u s e d a c o n c u r r e n t d e c r e a s e i n p h o t o s y n t h e t i c c a p a c i t y . E n r i c h e d l e a v e s c o n s i s t e n t l y showed h i g h e r s t a r c h c o n t e n t w i t h g e n e r a l l y a l o w e r p h o t o s y n t h e t i c c a p a c i t y t h a t c o n t r o l l e a v e s . F u r t h e r m o r e , an i n c r e a s e i n s i n k demand d i d not i n f l u e n c e t h e p h o t o s y n t h e t i c c a p a c i t y and s t a r c h c o n t e n t o f e n r i c h e d l e a v e s t o t h e e x t e n t of c o n t r o l l e a v e s . A h i g h l e a f s t a r c h c o n t e n t and l e a f d r y w e i g h t c o r r e l a t e d with a c c e l e r a t e d senescence of p l a n t s . C o n d i t i o n s such as t empera tu re , which f a c i l i t a t e d r e s u l t e d i n the most r a p i d and the pr imary l eaves of e n r i c h e d h i g h l i g h t i n t e n s i t y or reduced g r e a t e r s t a r c h a c c u m u l a t i o n , a l s o e x t e n s i v e senescence . i v T a b l e o f C o n t e n t s A b s t r a c t i i L i s t o f T a b l e s v i L i s t o f F i g u r e s v i i i L i s t of A b b r e v i a t i o n s x i Acknowledgement x i i i C h a p t e r I GENERAL INTRODUCTION 1 C h a p t e r II LITERATURE REVIEW 7 2.1 PLANT RESPONSE TO CARBON DIOXIDE ENRICHMENT 7 2.1.1 H i s t o r i c a l P e r s p e c t i v e 7 2.1.2 C a r b o n D i o x i d e E n r i c h m e n t And P l a n t Growth 8 2.1.3 D u r a t i o n Of E n r i c h m e n t 11 2.1.4 I n t e r a c t i o n W i t h P l a n t Age 12 2.1.5 I n t e r a c t i o n W i t h L i g h t I n t e n s i t y 13 2.1.6 I n t e r a c t i o n W i t h N u t r i e n t Regime 14 2.1.7 E f f e c t s On P a r t i t i o n i n g Of A s s i m i l a t e s 15 2.1.8 R e s p o n s e Of P h o t o s y n t h e t i c C a p a c i t y 17 2.1.9 L e a f M o r p h o l o g y " 23 2.1.10 L e a f C a r b o h y d r a t e s 23 2.2 SENESCENCE ' 24 2.2.1 G e n e r a l P h y s i o l o g y 25 2.2.2 Growth R e g u l a t o r s 27 2.2.3 E n v i r o n m e n t a l I n f l u e n c e s 29 2.2.4 C a r b o n D i o x i d e And S e n e s c e n c e 30 C h a p t e r I I I EFFECTS ON GROWTH AND PARTITIONING 32 3.1 INTRODUCTION 32 3.2 MATERIALS AND METHODS • 3 4 3.2.1 P l a n t C u l t u r e 34 3.2.2 C a r b o n D i o x i d e T r e a t m e n t Chamber D e s i g n 34 3.2.3 Growth A n a l y s i s 38 3.3 RESULTS AND DISCUSSION 42 3.3.1 E f f e c t s On Growth ' 42 3.3.2 E f f e c t s On P a r t i t i o n i n g 53 3.3.3 Summary 62 C h a p t e r IV EFFECTS ON NET CARBON DIOXIDE EXCHANGE 65 4.1 INTRODUCTION 65 4.2 MATERIALS AND METHODS 68 4.3 RESULTS AND DISCUSSION 73 4.3.1 G e n e r a l i z e d Response Of Net C a r b o n D i o x i d e E xchange C a p a c i t y 73 4.3.2 R e s p o n s e Of S p e c i f i c P a r a m e t e r s Of Net C a r b o n D i o x i d e Exchange C a p a c i t y 80 4.3.3 E f f e c t Of D u r a t i o n 84 4.3.4 E f f e c t Of P h o t o s y n t h e t i c P h o t o n F l u x D e n s i t y ....85 4.3.5 R e s p o n s e To S i n k M a n i p u l a t i o n 89 V 4.3.6 E f f e c t s On Dark R e s p i r a t i o n 102 4.3.7 E f f e c t s On Stomata 104 4.3.8 E f f e c t On Tomato 106 4.3.9 Summary 107 Chapter V EFFECTS ON LEAF SENESCENCE 111 5.1 INTRODUCTION 111 5.2 MATERIALS AND METHODS 113 5.3 RESULTS AND DISCUSSION 115 5.3.1 E f f e c t s Of Temperature 115 5.3.2 E f f e c t s On Stomata 120 5.3.3 E f f e c t s Of PPFD 125 5.3.4 E f f e c t s On Leaf Dry Weight 129 5.3.5 P r e l i m i n a r y Experiments With Detached Leaves ...129 5.3.6 Summary 136 Chapter VI GENERAL DISCUSSION 137 Chapter VII CONCLUSIONS 155 BIBLIOGRAPHY 157 APPENDIX A - PRIMARY DATA OF TRIAL 2 171 vi Lis t of Tables 3.1 Analysis of variance of the collective dry weight data of t r i a l 1 and t r i a l 2 43 3.2 Total dry weight, pod weight, and Harvest Index of plants grown at 340, 500, 800, 1200, 2000, or 3000 ul l " 1 carbon dioxide for 44 days 63 4.1a Net carbon dioxide exchange rate of the f i r s t and second t r i f o l i a t e s of control (340 ul 1 "1) and enriched (1400 ul l " 1 ) plants 74 4.1b Analysis of variance of the data of Table 4.1a 74 4.2 Oxygen sensit ivi ty of the net carbon dioxide exchange rate of the f i r s t t r i f o l i a t e of control (340 ul l " 1 ) and enriched (1400 ul l ' 1 ) plants treated for 13 days 78 4.3 Leaf thickness and intercellular air space volume of control (340 ul l " 1 ) and enriched (1400 ul plants 82 4.4 The effect of shading during growth on the net carbon dioxide exchange rate of the f i r s t t r i f o l i a t e of enriched (1400 ul l " 1 ) and control (340 ul l " 1 ) beans grown at 26 C 88 4.5 The effect of shading a l l but the f i r s t t r i f o l i a t e on starch content and net carbon dioxide exchange rate of the f i r s t t r i f o l i a t e of control (340 ul l " 1 ) and enriched (1400 and 3000 ul 1~1) plants 92 4.6 Effect of pot size on the net carbon dioxide exchange rate of the f i r s t t r i f o l i a t e of enriched (1400 ul l " 1 ) and control (340 ul l " 1 ) plants 100 4.7 Stomatal diffusive resistance of the primary leaf and f i r s t t r i f o l i a t e of enriched (1400 ul 1 *1) and control (340 ul l " 1 ) plants 105 4.8 Net carbon dioxide exchange rate of control (340 ul l " 1 ) and enriched (1000 ul l " 1 ) tomato plants at a PI of 5.0 108 vi i 5.1 Net carbon dioxide exchange rate and chlorophyll content of the primary leaves before and after 17 days of carbon dioxide enrichment at 32 or 20 C 116 5.2 Specific leaf weight and starch content of the primary leaves of control (340 ul I ' 1 ) and enriched (1400 ul 1"') plants at two temperatures and two levels of photosynthetic photon flux density 130 5.3 Water and osmotic potentials, and specific leaf weight of enriched (1400 ul I ' 1 ) and control (340 ul l ' 1 ) plants grown for 7 days at 24 C 135 v i i i List of Figures 3.1 Schematic diagram of one of the eight carbon dioxide treatment chambers 36 3.2 Leaf area over time of plants grown at 340, 500, 800, 1200, 2000, or 3000 ul l " 1 carbon dioxide 44 3.3 The dry weight over time of plants grown at 340, 500, 800, 1200, 2000, or 3000 ul l " 1 carbon dioxide 45 3.4 Absolute growth rate of dry weight over time of plants grown at 340, 500, 800, 1200, 2000, or 3000 ul l " 1 carbon dioxide 46 3.5 Relative growth rate over time of plants grown at 340, 500, 800, 1200, 2000, or 3000 ul 1 ' 1 carbon dioxide 47 3.6 Unit leaf rate over time of plants grown at 340, 500, 800, 1200, 2000, or 3000 ul l " 1 carbon dioxide 49 3.7 Leaf area ratio over time of plants grown at 340, 500, 800, 1200. 2000, or 3000 ul l " 1 carbon dioxide 51 3.8 Root-shoot ratio over time of plants grown at 340, 500, 800, 1200, and 3000 ul l ' 1 carbon dioxide 54 3.9 Leaf weight ratio over time of plants grown at 340, 500, 800, 1200, 2000, or 3000 ul l " 1 carbon dioxide 55 3.10 Leaf weight over time of plants grown at 340, 500, 800, 1200, 2000, or 3000 ul l " 1 carbon dioxide 56 3.11 Ratio of the growth rate of leaf weight to that of the remainder of the plant over time, in plants grown at 340, 500, 800, 1200, 2000, or 3000 ul 1"1 carbon dioxide 58 3.12 Specific leaf weight over time of plants grown at 340, 500, 800, 1200, 2000, or 3000 ul l " 1 carbon dioxide 59 ix 3.13 Specific leaf weight over time of the individual leaves of plants grown at 340 (broken line) or 1200 (solid line) ul 1 _ 1 carbon dioxide 61 4.1 Response of net carbon dioxide exchange rate of the f i r s t t r i f o l i a t e of enriched (1400 ul l " 1 ) and control (340 ul l " 1 ) plants (at PI=5.0) to intercellular space carbon dioxide concentration.. . .75 4.2 Response of the net carbon dioxide exchange rate of the f i r s t t r i f o l i a t e of enriched (1400 ul and control (340 ul l " 1 ) plants to photosynthetic photon flux density 79 4.3 Response of the net carbon dioxide exchange rate, expressed per unit chlorophyll , of the f i r s t t r i f o l i a t e of enriched (1400 ul 1 ~ 1 ) and control (340 ul l " 1 ) plants, to photosynthetic photon flux density 83 4.4 Response of net carbon dioxide exchange rate of the f i r s t t r i f o l i a t e of enriched (1400 ul l " 1 ) and control (340 ul 1 _ 1 ) plants (PI>5.0) to intercellular space carbon dioxide concentration.. . .86 4.5 Relationship between net carbon dioxide exchange rate and starch content of the f i r s t t r i f o l i a t e of enriched (1400 or 3000 ul l " 1 ) and control (340 ul l ' 1 ) plants 97 4.6 Time-course of the response of leaf water vapour and carbon dioxide exchange to a light-dark transition 103 5.1 Photograph of the primary leaves of enriched (1400 ul l " 1 ) and control (340 ul 1 " 1 ) plants grown for 20 days at 20 C 117 5.2 Time-course of net carbon dioxide exchange rate of primary leaves of enriched (1400 ul l " 1 ) (1400 ul I" 1) and control (340 ul l " 1 ) plants grown at 20 C 118 5.3 Photograph of the central leaflet of the f i r s t t r i f o l i a t e leaf of enriched (1400 ul l " 1 ) and control (340 ul l " 1 ) plants grown for 20 days at 20 C 121 5.4 Time-course of stomatal resistance of primary leaves after transfer to treatment chambers at 20 C 123 X 5.5 T ime-course of c h l o r o p h y l l con ten t and net carbon d i o x i d e exchange r a t e of shaded and unshaded pr imary l eaves of p l a n t s grown at 1400 u l l " 1 carbon d i o x i d e and at 20 C 126 5.6 C h l o r o p h y l l content of p r imary l eaves grown at 340 or 1400 u l l " 1 carbon d i o x i d e , and 90 or 370 umol m" 2 s " 1 p h o t o s y n t h e t i c photon f l u x d e n s i t y f o r 17 days 127 5.7 R e l a t i o n s h i p between s t a r c h conten t and s p e c i f i c l e a f weight of the pr imary l e aves 131 x i L i s t of A b b r e v i a t i o n s AbA . . . a b s c i s i c a c i d ADP adenos ine d iphospha te ATP adenos ine t r i p h o s p h a t e C i i n t e r c e l l u l a r carbon d i o x i d e concen t r a t ion Co carbon d i o x i d e c o n c e n t r a t i o n e x t e r n a l to the l e a f CER . . . n e t carbon d i o x i d e exchange r a t e Ch i c h l o r o p h y l l dLw change in l e a f weight dRw change i n the weight of the remainder of the p l a n t dt change i n t ime E u n i t l e a f r a t e F l e a f a rea r a t i o G a b s o l u t e growth r a t e J(H20) f l u x of water vapour J (C02) f l u x of carbon d i o x i d e kPa k i l o p a s c a l s La l e a f a rea LAD l e a f a rea d u r a t i o n Lw l e a f weight LWR l e a f weight r a t i o P i phosphate PI P l a s t o c h r o n Index PPFD p h o t o s y n t h e t i c photon f l u x d e n s i t y R r e l a t i v e growth r a t e x i i Rm mesophy l l r e s i s t a n c e RPP c y c l e . . . . r e d u c t i v e pentose phosphate c y c l e Rs s tomata l r e s i s t a n c e RuBP r ibu lose-1 ,5-bisphosphate SLW s p e c i f i c l e a f weight U D P - g l u c o s e . . u r i d i n e d iphosphate g lucose x i i i Acknowledgement A number of p e o p l e have c o n t r i b u t e d s i g n i f i c a n t l y t o t h i s s t u d y . I am g r a t e f u l t o my s u p e r v i s o r y c o m m i t t e e , namely D r . V.C. R u n e c k l e s , D r . F.B. H o l l , and D r . N.R. B u l l e y f o r t h e i r s u g g e s t i o n s and e n c o u r a g e m e n t . I w o u l d e s p e c i a l l y l i k e t o thank my a d v i s o r , D r . P.A. J o l l i f f e , f o r h i s h e l p f u l d i s c u s s i o n and m o r a l s u p p o r t . I am a l s o i n d e b t e d t o P e t e r G a r n e t t and B r i a n M c M i l l a n f o r t h e i r i n v a l u a b l e and e x p e d i t i o u s t e c h n i c a l a s s i s t a n c e , and t o J i m B e a t o n who h e l p e d i n t h e c o n s t r u c t i o n o f t h e e q u i p m e n t . I am a l s o v e r y g r a t e f u l t o t h e K i l l a m F o u n d a t i o n and t o t h e N a t u r a l S c i e n c e s and E n g i n e e r i n g R e s e a r c h C o u n c i l o f Canada f o r t h e i r g e n e r o u s f i n a n c i a l s u p p o r t i n t h e form o f a P r e d o c t o r a l F e l l o w s h i p and a P o s t g r a d u a t e S c h o l a r s h i p r e s p e c t i v e l y . L a s t l y , I would l i k e t o thank Anne E h r e t f o r h e r a s s i s t a n c e i n t h e p r e p a r a t i o n of t h e m a n u s c r i p t , and f o r h e r e n d l e s s s u p p p o r t . 1 I . GENERAL INTRODUCTION I t i s w e l l known t h a t a i r i s a f i n e e l a s t i c k f l u i d , w i t h p a r t i c l e s o f v e r y d i f f e r e n t n a t u r e s f l o a t i n g i n i t , whereby i t i s a d m i r a b l y f i t t e d by t h e g r e a t a u t h o r of n a t u r e , t o be t h e b r e a t h o f l i f e , o f v e g e t a b l e s , a s w e l l as of a n i m a l s , w i t h o u t w h i c h t h e y c an no more l i v e , nor t h r i v e t h a n a n i m a l s c a n . S t e p h e n H a l e s , 1727 Ca r b o n d i o x i d e i s , b i o l o g i c a l l y , one of t h e most i m p o r t a n t g a s e s i n t h e e a r t h ' s a t m o s p h e r e ; i t i s a l s o one of t h e l e a s t a b u n d a n t . O n l y c a r b o n d i o x i d e and i t s d i s s o l v e d i o n s c an be u t i l i z e d a s a s o u r c e of c a r b o n by a u t o t r o p h i c o r g a n i s m s , t h e p r i m a r y p r o d u c e r s i n t h e g l o b a l e c o s y s t e m . The a b i l i t y o f p h o t o s y n t h e t i c a u t o t r o p h s t o i n c o r p o r a t e l a r g e q u a n t i t i e s o f c a r b o n , d e r i v e d f r o m a t m o s p h e r i c c a r b o n d i o x i d e a t ambient c o n c e n t r a t i o n s o f 340 m i c r o l i t r e s p e r l i t r e 1 i s t r u l y , a r e m a r k a b l e e v o l u t i o n a r y a c h i e v e m e n t . C o n s i d e r i n g t h e i m p o r t a n c e of c a r b o n d i o x i d e t o p l a n t l i f e , i t s h o u l d n o t be s u r p r i s i n g t h a t c a r b o n d i o x i d e i n f l u e n c e s a range of p h y s i o l o g i c a l p r o c e s s e s i n h i g h e r p l a n t s . C a r b o n d i o x i d e c o n c e n t r a t i o n i n f l u e n c e s s e v e r a l a s p e c t s o f th e p h o t o s y n t h e t i c p r o c e s s . C a r b o n d i o x i d e i s a s u b s t r a t e of th e r e d u c t i v e p e n t o s e p h o s p h a t e (RPP) c y c l e o f p h o t o s y n t h e s i s . The c o n c e n t r a t i o n of c a r b o n d i o x i d e i s t h e r e f o r e a d e t e r m i n a n t 1 T h e n o t a t i o n u s e d t o e x p r e s s c a r b o n d i o x i d e c o n c e n t r a t i o n i n t h i s s t u d y i s m i c r o l i t r e s of c a r b o n d i o x i d e p e r l i t r e of a i r ( u l I " 1 ) . A t 25 C and 1 a t m o s p h e r e p r e s s u r e , t h e d e n s i t y of c a r b o n d i o x i d e i s 1.81 ug l " 1 . Thus 340 u l 1" 1=615 ug l " 1 . 2 of t h e r a t e of t r u e p h o t o s y n t h e s i s , w h i c h may be d e f i n e d a s p h o t o s y n t h e t i c c a r b o n a s s i m i l a t i o n i n t h e a b s e n c e o f p r o c e s s e s i n t h e l e a f w h i c h r e s u l t i n t h e e v o l u t i o n o f c a r b o n d i o x i d e . P h o t o r e s p i r a t i o n i s one s u c h p r o c e s s w h i c h c a u s e s r e l e a s e o f c a r b o n d i o x i d e i n t h e l i g h t , and t h e r e b y r e d u c e s t h e r a t e of n e t p h o t o s y n t h e s i s . However, c a r b o n d i o x i d e c o n c e n t r a t i o n a l s o i n f l u e n c e s n e t p h o t o s y n t h e s i s by r e d u c i n g t h e p e r c e n t a g e i n h i b i t i o n of t r u e p h o t o s y n t h e s i s by p h o t o r e s p i r a t i o n ( F o c k e t a l . , 1979). C a r b o n d i o x i d e n o t o n l y a c t s a s s u b s t r a t e i n p h o t o s y n t h e s i s , but a c t i v a t e s r i b u l o s e b i s p h o s p h a t e c a r b o x y l a s e (RuBP c a r b o x y l a s e ) , an i m p o r t a n t enzyme i n t h e RPP c y c l e ( L o r i m e r e t a l . , 1978), and i s needed as b i c a r b o n a t e i o n i n t h e o p e r a t i o n of P h o t o s y s t e m I I and oxygen e v o l u t i o n ( S t e m l e r and G o v i n d j e e , 1973; Khanna e t a l . , 1980). I t may even f u n c t i o n i n 1 i g h t - i n d u c e d c h l o r o p l a s t movements (Walczak and G a b r y s , 1981). Carbon d i o x i d e a l s o r e g u l a t e s s t o m a t a l a p e r t u r e ( R a s h c k e , 1975). C a r b o n d i o x i d e c a u s e s s t o m a t a l c l o s u r e and an i n c r e a s e i n s t o m a t a l d i f f u s i v e r e s i s t a n c e . The i n t e r c e l l u l a r c a r b o n d i o x i d e c o n c e n t r a t i o n a p p e a r s t o be more i m p o r t a n t t h a n t h e c a r b o n d i o x i d e c o n c e n t r a t i o n e x t e r n a l t o t h e l e a f i n r e g u l a t i n g s t o m a t a l a p e r t u r e . In many s p e c i e s , an i n c r e a s e d c a r b o n d i o x i d e c o n c e n t r a t i o n w i l l t e n d t o i n c r e a s e water use e f f i c i e n c y ( t h e r a t i o of c a r b o n a s s i m i l a t e d t o water t r a n s p i r e d ) . C a r b o n d i o x i d e i n t e r a c t s w i t h e t h y l e n e i n r e g u l a t i n g a s p e c t s o f p l a n t d e v e l o p m e n t , a l t h o u g h t h e n a t u r e o f t h e i n t e r a c t i o n i s s t i l l c o n t r o v e r s i a l . C a r b o n d i o x i d e i s known t o c o m p e t i t i v e l y i n h i b i t e t h y l e n e a c t i o n i n d e v e l o p m e n t a l p r o c e s s e s 3 s u c h as f l o r a l se-nescence, f r u i t r i p e n i n g , and s e e d g e r m i n a t i o n ( A b e l e s , 1973). C a r b o n d i o x i d e a l s o i n c r e a s e s e t h y l e n e r e l e a s e from i l l u m i n a t e d l e a v e s ( B a s s i and S p e n c e r , 1982). R e c e n t l y i t has been p r o p o s e d t h a t p h o t o s y n t h e s i s i s r e q u i r e d f o r e t h y l e n e s y n t h e s i s and i t s s u b s e q u e n t r e l e a s e from l e a v e s ( G r o d z i n s k i et. a l . , 1 9 8 3 ) . C a r b o n d i o x i d e c o u l d i n f l u e n c e t h i s p r o c e s s t h r o u g h an e f f e c t on p h o t o s y n t h e s i s r a t e . C a r b o n d i o x i d e a l s o i n f l u e n c e s d e v e l o p m e n t a l p r o c e s s e s s u c h as f l o w e r i n g ( B a s s i e t a l . , 1976; H i c k l e n t o n and J o l l i f f e , 1980a), and f r u i t r i p e n i n g ( A b e l e s , 1973). T h o s e e f f e c t s , a l o n g w i t h t h e i n h i b i t i o n of e t h y l e n e a c t i o n , a p p e a r t o be non-p h o t o s y n t h e t i c i n n a t u r e . In a d d i t i o n t o p h o t o s y n t h e t i c and d e v e l o p m e n t a l e f f e c t s , t h e r e i s e v i d e n c e t h a t h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n p r o m o t e s r e s p i r a t o r y gas exchange i n n o n - p h o t o s y n t h e t i c t i s s u e ( P e r e z - T r e j o e t a l . , 1981). I t s h o u l d be p o i n t e d out t h a t t h e c o n c e n t r a t i o n s of c a r b o n d i o x i d e r e q u i r e d t o i n f l u e n c e some of t h e s e p r o c e s s e s v a r y s u b s t a n t i a l l y . S t o m a t a l a p e r t u r e and n e t p h o t o s y n t h e s i s r e s p o n d t o c a r b o n d i o x i d e c o n c e n t r a t i o n s f r o m 0 t o a p p r o x i m a t e l y 2000 u l l " 1 , d e p e n d i n g on t h e l i g h t i n t e n s i t y ; d e v e l o p m e n t a l p r o c e s s e s , s u c h as i n h i b i t i o n o f e t h y l e n e a c t i o n , g e n e r a l l y r e q u i r e an o r d e r o f m a g n i t u d e g r e a t e r c a r b o n d i o x i d e c o n c e n t r a t i o n (10,000 t o 20,000 u l 1 " 1 ) . In s p i t e of t h e many and v a r i e d a c t i o n s o f c a r b o n d i o x i d e , s u r p r i s i n g l y l i t t l e work, p a r t i c u l a r l y o f a p h y s i o l o g i c a l n a t u r e , has been d i r e c t e d t o w a r d s t u d y i n g t h e e f f e c t s of l o n g 4 term e x p o s u r e o f p l a n t s t o h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n s . To d a t e , much o f t h e work has f o c u s s e d on t h e c o m m e r c i a l a p p l i c a t i o n s of i n c r e a s i n g g r o w t h o f h o r t i c u l t u r a l c r o p s t h r o u g h e n r i c h m e n t o f g r e e n h o u s e a t m o s p h e r e s w i t h c a r b o n d i o x i d e . As e a r l y as t h e 1930's, g r e e n h o u s e s i n E u r o p e , p a r t i c u l a r l y i n Germany, were u s i n g c a r b o n d i o x i d e on a r o u t i n e b a s i s ( W i t t w e r , 1967), w i t h w i d e - s p r e a d use becoming e v i d e n t i n N o r t h A m e r i c a and E u r o p e i n t h e e a r l y 1960's. In r e c e n t y e a r s , an i n c r e a s i n g number o f c a r b o n d i o x i d e e n r i c h m e n t s t u d i e s have been c o n d u c t e d on f i e l d c r o p s ( e g . H e s k e t h e t a l . , 1971; Wong, 1979). The i n c r e a s e d i n t e r e s t i n c a r b o n d i o x i d e c o n c e n t r a t i o n e f f e c t s on f i e l d c r o p s a r i s e s , i n p a r t , from t h e f a c t t h a t t h e a t m o s p h e r i c c a r b o n d i o x i d e c o n c e n t r a t i o n i s i n c r e a s i n g . L a r g e l y b e c a u s e o f t h e b u r n i n g of f o s s i l f u e l s , and t o a l e s s e r e x t e n t b e c a u s e of d e f o r e s t a t i o n of t r o p i c a l r a i n f o r e s t s , t h e p r e s e n t c a r b o n d i o x i d e c o n c e n t r a t i o n o f a p p r o x i m a t e l y 340 u l 1~ 1 i s e x p e c t e d t o d o u b l e by t h e e a r l y o r m i d d l e p a r t o f t h e n e x t c e n t u r y ( B a c h , 1980). Carbon d i o x i d e d e r i v e d f r o m f o s s i l f u e l s amounted t o 0.1 Gt i n 1860 and i s now 5 Gt p e r y e a r ( B a c h , 1980) ( 1 G t = 1 0 9 t o n ) . D e f o r e s t a t i o n i n f l u e n c e s a t m o s p h e r i c c a r b o n d i o x i d e c o n c e n t r a t i o n by r e d u c i n g t h e c a p a c i t y of t h e b i o s p h e r e t o a b s o r b c a r b o n ( t h r o u g h p h o t o s y n t h e s i s ) , and a l s o d i r e c t l y c o n t r i b u t e s c a r b o n d i o x i d e t o t h e a t m o s p h e r e by t h e o x i d a t i o n of s o i l o r g a n i c m a t t e r i n c l e a r e d l a n d . A b a l a n c e s h e e t o f c a r b o n i n p u t ( v i a p h o t o s y n t h e s i s ) and c a r b o n o u t p u t ( v i a r e s p i r a t i o n ) f o r t h e b i o s p h e r e s u g g e s t s t h a t t e r r e s t r i a l e c o s y s t e m s , on a g l o b a l s c a l e , a p p e a r t o be a c t i n g as a net s o u r c e of c a r b o n . 5 The o c e a n s , on t h e o t h e r hand, a r e a m a j o r s i n k f o r b o t h o r g a n i c and i n o r g a n i c forms o f c a r b o n ( t h e l a t t e r as d i s s o l v e d c a r b o n d i o x i d e and c a r b o n a t e s ) . The p r e s e n t day a t m o s p h e r i c c a r b o n d i o x i d e c o n c e n t r a t i o n i s i n c r e a s i n g b e c a u s e s o u r c e o u t p u t e x c e e d s t h e c a p a c i t y of s i n k s t o a b s o r b t h e c a r b o n . In a d d i t i o n t o t h e p o s s i b l e e f f e c t s of i n c r e a s e d a t m o s p h e r i c c a r b o n d i o x i d e c o n c e n t r a t i o n on p l a n t g r o w t h and p h y s i o l o g y , and h e n c e , a g r i c u l t u r a l p r o d u c t i v i t y , i n c r e a s i n g c a r b o n d i o x i d e may a l s o i n d i r e c t l y a f f e c t g l o b a l a g r i c u l t u r e . D r a m a t i c s h i f t s i n c l i m a t i c and a g r i c u l t u r a l z o n e s a r e e x p e c t e d t o o c c u r b e c a u s e o f t h e p r e d i c t e d warming o f t h e e a r t h ' s a t m o s p h e r e by 2 t o 3.5 C f o r a d o u b l i n g of c a r b o n d i o x i d e c o n c e n t r a t i o n ( s e e Hansen e t a l • , 1981), t h e s o - c a l l e d " g r e e n h o u s e e f f e c t " . An i n c r e a s i n g a t m o s p h e r i c c a r b o n d i o x i d e c o n c e n t r a t i o n i s a r e a l i t y of our t i m e . F o r t h i s r e a s o n a l o n e , i t i s i m p o r t a n t t o g a i n an u n d e r s t a n d i n g of how l i v i n g o r g a n i s m s r e s p o n d t o l o n g -t e r m e x p o s u r e t o e l e v a t e d c a r b o n d i o x i d e c o n c e n t r a t i o n s . We a r e a l r e a d y b e n e f i t i n g from s u c h s t u d i e s i n terms o f i n c r e a s e d c r o p p r o d u c t i o n i n g r e e n h o u s e s e n r i c h e d w i t h c a r b o n d i o x i d e . E q u a l l y i m p o r t a n t i s t h e i n s i g h t t o be g a i n e d f r o m t h o s e s t u d i e s i n u n d e r s t a n d i n g how p l a n t s f u n c t i o n . The work r e p o r t e d i n t h i s t h e s i s d e a l s w i t h t h e e f f e c t s of l o n g - t e r m e n r i c h m e n t w i t h c a r b o n d i o x i d e on t h r e e s e p a r a t e , but i n t e r r e l a t e d , a s p e c t s o f p l a n t f u n c t i o n . In C h a p t e r 3, i n v e s i g a t i o n s o f t h e g r o w t h o f p l a n t s and t h e p a r t i t i o n i n g of 6 c a r b o n among p l a n t p a r t s as a f f e c t e d by c o n t i n u o u s c a r b o n d i o x i d e e n r i c h m e n t w i l l be r e p o r t e d . T h o s e i n v e s t i g a t i o n s d e t a i l t h e g r o w t h r e s p o n s e of bush beans o v e r t h e c o u r s e of d e v e l o p m e n t from a v e r y young v e g e t a t i v e g r o w t h s t a g e , t o t h e n e a r - c o m p l e t i o n of r e p r o d u c t i v e g r o w t h . Growth a n a l y s i s was u s e d t o measure p l a n t g r o w t h r e s p o n s e t o 6 c a r b o n d i o x i d e c o n c e n t r a t i o n s r a n g i n g f r o m 340 t o 3000 u l 1" 1 . C h a p t e r 4 d e s c r i b e s t h e p h y s i o l o g y o f c a r b o n d i o x i d e e x change i n l e a v e s from bean p l a n t s grown a t d i f f e r e n t c a r b o n d i o x i d e c o n c e n t r a t i o n s . S i n c e p h o t o s y n t h e s i s i s s u c h a f u n d a m e n t a l a s p e c t of p l a n t g r o w t h and d e v e l o p m e n t , and i s known t o r e s p o n d t o c a r b o n d i o x i d e c o n c e n t r a t i o n , i t was p e r t i n e n t t o compare gas e xchange c h a r a c t e r i s t i c s o f p l a n t s .grown i n an e n r i c h e d c a r b o n d i o x i d e a t m o s p h e r e w i t h t h o s e grown i n u n e n r i c h e d c o n d i t i o n s . The c o n d i t i o n s and c i r c u m s t a n c e s o f t h e r e l a t i v e c h a n g e s i n gas e xchange c h a r a c t e r i s t i c s of e n r i c h e d and c o n t r o l p l a n t s were e x a m i n e d . L a s t l y , d u r i n g t h e c o u r s e of t h i s s t u d y , t h e o b s e r v a t i o n was made t h a t c a r b o n d i o x i d e e n r i c h m e n t c a u s e d e x t e n s i v e c h l o r o s i s o f s p e c i f i c l e a v e s . C h a p t e r 5 d e a l s w i t h some e n v i r o n m e n t a l and p h y s i o l o g i c a l a s p e c t s of l e a f s e n e s c e n c e i n e n r i c h e d p l a n t s . T h i s a s p e c t o f c a r b o n d i o x i d e e n r i c h m e n t has not been p r e v i o u s l y s t u d i e d i n a r i g o r o u s manner. As a f i r s t s t e p , i t was r e l e v e n t t o d e t e r m i n e t h e e n v i r o n m e n t a l c o n d i t i o n s u nder w h i c h t h i s e f f e c t of h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n o c c u r r e d . 7 I I . LITERATURE REVIEW 2.1 PLANT RESPONSE TO CARBON DIOXIDE ENRICHMENT 2.1.1 H i s t o r i c a l P e r s p e c t i v e Over t h r e e h u n d r e d y e a r s ago, J e a n - B a p t i s t e van Helmont c o n d u c t e d what w o u l d become a m i l e s t o n e e x p e r i m e n t i n t h e h i s t o r y of p l a n t s c i e n c e . He f o u n d t h a t a w i l l o w t r e e g r o w i n g f o r 5 y e a r s i n 200 pounds of s o i l i n c r e a s e d i n w e i g h t 164 pounds w h i l e s o i l w e i g h t c h a n g e d n e g l i g i b l y . He c o n c l u d e d t h a t t h e w e i g h t g a i n d e r i v e d from water t a k e n up from t h e s o i l and t h a t t h e s o i l i t s e l f c o n t r i b u t e d v e r y l i t t l e t o p l a n t d r y m a t t e r . S t e p h e n H a l e s i n 1727 s u r m i s e d t h a t some of t h e w e i g h t g a i n d e r i v e d f r o m t h e a i r . T h i s i d e a was d e v e l o p e d by S e n e b i e r and I n g e n h o u s z ( R a b i n o w i t c h and G o v i n d j e e , 1969); t h e component of t h e a i r w h i c h p l a n t s i n c o r p o r a t e was d e s c r i b e d i n t e r m s of t h e new c h e m i s t r y of L a v o u s i e r as c a r b o n d i o x i d e ( R a b i n o w i t c h and G o v i n d j e e , 1969). Brown and Escombe (1902) were p r o b a b l y t h e f i r s t t o e x p l o r e t h e e f f e c t s o f h i g h e r t h a n ambient c o n c e n t r a t i o n s of c a r b o n d i o x i d e on p l a n t s . They s u b j e c t e d s e v e r a l h o r t i c u l t u r a l s p e c i e s t o 1147 u l l " 1 c a r b o n d i o x i d e and f o u n d some r a t h e r d r a m a t i c n e g a t i v e e f f e c t s . P l a n t s r e s p o n d e d t o c a r b o n d i o x i d e by s h e d d i n g f l o w e r buds p r e m a t u r e l y , and by t h e s h e d d i n g and c u r l i n g o f l e a v e s . A c c o r d i n g t o Demoussey ( s e e W i t t w e r and Robb, 1964) t h i s was p e r h a p s due t o i m p u r i t i e s i n t h e a i r . In 1902-04, u s i n g a d i f f e r e n t t e c h n i q u e t o p r o d u c e c a r b o n d i o x i d e , Demoussey ( s e e W i t t w e r and Robb, 1964) f o u n d l a r g e i n c r e a s e s i n 8 p l a n t d r y w e i g h t v a r y i n g f r o m 97% f o r f u c h s i a t o 262% f o r g e r a n i u m , a t h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n s . By t h e 1920's and 30's, work was b e i n g done w i t h i n c r e a s i n g e m p h a s i s on t h e u s e s o f c a r b o n d i o x i d e i n g l a s s h o u s e c r o p p r o d u c t i o n ( W h i t e , 1930; S m a l l and W h i t e , 1930; B o l a s and M e l v i l l e , 1935). Work of t h e 1950's and 60's was s p u r r e d by t h e c l a s s i c s t u d i e s by G a a s t r a (1963) and by Waggoner, Moss and H e s k e t h (1963) i n t o t h e e f f e c t s of c a r b o n d i o x i d e c o n c e n t r a t i o n on t h e ne t r a t e of p h o t o s y n t h e s i s . To d a t e , a g r e a t d e a l of i n f o r m a t i o n has become a v a i l a b l e d e s c r i b i n g t h e e f f e c t s of c a r b o n d i o x i d e on p h o t o s y n t h e s i s and t h e e f f e c t s of c a r b o n d i o x i d e e n r i c h m e n t on p l a n t p r o d u c t i v i t y . However, r e l a t i v e l y l i t t l e i s known about t h e p h y s i o l o g y o f p l a n t s grown i n c a r b o n d i o x i d e e n r i c h e d a t m o s p h e r e s . 2.1.2 C a r b o n D i o x i d e E n r i c h m e n t And P l a n t G rowth In v i e w of t h e c e n t r a l r o l e of c a r b o n d i o x i d e i n p h o t o s y n t h e s i s , and hence d r y m a t t e r p r o d u c t i o n , .numerous s t u d i e s have been u n d e r t a k e n t o d e s c r i b e t h e e f f e c t s of c a r b o n d i o x i d e e n r i c h m e n t on y i e l d of p l a n t s . Much o f t h a t work has been done i n r e l a t i o n t o e n r i c h m e n t of g r e e n h o u s e c r o p s b e c a u s e of t h e p o t e n t i a l b e n e f i t s o f c o m m e r c i a l c a r b o n d i o x i d e a p p l i c a t i o n s . Work on tomato ( W i t t w e r and Robb, 1964; Madsen, 1973a, 1973b, 1974; C a l v e r t and S l a c k , 1976; H i c k l e n t o n and J o l l i f f e , 1978), l e t t u c e ( W i t t w e r and Robb, 1964), and cucumber ( A o k i and Y a b u k i , 1977) has been done on t h e e f f e c t of c a r b o n d i o x i d e e n r i c h m e n t on d r y w e i g h t a c c u m u l a t i o n . S i m i l a r l y , f i e l d 9 c r o p s s u c h as soybean (Hardman and B r u n , 1971; Mauney e t a l . , 1978) , c o t t o n (Mauney e t a l . , 1978; Wong, 1979), s u g a r b e e t (Wyse, 1980), wheat ( G i f f o r d , 1977; S i o n i t e t a l . , 1981a), c o r n (Wong, 1979), and r i c e (Imai and M u r a t a , 1976, 1977, 1978b) have been s t u d i e d . B o t h t o t a l d r y w e i g h t and a g r o n o m i c y i e l d a r e u s u a l l y i n c r e a s e d by c a r b o n d i o x i d e e n r i c h m e n t . F o l l o w i n g a r e a few e x a m p l e s . I n c r e a s e s i n tomato d r y w e i g h t o f 100% o c c u r r e d a f t e r 42 d a y s i n 800 t o 2000 u l l " 1 c a r b o n d i o x i d e ( W i t t w e r and Robb, 1964). Y i e l d s were i n c r e a s e d 29% t o 71% and t h e q u a l i t y of f r u i t was i m p r o v e d . H i c k l e n t o n and J o l l i f f e (1978) a l s o f o u n d t o m a t o e s grown a t 900 u l 1~ 1 c a r b o n d i o x i d e p r o d u c e d 30% more m a r k e t a b l e f r u i t . In s u g a r b e e t , d r y w e i g h t i n c r e a s e d 180% i n h i g h c a r b o n d i o x i d e (Wyse, 1980). E n r i c h m e n t t o 500 u l 1~ 1 i n c r e a s e d d r y . w e i g h t 32% i n wheat; a d e c r e a s e i n c a r b o n d i o x i d e t o 150 u l l " 1 r e d u c e d d r y w e i g h t 43% ( G i f f o r d , 1977). G r a i n y i e l d i n c r e a s e d 43% and d e c r e a s e d 44% i n t h e s e two t r e a t m e n t s r e s p e c t i v e l y . C o t t o n showed a 2 - f o l d i n c r e a s e i n d r y w e i g h t a f t e r 40 d a y s a t 640 u l l " 1 c a r b o n d i o x i d e w h i l e c o r n showed o n l y a 20% i n c r e a s e a t t h e same l e v e l of e n r i c h m e n t (Wong, 1979) . In a d d i t i o n t o t o t a l y i e l d and r e p r o d u c t i v e y i e l d , v a r i o u s o t h e r f e a t u r e s of p l a n t g r o w t h have been a n a l y s e d i n r e l a t i o n t o a t m o s p h e r i c c a r b o n d i o x i d e e n r i c h m e n t . R e l a t i v e growth r a t e (R) i n c r e a s e d i n r e s p o n s e t o c a r b o n d i o x i d e e n r i c h m e n t i n c o t t o n (Mauney e t a l . , 1 9 7 8 ) , wheat ( N e a l e s and N i c h o l l s , 1978), and tomato ( H i c k l e n t o n and J o l l i f f e , 1978). 10 U n i t l e a f r a t e (E) a l s o i n c r e a s e s as a r e s u l t o f c a r b o n d i o x i d e e n r i c h m e n t . I n c r e a s e s have been o b s e r v e d i n cucumber ( A o k i and Y a b u k i , 1977), tomato ( H i c k l e n t o n and J o l l i f f e , 1978), s o y b e a n ( P a t t e r s o n and F l i n t , 1980, 1982; G u s t a f s o n and B r e e n , 1982), wheat ( N e a l e s and N i c h o l l s , 1978), c o t t o n (Wong 1979), sorghum (Mauney e t a l . , 1978), s u n f l o w e r (Mauney e t a l . , 1978), and Desmodium ( W u l f f and S t r a i n , 1982). In one i n s t a n c e , E was f o u n d t o d e c r e a s e i n e n r i c h e d p l a n t s (Raper and P e e d i n , 1978). As w i l l be d i s c u s s e d i n a l a t e r s e c t i o n , t h i s l a t t e r c a s e i s p r o b a b l y a f u n c t i o n of p l a n t age o r t h e d u r a t i o n o f t r e a t m e n t . T h e r e i s g e n e r a l l y a good c o r r e l a t i o n between t h e e f f e c t s o f c a r b o n d i o x i d e e n r i c h m e n t on u n i t l e a f r a t e and r e l a t i v e g r o w t h r a t e . ( A o k i and Y a b u k i , 1977; N e a l e s and N i c h o l l s , 1978; P a t t e r s o n and F l i n t , 1980, 1982; G u s t a f s o n and B r e e n , 1982; W u l f f and S t r a i n , 1982). However, i t must be k e p t i n mind t h a t s i n c e R i s a f u n c t i o n o f b o t h E and l e a f a r e a r a t i o ( F ) , an i n c r e a s e i n E a t h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n does not n e c e s s a r i l y t r a n s l a t e i n t o a p r o p o r t i o n a t e l y g r e a t e r R; c h a n g e s i n l e a f a r e a must a l s o be c o n s i d e r e d . F o r example, Hurd (1968) f o u n d t h a t e n r i c h e d t o m a t o e s i n i t i a l l y showed i n c r e a s e d R and E, but s u b s e q u e n t l y showed no d i f f e r e n c e i n R w i t h r e s p e c t t o c o n t r o l s , d e s p i t e a c o n t i n u a l l y g r e a t e r E. H e r e , R was b e i n g i n f l u e n c e d by a d e c r e a s i n g F. An i n c r e a s e i n t o t a l l e a f a r e a p e r p l a n t due t o e n r i c h m e n t ( F o r d and T h o r n e , 1967; Mauney e t a l . , 1978; Wong, 1979; Wyse, 1980; P a t t e r s o n and F l i n t , 1982; W u l f f and S t r a i n , 1982) and l e a f number p e r p l a n t ( F o r d and T h o r n e , 1967; G i f f o r d , 1977) 11 r e f l e c t s a change i n a b s o l u t e g r o w t h r a t e , and as s u c h i s n o t a r e l i a b l e measure o f t h e l e a f a r e a i n r e l a t i o n t o t h e s i z e o f t h e p l a n t . L e a f a r e a r a t i o i s a b e t t e r e s t i m a t e , and i s f o u n d t o d e c r e a s e i n r e s p o n s e t o e n r i c h m e n t ( F o r d and T h o r n e , 1967; Hughes and Freeman, 1967; T o g n o n i e t a l . , 1967; N e a l e s and N i c h o l l s , 1978; P a t t e r s o n and F l i n t , 1982; W u l f f and S t r a i n , 1982). T h e r e i s much e v i d e n c e t h e n , t h a t c a r b o n d i o x i d e e n r i c h m e n t r e s u l t s i n an i n c r e a s e i n t o t a l d r y w e i g h t , R, and E. However, p l a n t r e s p o n s e s t o e n r i c h m e n t a r e somewhat more complex t h a n w o u l d be a n t i c i p a t e d b a s e d on t h i s i n f o r m a t i o n a l o n e . Some i m p o r t a n t a d d i t i o n a l f a c t o r s w i l l now be c o n s i d e r e d . 2.1.3 D u r a t i o n Of E n r i c h m e n t T h e r e i s some e v i d e n c e t h a t t h e l e n g t h o f t i m e t h e p l a n t i s e x p o s e d t o e l e v a t e d c a r b o n d i o x i d e a f f e c t s t h e r e s p o n s e . F o r example, g r o w t h o f cucumber under c a r b o n d i o x i d e e n r i c h m e n t was 1.5 t o 1.8 t i m e s g r e a t e r t h a n t h a t of c o n t r o l s ( A o k i and Y a b u k i , 1977), but as t h e t r e a t m e n t d u r a t i o n i n c r e a s e d , t h e most e f f e c t i v e c o n c e n t r a t i o n f o r g r o w t h d e c r e a s e d . A l s o , r a t e s o f n e t c a r b o n d i o x i d e a s s i m i l a t i o n a t t h e h i g h e r c a r b o n d i o x i d e c o n c e n t r a t i o n s (2400 and 5500 u l I " 1 ) d e c r e a s e d t o l e v e l s below t h a t of c o n t r o l s . B e c a u s e o l d e r l e a v e s e x p e r i e n c e d a l o n g e r p e r i o d of e n r i c h m e n t t h a n y o u n g e r l e a v e s , i t i s r e a s o n a b l e t o e x p e c t t h a t t h e o l d e r l e a v e s showed t h e major d e c r e a s e i n n e t a s s i m i l a t i o n . The d a i l y d u r a t i o n of e n r i c h m e n t a l s o a f f e c t s t h e g r o w t h r e s p o n s e . Tomatoes showed i n c r e a s e s i n y i e l d r o u g h l y 1 ? proportional to the increase in enrichment time per day (Calvert and Slack, 1976). 2.1.4 Interaction With Plant Age Plant response to enrichment is influenced by developmental stage. R and E were higher during the juvenile stage (10 to 30 days) in enriched cotton, soybean, sunflower and sorghum (Mauney et a l . , 1978), but were similar to control plants at the reproductive stage. Sunflower and soybean had a decreased E r e l a t i v e to controls l a t e r , during the maturation stage of development, but s t i l l maintained greater t o t a l dry weight after 100 days of enrichment (due largely to increases in leaf area). Because the effect of enrichment is influenced by the duration of exposure, i t i s d i f f i c u l t to separate plant age e f f e c t s from duration e f f e c t s in many of the c i t e d studies. In a more d e f i n i t i v e study by Neales and Nicholls (1978), wheat plants of various ages were exposed to high carbon dioxide concentrations for the same length of time. R and E of 10 day old plants were increased 35% and 55% respectively, but were reduced 44 and 16% respectively in 24 day old plants r e l a t i v e to controls (Neales and N i c h o l l s , 1978). Although t h i s seems to indicate that young plants responded better to enrichment, Hardman and Brun (1971) found no l a s t i n g e f f e c t s of enrichment administered during the preflowering stage on the mature plant. Only enrichment during the flowering and post-flowering stages had any e f f e c t on dry weight or vegetative and reproductive characters. Krizek et a l • (1971) however, found that the stimulatory e f f e c t s of the enrichment of crabapple seedlings for 1 3 4 weeks p e r s i s t e d f o r 2 t o 3 months a f t e r t r a n s f e r t o ambient c a r b o n d i o x i d e . 2.1.5 I n t e r a c t i o n W i t h L i g h t I n t e n s i t y C a r b o n d i o x i d e e n r i c h m e n t i n c r e a s e d p l a n t g r o w t h and u n i t l e a f r a t e i n g r e e n h o u s e t o m a t o e s i n m i d - w i n t e r when l i g h t i n t e n s i t y was low ( W i t t w e r and Robb, 1964). Hopen and R i e s (1962) f o u n d t h a t h i g h c a r b o n d i o x i d e c o u l d compensate f o r low l i g h t i n t e n s i t y i n m a i n t a i n i n g g r o w t h of cucumber. S i m i l a r l y , H u r d (1968) showed t h a t a c o m b i n a t i o n o f h i g h c a r b o n d i o x i d e / l o w l i g h t p r o d u c e d t h e same growth as low c a r b o n d i o x i d e / h i g h l i g h t . The m a g n i t u d e of t h e r e s p o n s e t o e n r i c h m e n t i s d e p e n d e n t on t h e l i g h t i n t e n s i t y d u r i n g g r o w t h . Hopen and R i e s ( 1 9 6 2 ) , F o r d and T h o r n e ( 1 9 6 7 ) , and Hughes and C o c k s h u l l (1971) f o u n d t h a t e n r i c h m e n t had a p r o g r e s s i v e l y g r e a t e r e f f e c t on g r o w t h as l i g h t i n t e n s i t y i n c r e a s e d . However, o t h e r s have f o u n d a g r e a t e r s t i m u l a t i o n o f g r o w t h by c a r b o n d i o x i d e under c o n d i t i o n s of low l i g h t t h a n h i g h l i g h t ( M a c d o w a l l , 1972; G i f f o r d , 1977). M a c d o w a l l i n t e r p r e t e d t h i s as i n d i c a t i n g t h a t c a r b o n d i o x i d e i n c r e a s e d t h e e f f i c i e n c y o f n e t p h o t o s y n t h e s i s ( w h i c h would g i v e g r e a t e r n e t p h o t o s y n t h e s i s t h a n c o n t r o l s under l i m i t i n g l i g h t ) , b u t had l i t t l e o r no e f f e c t on t h e c a p a c i t y of p h o t o s y n t h e s i s (no d i f f e r e n t t h a n c o n t r o l s a t l i g h t s a t u r a t i o n ) . However, he d i d not c o n s i d e r t h e f a c t t h a t a t e l e v a t e d c a r b o n d i o x i d e , a n o r m a l l i g h t r e s p o n s e c u r v e f o r C3 p l a n t s would e x h i b i t g r e a t e r e f f i c i e n c y and g r e a t e r c a p a c i t y . T h e r e f o r e , i f p l a n t s grown a t h i g h c a r b o n d i o x i d e show the same p h o t o s y n t h e t i c c a p a c i t y as c o n t r o l s when measured under growth c a r b o n d i o x i d e 1 4 c o n c e n t r a t i o n s , t h e y p r o b a b l y have d e p r e s s e d p h o t o s y n t h e t i c c a p a c i t y i f compared t o c o n t r o l p l a n t s under t h e same m e a s u r i n g c o n d i t i o n s o f l i g h t i n t e n s i t y and c a r b o n d i o x i d e c o n c e n t r a t i o n . 2.1.6 I n t e r a c t i o n W i t h N u t r i e n t Regime A l t h o u g h an i n c r e a s e d n u t r i e n t l e v e l g e n e r a l l y i n c r e a s e s d r y m a t t e r p r o d u c t i o n , i t s i n f l u e n c e on p l a n t r e s p o n s e t o c a r b o n d i o x i d e e n r i c h m e n t i s e q u i v o c a l , and may depend on s p e c i e s . R i c e ( I m a i and M u r a t a , 1978a) and c o t t o n (Wong, 1979) showed p r o p o r t i o n a t e l y g r e a t e r d r y w e i g h t a c c u m u l a t i o n i n r e s p o n s e t o c a r b o n d i o x i d e a t h i g h e r n i t r o g e n l e v e l s ; c o r n (Wong, 1979), wheat ( S i o n i t e t a l . , 1981a), and soybean ( P a t t e r s o n and F l i n t , 1982) d i d n o t show c l e a r i n c r e a s e s i n r e s p o n s e t o n i t r o g e n . The i n c r e a s e i n E due t o c a r b o n d i o x i d e e n r i c h m e n t shows no d e p e n d e n c e on n i t r o g e n l e v e l i n r i c e o r J a p a n e s e m i l l e t (Imai and M u r a t a , 1978a), a n e g a t i v e r e l a t i o n s h i p w i t h n u t r i e n t l e v e l i n s o y b e a n ( P a t t e r s o n and F l i n t , 1982), and a p o s i t i v e r e l a t i o n s h i p w i t h n i t r o g e n l e v e l i n c o t t o n and c o r n (Wong, 1979). T h e r e a r e s e v e r a l o t h e r f a c t o r s w h i c h a c c o u n t f o r a p r o p o r t i o n a t e l y g r e a t e r i n c r e a s e i n d r y w e i g h t due t o e n r i c h m e n t a t h i g h e r n u t r i e n t l e v e l s . N u t r i e n t r e g i m e a l s o a f f e c t s t h e r e s p o n s e o f l e a f a r e a d u r a t i o n (LAD) t o c a r b o n d i o x i d e , i n c r e a s i n g i t i n s o y b e a n ( P a t t e r s o n and F l i n t , 1982) and t o some e x t e n t i n wheat ( S i o n i t e t a l . , 1981a). An i n c r e a s e d LAD i n s o y b e a n a t h i g h n u t r i e n t and c a r b o n d i o x i d e l e v e l s c ompensated f o r a d e c r e a s e d E and c o n t r i b u t e d t o g r e a t e r d r y w e i g h t a c c u m u l a t i o n ( P a t t e r s o n and F l i n t , 1982). In r i c e , d e s p i t e no 1 5 p r o p o r t i o n a l change i n E w i t h n u t r i e n t l e v e l , t i l l e r i n g was p r o m o t e d i n e n r i c h e d p l a n t s a t h i g h n u t r i e n t l e v e l s , and t h e r e b y c o n t r i b u t e d t o p r o p o r t i o n a l y g r e a t e r d r y w e i g h t i n c r e a s e (Imai and M u r a t a , 1976). 2.1.7 E f f e c t s On P a r t i t i o n i n g Of A s s i m i l a t e s C a r b o n d i o x i d e e n r i c h m e n t has a v a r i e t y of e f f e c t s on p l a n t d e v e l o p m e n t , due i n p a r t t o c h a n g i n g p a t t e r n s of a s s i m i l a t e p a r t i t i o n i n g . Many of t h e c h a n g e s i n t h e growth of p l a n t s or p l a n t p a r t s a r e a r e s u l t of an i n c r e a s e i n a b s o l u t e and r e l a t i v e g r o w t h r a t e s , and do not n e c e s s a r i l y r e f l e c t c h a n g e s i n p a r t i t i o n i n g . However, some g r o w t h p a r a m e t e r s do r e s p o n d d i f f e r e n t i a l l y t o c a r b o n d i o x i d e . F o r example, d e c r e a s e s i n F have 'a l r e a d y been m e n t i o n e d . LAD i s i n c r e a s e d by e n r i c h m e n t i n soybean ( P a t t e r s o n and F l i n t , 1982), wheat ( S i o n i t e t a l . , 1981a), and Desmodium ( W u l f f and S t r a i n , 1982). S p e c i f i c l e a f a r e a (SLA) i s d e c r e a s e d a t h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n s ( F o r d and T h o r n e , 1967; Hurd, 1968; N e a l e s and N i c h o l l s , 1978; W u l f f and S t r a i n , 1982), i n d i c a t i n g t h a t a g r e a t e r p r o p o r t i o n of f i x e d c a r b o n i s r e t a i n e d i n t h e l e a v e s under c o n d i t i o n s of h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n s . H i g h c a r b o n d i o x i d e c o n c e n t r a t i o n s g e n e r a l l y r e s u l t i n an i n c r e a s e i n t i l l e r i n g i n c e r e a l s ( F o r d and T h o r n e , 1967; Imai and M u r a t a , 1976; G i f f o r d , 1977; N e a l e s and N i c h o l l s , 1978). A g r e a t e r p r o p o r t i o n of a s s i m i l a t e i s a l s o d i r e c t e d t o r o o t g r o w t h as shown by an i n c r e a s e d r o o t / s h o o t r a t i o ( F o r d and T h o r n e , 1967; K r i e d e m a n n e t a l . , 1976, T o g n o n i , e t a l . , 1967; W i t t w e r , 1967; Wyse, 1980). In legumes, r o o t n o d u l e s a r e a major .sink f o r 1 6 p h o t o s y n t h e t i c m e t a b o l i t e s , - and a v a i l a b i l i t y of p h o t o s y n t h a t e i s a m a j o r f a c t o r i n d e t e r m i n i n g r a t e s of n i t r o g e n f i x a t i o n . ' C a r b o n d i o x i d e e n r i c h m e n t i n c r e a s e d t h e t o t a l n i t r o g e n f i x e d and t h e p e r c e n t a g e o f n i t r o g e n d e r i v e d f r o m a t m o s p h e r i c n i t r o g e n i n s o y b e a n and pea (Hardy and H a v e l k a , 1977), w h i c h r e s u l t e d f r o m an i n c r e a s e d n o d u l e mass and d e l a y e d n o d u l e s e n e s c e n c e . In a d d i t i o n , t h e s p e c i f i c n o d u l e a c t i v i t y was i n c r e a s e d . P h i l l i p s e t a l . (1976) f o u n d an i n c r e a s e i n s p e c i f i c n o d u l e a c t i v i t y o n l y a f t e r s h o r t t e r m c a r b o n d i o x i d e e n r i c h m e n t (6 h o u r s ) , but not a f t e r l o n g t e r m e n r i c h m e n t (4 w e e k s ) . They c o n c l u d e d t h a t a s h o r t t e r m i n c r e a s e i n c u r r e n t p h o t o s y n t h e s i s t h r o u g h a s h o r t t e r m i n c r e a s e i n c a r b o n d i o x i d e c o n c e n t r a t i o n i n c r e a s e d n i t r o g e n f i x a t i o n by a f f e c t i n g n o d u l e f u n c t i o n i n g , and t h a t l o n g t e r m c a r b o n d i o x i d e t r e a t m e n t s i n c r e a s e d n i t r o g e n f i x a t i o n by i n c r e a s i n g n o d u l e mass. However, F i n n and Brun (1982) f o u n d no i n c r e a s e i n s p e c i f i c n o d u l e a c t i v i t y i n s h o r t or l o n g t e r m c a r b o n d i o x i d e t r e a t m e n t s of s o y b e a n s . The f a c t t h a t c a r b o n d i o x i d e c a u s e s an i n c r e a s e i n n i t r o g e n f i x a t i o n o v e r t h e l o n g t e r m t h r o u g h an i n c r e a s e i n n o d u l e mass does not s u g g e s t , however, t h a t p a r t i t i o n i n g o f a s s i m i l a t e s t o n o d u l e s has been a f f e c t e d by e n r i c h m e n t . In t h i s r e s p e c t , P h i l l i p s (1976) f o u n d t h a t c a r b o n d i o x i d e d i d not a l t e r t h i s p a r t i t i o n i n g . 17 2.1.8 Response Of P h o t o s y n t h e t i c C a p a c i t y Net p h o t o s y n t h e s i s o f p l a n t s may i n c r e a s e d u r i n g e x p o s u r e t o h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n , but as m e n t i o n e d p r e v i o u s l y , t h e e f f e c t i s not c o n s i s t e n t t h r o u g h age, d e v e l o p m e n t a l s t a g e , or d u r a t i o n o f e x p o s u r e . Growth a n a l y s i s y i e l d s i n f o r m a t i o n on E o v e r d a i l y o r even w e e k l y p e r i o d s , but i s n o t w e l l s u i t e d f o r d e t e r m i n i n g s p e c i f i c c h a r a c t e r i s t i c s o f p h o t o s y n t h e s i s . R a t h e r , i t s i m p o r t a n c e l i e s i n i t s a b i l i t y t o d e s c r i b e g r o w t h c h a r a c t e r i s t i c s ( f o r example, R, E, and F) w h i c h a p p l y t o t h e p l a n t o v e r e x t e n d e d p e r i o d s . However, d e s p i t e t h e s e l i m i t a t i o n s , N e a l e s and N i c h o l l s (1978) u s e d g r o wth a n a l y s i s d a t a t o c a l c u l a t e t h a t an i n c r e a s e i n g r o w t h due t o c a r b o n d i o x i d e e n r i c h m e n t s h o u l d l e a d t o much h i g h e r i n c r e m e n t s i n d r y w e i g h t t h a n were a c t u a l l y o b s e r v e d . T h i s would r e q u i r e a s u s t a i n e d i n c r e a s e i n R, w h i c h i n f a c t d i d n o t o c c u r . R d i m i n i s h e d w i t h t i m e b e c a u s e of a d e c r e a s e i n F; a c o n c u r r e n t i n c r e a s e i n SLW was a l s o s u g g e s t e d t o c o n t r i b u t e t o a f a i l u r e t o m a i n t a i n f u l l p h o t o s y n t h e t i c p o t e n t i a l by d e c r e a s i n g l e a f a s s i m i l a t i o n r a t e s . The r e l a t i o n s h i p between SLW and net c a r b o n d i o x i d e exchange r a t e (CER), p a r t i c u l a r l y as a f f e c t e d by s t a r c h c o n t e n t , w i l l be d i s c u s s e d more f u l l y i n a l a t e r s e c t i o n . Gas e xchange a n a l y s i s , w h i l e l a c k i n g t h e i n t e g r a t e d a p p r o a c h of g r o w t h a n a l y s i s , a l l o w s measurements o f p h o t o s y n t h e t i c e f f i c i e n c y and c a p a c i t y . I t c a n d e s c r i b e n e t p h o t o s y n t h e s i s i n terms o f i t s component p r o c e s s e s . A l t h o u g h E may i n c r e a s e due t o e n r i c h m e n t , s i m p l y b e c a u s e o f t h e g r e a t e r c a r b o n d i o x i d e c o n c e n t r a t i o n a v a i l a b l e as s u b s t r a t e f o r 18 p h o t o s y n t h e s i s , t h i s does not r e f l e c t t h e i n n a t e p h o t o s y n t h e t i c c a p a c i t y of t h o s e p l a n t s compared t o u n e n r i c h e d p l a n t s . To e s t i m a t e r e l a t i v e c a p a c i t i e s , p h o t o s y n t h e s i s must be m e asured under t h e same c o n d i t i o n s of c a r b o n d i o x i d e , l i g h t , and t e m p e r a t u r e . Hence, i n my s t u d y , c a r b o n d i o x i d e exchange c a p a c i t y i s d e f i n e d as t h e a b i l i t y o f e n r i c h e d p l a n t s t o c a r r y out n e t c a r b o n d i o x i d e exchange compared t o c o n t r o l s when measured under t h e same c o n d i t i o n s o f c a r b o n d i o x i d e c o n c e n t r a t i o n , p h o t o s y n t h e t i c p h o t o n f l u x d e n s i t y (PPFD), and t e m p e r a t u r e . Gas e x c h a n g e measurements made a t t h e same c o n c e n t r a t i o n s of c a r b o n d i o x i d e show d e c r e a s e d n e t p h o t o s y n t h e s i s p e r u n i t l e a f a r e a i n e n r i c h e d p l a n t s (Ho, 1977; Mauney e t a l . , 1979; C l o u g h e t a l . , 1981; W u l f f and S t r a i n , 1982). T h i s i n d i c a t e s a r e d u c e d p h o t o s y n t h e t i c c a p a c i t y i n e n r i c h e d p l a n t s . The r e a s o n f o r t h e d e c r e a s e has been a t t r i b u t e d t o s t o m a t a l c l o s u r e i n r i c e (Imai and M u r a t a , 1978c). However, t h i s i s n o t t h e o n l y p o s s i b l e e x p l a n a t i o n . A o k i and Y a b u k i (1977) f o u n d a d u r a t i o n -d e p e n d e n t i n c r e a s e i n s t o m a t a l r e s i s t a n c e and a d e c r e a s e i n m e s o p h y l l r e s i s t a n c e i n e n r i c h e d cucumber. S i m i l a r l y , H o f s t r a and H e s k e t h (1975) showed r e d u c e d r a t e s of p h o t o s y n t h e s i s i n s o y b e a n s , p a r t i c u l a r l y i n o l d e r l e a v e s , w h i c h c o u l d a l s o be a f u n c t i o n o f d u r a t i o n of e x p o s u r e . F r y d r y c h (1976) f o u n d a r e d u c e d p h o t o s y n t h e t i c c a p a c i t y i n e n r i c h e d cucumber, b u t o n l y when measured a t h i g h l i g h t i n t e n s i t y . However, t h e measurements made a t low l i g h t i n t e n s i t y (47.5 W m~ 2) were p r o b a b l y l i g h t - l i m i t e d and d i f f e r e n c e s i n p h o t o s y n t h e t i c 19 c a p a c i t y would n o t be e x p e c t e d . W u l f f and S t r a i n ( 1 9 8 2 ) , on t h e o t h e r hand, f o u n d r e d u c e d p h o t o s y n t h e s i s a t a l l l i g h t i n t e n s i t i e s up t o 1300 umol n r 2 s " 1 . T h i s i s p r o b a b l y due t o r e d u c e d c h l o r o p h y l l l e v e l s i n e n r i c h e d p l a n t s . In f a c t , i f p h o t o s y n t h e s i s was p l o t t e d v e r s u s c h l o r o p h y l l c o n t e n t , e n r i c h e d p l a n t s showed h i g h e r r a t h e r t h a n l o w e r p h o t o s y n t h e t i c c a p a c i t y , p e r h a p s due t o c h a n g e s i n p h o t o s y n t h e t i c u n i t s i z e . C l o u g h e t a l . (1981) compared p h o t o s y n t h e t i c r a t e s of podded ( h i g h s i n k ) and depodded (low s i n k ) s o y b e a n s under two c a r b o n d i o x i d e r e g i m e s . Low s i n k p l a n t s had l o w e r p h o t o s y n t h e t i c r a t e s t h a n h i g h s i n k p l a n t s and low c a r b o n d i o x i d e t r e a t m e n t s had h i g h e r r a t e s t h a n h i g h c a r b o n d i o x i d e t r e a t m e n t s , when measured under t h e same c a r b o n d i o x i d e c o n c e n t r a t i o n . K r i e d e m a n n e t . a l . (1976) f o u n d t h a t e n r i c h e d V i t i s p l a n t s had r e d u c e d p h o t o s y n t h e s i s r a t e s a t low measurement c o n c e n t r a t i o n s o f c a r b o n d i o x i d e , r e l a t i v e t o c o n t r o l s , but h i g h e r r a t e s a t h i g h measurement c o n c e n t r a t i o n s . T h i s u n u s u a l r e s u l t can p e r h a p s be e x p l a i n e d by r e f e r r i n g t o t h e l e a f anatomy o f V i t i s . E n r i c h e d l e a v e s were t h i c k e r . Hence, t h e d e c r e a s e d r a t e of p h o t o s y n t h e s i s i n e n r i c h e d p l a n t s was p r o b a b l y n o t due t o a r e d u c e d i n t e r n a l s u r f a c e a r e a , but due t o a r e d u c e d c a p a c i t y t o p h o t o s y n t h e s i z e . How t h e n d o e s one e x p l a i n g r e a t e r p h o t o s y n t h e s i s a t h i g h e r c a r b o n d i o x i d e c o n c e n t r a t i o n s ? The a u t h o r s s u g g e s t e d t h a t b e c a u s e g r a p e l e a v e s were h y p o s t o m a t o u s and e n r i c h e d l e a v e s were t h i c k e r , c a r b o n d i o x i d e a t low c o n c e n t r a t i o n s c o u l d be p r o p o r t i o n a t e l y more l i m i t i n g i n e n r i c h e d l e a v e s i n r e g i o n s f u r t h e s t away from t h e s t o m a t a . At 20 high carbon dioxide concentrations, those regions were no longer limited by carbon dioxide, and contributed proportionately more to net photosynthesis per unit leaf area. Photosynthetic capacity appeared to increase, but was in fact s t i l l less than controls. Most investigations have therefore found that there appears to be a reduction in photosynthetic capacity in enriched plants, the extent of which i s dependent on various developmental and environmental factors. Several authors, in fact, have found no ef f e c t of high carbon dioxide concentration on photosynthetic capacity (Ford and Thorne, 1967; G i f f o r d , 1977), but t h i s does not preclude the p o s s i b i l i t y of an effect occurring under di f f e r e n t conditions or at a d i f f e r e n t age. Somewhat more d i f f i c u l t to reconcile is the work of several authors - (Bishop and Whittingham, 1.968; Madsen, 1975; Hicklenton and J o l l i f f e , 1980b) who found that tomato plants grown in enriched atmospheres exhibited an increase in photosynthetic capacity. Hicklenton and J o l l i f f e (1980b) have provided the most detailed study to date. The carbon dioxide effect on photosynthesis was not caused by stomatal responses. Rather, the photosynthetic machinery per se was being affected. Enrichment caused a decrease in both the carbon dioxide and l i g h t compensation points, greater net photosynthesis at a given l i g h t i ntensity and carbon dioxide concentration, and decreased oxygen i n h i b i t i o n of net photosynthesis. How does one explain an increase in photosynthetic capacity in some studies of tomato, and a decrease in capacity in 21 v i r t u a l l y a l l o t h e r s p e c i e s s t u d i e d ? I t i s not b e c a u s e tomato r e s p o n d s d i f f e r e n t l y t o e n r i c h m e n t ; Ho (1977) f o u n d d e c r e a s e d p h o t o s y n t h e t i c c a p a c i t y i n t h i s s p e c i e s . I t may r e l a t e t o t h e e n v i r o n m e n t a l c o n d i t i o n s d u r i n g g r o w t h . Growth t e m p e r a t u r e , however, d o e s not seem t o be a f a c t o r . E n r i c h e d t omato p l a n t s grown a t a r e l a t i v e l y h i g h t e m p e r a t u r e of 30 t o 35 C ( H i c k l e n t o n and J o l l i f f e , 1980b), o r a t a l o w e r t e m p e r a t u r e of 18 C ( B i s h o p and W h i t t i n g h a m , 1968) b o t h show an i n c r e a s e i n p h o t o s y n t h e t i c c a p a c i t y . Hence, y e t o t h e r e n v i r o n m e n t a l or d e v e l o p m e n t a l c o n d i t i o n s may be r e q u i r e d t o e l i c i t t h i s r e s p o n s e . Changes i n p h o t o s y n t h e t i c c a p a c i t y due t o h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n c o r r e l a t e w i t h c h a n g e s i n s e v e r a l p h o t o s y n t h e t i c enzymes. A d e c r e a s e i n c a p a c i t y c o r r e l a t e s w i t h d e c r e a s e d RuBP c a r b o x y l a s e a c t i v i t y ( K r i e d e m a n n e t a l . , 1976; Wong 1979), and an i n c r e a s e i n c a p a c i t y c o i n c i d e s w i t h i n c r e a s e d a c t i v i t y o f t h i s enzyme ( H i c k l e n t o n and J o l l i f f e , 1980b). RuBP c a r b o x y l a s e was a l s o r e p o r t e d t o i n c r e a s e i n e n r i c h e d b a r l e y l e a v e s ( F a i r e t a l . , 1973) a l t h o u g h p h o t o s y n t h e s i s r a t e s were not m easured. G l y c o l a t e o x i d a s e a c t i v i t y has been r e p o r t e d t o d e c r e a s e a t h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n ( F a i r e t a l . , 1973; H i c k l e n t o n and J o l l i f f e , 1980b), w h i c h may a c c o u n t f o r t h e r e d u c e d p h o t o r e s p i r a t i o n i n e n r i c h e d p l a n t s ( H i c k l e n t o n and J o l l i f f e , 1980b). A l s o , a l o w e r n i t r a t e r e d u c t a s e a c t i v i t y has been shown t o o c c u r ( F a i r e t a l . , 1973; K r i e d e m a n n , e t a l . , 1976), w h i c h c o u l d a l s o c o n t r i b u t e t o l o w e r p h o t o r e s p i r a t i o n . In a s t u d y o f r o o t zone e n r i c h m e n t o f p o t a t o ( A r t e c a and P o o v a i a h , 1982), PEP c a r b o x y l a s e a c t i v i t y of r o o t s i n c r e a s e d , 22 but b o t h PEP c a r b o x y l a s e and RuBP c a r b o x y l a s e a c t i v i t y of l e a v e s r e m a i n e d unchanged. C a r b o n d i o x i d e e n r i c h m e n t c o u l d a l s o i n f l u e n c e p h o t o s y n t h e s i s r a t e s t h r o u g h c h a n g e s i n c h l o r o p h y l l c o n t e n t . Madsen (1968) f o u n d no e f f e c t on c h l o r o p h y l l c o n t e n t p e r u n i t l e a f a r e a but showed lower c h l o r o p h y l l c o n t e n t p e r u n i t f r e s h w e i g h t i n e n r i c h e d p l a n t s . Cave e t a l . (1981) f o u n d a l o w e r mean c h l o r o p h y l l c o n t e n t p e r u n i t l e a f a r e a i n b o t h immature and m ature l e a v e s o f e n r i c h e d T r i f o l i u m s u b t e r r a n e u m , but t h i s r e s p o n s e was n o t s t a t i s t i c a l l y s i g n i f i c a n t . C h l o r o p h y l l c o n t e n t p e r u n i t d r y w e i g h t was s i g n i f i c a n t l y l o w e r i n e n r i c h e d immature l e a v e s , but n o t s i g n i f i c a n t l y l o w e r i n mature l e a v e s . S i m i l a r l y , c h l o r o p h y l l c o n t e n t i n e n r i c h e d c o t t o n was n o t s i g n i f i c a n t l y l o w e r t h a n c o n t r o l s (Wong, 1979). In s o y b e a n , s i c k l e p o d , and showy c r o t a l a r i a e x a m i n e d a t two n u t r i e n t l e v e l s , c h l o r o p h y l l c o n t e n t was g e n e r a l l y , a l t h o u g h not a l w a y s , s i g n i f i c a n t l y l o w e r i n e n r i c h e d " p l a n t s ( P a t t e r s o n and F l i n t , 1982). Hence, e n r i c h m e n t a p p e a r s t o c a u s e a somewhat l o w e r c h l o r o p h y l l c o n t e n t , a l t h o u g h t h e d i f f e r e n c e may n o t a l w a y s be s u b s t a n t i a l . A s i g n i f i c a n t d e c r e a s e i n t h e c h l o r o p h y l l a:b r a t i o i n e n r i c h e d p l a n t s (Cave e t a l . , 1981) s u g g e s t s t h a t a change i n t h e c o m p o s i t i o n o f t h e p h o t o s y n t h e t i c p i g m e n t complex o c c u r s . 23 2.1.9 L e a f M o r p h o l o g y A number of s t u d i e s have e x p l o r e d t h e d e v e l o p m e n t a l a s p e c t s of c a r b o n d i o x i d e e n r i c h m e n t . In a d d i t i o n t o t h e c h a n g e s i n p l a n t g r o w t h and d e v e l o p m e n t w h i c h have a l r e a d y been d i s c u s s e d , c h a n g e s i n l e a f s t r u c t u r e a r e a l s o known t o o c c u r . The number o f s t o m a t a p e r u n i t l e a f a r e a has been shown t o d e c r e a s e i n tomato (Madsen, 1973b), i n r e s p o n s e t o c a r b o n d i o x i d e e n r i c h m e n t , but i s not a f f e c t e d i n b a r l e y , k a l e , or m a i z e ( F o r d and T h o r n e , 1967). G e n e r a l l y l e a f t h i c k n e s s i n c r e a s e s i n r e s p o n s e t o h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n (Madsen, 1973b; H o f s t r a and H e s k e t h , 1975; Kriedemann e t a l . , 1976), w h i c h c o u l d have an e f f e c t on p h o t o s y n t h e t i c measurements as p r e v i o u s l y d i s c u s s e d . The t h i c k n e s s o f t h e p a l i s a d e l a y e r i n c r e a s e s due t o i n c r e a s e s i n c e l l volume i n e n r i c h e d tomato (Madsen, 1973) and c e l l number i n s o y b e a n ( H o f s t r a and H e s k e t h , 1975). 2.1.10 L e a f C a r b o h y d r a t e s T h e r e a r e numerous r e p o r t s of i n c r e a s e s i n s t a r c h c o n t e n t i n l e a v e s of e n r i c h e d p l a n t s (Madsen, 1968; H o f s t r a and H e s k e t h , 1975; Ho, 1977; Mauney e t a l . , 1979; Cave e t a l . 1981; F i n n and B r u n , 1982).. T h e s e d i f f e r e n c e s a r e g r e a t enough t o be seen i n l i g h t m i c r o g r a p h s ( H o f s t r a and H e s k e t h , 1975; K r i e d e m a n n e t a l . , 1976) o r i n e l e c t r o n m i c r o g r a p h s of c h l o r o p l a s t s , where e x t r e m e l y l a r g e s t a r c h d e p o s i t s c a n be seen t o d i s r u p t n o r m a l g r a n a s t r u c t u r e (Cave e t a l . , 1981; W u l f f and S t r a i n , 1982). T h e s e d r a m a t i c c h a n g e s i n s t a r c h c o n t e n t p r e s u m a b l y a r e a major 24 c o n t r i b u t o r t o t h e i n c r e a s e s i n SLW t h a t a r e o b s e r v e d i n e n r i c h e d p l a n t s . S u c r o s e c o n c e n t r a t i o n t e n d e d t o i n c r e a s e i n e n r i c h e d tomato l e a v e s ( B i s h o p and W h i t t i n g h a m , 1968; Madsen, 1968; Ho, 1977). However, t o t a l s o l u b l e s u g a r s ( i n c l u d i n g s u c r o s e ) d e c r e a s e d i n e n r i c h e d s o y b e a n ( H o f s t r a and H e s k e t h , 1975) and were u n a f f e c t e d i n c o t t o n , s o y b e a n , s u n f l o w e r , and sorghum a c c o r d i n g t o Mauney e t a l . ( 1 9 7 9 ) . 2.2 SENESCENCE Be c a u s e o f t h e i n t e r e s t i n g o b s e r v a t i o n made d u r i n g t h e c o u r s e o f my s t u d y t h a t h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n s a p p e a r e d t o promote l e a f s e n e s c e n c e , an o v e r v i e w of s e n e s c e n c e p h y s i o l o g y i s p r o v i d e d as b a c k g r o u n d f o r t h e r e a d e r . S e n e s c e n c e can be c h a r a c t e r i z e d as a c o l l e c t i o n o f d e t e r i o r a t i v e p r o c e s s e s w h i c h c u l m i n a t e i n d e a t h (Nooden and L e o p o l d , 1978). S e n e s c e n c e may o c c u r a t v i r t u a l l y any l e v e l of o r g a n i z a t i o n , f r o m i n d i v i d u a l c e l l s , t o t i s s u e s , t o t h e e n t i r e o r g a n i s m . Whether r e s p o n d i n g t o e n v i r o n m e n t a l or i n t r i n s i c s i g n a l s , s e n e s c e n c e i s a l w a y s a h i g h l y r e g u l a t e d and d i r e c t e d s e q u e n c e o f e v e n t s . In many, but n o t a l l c a s e s , s e n e s c e n c e i s r e l a t e d t o a g i n g , w h i c h may be d e f i n e d as a g r a d u a l a c c u m u l a t i o n o f p h y s i o l o g i c a l c h a n g e s w i t h t i m e (Nooden and L e o p o l d , 1978). N e i t h e r a g i n g nor s e n e s c e n c e i n p l a n t s seems t o be due t o t h e a c c u m u l a t i o n o f e r r o r s i n t r a n s c r i p t i o n , as s u g g e s t e d i n work on a n i m a l s e n e s c e n c e . F o r example, l e a v e s may u n d e r g o c o m p l e t e s e n e s c e n c e i n l e s s t h a n 4 d a y s . C o n v e r s e l y , t i s s u e f r o m a n n u a l p l a n t s can be c u l t u r e d f o r y e a r s , when, i f l e f t on t h e p l a n t , 25 w o u l d s e n e s c e a f t e r one f l o w e r i n g s e a s o n ( m o n o c a r p i c s e n e s c e n c e ) . A g i n g need n o t a l w a y s r e s u l t i n d e a t h , or v i c e  v e r s a . M o n o c a r p i c s e n e s c e n c e f o r example, i s n o t due t o a g i n g as d e f i n e d p r e v i o u s l y . -2.2.1 G e n e r a l P h y s i o l o g y A number of c h a r a c t e r i s t i c p h y s i o l o g i c a l c h a n g e s a r e a s s o c i a t e d w i t h l e a f s e n e s c e n c e . A l t h o u g h most of t h e s e a r e c e r t a i n l y not a c a u s e of s e n e s c e n c e , t h e y c a n be u s e f u l i n d i c a t o r s of t h e e x t e n t and k i n e t i c s of t h e p r o c e s s . C h l o r o p h y l l l o s s , one of t h e most d r a m a t i c and v i s i b l e c h a n g e s t o o c c u r , i s f r e q u e n t l y u s e d as a measure of s e n e s c e n c e . T h i s g e n e r a l l y c o r r e l a t e s w i t h a d e c r e a s e i n p h o t o s y n t h e t i c r a t e . However, a d e c l i n e i n p h o t o s y n t h e s i s may p r e c e e d a l o s s of c h l o r o p h y l l ( H a r d w i c k e t a l . , 1968), and i n a mutant o f f e s c u e , s e n e s c e n c e , measured as p r o t e i n breakdown, o c c u r s w i t h o u t c h l o r o p h y l l d e g r a d a t i o n (Thomas and S t o d d a r t , 1975). L o s s of c h l o r o p h y l l need not t h e n be c l o s e l y a s s o c i a t e d w i t h t h e o n g o i n g p r o c e s s of s e n e s c e n c e . D e c l i n e i n p h o t o s y n t h e s i s i s a major i n i t i a l e v e n t i n s e n e s c e n c e , a l t h o u g h t h e p r i m a r y e v e n t s c a u s i n g p h o t o s y n t h e t i c r e d u c t i o n a r e n o t known. The d e c l i n e i s a c c o m p a n i e d by t h e l o s s of RuBP c a r b o x y l a s e ( K a n n a n g a r a and Woolhouse, 1968; F r i e d r i c h and H u f f a k e r , 1980) and some o t h e r enzymes o f p h o t o s y n t h e t i c c a r b o n a s s i m i l a t i o n ( B a t t and Woolhouse, 1975). W i t t e n b a c h e_t a l . (1980) however, f o u n d t h a t t h e i n i t i a l d e c l i n e i n p h o t o s y n t h e s i s was n o t due t o a d e c r e a s e i n RuBP c a r b o x y l a s e a c t i v i t y o r l e v e l . R a t h e r , t h e d e c l i n e was p r e c e e d e d by a 26 s w e l l i n g of c h l o r o p l a s t s and d i s r u p t i o n of t h y l a k o i d s , w h i c h c o u l d r e s u l t from an o b s e r v e d i n c r e a s e i n s t a r c h d e p o s i t i o n . L a r g e numbers o f o s m i o p h i l i c g r a n u l e s a l s o a p p e a r i n t h e c h l o r o p l a s t s , as has a l s o been o b s e r v e d by B a r t o n ( 1 9 6 6 ) . M i t o c h o n d r i a on t h e o t h e r hand, remain s t r u c t u r a l l y i n t a c t u n t i l t h e f i n a l s t a g e s of s e n e s c e n c e (Rhodes, 1980). R e s p i r a t i o n r a t e g e n e r a l l y does not change u n t i l l a t e i n s e n e c e n c e , when i t d e c r e a s e s r a p i d l y (Rhodes, 1980). A t r a n s i e n t r i s e i n r e s p i r a t i o n , p a r t i c u l a r l y i n d e t a c h e d l e a v e s has been r e p o r t e d ( H a r d w i c k e t a l . , 1968; T e t l e y and Thimann, 1974), w h i c h may r e s u l t from an i n c r e a s e i n r e s p i r a t i o n of amino a c i d s , o r an u n c o u p l i n g of r e s p i r a t i o n ( T e t l e y and Thimann, 1974). The f a c t t h a t r e s p i r a t i o n r e m a i n s r e l a t i v e l y h i g h t h r o u g h o u t most of s e n e s c e n c e s u g g e s t s t h a t i t may be needed i n e n e r g y p r o d u c t i o n f o r t h e s y n t h e s i s of d e g r a d a t i v e enzymes (Rhodes, 1980). P r o t e i n c o m p o s i t i o n c h a n g e s d r a m a t i c a l l y d u r i n g s e n e s c e n c e . P r o t e o l y s i s s e r v e s t h e d u a l f u n c t i o n of t u r n i n g o f f s y n t h e s i s of unneeded m e t a b o l i t e s , as w e l l as l i b e r a t i n g amino a c i d s f o r e x p o r t t o n o n - s e n e s c e n t p l a n t p a r t s . Not a l l p r o t e i n s d e c r e a s e however. D e g r a d a t i v e enzymes s u c h as RNase and p r o t e o l y t i c enzymes i n c r e a s e d u r i n g s e n e s c e n c e ( M a t i l e and W i n kenbach, 1971). RNase i s r e s p o n s i b l e f o r t h e d e c r e a s e i n RNA and r i b o s o m e s o b s e r v e d d u r i n g s e n e s c e n c e (Thimann, 1980). 27 2.2.2 Growth R e g u l a t o r s As i n v i r t u a l l y e v e r y o t h e r a s p e c t o f p l a n t d e v e l o p m e n t , g r o w t h r e g u l a t o r s a r e i n v o l v e d i n , o r a t l e a s t a f f e c t , l e a f s e n e s c e n c e . The most e f f e c t i v e g r o w t h r e g u l a t o r t o d e l a y s e n e s c e n c e i s c y t o k i n i n and i t s s y n t h e t i c a n a l o g s . K i n e t i n e f f e c t i v e l y d e l a y s c h l o r o p h y l l l o s s and p r o t e o l y s i s i n t h e d a r k , and t o a l e s s e r e x t e n t i n t h e l i g h t . I f a p p l i e d t o a s m a l l a r e a of a l e a f , i t d e l a y s s e n e s c e n c e i n o n l y t h a t l o c a l i t y , and i n d u c e s t r a n s p o r t o f s o l u t e s from n o n - t r e a t e d a r e a s (Thimann, 1980). K i n e t i n has i t s most p r o n o u n c e d e f f e c t on d e t a c h e d l e a f t i s s u e , w i t h l i t t l e o r no e f f e c t on an i n t a c t s y s t e m . The l a c k o f e f f e c t i n i n t a c t s y s t e m s i s p e r h a p s due t o h i g h endogenous l e v e l s of c y t o k i n i n a l r e a d y i n t h e t i s s u e , p e r h a p s t r a n s p o r t e d from t h e r o o t s . K i n e t i n a l s o has t h e a b i l i t y t o i n d u c e r e g r e e n i n g a f t e r s e n e s c e n c e has been i n i t i a t e d (Nooden and L e o p o l d , 1978). T h i s i s s i m i l a r t o t h e d e l a y i n g e f f e c t s of d e b u d d i n g or d e c a p i t a t i o n on l e a f s e n e s c e n c e , s u g g e s t i n g t h a t a r e d i s t r i b u t i o n of c y t o k i n i n o c c u r s when s i n k s f o r t h e hormone a r e removed. A b s c i s i c a c i d (AbA) i s known t o i n d u c e s e n e s c e n c e , o r a t l e a s t promote t h e v a r i o u s p r i m a r y o r s e c o n d a r y p h y s i o l o g i c a l p r o c e s s e s a s s o c i a t e d w i t h i t . As i s t h e c a s e w i t h c y t o k i n i n , e xogenous AbA has i t s g r e a t e s t e f f e c t on d e t a c h e d t i s s u e . Endogenous l e v e l s o f AbA i n c r e a s e d u r i n g s e n e s c e n c e . F o r example, water d e f i c i t s , w h i c h a r e known t o i n d u c e s e n e s c e n c e , a l s o c a u s e h i g h e r AbA l e v e l s ( s e e Nooden and L e o p o l d , 1978). The o t h e r major g r o u p s of p l a n t hormones do n o t have t h e 28 c l e a r l y d e f i n e d e f f e c t s o f AbA and c y t o k i n i n . A u x i n s have o n l y a s m a l l e f f e c t i n d e l a y i n g s e n e s c e n c e , and g i b b e r e l l i c a c i d s a r e e f f e c t i v e i n o n l y a few s p e c i e s , f o r example, Taraxacum and Rumex. The r e l a t i o n s h i p s between g i b b e r e l l i c a c i d s and c y t o k i n i n - i n d u c e d d e l a y of s e n e s c e n c e a r e not c l e a r . A l t h o u g h e t h y l e n e i s c l e a r l y i n v o l v e d i n p r o m o t i n g f l o w e r s e n e s c e n c e and l e a f a b s c i s s i o n ( A b e l e s 1973), i t s r o l e i n l e a f s e n e s c e n c e i s e q u i v o c a l . C o n f l i c t i n g i n t e r p r e t a t i o n s may a r i s e i f t h e r e s p o n s i v e n e s s of t h e t i s s u e t o e t h y l e n e v a r i e s i n s u c h a way t h a t i t r e s p o n d s a t one age o r under one s e t o f c o n d i t i o n s b u t n o t a n o t h e r . F o r example, e t h y l e n e s y n t h e s i s may i n c r e a s e , but i f t h e t i s s u e i s n o t r e s p o n s i v e , t h e o b s e r v e d i n c r e a s e i s of l i t t l e c o n s e q u e n c e . One must t h e r e f o r e be c o g n i z a n t of t h e d i s t i n c t i o n between e t h y l e n e l e v e l and e t h y l e n e a c t i o n . E t h y l e n e has 'been o b s e r v e d t o i n c r e a s e d u r i n g l e a f s e n e s c e n c e ( s e e Thimann, 1980). However, l e v e l s of e t h y l e n e i n s e n e s c i n g l e a v e s of beans and X a n t h i u m a r e t h e same as t h a t of mature, n o n - s e n e s c i n g l e a v e s ( s e e O s b o r n e , 1978). Work of A h a r o n i and L i e b e r m a n ( 1 9 7 9 ) , however, s u p p o r t s t h e i d e a o f a r o l e of e t h y l e n e i n s e n e s c e n c e . A l t h o u g h e t h y l e n e r e l e a s e i s a c t u a l l y d e c r e a s e d d u r i n g t h e i n i t i a l s t a g e s of s e n e s c e n c e i n t h e d a r k , i n h i b i t o r s of e t h y l e n e s y n t h e s i s , s u c h as Ag+ and a m i n o e t h o x y v i n y l g l y c i n e d e l a y c h l o r o p h y l l l o s s . C a r b o n d i o x i d e i s know t o a n t a g o n i z e e t h y l e n e a c t i o n i n many h i g h e r p l a n t s ( A b e l e s , 1973), a l t h o u g h t h e c o n c e n t r a t i o n s needed a r e r e l a t i v e l y h i g h (>10,000 u l l " 1 ) . H i g h c a r b o n d i o x i d e c o n c e n t r a t i o n (5000 u l l " 1 ) i s a l s o known t o a c c e l e r a t e 29 e t h y l e n e r e l e a s e from l e a f t i s s u e a l t h o u g h t h e e f f e c t i s o n l y t e m p o r a r y ( B a s s i and S p e n c e r , 1982). I t i s now g e n e r a l l y b e l i e v e d t h a t c a r b o n d i o x i d e a n t a g o n i z e s e t h y l e n e a c t i o n by d i s p l a c i n g e t h y l e n e f r o m i t s b i n d i n g s i t e . V e r y few s t u d i e s have l o o k e d a t t h e i n t e r a c t i o n o f c a r b o n d i o x i d e and e t h y l e n e i n s e n e s c e n c e . A h a r o n i and L i e b e r m a n (1979) f o u n d t h a t 50,000 t o 10,000 u l 1~ 1 c a r b o n d i o x i d e s t i m u l a t e d e t h y l e n e p r o d u c t i o n by t o b a c c o l e a f d i s k s s e n e s c i n g i n t h e d a r k , b ut i n f a c t d e l a y e d s e n e s c e n c e . 2.2.3 E n v i r o n m e n t a l I n f l u e n c e s L e a f s e n e s c e n c e can be i n i t i a t e d by a number o f e n v i r o n m e n t a l s i g n a l s , w h i c h i n some c a s e s a t l e a s t , a r e r e l a t e d t o c h a n g e s i n l e v e l s of growth r e g u l a t o r s . L e a f s e n e s c e n c e , f o r example, i s a c c e l e r a t e d by low. l e a f water p o t e n t i a l s ( s e e Thimann, 1980), w h i c h a l s o c a u s e an i n c r e a s e i n l e a f AbA c o n t e n t ( W r i g h t and H i r o n , 1969). I n t e r e s t i n g l y , Ludlow (1975) f o u n d t h a t low water p o t e n t i a l s a p p e a r e d t o s u s p e n d t h e a g i n g p r o c e s s i n t h o s e l e a v e s of Panicum w h i c h d i d not d i e f r o m t h e d i r e c t e f f e c t s of water s t r e s s . I t i s a l s o p o s s i b l e t h a t r e p e a t e d c y c l e s of low water p o t e n t i a l may r e n d e r young l e a v e s l e s s s u s c e p t a b l e t o t h e s t r e s s - i n d u c i n g c o n d i t i o n s ( s e e Thomas and S t o d d a r t , 1980). T e m p e r a t u r e e x t r e m e s a r e a l s o known t o i n i t i a t e l e a f c h l o r o s i s , p r e s u m a b l y as a r e s u l t of m e t a b o l i c damage (Thomas and S t o d d a r t , 1980). C o o l t e m p e r a t u r e may i n t e r a c t w i t h l i g h t i n t e n s i t y t o b r i n g a b o u t a c c e l e r a t e d s e n e s c e n c e ( T a y l o r and C r a i g , 1971). 30 L i g h t i s g e n e r a l l y known t o d e l a y s e n e s e n c e of l e a f t i s s u e ( G o l d t h w a i t e and L a e t s c h , 1967; Thomas and S t o d d a r t , 1980) and i s t h o u g h t t o a c t t h r o u g h e f f e c t s on p h o t o s y n t h e s i s . M i n e r a l d e f i c i e n c y i s a m a j o r c a u s e of l e a f s e n e s c e n c e ; o f t e n low n u t r i e n t a v a i l a b i l i t y w i l l r e s u l t i n m o b i l e n u t r i e n t s b e i n g t r a n s p o r t e d f r o m t h e o l d e r l e a v e s . T h i s may be one r e a s o n why s e n e s c e n c e symptoms a r e f i r s t e v i d e n t i n t h o s e l e a v e s . In c o n c l u s i o n , i t seems t h a t , u n l i k e m o n o c a r p i c s e n e s c e n c e , w h i c h i s i n i t i a t e d by a d e v e l o p m e n t a l p r o g r a m , s e n e s c e n c e i n d u c e d by t h e e n v i r o n m e n t i s , i n l a r g e p a r t , a f u n c t i o n of t i s s u e s u s c e p t a b i l i t y . S u s c e p t a b i l i t y o f l e a v e s t o a s t r e s s seems t o depend on s u c h f a c t o r s as l e a f age, p o s t i o n on t h e p l a n t , and p r e v i o u s s t r e s s h i s t o r y . 2.2.4 C a r b o n D i o x i d e And S e n e s c e n c e The p r o s p e c t o f c a r b o n d i o x i d e a c t i n g as a s e n e s c e n c e -i n d u c i n g a g e n t i n p l a n t s has r e c e i v e d v e r y l i t t l e a t t e n t i o n , even t h o u g h as e a r l y a s 1902, n e g a t i v e e f f e c t s of c a r b o n d i o x i d e e n r i c h m e n t were o b s e r v e d i n p l a n t s (Brown and Escombe, 1902). T h i s was l a t e r a s c r i b e d t o i m p u r i t i e s i n t h e a i r . In more r e c e n t t i m e s , l e a v e s of e n r i c h e d cucumbers have been o b s e r v e d t o have y e l l o w s p o t t i n g , e v e n t u a l l y f o l l o w e d by n e c r o s i s a t c a r b o n d i o x i d e c o n c e n t r a t i o n s above 1000 u l 1" 1 ( W i t t w e r , 1967). H e s k e t h e t a l . (1971) f o u n d t h a t l e a v e s o f e n r i c h e d c o t t o n and s o y b e a n p l a n t s became v e r y c h l o r o t i c . T h i s was a s s o c i a t e d w i t h l a r g e s t a r c h a c c u m u l a t i o n s i n t h e l e a v e s . C h l o r o s i s o c c u r r e d a t 31 C but n o t 35 C. In a p a p e r r e l a t i n g 31 c a r b o n d i o x i d e e n r i c h m e n t t o s e n e s c e n c e of c o t t o n l e a v e s , Chang (1975) f o u n d t h a t 1000 u l 1~ 1 c a r b o n d i o x i d e c a u s e d d e c r e a s e d c h l o r o p h y l l and p r o t e i n l e v e l s a f t e r 30 d a ys of e n r i c h m e n t . A d e c r e a s e i n p h o t o p h o s p h o r y l a t i o n and H i l l r e a c t i o n a c t i v i t y a l s o o c c u r r e d when measured i n v i t r o i n t h e p r e s e n c e of b i c a r b o n a t e . C a r b o n i c a n h y d r a s e , an enzyme c a t a l y z i n g t h e h y d r a t i o n of c a r b o n d i o x i d e , a l s o d e c r e a s e d i n a c t i v i t y . I t i s not c l e a r , however, whether t h o s e c h a n g e s r e l a t e d t o t h e c a u s e of c a r b o n d i o x i d e -i n d u c e d s e n e s c e n c e . 32 I I I . EFFECTS ON GROWTH AND PARTITIONING 3.1 INTRODUCTION Enh a n c e d p l a n t g r o w t h o c c u r s a t e l e v a t e d c a r b o n d i o x i d e c o n c e n t r a t i o n s ( e g . W i t t w e r and Robb, 1964). However, t h e r e i s some i n d i c a t i o n t h a t t h e f u l l p o t e n t i a l f o r i n c r e a s e d g r o w t h , b a s e d on a s i m p l e model of i n c r e a s e d s u b s t r a t e f o r n e t a s s i m i l a t i o n i s n o t a c h i e v e d ( N e a l e s and N i c h o l l s , 1978). F u r t h e r m o r e , t h e d e g r e e o f g r o w t h enhancement due t o c a r b o n d i o x i d e e n r i c h m e n t i s h i g h l y v a r i a b l e among s p e c i e s (Wong, 1979), o v e r t i m e ( A o k i and Y a b u k i , 1977), d e v e l o p m e n t a l s t a g e , (Mauney e t a l . , 1978), and e n v i r o n m e n t a l c o n d i t i o n s ( F o r d and T h o r n e , 1967). As a f i r s t s t e p i n e l u c i d a t i n g t h e p h y s i o l o g i c a l r e s p o n s e s of beans t o c a r b o n d i o x i d e e n r i c h m e n t , i t was e s s e n t i a l to-d e s c r i b e t h e growth t r e n d s and r e l a t i o n s h i p s o f t h e whole p l a n t under c o n t r o l l e d c o n d i t i o n s . The p r i m a r y o b j e c t i v e s of t h e work r e p o r t e d i n t h i s c h a p t e r , t h e r e f o r e , were t o answer t h e f o l l o w i n g q u e s t i o n s : 1. When and t o what d e g r e e does c a r b o n d i o x i d e e n r i c h m e n t have an e f f e c t on o v e r a l l p l a n t growth? 2. How i s g r o w t h p a r t i t i o n e d w i t h i n p l a n t s i n r e l a t i o n t o c a r b o n d i o x i d e e n r i c h m e n t ? 3. What f a c t o r s a r e r e s p o n s i b l e f o r t h e c h a n g e s i n g r o w t h a t d i f f e r e n t c a r b o n d i o x i d e l e v e l s ? F o r example, how d o e s net a s s i m i l a t i o n r a t e change i n r e s p o n s e t o e n r i c h m e n t and i s i t c o r r e l a t e d w i t h c h a n g e s i n growth? 33 These q u e s t i o n s can, i n p a r t , be answered using techniques of p l a n t growth a n a l y s i s . V a r i o u s i n d i c e s of p l a n t performance, such as r e l a t i v e growth rate and u n i t l e a f r a t e , as w e l l as i n d i c e s of p a r t i t i o n i n g such as l e a f area r a t i o , and s p e c i f i c l e a f weight, were used to e s t a b l i s h the e f f e c t of long-term carbon d i o x i d e enrichment on the growth of bean p l a n t s from the s e e d l i n g to the r e p r o d u c t i v e stage. Note that a study of r e p r o d u c t i v e development per se was not intended, although the general f e a t u r e s of f l o w e r i n g and pod formation were observed d u r i n g the study. 34 3.2 MATERIALS AND METHODS 3.2.1 P l a n t C u l t u r e U n l e s s o t h e r w i s e i n d i c a t e d , t h e f o l l o w i n g p r e - t r e a t m e n t c u l t u r a l p r o c e d u r e s were u s e d i n a l l e x p e r i m e n t s d e s c r i b e d i n t h i s t h e s i s . T h r e e bush bean ( P h a s e o l u s v u l g a r i s L. c v . , P u r e G o l d Wax) p l a n t s were s e e d e d p e r 5 i n c h p l a s t i c p o t ( s o i l volume was 0.85 1) i n sandy loam s o i l , and were t h i n n e d t o one p l a n t pe r p o t a t 10 t o 14 d a y s a f t e r p l a n t i n g . D u r i n g t h i s p e r i o d , p l a n t s were i n a P e r c i v a l (Model PG.78) g r o w t h chamber. The t e m p e r a t u r e r e g i m e was 26 C d a y / 20 C n i g h t , p h o t o p e r i o d was 16 h o u r s , and l i g h t i n t e n s i t y i n c i d e n t on t h e p r i m a r y l e a v e s (measured w i t h a L i - C o r model L I - 1 8 5 quantum m e t e r ) was 350 umol n r 2 s " 1 p h o t o s y n t h e t i c p h o t o n f l u x d e n s i t y ( P P F D K Note t h a t 1 mole m"2 s~ 1 i s e q u a l t o 1 E i n s t e i n - n r 2 s _ 1 , o r A v o g a d r o ' s Number of p h o t o n s nr 2 s ~ 1 . P l a n t s were w a t e r e d d a i l y w i t h t a p w a t e r . At 10 t o 14 d a y s , t h e p l a n t s were t r a n s f e r r e d t o s p e c i a l l y c o n s t r u c t e d g r o w t h chambers f o r c a r b o n d i o x i d e t r e a t m e n t . 3.2.2 C a r b o n D i o x i d e T r e a t m e n t Chamber D e s i g n C o n v e n t i o n a l p l a n t g r o w t h chambers do n o t p e r m i t e a s y c o n t r o l of c a r b o n d i o x i d e l e v e l s . Chambers a r e f r e q u e n t l y d e s i g n e d as s e m i - c l o s e d s y s t e m s , i n w h i c h a f e e d b a c k l o o p c o n t r o l s chamber c a r b o n d i o x i d e l e v e l s . In t h i s s i t u a t i o n , i f th e chamber a i r volume i s l a r g e , i t c a n be d i f f i c u l t t o m a i n t a i n a c o n s t a n t c a r b o n d i o x i d e c o n c e n t r a t i o n b e c a u s e of e x t e n d e d l a g t i m e s i n a d m i n i s t e r i n g c a r b o n d i o x i d e . F u r t h e r m o r e , t h e p l a n n e d 35 s t u d y of c a r b o n d i o x i d e e n r i c h m e n t e f f e c t s r e q u i r e d t h a t many chambers be u s e d s i m u l t a n e o u s l y , and a p p r o p r i a t e f a c i l i t i e s were no t i n i t i a l l y a v a i l a b l e . E i g h t g r o w t h chambers were t h e r e f o r e b u i l t w h i c h were i n e x p e n s i v e , b u t p r o v i d e d e a s y m a i n t e n a n c e o f c a r b o n d i o x i d e l e v e l s . The chambers were c o n s t r u c t e d i n two s e t s of f o u r e a c h . In e a c h s e t , f o u r i n d i v i d u a l P l e x i g l a s chambers, e a c h 0.45m x 0.45m x 0.80m (L,W,H) and volume o f 0.16 m 3, s h a r e d a common s u p p o r t i n g framework, c o o l i n g s y s t e m , and l i g h t s o u r c e . The l i g h t i n g s y s t e m f o r e a c h s e t c o n s i s t e d of a bank of 25 v e r y h i g h o u t p u t (VHO) c o o l - w h i t e f l u o r e s c e n t t u b e s , and 13 60-watt i n c a n d e s c e n t b u l b s , w h i c h was s u s p e n d e d 0.10 m above t h e t o p of t h e chambers. W i t h i n e a c h chamber, p l a n t s were p l a c e d on a movable p l a t f o r m , w h i c h a l s o f u n c t i o n e d a s a h e a t e x c h a n g e r t o m a i n t a i n chamber t e m p e r a t u r e ( F i g . 3.1 ). The p l a t f o r m was s u s p e n d e d by a c a b l e a t e a c h c o r n e r . The c a b l e s e x i t e d e a c h chamber t h r o u g h one i n c h d i a m e t e r p o r t s i n t h e chamber t o p and were s e c u r e d t o t h e chamber framework. T w o - i n c h t h i c k s t y r o f o a m s h e e t s , w h i c h s e r v e d a s i n s u l a t i o n , were p o s i t i o n e d a d j a c e n t t o t h e o u t s i d e of th e P l e x i g l a s w a l l s of e a c h s e t of 4 chambers and were h e l d i n p l a c e w i t h s h e e t s of one q u a r t e r - i n c h p l y w o o d . Chamber t e m p e r a t u r e was m a i n t a i n e d by c i r c u l a t i n g c o o l a n t f i r s t t h r o u g h a r e f r i g e r a t e d h e a t e x c h a n g e r , and t h e n t h r o u g h a s e c o n d a r y c o p p e r h e a t e x c h a n g e r w h i c h c o m p r i s e d t h e m a t r i x of t h e chamber p l a t f o r m . T e m p e r a t u r e was r e g u l a t e d by a d j u s t i n g t h e s u p p l y o f c o o l a n t by means o f v a l v e s . Chamber h e a t i n g was 36 0.1 m A I R I N L E T F I G U R E 3 .1 S c h e m a t i c d i a g r a m o f o n e o f t h e e i g h t c a r b o n d i o x i d e t r e a t m e n t c h a m b e r s . S e e t e x t f o r d e t a i l s . 37 u n n e c e s s a r y , s i n c e d a y t i m e t e m p e r a t u r e s o f 38 C c o u l d be a c h i e v e d s i m p l y t h r o u g h h e a t o u t p u t from t h e l i g h t s when c o o l a n t f l o w was s t o p p e d . Chamber t e m p e r a t u r e was m o n i t o r e d w i t h t h e r m o c o u p l e s c o n n e c t e d t o a d i g i t a l thermometer (Omega, model 199) . The chambers were d e s i g n e d as open s y s t e m s , w i t h c o n t i n u o u s , r a p i d t u r n o v e r o f a i r . The t u r n o v e r t i m e of chamber a i r was a p p r o x i m a t e l y 2 m i n u t e s . F r e s h ambient a i r was p u l l e d i n f r o m o u t s i d e t h e b u i l d i n g t h r o u g h g a l v a n i z e d z i n c d u c t i n g by means of an i n t a k e f a n ( T o r i n , model TA300) i n t h e b o t t o m of e a c h chamber. A i r e x i t e d from e a c h chamber t h r o u g h t h e f o u r p o r t s i n t h e t o p ( t h r o u g h w h i c h t h e p l a t f o r m c a b l e s a l s o p a s s e d ) . In a d d i t i o n t o t h e i n t a k e f a n , f o u r c i r c u l a t i n g f a n s ( T o r i n , model TA300) were s u s p e n d e d from e a c h p l a t f o r m and mixed t h e • chamber a i r . A c c o r d i n g t o t h e m a n u f a c t u r e r ' s s p e c i f i c a t i o n s , t h e r a t e d o u t p u t of e a c h f a n was 1250 1 m i n - 1 a g a i n s t z e r o s t a t i c p r e s s u r e . Flow r a t e t h r o u g h e a c h chamber was 160 1 min-1, as measured w i t h a hot w i r e anemometer ( e i t h e r D a t a m e t r i c s , model 800-VTP, o r Weather Measure C o r p . , model W-141-A1/5). F o r e a c h measurement, t h e a i r v e l o c i t y t h r o u g h e a c h p o r t was d e t e r m i n e d i n f e e t p e r m i n u t e , and c o n v e r t e d t o a volume o f a i r u s i n g t h e f o l l o w i n g e q u a t i o n : SCFH = .327 ( F P M ) ( D 2 ) where: SCFH = s t a n d a r d c u b i c f e e t p e r h o u r (1 SCFH= 7.8 ml s" 1 ) D = p o r t d i a m e t e r i n i n c h e s 38 FPM = f e e t p e r m i n u t e (1FPM= 5.08 mm s " 1 ) P u r e c a r b o n d i o x i d e from c o m p r e s s e d gas c y l i n d e r s was b l e d i n t o e a c h chamber t h r o u g h a P l e x i g l a s tube above t h e i n t a k e f a n , w h i c h m i x e d t h e c a r b o n d i o x i d e w i t h ambient a i r . The r a t e of c a r b o n d i o x i d e a d d i t i o n was r e g u l a t e d w i t h f l o w m e t e r s (Matheson 603 or 6 1 0 ) . The c a r b o n d i o x i d e c o n c e n t r a t i o n i n t h e a i r l e a v i n g t h e e x i t p o r t s was m o n i t o r e d w i t h an i n f r a - r e d gas a n a l y s e r (Beckman, model 8 6 4 ) . The chamber c a r b o n d i o x i d e c o n c e n t r a t i o n s were 340±10, 5 0 0 ± 1 0 , 800±20,. 1200±20, 2 0 0 0 ± 4 0 , and 3000±40 u l 1 " 1 . Over t h e c o u r s e of t h e c a r b o n d i o x i d e t r e a t m e n t p e r i o d , p l a n t s were m a i n t a i n e d a t 26/20 C d a y / n i g h t t e m p e r a t u r e , PPFD o f 350 umol n r 2 s " 1 a t t h e t o p of t h e p l a n t canopy, and a p h o t o p e r i o d o f 15 h o u r s . P l a n t s were w a t e r e d t w i c e weekly w i t h h a l f - s t r e n g t h H o a g l a n d ' s s o l u t i o n and w i t h d i s t i l l e d water when needed. When a c c e s s t o t h e p l a n t s was r e q u i r e d , t h e chamber d o o r was opened f o r , a t most, 2 m i n u t e s . 3.2.3 Growth A n a l y s i s F o r g r o w t h a n a l y s i s , 4 r e p l i c a t e p l a n t s i n e a c h of t h e 6 c a r b o n d i o x i d e r e g i m e s were h a r v e s t e d 11 d a y s a f t e r p l a n t i n g ( t i m e z e r o f o r e n r i c h m e n t ) , o r a t 22, 32, 42, o r 55 d a y s of c o n t i n u o u s e n r i c h m e n t . B e c a u s e o f s p a c e l i m i t a t i o n s i n t h e t r e a t m e n t chambers, t h e i n t r o d u c t i o n and r e m o v a l o f p l a n t s i n t h e chambers had t o be s t a g g e r e d ; enough p l a n t s f o r 2 c o m p l e t e h a r v e s t s ( i . e . 8 p l a n t s ) were p l a c e d i n t h e chambers a t one 39 t i m e . At e a c h h a r v e s t d a t e , t h e a r e a of e a c h l e a f was measured w i t h a l e a f a r e a meter ( L i - C o r model L I - 3 0 0 0 ) , and i n d i v i d u a l l y r e c o r d e d . Dry w e i g h t s of r o o t s , i n d i v i d u a l l e a v e s , stems, and pods were d e t e r m i n e d a f t e r oven d r y i n g t o c o n s t a n t w e i g h t a t 150 C. P r i m a r y d a t a were l o g - t r a n s f o r m e d , and f i t t e d c u r v e s d e s c r i b i n g t h e i r t i m e t r e n d s were g e n e r a t e d u s i n g a c u b i c s p l i n e r e g r e s s i o n p r o c e d u r e . T h i s p r o c e d u r e ( R e i n s c h , 1971) i n v o l v e d t h e F o r t r a n s u b r o u t i n e s DSPLFT and DSPLN a v a i l a b l e t h r o u g h t h e U n i v e r s i t y of B r i t i s h C o l u m b i a Computing C e n t r e . The p r o c e d u r e a l s o g e n e r a t e d f i t t e d c u r v e s f o r t h e f i r s t d e r i v a t i v e and t h e f i r s t d e r i v a t i v e of t h e l o g a r i t h m ( i . e . t h e r e l a t i v e g r o w t h r a t e ) o f e a c h v a r i a t e . S m o o t h i n g o f t h e s p l i n e r e g r e s s i o n s was d e t e r m i n e d e n t i r e l - y by t h e computer program a c c o r d i n g t o t h e s t a n d a r d d e v i a t i o n s of t h e v a r i a t e s a t e a c h h a r v e s t . The f i t t e d c u r v e s were t h e n u s e d t o g e n e r a t e t h e i n d i c e s of p l a n t g r o w t h d e f i n e d below: R e l a t i v e Growth R a t e : t h e r a t e of change i n d r y m a t t e r p e r u n i t of d r y m a t t e r a l r e a d y e x i s t i n g . The i n s t a n t a n e o u s v a l u e was c a l c u l a t e d a s : R= 1/W (dW/dT) = d ( l n W)/dt R e l a t i v e g r o w t h r a t e has u n i t s of d r y w e i g h t « d r y w e i g h t " 1 t ime" 1 . L e a f A r e a R a t i o : t h e r a t i o of t o t a l l e a f a r e a t o t o t a l p l a n t d r y w e i g h t . The i n s t a n t a n e o u s v a l u e was c a l c u l a t e d a s : 4 0 F= La/W Leaf area r a t i o has u n i t s of l e a f area«dry w e i g h t " 1 . U n i t Leaf Rate: a measure of the r a t e of change of dry matter per u n i t l e a f area. The instantaneous value was c a l c u l a t e d as: E= 1/La (dW/dT) The u n i t s are dry weight-leaf a r e a " 1 t i m e " 1 . U n i t l e a f r a t e was c a l c u l a t e d by d i v i d i n g R by F. S p e c i f i c Leaf Weight: the r a t i o of l e a f dry weight to l e a f area.- I t was c a l c u l a t e d f o r the e n t i r e p l a n t or f o r each l e a f i n d i v i d u a l l y . The instantaneous Value was c a l c u l a t e d as: SLW= Lw/La S p e c i f i c l e a f weight has u n i t s of dry weighty l e a f a r e a " 1 . Leaf weight r a t i o : the r a t i o of l e a f dry weight to t o t a l p l a n t dry weight. The instantaneous value was c a l c u l a t e d as: LWR= Lw/W The r a t i o i s d i m e n s i o n l e s s . Absolute growth r a t e : the rate of change of a v a r i a b l e , such as t o t a l p l a n t dry weight. The instantaneous value was c a l c u l a t e d as: G=dW/dt The u n i t s i n t h i s case are dry weight t i m e " 1 . Absolute growth rate i s a f u n c t i o n of l e a f area times u n i t l e a f r a t e . R e l a t i v e growth r a t e , however, i s a f u n c t i o n of l e a f 41 a r e a r a t i o t i m e s u n i t l e a f r a t e . The e x p e r i m e n t was r e p e a t e d a p p r o x i m a t e l y 1 y e a r a f t e r c o m p l e t i o n of t h e f i r s t t r i a l . I n t h e s e c o n d t r i a l , however, o n l y c a r b o n d i o x i d e c o n c e n t r a t i o n s o f 340, 500, 1200, and 3000 u l l " 1 were u s e d . A n a l y s i s o f v a r i a n c e was p e r f o r m e d on t h e c o l l e c t i v e d a t a of t h o s e 4 c a r b o n d i o x i d e t r e a t m e n t s from b o t h t r i a l s . The two t r i a l s were t r e a t e d as two b l o c k s i n t h e a n a l y s i s . A m u l t i f a c t o r i a l a n a l y s i s o f v a r i a n c e p a c k a g e (MFAV), p r o v i d e d by t h e U n i v e r s i t y of B r i t i s h C o l u m b i a Computing C e n t r e , was u s e d t o p e r f o r m t h e a n a l y s i s . 42 3.3 RESULTS AND DISCUSSION 3.3.1 E f f e c t s On Growth B e c a u s e o f s i g n i f i c a n t b l o c k e f f e c t s i n t h e a n a l y s i s of v a r i a n c e of t o t a l d r y w e i g h t ( T a b l e 3.1), d a t a o f t h e two b l o c k s were not p o o l e d . H e n c e f o r t h , f o r s i m p l i c i t y , o n l y t h e d a t a of t h e f i r s t b l o c k a r e p r e s e n t e d ; t h e p r i m a r y d a t a of t h e s e c o n d b l o c k showed s i m i l a r t r e n d s i n t r e a t m e n t e f f e c t s ( A p p e n d i x A ) . The p r i m a r y d a t a and f i t t e d c u r v e s f o r l e a f a r e a a r e g i v e n i n F i g u r e 3.2. D i f f e r e n c e s i n l e a f a r e a were most p r o n o u n c e d a t t h e l a s t h a r v e s t d a t e , w i t h e n r i c h e d p l a n t s g e n e r a l l y h a v i n g more t o t a l l e a f a r e a . The d i f f e r e n c e s among t r e a t m e n t s i n t o t a l d r y w e i g h t ( F i g . 3.3) were g r e a t e r t h a n t h e d i f f e r e n c e s i n l e a f a r e a and were a l s o p r o g r e s s i v e l y g r e a t e r w i t h t i m e . F o r example, p l a n t s grown a t 1200 u l l " 1 .were 1.7 t i m e s l a r g e r t h a n t h o s e a t 340 u l 1-1 ( c o n t r o l s ) . Note, however, t h a t an a d d i t i o n a l i n c r e a s e i n c a r b o n d i o x i d e c o n c e n t r a t i o n t o 3000 u l l " 1 d i d not f u r t h e r i n c r e a s e d r y w e i g h t . The g r o w t h r e s p o n s e of b eans, under t h e p r e v a i l i n g e n v i r o n m e n t a l c o n d i t i o n s , a p p e a r e d t o have s a t u r a t e d by 1200 u l 1~ 1. A b s o l u t e growth r a t e (G) was a l s o h i g h e r a t e l e v a t e d c a r b o n d i o x i d e c o n c e n t r a t i o n s ( F i g . 3 . 4 ) . A b s o l u t e g r o w t h r a t e a l s o i n c r e a s e d w i t h t i m e , i n p a r t a r e s u l t o f an e n l a r g i n g t o t a l l e a f a r e a ( F i g . 3 . 2 ) . R e l a t i v e g r o w t h r a t e ( R ) , w h i c h n o r m a l i z e s g r o w t h r a t e on a u n i t t o t a l p l a n t d r y w e i g h t b a s i s , d e c r e a s e d i n a l l t r e a t m e n t s f o r t h e f i r s t 19 d a y s a f t e r t r a n s f e r ( F i g . 3 . 5 ) . The r e l a t i v e g r o w t h r a t e o f e n r i c h e d p l a n t s , w h i c h was i n i t i a l l y 43 TABLE 3 . 1 . A n a l y s i s of dry weight v a r i a n c e of da ta of t r i a l the c o l l e c t i v e 1 and t r i a l 2 . 1 Source df Sum Sq. Mean Sq . E r r o r F Requ i red F 2 B l o c k 3 1 140.1 140.1 1 5.54 3.92 Harvest 3 9745.2 3248.4 360.37 2.68 T r e a t . 3 474.3 1 58.1 17.54 2.68 E r r o r 120 1081.7 9.0 T o t a l 127 11441.0 The t reatment carbon d i o x i d e c o n c e n t r a t i o n s were 340, 500, 1200, and 3000 u l l ' 1 At 5% l e v e l of p r o b a b i l i t y T r i a l 1 and t r i a l 2 were t r e a t e d as b l o c k s 44 a in O FIGURE 3.2 Leaf area over time of plants grown at 340, 500, 800, 1200, 2000, or 3000 ul l " 1 carbon dioxide. Each value is the mean of 4 plants, with standard deviation. Curves are cubic spline functions f i t ted to data for each carbon dioxide treatment. 45 o i n -i tn 10.0 15.0 2D.D 25.D 30.0 35.D 40.0 45.0 50.D 55.0 TINE AFTER PLANTING IN DAYS FIGURE 3.3 Dry weight over time of plants grown at 340, 500, 800, 1200, 2000, or 3000 ul l " 1 carbon dioxide. Each value is the mean of 4 plants, with standard deviation. Curves are cubic spine functions f i t ted to data for each carbon dioxide treatment. 46 o 12 00 3 0 0 0 8 0 0 2 0 0 0 5 0 0 340 l 1 I D . 0 1 5 . D 20.0 25.0 30.0 35.0 40.0 45.0 50.0 TIME AFTER PLANTING IN DAYS 5 5 . 0 FIGURE 3.4 Absolute growth rate of dry weight over time of plants grown at 340, 500, 800, 1200, 2000, or 3000 ul l " 1 carbon dioxide. Absolute growth rate is expressed in in units of g day" 1 . 47 FIGURE 3.5 Relative growth rate over time of plants grown at 340, 500, 800, 1200, 2000, or 3000 ul l " 1 carbon dioxide. R i s expressed in units of g g" 1 day" 1. 48 h i g h e r t h a n c o n t r o l s , d e c r e a s e d t h e most r a p i d l y . By 22 d a y s of age, e n r i c h e d p l a n t s had a p p r o x i m a t e l y t h e same R as c o n t r o l s and t h i s was m a i n t a i n e d f o r t h e r e m a i n d e r of t h e e x p e r i m e n t . R e l a t i v e g r o w t h r a t e i s t h e p r o d u c t of u n i t l e a f r a t e (E) and l e a f a r e a r a t i o ( F ) , e i t h e r one of w h i c h c o u l d c hange and t h e r e b y change R. U n i t l e a f r a t e was i n c r e a s e d by e n r i c h m e n t d u r i n g e a r l y d e v e l o p m e n t , w i t h a maximum a t a b o u t day 18 ( F i g . 3 . 6 ) . By day 30, i t had d e c r e a s e d t o n e a r l y c o n t r o l r a t e s i n a l l t r e a t m e n t s . The c h a n g e s i n E were s i m i l a r t o t h o s e of R a f t e r day 25, s u g g e s t i n g t h a t c h a n g e s i n E c o n t r i b u t e d t o t h e c h a n g e s i n R. T h i s l o s s o f e f f e c t of c a r b o n d i o x i d e e n r i c h m e n t on E a f t e r p r o l o n g e d t r e a t m e n t s has a l s o been o b s e r v e d i n cucumber ( A o k i and Y a b u k i , 1977), s u n f l o w e r , and s o y b e a n (Mauney e t a l . , 1978). A g r e a t e r u n i t l e a f r a t e a t h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n does not n e c e s s a r i l y t r a n s l a t e i n t o a g r e a t e r g r o w t h r a t e . The t o t a l l e a f s u r f a c e a r e a i s a l s o an i m p o r t a n t d e t e r m i n a n t of o v e r a l l d r y m a t t e r p r o d u c t i o n . L e a f a r e a i s a d e t e r m i n a n t of b o t h a b s o l u t e g r o w t h r a t e and r e l a t i v e g r o w t h r a t e . The i m p o r t a n c e of l e a f a r e a i n d e t e r m i n i n g g r o w t h i s p o i n t e d o u t i n a s t u d y of wheat ( G i f f o r d , 1977).. On a u n i t l e a f a r e a b a s i s , e n r i c h e d p l a n t s showed 15 t o 20% l e s s a s s i m i l a t i o n t h a n c o n t r o l s a t ambient c a r b o n d i o x i d e , but on a u n i t g r o u n d a r e a b a s i s , e n r i c h e d p l a n t s had t w i c e t h e a s s i m i l a t i o n r a t e o f c o n t r o l s a t h i g h i r r a d i a n c e ( G i f f o r d , 1977). T h i s was due t o a g r e a t e r l e a f a r e a i n e n r i c h e d p l a n t s . The m a j o r i n f l u e n c e o f e n r i c h m e n t on E was e a r l y i n 49 FIGURE 3.6 U n i t l e a f r a t e over time of p l a n t s grown a t 340, 500, 800, 1200, 2000, or 3000 u l l ' 1 carbon d i o x i d e . E i s e x p r e s s e d i n u n i t s of g n r 2 d a y - 1 x ! 0 " 3 50 d e v e l o p m e n t ( F i g . 3 . 6), w h i l e l e a f a r e a r e s p o n d e d l a t e i n d e v e l o p m e n t ( F i g . 3 . 2 ) . Hence, e a r l y i n d e v e l o p m e n t , t h e g r e a t e r a b s o l u t e g r o w t h r a t e of e n r i c h e d p l a n t s was d e t e r m i n e d p r i m a r i l y by i n c r e a s e s i n E, w h i l e l a t e r i n d e v e l o p m e n t , a somewhat g r e a t e r l e a f a r e a a l s o c o n t r i b u t e d . L e a f a r e a r a t i o was p r o g e s s i v e l y l o w e r a t h i g h e r c a r b o n d i o x i d e c o n c e n t r a t i o n s ( F i g . 3.7) and t h i s r e s p o n s e p e r s i s t e d t h r o u g h t i m e . Thus, e n r i c h e d p l a n t s a c c u m u l a t e d g r e a t e r d r y w e i g h t p e r u n i t of l e a f s u r f a c e a r e a t h a n c o n t r o l s . Note t h a t a change i n u n i t l e a f r a t e c an i n f l u e n c e t h e l e a f a r e a r a t i o . The d e c r e a s e d F i n e n r i c h e d p l a n t s was i n p a r t due t o a g r e a t e r E ( a s s u m i n g l e a f a r e a t o be c o n s t a n t , an i n c r e a s e i n E w i l l c a u s e a c c u m u l a t i o n o f g r e a t e r d r y w e i g h t , t h e r e b y l o w e r i n g F; F i g u r e 3.2 showed t h a t l e a f a r e a of e n r i c h e d p l a n t s was e i t h e r e q u a l t o or g r e a t e r t h a n t h a t of c o n t r o l s , t h e r e f o r e a d e c r e a s e i n F i n e n r i c h e d p l a n t s must have been due t o a g r e a t e r E ) . S i n c e e n r i c h m e n t i n f l u e n c e d r e l a t i v e g r o w t h r a t e p r i m a r i l y d u r i n g e a r l y g r o w t h , t h e q u e s t i o n a r i s e s a s t o whether s u s t a i n e d e n r i c h m e n t beyond t h a t s t a g e i s a d v a n t a g e o u s . In t h i s r e g a r d , i t i s i n t e r e s t i n g t o n o t e t h a t , i r r e s p e c t i v e of c a r b o n d i o x i d e t r e a t m e n t , R was s i m i l a r a f t e r 30 d a y s of age, but t h e a b s o l u t e g r o w t h r a t e was c o n s i s t e n t l y h i g h e r w i t h e l e v a t e d c a r b o n d i o x i d e c o n c e n t r a t i o n d u r i n g g r o w t h . T h a t i s , l a t e r i n d e v e l o p m e n t , e n r i c h e d p l a n t s were l a r g e r , but d i d n o t show d i f f e r e n c e s i n R. T h e r e f o r e , t h e g r e a t e r s i z e of e n r i c h e d p l a n t s i n l a t e r d e v e l o p m e n t p r o b a b l y d e r i v e d f r o m (1) a s u s t a i n e d g r o w t h a d v a n t a g e i n c u r r e d d u r i n g e a r l y d e v e l o p m e n t , when R was v e r y 51 10.0 15.0 20.0 25.0 30.0 35.0 40.0 45.0 50.0 55.0 TIME AFTER PLANTING IN DAYS FIGURE 3.7 Leaf area r a t i o over time of plants grown at 340, 500, 800, 1200, 2000, or 3000 ul l " 1 carbon dioxide. F i s expressed in units of m g - 1 X10" 52 h i g h , (2) a m a r g i n a l l y h i g h e r E; even t h o u g h t h e a b s o l u t e u n i t l e a f r a t e s were q u i t e low, e n r i c h e d p l a n t s s t i l l showed a l a r g e p e r c e n t a g e i n c r e a s e i n E ( t h i s w o u l d n o t i n f l u e n c e R b e c a u s e of t h e c o m p e n s a t i n g e f f e c t o f a l o w e r F v a l u e ) , and (3) a s l i g h t l y g r e a t e r l e a f a r e a . Hence, a c o m b i n a t i o n of c u m u l a t i v e i n c r e a s e s i n s i z e , as w e l l as an i n c r e a s e i n c u r r e n t g r o w t h may have c o n t r i b u t e d t o l a r g e r e n r i c h e d p l a n t s d u r i n g t h e t i m e when r e l a t i v e g r o w t h r a t e was a p p a r e n t l y no g r e a t e r t h a n i n c o n t r o l s . However, whether a g r o w t h a d v a n t a g e i s m a i n t a i n e d i f e n r i c h m e n t i s d i s c o n t i n u e d may depend on t h e s p e c i e s . K r i z e k e t a l . (1971) f o u n d t h a t t h e s t i m u l a t o r y e f f e c t o f 4 weeks of e n r i c h m e n t of c r a b a p p l e s e e d l i n g s p e r s i s t e d f o r 2 t o 3 months. Hardman and Brun ( 1 9 7 1 ) , however, f o u n d no l a s t i n g e f f e c t of e n r i c h m e n t on s o y b e a n a t m a t u r i t y , when c a r b o n d i o x i d e was a d m i n i s t e r e d d u r i n g t h e f - i r s t 5 weeks of v e g e t a t i v e g r o w t h . . D a v i s and P o t t e r (1983) f o u n d t h a t o f 5 o r n a m e n t a l s p e c i e s w h i c h r e s p o n d e d t o e n r i c h m e n t , o n l y one (• P e p e r o m i a ) r e t a i n e d t h e s i z e d i f f e r e n t i a l a f t e r an a d d i t i o n a l 4 weeks a t a m b i e n t c a r b o n d i o x i d e . C l e a r l y , more work i s n e e ded t o d e t e r m i n e t h e l o n g -t e r m e f f e c t i v e n e s s o f s h o r t - t e r m e n r i c h m e n t . A l t h o u g h c a r b o n d i o x i d e e n r i c h m e n t i n c r e a s e d g r o w t h r a t e , i t had no d i s c e r n a b l e e f f e c t on r a t e of d e v e l o p m e n t . P l a n t s i n a l l c a r b o n d i o x i d e t r e a t m e n t s f l o w e r e d a t a p p r o x i m a t e l y 35 d a y s of age, and began t o s e t pods a t a p p r o x i m a t e l y 42 d a y s of age. T h i s c o n t r a s t s t o t h e work o f H i c k l e n t o n and J o l l i f f e (1978) who f o u n d t h a t f l o w e r i n g i n e n r i c h e d tomato was a d v a n c e d 3 d a y s . The d i f f e r e n c e i n r e s u l t s between my work and t h a t of H i c k l e n t o n 53 and J o l l i f f e (1978) may r e l a t e t o s p e c i e s d i f f e r e n c e s . 3.3.2 E f f e c t s On P a r t i t i o n i n g A change i n t h e p a r t i t i o n i n g of c a r b o n among p l a n t p a r t s may o c c u r as a r e s u l t o f p h y s i o l o g i c a l a d j u s t m e n t s of p l a n t s t o d i f f e r e n t e n v i r o n m e n t a l s i t u a t i o n s . A l t h o u g h t h e r e was some v a r i a b l i t y i n t h e e f f e c t of c a r b o n d i o x i d e c o n c e n t r a t i o n on t h e r o o t - s h o o t r a t i o (R/S) e a r l y i n d e v e l o p m e n t ( F i g . 3.8), a f t e r 30 d a y s o f age t h e R/S was c o n s i s t e n t l y l o w e r i n e n r i c h e d p l a n t s . O t h e r w o r k e r s , however, have r e p o r t e d a g r e a t e r R/S w i t h e n r i c h m e n t i n V i t i s ( K r i e d e m a n n e t a l . , 1976), tomato ( T o g n o n i e t a l . , 1967), b a r l e y , k a l e ( F o r d and T h o r n e , 1967), and s u g a r b e e t ( F o r d and T h o r n e , 1967; Wyse, 1980). I t may be s i g n i f i c a n t t h a t a l l t h e s e a r e non-legumes. Desmodium, a legume, showed a d e c r e a s e d R/S ( W u l f f and S t r a i n , 1982) as d i d t h e bean p l a n t s i n t h i s s t u d y ( a l t h o u g h , i n a n o t h e r legume s t u d y , T o g n o n i e t a l . (1967) f o u n d an i n c r e a s e d R/S i n bush b e a n s ) . . L e a f w e i g h t r a t i o (LWR) was p r o g e s s i v e l y h i g h e r w i t h i n c r e a s e d c a r b o n d i o x i d e e n r i c h m e n t ; t h i s e f f e c t was p e r s i s t e n t o v e r t i m e ( F i g . 3 . 9 ) . The i n c r e a s e i n LWR s u g g e s t s t h a t h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n c a u s e d a g r e a t e r p r o p o r t i o n o f c a r b o n t o be p a r t i t i o n e d a s l e a f w e i g h t . T h i s o b s e r v a t i o n i s s u p p o r t e d by t h e f a c t t h a t b o t h l e a f w e i g h t ( F i g . 3.10) and l e a f a r e a ( F i g . 3.2) i n c r e a s e d w i t h e n r i c h m e n t . T h e r e a r e two a s p e c t s of c a r b o n p a r t i t i o n i n g w h i c h a r e p a r t i c u l a r l y n o t e w o r t h y . T h o s e c o n c e r n t h e p a r t i t i o n i n g o f c a r b o n r e s o u r c e s i n t o new l e a f a r e a p r o d u c t i o n , and i n t o l e a f 54 PLANT AGE , d a y s FIGURE 3.8. R o o t - s h o o t r a t i o o v e r t i m e of p l a n t s grown a t 340, 500, 1200, and 3000 u l l " 1 c a r b o n d i o x i d e . 55 FIGURE 3.9 Leaf weight r a t i o over time of p l a n t s grown a t 340, 500, 800, 1200, 2000, or 3000 u l l " 1 carbon d i o x i d e . 56 a a CO o O CO to az t r CD a {-< ^ r-, U J ^ C T o b. 3 0 0 0 340 10.0 15.0 20.0 25.0 30.0 35.0 40.0 45.0 50.0 TIME AFTER PLANTING IN DAYS 55.0 FIGURE 3.10 Leaf weight over time of plants grown at 340, 500, 800, 1200, 2000, or 3000 ul l " 1 carbon dioxide. Each value i s the mean of 4 plants, with standard deviation. Curves are cubic spine functions of f i t t e d data for each carbon dioxide treatment. 57 w e i g h t p r o d u c t i o n , r e s p e c t i v e l y . From t h e l e a f a r e a r a t i o d a t a ( F i g . 3 . 7 ) , i t was a p p a r e n t t h a t f o r a g i v e n p l a n t s i z e ( s i z e b e i n g e q u a t e d w i t h p l a n t w e i g h t ) , e n r i c h e d p l a n t s a p p e a r e d t o p a r t i t i o n l e s s c a r b o n i n t o l e a f a r e a p r o d u c t i o n t h a n d i d t h e c o n t r o l s . T h e r e f o r e , d e s p i t e a g r e a t e r t o t a l l e a f a r e a i n e n r i c h e d p l a n t s , t h e l e a f a r e a r e l a t i v e t o p l a n t s i z e was a p p a r e n t l y r e d u c e d . A n o t h e r r e l e v a n t a s p e c t of p a r t i t i o n i n g c o n c e r n s t h e r a t e of c a r b o n i n p u t i n t o l e a f w e i g h t . To c h a r a c t e r i z e t h i s p a r t i t i o n i n g , t h e a b s o l u t e g r o w t h r a t e of l e a f w e i g h t (dLw/dt) was compared t o t h a t of t h e r e m a i n d e r of t h e p l a n t (dRw/dt). C a r b o n d i o x i d e e n r i c h m e n t i n i t i a l l y c a u s e d a l a r g e i n c r e a s e i n t h e r a t i o of t h e two g r o w t h r a t e s (dLw/dRw) ( F i g . 3 .11). Hence, a t l e a s t e a r l y i n g r o w t h , a g r e a t e r p r o p o r t i o n o f t h e e n h a n c e d d r y m a t t e r , a c c u m u l a t i o n i n e n r i c h e d p l a n t s was p a r t i t i o n e d i n t o l e a f w e i g h t . T h i s i n i t i a l i n c r e a s e i n t h e r a t e of g r o w t h of l e a f w e i g h t c o u l d have r e s u l t e d from e i t h e r an i n c r e a s e i n t o t a l l e a f a r e a , an i n c r e a s e i n l e a f w e i g h t p e r u n i t a r e a , o r a c o m b i n a t i o n o f t h e two. F o r example, t h e v e r y l a r g e i n c r e a s e i n s p e c i f i c l e a f w e i g h t (SLW) d u r i n g e n r i c h m e n t ( F i g . 3.12) s u g g e s t s t h a t t h e i n c r e a s e i n l e a f w e i g h t was, i n p a r t , due t o an i n c r e a s e i n t h e amount of c a r b o n a c c u m u l a t e d p e r u n i t l e a f a r e a . T h i s c o r r o b o r a t e d s t u d i e s by F o r d and T h o r n e ( 1 9 6 7 ) , Hurd ( 1 9 6 8 ) , N e a l e s and N i c h o l l s ( 1 9 7 8 ) , and W u l f f and S t r a i n ( 1 9 8 2 ) . B e c a u s e t o t a l l e a f a r e a ( F i g . 3.2) d i d not change s i g n i f i c a n t l y i n e n r i c h e d p l a n t s compared t o c o n t r o l s o v e r t h e p e r i o d i n w h i c h 58 O O 3000 CO 2000' ar or UJ ar or r2 o or CD 800 500, 340 a a . 10.0 15.0 2D.D 25.0 30.0 35.• 40.0 45.0 50.0 TIME AFTER PLANTING IN DAYS 55.0 FIGURE 3.11 Ratio of the growth rate of leaf weight to that of the remainder of the plant over time, in plants grown at 340, 500, 800, 1200, 2000, or 3000 ul l " 1 carbon dioxide. 59 FIGURE 3.12 Specific leaf weight over time of plants grown at 340, 500, 800, 1200, 2000, or 3000 ul l " 1 carbon dioxide SLW i s expressed in units of g n r 2 X10"3. 60 a l a r g e change i n dLw/dRw o c c u r r e d ( F i g . 3.11), i t i s r e a s o n a b l e t o c o n c l u d e t h a t t h e change i n p a r t i t i o n i n g was due p r i m a r i l y t o an i n c r e a s e i n l e a f w e i g h t p e r u n i t l e a f a r e a (SLW). An i n c r e a s e i n SLW a p p a r e n t l y r e s u l t e d f r o m an i n c r e a s e i n t h e s t o r a g e f u n c t i o n o f e n r i c h e d l e a v e s , and s u g g e s t s t h a t t h e p a r t i t i o n i n g of c a r b o n between t h e amount t r a n s p o r t e d from l e a v e s , and t h e amount s t o r e d i n l e a v e s , was a l t e r e d due t o e n r i c h m e n t . The major d e c r e a s e i n SLW o c c u r r e d a t a p p r o x i m a t e l y t h e t i m e of r e p r o d u c t i v e g r o w t h , w h i c h c o u l d r e f l e c t a m o b i l i z a t i o n of l e a f c a r b o n r e s e r v e s . However, a t t h e l a t e r s t a g e s o f d e v e l o p m e n t where b o t h l e a f a r e a and SLW had c h a n g e d , t h e r e l a t i v e p a r t i t i o n i n g of c a r b o n i n t o t h e p r o d u c t i o n of new l e a f a r e a , t r a n s p o r t , or s t o r a g e , c o u l d n ot be e v a l u a t e d . When o v e r a l l p l a n t SLW was s u b - d i v i d e d i n t o component SLW f o r e a c h l e a f , an i n t e r e s t i n g p a t t e r n emerged ( F i g . 3 .13). The i n d i v i d u a l l e a v e s of e n r i c h e d p l a n t s i n i t i a l l y showed an i n c r e a s i n g SLW. The p r i m a r y l e a f had t h e l a r g e s t r e s p o n s e , w h i c h was p r o b a b l y a f u n c t i o n of t h e g r e a t e r E a t t h a t s t a g e o f d e v e l o p m e n t (11-22 d a y s ) . The t r i f o l i a t e s d i d n o t a c h i e v e t h e h i g h SLW of t h e p r i m a r y l e a v e s i n e i t h e r c o n t r o l or e n r i c h e d p l a n t s , p e r h a p s b e c a u s e s i n k demand, and hence t r a n s p o r t r a t e s from s o u r c e l e a v e s , i n c r e a s e d w i t h p l a n t a ge. The SLW o f o l d e r l e a v e s ( p r i m a r y and f i r s t t r i f o l i a t e s ) d e c r e a s e d w i t h a c o n c u r r e n t i n c r e a s e i n SLW of you n g e r l e a v e s ( f o u r t h t r i f o l i a t e ) . T h i s may be due t o a p r o g r e s s i v e l y r e d u c e d E i n o l d e r l e a v e s ( s e e C h a p t e r 6 ) . In a d d i t i o n t o o v e r a l l p l a n t g r o w t h r a t e s , t h e p a r t i t i o n i n g 61 PLANT AGE, days FIGURE 3.13 S p e c i f i c l e a f w e i g h t o v e r t i m e o f t h e i n d i v i d u a l l e a v e s of p l a n t s grown a t 340 ( b r o k e n l i n e ) o r 1200 ( s o l i d l i n e ) u l l " 1 c a r b o n d i o x i d e . D a t a a r e p r e s e n t e d f o r t h e p r i m a r y l e a v e s ( P ) , t h e f i r s t t r i f o l i a t e s ( 1 ) , and t h e f o u r t h t r i f o l i a t e s ( 4 ) . 62 of d r y w e i g h t t o t h e pods i s an i m p o r t a n t f a c t o r i n d e t e r m i n i n g y i e l d . D e s p i t e t h e f a c t t h a t pod y i e l d i n t e r m s o f d r y w e i g h t was g r e a t e r i n e n r i c h e d p l a n t s ( T a b l e 3.2), t h e H a r v e s t Index was t h e same, o r somewhat l e s s t h a n t h a t o f c o n t r o l s . E n r i c h m e n t , t h e r e f o r e , d i d not c a u s e a g r e a t e r p a r t i t i o n i n g of c a r b o n i n t o r e p r o d u c t i v e g r o w t h . O v e r a l l t h e n , i t seems t h a t t h e p a r t i t i o n i n g o f a s s i m i l a t e s w i t h i n t h e p l a n t was i n f l u e n c e d by e n r i c h m e n t m a i n l y t o t h e e x t e n t t h a t w i t h g r e a t e r E, a g r e a t e r p r o p o r t i o n o f f i x e d c a r b o n r e m a i n e d i n t h e l e a v e s . 3.3.3 Summary C o n t i n u o u s c a r b o n d i o x i d e e n r i c h m e n t o f b u s h beans c a u s e d g r e a t e r g r o w t h , t h a t i s l a r g e r p l a n t s , but' d i d n o t i n f l u e n c e r a t e of. d e v e l o p m e n t . A l t h o u g h e n r i c h e d p l a n t s were c o n s i s t e n t l y l a r g e r t h a n c o n t r o l s , t h e r e l a t i v e g r o w t h r a t e was g r e a t e r o n l y d u r i n g e a r l y g r o w t h . The t r e n d i n r e l a t i v e g r o w t h r a t e was, a t l e a s t i n p a r t , a r e s u l t of c h a n g e s i n u n i t l e a f r a t e , w h i c h was a l s o h i g h e r i n e n r i c h e d p l a n t s o n l y d u r i n g e a r l y g r o w t h . The d a t a s u p p o r t o b s e r v a t i o n s made i n o t h e r s p e c i e s t h a t c o n t i n u o u s c a r b o n d i o x i d e e n r i c h m e n t was e f f e c t i v e i n i n c r e a s i n g r e l a t i v e g r o w t h r a t e and u n i t l e a f r a t e f o r o n l y a l i m i t e d t i m e . The i n c r e a s e d u n i t l e a f r a t e i n e n r i c h e d p l a n t s was p r o b a b l y r e s p o n s i b l e f o r t h e d e c r e a s e d l e a f a r e a r a t i o and i n c r e a s e d s p e c i f i c l e a f w e i g h t compared t o c o n t r o l s ; t h o s e c h a n g e s have a l s o been o b s e r v e d i n o t h e r s p e c i e s . The change i n l e a f w e i g h t p a r t i t i o n i n g , d e t e r m i n e d from th e r a t i o o f t h e g r o w t h r a t e o f l e a f w e i g h t t o t h e g r o w t h r a t e of 63 TABLE 3.2. Total dry weight, pod weight, and Harvest Index of plants grown at 340, 500, 800, 1200, 2000, or 3000 ul l " 1 carbon dioxide for 44 days. 1 Treatment Pod Weight Total Weight Harvest Index (g) (g) 340 ul I ' 1 7.54 ±0 .28 18.30 ± 0 . 4 8 0.41±0.01 500 ul I" 1 9.30 ±0 .20 22.64 ±0 .87 0 .41±0 .01 800 ul I" 1 1 0.58 ±0 .10 26.28 ± 0 . 5 2 0 .39±0 .01 1 200 ul I" 1 13.28 ± 0 . 6 6 31.21 ±0 .67 0 .43±0 .01 2000 ul I ' 1 1 1 .97 ± 0 . 5 2 29.86 ±1 .37 0 . 4 0 ± 0 . 0 2 3000 ul I" 1 1 1 .05 ± 0 . 4 3 31 .02 ± 0 . 6 0 0 . 3 6 ± 0 . 0 2 Values are the means of 4 plants, with standard error 6 4 t h e r e m a i n d e r of t h e p l a n t , i s e v i d e n c e f o r a change i n p a r t i t i o n i n g o f d r y m a t t e r t o t h e l e a v e s . At l e a s t e a r l y i n d e v e l o p m e n t , t h i s p a r t i t i o n i n g was due e n t i r e l y t o an i n c r e a s e i n t h e s t o r a g e f u n c t i o n o f e n r i c h e d l e a v e s . To d a t e , t h e c o n t r i b u t i o n t o o v e r a l l s p e c i f i c l e a f w e i g h t of t h a t of i n d i v i d u a l l e a v e s of e n r i c h e d p l a n t s has not been r e p o r t e d . The s p e c i f i c l e a f w e i g h t of r e p r e s e n t a t i v e l e a v e s of e n r i c h e d and c o n t r o l p l a n t s was t r a c k e d t h r o u g h d e v e l o p m e n t . T h i s o f f e r e d a more d e t a i l e d d e s c r i p t i o n o f d r y m a t t e r a l l o c a t i o n i n r e p r e s e n t a t i v e l e a v e s of e n r i c h e d and c o n t r o l p l a n t s t h a n d i d a g e n e r a l i z e d s p e c i f i c l e a f w e i g h t . The r e s u l t s showed d i s t i n c t i v e c h a n g e s i n s p e c i f i c l e a f w e i g h t f o r t h o s e l e a v e s t h r o u g h t i m e , p a r t i c u l a r l y i n e n r i c h e d p l a n t s . 65 IV. EFFECTS ON NET CARBON DIOXIDE EXCHANGE 4.1 INTRODUCTION In view o f t h e d r a m a t i c c h a n g e s i n E t h r o u g h t i m e and among c a r b o n d i o x i d e t r e a t m e n t s , and b e c a u s e of t h e i m p o r t a n c e of c a r b o n d i o x i d e a s s i m i l a t i o n i n g r o w t h , i t . was p e r t i n e n t t o d e t e r m i n e i n a more d e t a i l e d manner t h e p h o t o s y n t h e t i c p h y s i o l o g y of e n r i c h e d p l a n t s . The few i n v e s t i g a t i o n s w h i c h have been made of t h e p h o t o s y n t h e t i c gas exchange o f i n d i v i d u a l l e a v e s of e n r i c h e d p l a n t s have y i e l d e d c o n f l i c t i n g r e s u l t s . Some have r e p o r t e d i n c r e a s e d p h o t o s y n t h e t i c c a p a c i t y ( B i s h o p and W h i t t i n g h a m , 1968; Madsen, 1975; H i c k l e n t o n and J o l l i f f e , 1980b), no e f f e c t ( F o r d and T h o r n e , 1967;. G i f f o r d , 1 9 7 7 ) , or d e c r e a s e d c a p a c i t y ( f o r example, H o f s t r a and H e s k e t h , 1975; Ho, 1977; Mauney e t a l . , 1979; C l o u g h - e t a l . , 1981; W u l f f and S t r a i n , 1982). I t i s n o t a t a l l c l e a r why t h e change i n c a p a c i t y w i t h e n r i c h m e n t o c c u r s ; t h e r e i s e v i d e n c e t h a t a change i n s t o m a t a l r e s i s t a n c e ( A o k i and Y a b u k i , 1977; Imai and M u r a t a , 1978c) and m e s o p h y l l r e s i s t a n c e ( A o k i and Y a b u k i , 1977) may be i n v o l v e d , but i t i s not known why e i t h e r of t h o s e c h a n g e . However, l a r g e a c c u m u l a t i o n s o f s t a r c h have been o b s e r v e d i n e n r i c h e d l e a v e s ( e g . H o f s t r a and H e s k e t h , 1975; W u l f f and S t r a i n , 1982), and N a f z i g e r and R o l l e r (1976) have shown t h a t l e a f s t a r c h c o n t e n t was n e g a t i v e l y c o r r e l a t e d w i t h p h o t o s y n t h e s i s r a t e . T h e r e f o r e , a l a r g e s t a r c h c o n c e n t r a t i o n i n l e a v e s of e n r i c h e d p l a n t s may be r e l a t e d t o t h e r e d u c t i o n i n p h o t o s y n t h e t i c c a p a c i t y of t h o s e 66 l e a v e s . In a d d i t i o n , i t i s not known i f the response to enrichment i s i n f l u e n c e d by other environmental c o n d i t i o n s . With t h i s i n mind, the o b j e c t i v e s of t h i s chapter were to answer the f o l l o w i n g p e r t i n e n t q u e s t i o n s : 1. Does carbon d i o x i d e enrichment change the net carbon d i o x i d e exchange c a p a c i t y of bean p l a n t s ? 2. If so, i s the apparent change i n carbon d i o x i d e exchange c a p a c i t y due to a change i n p r o p e r t i e s of the l e a f which i n f l u e n c e CER? For example, changes i n stomatal r e s i s t a n c e , or changes i n l e a f t h i c k n e s s or c h l o r o p h y l l content may i n f l u e n c e the measurement of carbon d i o x i d e exchange c a p a c i t y . 3. Can the change in carbon d i o x i d e exchange c a p a c i t y of e n r i c h e d p l a n t s be' a t t r i b u t e d to a change in p h o t o r e s p i r a t ion rate? 4. Is dark r e s p i r a t i o n rate a f f e c t e d by enrichment? 5. Is the change i n carbon d i o x i d e exchange c a p a c i t y of en r i c h e d p l a n t s m o d i f i e d by other growth c o n d i t i o n s , such as l i g h t i n t e n s i t y ? 6. Is the change i n carbon d i o x i d e exchange c a p a c i t y of an i n d i v i d u a l l e a f c o n s i s t e n t through time, and how would t h i s r e l a t e to changing environmental c o n d i t i o n s (such as shading)? 7. Does s t a r c h content of e n r i c h e d leaves change, and i f so, i s there a r e l a t i o n s h i p between t h i s change and the carbon d i o x i d e exchange c a p a c i t y ? 8. Is the change i n carbon d i o x i d e exchange c a p a c i t y of e n r i c h e d p l a n t s r e l a t e d to a change i n the so u r c e / s i n k r e l a t i o n s h i p s of the p l a n t , and can the change in c a p a c i t y be 67 a l l e v i a t e d by c h a n g i n g s i n k demand? Answers t o t h e s e q u e s t i o n s a r e p e r t i n e n t t o an u n d e r s t a n d i n g o f how c a r b o n d i o x i d e e n r i c h m e n t a f f e c t s t h e p h y s i o l o g y of l e a v e s and u l t i m a t e l y p l a n t g r o w t h and p r o d u c t i v i t y . 68 4.2 MATERIALS AND METHODS Seeds were germinated, and before transfer to the carbon dioxide treatment chambers, grown according to the conditions previously outlined (section 3.2.1). In one instance, plants were also grown in 3 inch pots (one plant per pot; s o i l volume was 0.15 1). At 12 days after planting, 4 to 8 bean plants were transferred to each chamber at either 340±10 ul l " 1 (control) or 1400±20 ul l " 1 (enriched) carbon dioxide, unless otherwise indicated. Chamber temperature was 32/25 C day/night and photosynthetic photon flux density (PPFD) incident on the f i r s t t r i f o l i a t e was approximately 350 umol n r 2 s~ 1, unless otherwise indicated. In addition to bean, tomato' plants ( Lycopersicon  esculentum L., cv. Vendor) were used in some experiments, and were transferred at 19 days to 340±10 or 1000±20 ul l " 1 carbon dioxide. Plastochron Index. (Erickson, 1957) was used to establish physiological age. The Plastochron Index uses leaf number as an index of development; plants of the same leaf number were assumed to be at the same developmental stage, despite possible differences in growth rate. In e f f e c t , this allowed separation of growth from developmental effects of the environment. The Plastochron Index was obtained by counting a l l leaves over a designated length. Fractional values resulted when the smallest leaf length was a fraction of the reference length. My reference length was 10mm with the f i r s t t r i f o l i a t e being the f i r s t plastochron. At the time of transfer, PI=0.5. 69 Measurements of net c a r b o n d i o x i d e exchange r a t e (CER, e x p r e s s e d as umoles of c a r b o n d i o x i d e l e a f • a r e a * 1 t i m e " 1 ) , t r a n s p i r a t i o n r a t e s , and d a r k r e s p i r a t i o n r a t e s were o b t a i n e d i n th e f i r s t and s e c o n d t r i f o l i a t e s . A semi-open gas e x c h a n g e s y s t e m was b u i l t s p e c i f i c a l l y f o r t h i s p u r p o s e ( E h r e t and J o l l i f f e , 1983). The s y s t e m p e r m i t t e d s t e a d y - s t a t e measurements of b o t h water v a p o u r and c a r b o n d i o x i d e gas e x c h a n g e s of a l e a f e n c l o s e d i n a " t r a p " t y p e c u v e t t e . C o n d i t i o n s o f t e m p e r a t u r e , l i g h t i n t e n s i t y , h u m i d i t y and gas c o m p o s i t i o n i n t h e c u v e t t e v a r i e d a c c o r d i n g t o t h e e x p e r i m e n t , and w i l l be o u t l i n e d where needed. Measurement of water v a p o u r f l u x ( t r a n s p i r a t i o n ) p e r m i t t e d c a l c u l a t i o n of s t o m a t a l r e s i s t a n c e , w h i c h i s a measure of t h e a b i l i t y of s t o m a t a t o a c t as p o r t a l s f o r t h e n e t d i f f u s i o n o f water v a p o u r . S t o m a t a l r e s i s t a n c e was c a l c u l a t e d f r o m t h e f o l l o w i n g e q u a t i o n d e r i v e d f r o m Ohm's Law, where n e t f l u x i s e q u a l t o a d r i v i n g f o r c e d i v i d e d by a r e s i s t a n c e : X i = a b s o l u t e c o n c e n t r a t i o n o f w a t e r v a p o u r a t t h e s i t e of e v a p o r a t i o n i n t h e l e a f (mg m" 3) Xo = a b s o l u t e c o n c e n t r a t i o n of water v a p o u r i n t h e e x t e r n a l a i r (mg m" 3) Rs(H20)= s t o m a t a l r e s i s t a n c e t o t h e J ( H 2 0 ) X i - Xo where: J ( H 2 0 ) Rs(H20)+ Ra(H20) f l u x of water v a p o u r (mg m"2 s " 1 ) d i f f u s i o n of water v a p o u r (s m" 1) Ra(H20) boundary l a y e r r e s i s t a n c e t o t h e d i f f u s i o n of water v a p o u r (s m" 1) 70 Ra was determined using b l o t t i n g paper in place of the leaf in the chamber (Ra(H2O) = 50 s ITT 1 ) . Hence, a l l components except Rs were known, and Rs could be determined by solving the above equat ion. The same concept applies to the flux of carbon dioxide. The resistance to d i f f u s i o n of carbon dioxide to the mesophyll c e l l s is again Rs + Ra. However, the d i f f u s i v i t y of carbon dioxide is less than that of water vapour, so in e f f e c t , the resistance to transfer of carbon dioxide i s greater by a factor of 1.56. The following equation was used to solve for the concentration of carbon dioxide in the i n t e r c e l l u l a r spaces: J(C02) = Co - Ci  Rs(C02) + Ra(C02) where: J(C02) = net flux of carbon dioxide (CER, in mg nr 2 s~ 1) Co = external concentration of carbon dioxide (mg nr 3) Ci = i n t e r c e l l u l a r space carbon dioxide concentration (mg nr 3) Rs(C02) = stomatal resistance to the d i f f u s i o n of carbon dioxide (s nr 1) Ra(C02) = boundary layer resistance to the di f f u s i o n of carbon dioxide (s nr 1) In some experiments, the starch content of leaf tissue was determined using a modification of the method of Huber and Israel (1982). Samples (200 to 500 mg fresh weight, equivalent to about 4 one-cm leaf disks) were extracted in 80% ethanol at 80 C u n t i l the tissue was pigment-free. This step removed 71 s o l u b l e sugars. Samples were then homogenized i n 2 to 5 ml of 0.2N KOH c o n t a i n i n g 100 mM ethanol and heated f o r 2 hours at 80 C to s o l u b i l i z e the s t a r c h . A f t e r c o o l i n g , the pH was a d j u s t e d to 5.5 with 1M a c e t i c a c i d . An equal volume of amyloglucosidase (Sigma, A-7255, obtained from Rhizopus ) s o l u t i o n (400 u n i t s ml" 1 i n 0.1M c i t r a t e b u f f e r , pH 5.5) was added and the s o l u t i o n was then incubated f o r 4 hours at 45 C. T h i s converted s t a r c h to g l u c o s e . P l a c i n g the r e a c t i o n v e s s e l i n b o i l i n g water f o r 1 minute terminated the r e a t i o n . A f t e r a 2 0 - f o l d d i l u t i o n , a 0.5 ml a l i q u o t was taken f o r glucose d e t e r m i n a t i o n . Glucose was assayed i n a two-enzyme sequence, using Sigma glucose k i t 510-A (see a l s o Sigma t e c h n i c a l b u l l e t i n 510). In t h i s procedure, glucose was f i r s t converted to g l u c u r o n i c a c i d u s ing glucose oxidase, and t h i s step l i b e r a t e d hydrogen pe r o x i d e . Then, i n a p e r o x i d a s e - c a t a l y z e d r e a c t i o n , hydrogen peroxide o x i d i z e d o-d i a n i s i d i n e , r e s u l t i n g i n c o l o u r development. T h i s was measured at 450 nm i n a spectrophotometer. C h l o r o p h y l l content was measured in 4 one-cm d i s k s of i n t e r v e i n a l t i s s u e taken from the leaves which was used i n the gas exchange s t u d i e s . Samples were s t o r e d at -20 C u n t i l needed. C h l o r o p h y l l was q u a n t i f i e d using a m o d i f i e d v e r s i o n of the technique of Bruinsma (1963). Disks were homogenized in 30 ml of c o l d 80% acetone f o r one minute in a blender ( O s t e r i z e r , model C y c l o t r o l 8 ). A l l steps of the e x t r a c t i o n were c a r r i e d out i n dim l i g h t using c o l d reagents and v e s s e l s . The homogenate was vacuum f i l t e r e d on Whatman #1 f i l t e r paper. The e x t r a c t was assayed f o r c h l o r o p h y l l by measuring absorbance at 72 645, 652, and 663 nm. C h l o r o p h y l l s a and b and t h e i r sum were t h e n c a l c u l a t e d as f o l l o w s ( i n mg l " 1 ) : C h i a = 12.7 A663 - 2.7 A645 C h i b = 22.9 A645 - 4.7 A663 C h i a + b = 20.2 A645 + 8 A663 = 27.8 A652 In some e x p e r i m e n t s , l e a f t h i c k n e s s was d e t e r m i n e d by d i v i d i n g l e a f a r e a by l e a f volume. L e a f a r e a was measured w i t h a l e a f a r e a meter ( L i - C o r model L I - 3 0 0 0 ) , and l e a f volume was d e t e r m i n e d u s i n g a l i q u i d d i s p l a c e m e n t t e c h n i q u e ( H u x l e y , 1971). L e a f i n t e r c e l l u l a r a i r volume was a p p r o x i m a t e d by d i v i d i n g l e a f f r e s h w e i g h t by l e a f volume. As s u m i n g w a t e r t o be t h e m a j o r component of l e a f f r e s h w e i g h t , and knowing t h e d e n s i t y of w a t e r t o be 1, t h e d i f f e r e n c e i n measured l e a f volume and t h e e x p e c t e d l e a f volume ( b a s e d on t h e d e n s i t y o f w a t e r ) was t h e volume o f t h e i n t e r c e l l u l a r a i r s p a c e s . ' • Where r e q u i r e d , PPFD was d e t e r m i n e d w i t h a quantum meter ( L i - C o r , model L I - 1 8 5 ) . Where a p p r o p r i a t e , two r e g r e s s i o n p a c k a g e s p r o v i d e d by t h e U n i v e r s i t y of B r i t i s h C o l u m b i a Computing S e r v i c e s C e n t r e were use d t o f i t d a t a t o a c u r v e . An a s y m p t o t i c r e g r e s s i o n p a c k a g e was u s e d t o f i t c a r b o n d i o x i d e r e s p o n s e c u r v e s , and a t r i a n g u l a r r e g r e s s i o n p a c k a g e (TRP) was u s e d t o f i t t h e d a t a o f s t a r c h c o n t e n t v e r s u s CER t o a t h i r d - o r d e r p o l y n o m i a l r e g r e s s i o n . 73 4.3 RESULTS AND DISCUSSION 4.3.1 G e n e r a l i z e d Response Of Net C a r b o n D i o x i d e Exchange  Capac i t y A l t h o u g h c a r b o n d i o x i d e e n r i c h e d beans showed en h a n c e d u n i t l e a f r a t e s compared t o c o n t r o l s e a r l y i n d e v e l o p m e n t ( s e e s e c t i o n 3.3.1), t h e c a p a c i t y o f i n d i v i d u a l l e a v e s t o exchange c a r b o n d i o x i d e was r e d u c e d ( T a b l e 4.1a and 4.1b). The CER of e n r i c h e d p l a n t s measured a t 340 u l 1~ 1 c a r b o n d i o x i d e a f t e r 15 t o 19 d a y s of t r e a t m e n t was r e d u c e d 19% and 27% i n t h e f i r s t and s e c o n d t r i f o l i a t e s , r e s p e c t i v e l y . An i m p o r t a n t d i s t i n c t i o n i n t e r m i n o l o g y must be r e i n f o r c e d a t t h i s p o i n t . Measurement o f CER of e n r i c h e d and c o n t r o l p l a n t s a t t h e i r r e s p e c t i v e c a r b o n d i o x i d e c o n c e n t r a t i o n s i s n o t an a d e q u a t e measure o f t h e c o m p a r a t i v e a b i l i t y o f t h o s e p l a n t s t o exchange c a r b o n d i o x i d e w i t h t h e a t m o s p h e r e . The measurement of CER i n t h i s c a s e i s o b t a i n e d a t two d i f f e r e n t c a r b o n d i o x i d e c o n c e n t r a t i o n s , and o n l y shows t h a t t h e r a t e s o f c a r b o n d i o x i d e e x c h a n g e may be d i f f e r e n t . T h e r e f o r e , c a r b o n d i o x i d e exchange c a p a c i t y i s d e f i n e d as t h e a b i l i t y of e n r i c h e d p l a n t s t o c a r r y o u t n e t c a r b o n d i o x i d e exchange (measured as CER) compared t o c o n t r o l s under t h e same measurement c o n d i t i o n s . T h r o u g h o u t t h i s c h a p t e r , a change i n c a p a c i t y was f r e q u e n t l y e x p r e s s e d i n t e r m s of a p e r c e n t o f c o n t r o l CER v a l u e s . The r e d u c t i o n i n CER of e n r i c h e d l e a v e s was o b s e r v e d a t a l l measurement c a r b o n d i o x i d e c o n c e n t r a t i o n s examined ( F i g . 4 . 1 ) . The CER i n F i g u r e 4.1 was e x p r e s s e d i n terms o f i n t e r c e l l u l a r 74 TABLE 4 . 1 a . Net carbon d i o x i d e exchange ra te of the f i r s t and second t r i f o l i a t e s of c o n t r o l (340 u l I ' M and e n r i c h e d (1400 u l l " 1 ) p l a n t s . 1 Treatment Leaf CER 2 (umol nr 2 s " 1 ) 340 F i r s t T r i f o l i a t e 3 17.8 ±0.9 u l l " 1 Second T r i f o l i a t e " 16.7 ±1.2 1400 F i r s t T r i f o l i a t e 3 14.3 ±1.5 u l I"1 Second T r i f o l i a t e " 12.3 ±0.8 Va lues are the means of 3 measurements made on d i f f e r e n t p l a n t s , w i th s tandard e r r o r CER measured at 1 000 umol n r 2 s " 1 PPFD and 340 u l l " 1 carbon d i o x i d e Measured a f t e r 15 days of t reatment Measured a f t e r 19 days of t reatment TABLE 4 .1b . A n a l y s i s of v a r i a n c e of the data of TABLE 4 . 1 a . Source df Sum Sq. Mean Sq . E r r o r F Requ i red F 1 Treatment 1 161.29 161.29 11.30 4.67 T r i f o l i a t e 1 21.62 21.62 1.51 4.67 E r r o r 13 185.59 14.28 T o t a l 15 368.50 1 At 5% l e v e l of p r o b a b i l i t y 75 FIGURE 4.1 Response of n e t c a r b o n d i o x i d e exchange r a t e of t h e f i r s t t r i f o l i a t e o f e n r i c h e d (1400 u l 1"') and c o n t r o l (340 u l l " 1 ) p l a n t s ( a t PI=5.0) t o i n t e r c e l l u l a r s p a c e c a r b o n d i o x i d e c o n c e n t r a t i o n . Measurement PPFD was 1000 umol m"" s * 1 . P l a n t s were i n t h e t r e a t m e n t chambers f o r 18 day b e f o r e CER measurement. PPFD i n c i d e n t on t h e f i r s t t r i f o l i a t e d u r i n g g r o w t h was 300 t o 350 umol m~ * s"'. Measurements were o b t a i n e d f r o m 3 p l a n t s i n e a c h t r e a t m e n t . E n r i c h e d l e a v e s , c l o s e d c i r c l e s ; c o n t r o l l e a v e s , open c i r c l e s . C u r v e s were f i t t e d t o an a s y m p t o t i c e q u a t i o n u s i n g 13 and 14 v a l u e s f o r c o n t r o l and e n r i c h e d t r e a t m e n t s r e s p e c t i v e l y . 76 c a r b o n d i o x i d e c o n c e n t r a t i o n , e s t a b l i s h i n g t h a t t h e r e d u c t i o n i n c a r b o n d i o x i d e exchange c a p a c i t y was not due t o a r e d u c t i o n i n s t o m a t a l a p e r t u r e . A knowledge o f s t o m a t a l a p e r t u r e was an i m p o r t a n t c o n s i d e r a t i o n , s i n c e , f o r example, a t r e a t m e n t - i n d u c e d change i n s t o m a t a l a p e r t u r e c o u l d have c h a n g e d t h e c a r b o n d i o x i d e c o n c e n t r a t i o n w h i c h was a v a i l a b l e f o r a s s i m i l a t i o n i n t h e l e a f , and t h e r e b y i n f l u e n c e d t h e measured CER v a l u e . The s u b s t a n t i a l i n f l u e n c e of s t o m a t a l a p e r t u r e on t h e measurement of CER i s not t a k e n i n t o c o n s i d e r a t i o n i n many s t u d i e s o f c a r b o n d i o x i d e e n r i c h m e n t e f f e c t s on p l a n t s . E x t r a p o l a t i o n o f t h e c u r v e s t o z e r o n e t c a r b o n d i o x i d e e xchange s u g g e s t e d t h a t b o t h t r e a t m e n t s had s i m i l a r c a r b o n d i o x i d e c o m p e n s a t i o n p o i n t s of a b o u t 50 u l 1 ~ 1 . The c a r b o n d i o x i d e c o m p e n s a t i o n p o i n t i s f r e q u e n t l y u s e d t o c h a r a c t e r i z e t h e r e l a t i o n s h i p between p h o t o s y n t h e s i s and r e s p i r a t i o n i n t h e l i g h t , t h e l a t t e r b e i n g p r i m a r i l y p h o t o r e s p i r a t i o n ( C a n v i n , 1979). At t h e c a r b o n d i o x i d e c o m p e n s a t i o n p o i n t , t h e r a t e of r e s p i r a t o r y c a r b o n d i o x i d e r e l e a s e i s e q u a l t o t h e r a t e of p h o t o s y n t h e t i c c a r b o n d i o x i d e u p t a k e ; a change i n t h e c o m p e n s a t i o n p o i n t i s i n d i c a t i v e of a change i n t h e r e l a t i v e r a t e s of t h o s e two p r o c e s s e s . T h e r e f o r e , a s i m i l a r c a r b o n d i o x i d e c o m p e n s a t i o n p o i n t i m p l i e s t h a t t h e r e d u c e d c a r b o n d i o x i d e e x c h a n g e c a p a c i t y of e n r i c h e d p l a n t s was p r o b a b l y n ot due t o a p r o p o r t i o n a l l y g r e a t e r p h o t o r e s p i r a t i o n r a t e . The c o n c l u s i o n t h a t t h e r e was no d i f f e r e n c e i n p h o t o r e s p i r a t i o n r a t e between c a r b o n d i o x i d e t r e a t m e n t s when measured under t h e same c o n d i t i o n s was s u p p o r t e d by t h e f a c t t h a t b o t h t r e a t m e n t s showed 77 th e same d e g r e e of oxygen i n h i b i t i o n o f CER ( T a b l e 4 . 2 ) . Oxygen i n h i b i t i o n o f CER has been shown t o be r e l a t e d t o p h o t o r e s p i r a t i o n r a t e ( J o l l i f f e and T r e g u n n a , 1968). C o n t i n u o u s e x p o s u r e of bean p l a n t s t o h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n s i n my s t u d y a p p a r e n t l y d i d not change t h e r e l a t i v e r a t e s o f t r u e c a r b o n d i o x i d e exchange ( c a r b o n d i o x i d e e xchange i n t h e a b s e n c e of p h o t o r e s p i r a t i o n ) and p h o t o r e s p i r a t o r y c a r b o n " d i o x i d e exchange when measured under t h e same c o n d i t i o n s . Even t h o u g h e n r i c h e d p l a n t s p r e s u m a b l y had r e d u c e d p h o t o r e s p i r a t i o n r a t e s compared t o c o n t r o l s , i f measured a t t h e i r r e s p e c t i v e g r o w t h c a r b o n d i o x i d e c o n c e n t r a t i o n s , t h e a b i l i t y of e n r i c h e d p l a n t s t o p h o t o r e s p i r e d i d n o t d i f f e r from c o n t r o l s . G i f f o r d (1977) f o u n d no d i f f e r e n c e i n t h e c a r b o n d i o x i d e c o m p e n s a t i o n p o i n t among p l a n t s grown a t d i f f e r e n t c a r b o n d i o x i d e c o n c e n t r a t i o n s , w h i c h i s not s u r p r i s i n g s i n c e - no d i f f e r e n c e i n p h o t o s y n t h e t i c r e s p o n s e was r e p o r t e d . H i c k l e n t o n and J o l l i f f e ( 1 9 8 0 b ) , on t h e o t h e r hand, f o u n d t h a t t o g e t h e r w i t h an e n h a n c e d c a r b o n d i o x i d e e xchange c a p a c i t y i n p l a n t s grown a t 1000 u l 1" 1 c a r b o n d i o x i d e , a d e c r e a s e d c a r b o n d i o x i d e c o m p e n s a t i o n p o i n t and r e d u c e d oxygen s e n s i t i v i t y a l s o o c c u r r e d . The s i g n i f i c a n c e o f t h o s e r e s u l t s w i l l be d i s c u s s e d l a t e r ( s e c t . 4.3.8) . The r e s p o n s e of t h e f i r s t t r i f o l i a t e l e a f o f e n r i c h e d p l a n t s t o PPFD was d e t e r m i n e d . A t b o t h 340 and 1000 u l 1-1 measurement c a r b o n d i o x i d e c o n c e n t r a t i o n s and a t PPFD a p p r o a c h i n g s a t u r a t i o n , e n r i c h e d p l a n t s showed r e d u c e d CER compared t o c o n t r o l s ( F i g . 4 . 2 ) . The r e d u c e d CER o f e n r i c h e d 78 TABLE 4 . 2 . Oxygen s e n s i t i v i t y of the net carbon d i o x i d e exchange r a t e of the f i r s t t r i f o l i a t e of c o n t r o l (340 u l l " 1 ) and en r i ched (1400 u l l " 1 ) p l a n t s t r e a t e d f o r 13 d a y s . 1 Treatment CER 2 CER 2 % I n h i b i t i o n 3 ( 1 % oxygen) (21% oxygen) (umol m~2 s " 1 ) (umol m~2 s " 1 ) 340 24.1±1.3 16.410.5 31 .911 .6 u l I'1 1400 18.111.7 12.211.2 32.6+3.2 u l 1- 1 Va lues are the means of 3 p l a n t s , w i th s tandard e r r o r CER measured at 1000 umol m" 2 s " 1 PPFD D i f f e r e n c e between CER measured at 21% and 1% oxygen w i t h i n a carbon d i o x i d e t r ea tment , expressed as a percentage of the 1% oxygen va lue 79 FIGURE 4.2 Response of t h e n e t c a r b o n d i o x i d e exchange r a t e of t h e f i r s t t r i f o l i a t e of e n r i c h e d (1400 u l l " 1 ) and c o n t r o l (340 u l 1"') p l a n t s t o p h o t o s y n t h e t i c p h o t o n f l u x d e n s i t y . L e a v e s measured a t 1000 u l 1"' ( s o l i d l i n e s ) and 340 u l ( b r o k e n l i n e s ) c a r b o n d i o x i d e c o n c e n t r a t i o n s were from d i f f e r e n t s e t s of p l a n t s . P l a n t s i n a l l c a s e s were t r e a t e d f o r 18 days a t t h e c a r b o n d i o x i d e c o n c e n t r a t i o n s i n d i c a t e d i n t h e f i g u r e . E n r i c h e d l e a v e s , c l o s e d c i r c l e s ; c o n t r o l l e a v e s , open c i r c l e s . E a c h v a l u e i s t h e mean o f 3 l e a v e s , w i t h s t a n d a r d e r r o r . 80 plants was again not due to a reduction in stomatal aperture since i n t e r c e l l u l a r carbon dioxide concentrations were similar in both treatments at a given l e v e l of PPFD (data not presented). At r a t e - l i m i t i n g PPFD there was no difference between treatments, indicating an equivalent quantum e f f i c i e n c y and l i g h t compensation point. Quantum e f f i c i e n c y decreases with an increase in oxygen s e n s i t i v i t y (Ehleringer and Bjorkman, 1977). The fact that quantum e f f i c i e n c y was not affected by enrichment again points out the lack of enrichment e f f e c t s on photorespiration rate when measured under the same conditions in both treatments. In summary, the gas exchange data supported the general trend of reduced net carbon dioxide exchange capacity in enriched plants which has been observed by many others (Hofstra and Hesketh, 1975; Frydrych, 1976; Aoki and Yabuki", 1977; Ho, 1977; Mauney et a l . , 1979; Wong, 1979; Clough et a l . , 1981; Wulff and Strain, 1982), and also suggested that the effect was not caused by a change in photorespiration rate. 4.3.2 Response Of Specific Parameters Of Net Carbon Dioxide  Exchange Capacity Many reference points commonly used to express CER may change when plants are grown under d i f f e r e n t conditions. For example, chlorophyll content, dry or fresh weight, or protein content, a l l of which are used as a basis for comparing photosynthesis rates, may themselves change in response to carbon dioxide treatment. It was therefore necessary to establish that the measured changes in CER were independent of 81 c h a n g e s i n t h o s e p a r a m e t e r s . P o s s i b l e e f f e c t s o f c a r b o n d i o x i d e t r e a t m e n t s on l e a f t h i c k n e s s o r t o t a l c e l l volume would d i m i n i s h t h e u s e f u l n e s s of l e a f a r e a as a b a s i s o f e x p r e s s i n g CER. L e a f t h i c k n e s s of t h e f i r s t and s e c o n d t r i f o l i a t e s , however, was not a f f e c t e d by c a r b o n d i o x i d e e n r i c h m e n t ( T a b l e 4 . 3 ) . Hence, e x p r e s s i o n o f p h o t o s y n t h e s i s on a u n i t l e a f a r e a b a s i s was a r e a s o n a b l e a p p r o a c h i n t h i s s t u d y . The p e r c e n t a g e of l e a f volume o c c u p i e d by a i r , a p a r a m e t e r w h i c h c o u l d a f f e c t CER t h r o u g h c h a n g e s i n t h e c h a r a c t e r i s t i c s of t h e i n t e r n a l d i f f u s i o n of c a r b o n d i o x i d e , was a l s o f o u n d not t o be i n f l u e n c e d by e n r i c h m e n t ( T a b l e 4 . 3 ) . Net c a r b o n d i o x i d e e xchange r a t e i s o f t e n e x p r e s s e d on a u n i t c h l o r o p h y l l b a s i s , even t h o u g h c h l o r o p h y l l l e v e l d o e s not a l w a y s c o r r e l a t e w i t h p h o t o s y n t h e s i s r a t e ( H e s k e t h e t a l . 1981). C a r b o n d i o x i d e e n r i c h m e n t had no e f f e c t on c h l o r o p h y l l l e v e l s i n mature t r i f o l i a t e s ( e g . F i g . 4.3), a l t h o u g h c h l o r o p h y l l l e v e l s i n young l e a v e s were o f t e n r e d u c e d ( p e r s o n a l o b s e r v a t i o n ; Cave e t a l . , 1981). T h i s c o u l d r e p r e s e n t an i n f l u e n c e o f c a r b o n d i o x i d e on t h e r a t e of g r e e n i n g . R a t e s of n e t c a r b o n d i o x i d e exchange g i v e n i n F i g u r e 4.2 (measured a t 1000 u l l " 1 c a r b o n d i o x i d e ) were c o n v e r t e d t o a u n i t c h l o r o p h y l l b a s i s i n F i g u r e 4.3. C a r b o n d i o x i d e e n r i c h m e n t c a u s e d r e d u c e d p h o t o s y n t h e t i c r a t e s not a s s o c i a t e d w i t h c h a n g e s i n l e a f c h l o r o p h y l l c o n t e n t , an o b s e r v a t i o n i n agreement w i t h work on c o t t o n by Wong (1 9 7 9 ) , but i n c o n s i s t e n t w i t h t h e d a t a of W u l f f and S t r a i n (1982) on Desmodium. The l a t t e r a u t h o r s f o u n d e n r i c h e d p l a n t s had r e d u c e d c h l o r o p h y l l l e v e l s , and t h a t 82 TABLE 4 . 3 . Leaf t h i c k n e s s and i n t e r c e l l u l a r a i r space volume of c o n t r o l (340 u l I"1) and e n r i c h e d (1400 u l 1" 1) p l a n t s . 1 Treatment Leaf Th i ckness A i r Volume (mm) (%) 340 0.30 ±0.01 16.5 ±2.1 u l I" 1 1400 0.31 ±0.01 16.4 ±0.8 u l l " 1 1 Va lues are the means of 3 f i r s t t r i f o l i a t e s and 3 second t r i f o l i a t e s , w i th s tandard e r r o r 83 FIGURE 4.3 Response o f t h e n e t c a r b o n d i o x i d e e xchange rate', e x p r e s s e d p e r u n i t c h l o r o p h y l l , o f t h e f i r s t t r i f o l i a t e of e n r i c h e d (1400 u l 1~ 1) and c o n t r o l (340 u l 1~ 1) p l a n t s , t o p h o t o s y n t h e t i c p h o t o n f l u x d e n s i t y . The d a t a were o b t a i n e d from t h e same p l a n t s t h a t were u s e d i n F i g u r e 2.2 ( s o l i d l i n e s ) . CER was measured a t 1000 u l 1~ 1 c a r b o n d i o x i d e c o n c e n t r a t i o n . The c h l o r o p h y l l c o n t e n t was 0.646±0.180 and 0.647 + 0.240 g n r 2 i n c o n t r o l and e n r i c h e d p l a n t s r e s p e c t i v e l y . E n r i c h e d l e a v e s , c l o s e d c i r c l e s ; c o n t r o l l e a v e s , open c i r c l e s . E a c h v a l u e i s t h e mean of 3 l e a v e s , w i t h s t a n d a r d e r r o r . 8 4 a l t h o u g h c a r b o n d i o x i d e e x c h a n g e c a p a c i t y p e r u n i t l e a f a r e a w a s r e d u c e d b y e n r i c h m e n t t o 1 0 0 0 u l l " 1 c a r b o n d i o x i d e , t h e p h o t o s y n t h e t i c c a p a c i t y w a s h i g h e r o n a u n i t c h l o r o p h y l l b a s i s . T h e y f o u n d e n r i c h e d p l a n t s t o h a v e r e d u c e d c h l o r o p h y l l l e v e l s . A n u m b e r o f s t u d i e s ( s e e H e s k e t h e t a l . , 1 9 8 3 ) h a v e s h o w n t h a t 9 0 % o f m a x i m u m q u a n t u m e f f i c i e n c y w a s a c h i e v e d a t c h l o r o p h y l l c o n c e n t r a t i o n s o f 4 5 0 t o 6 0 0 mg m " 2 . M e a s u r e m e n t s o f p h o t o s y n t h e s i s r a t e p e r u n i t c h l o r o p h y l l i n p l a n t s n o t l i m i t e d b y c h l o r o p h y l l a r e u n d e r e s t i m a t e s a n d s o m e w h a t m i s l e a d i n g . W u l f f a n d S t r a i n ( 1 9 8 2 ) r e p o r t a v e r a g e c h l o r o p h y l l c o n t e n t v a l u e s o f 4 6 6 a n d 9 2 7 mg n r 2 i n e n r i c h e d a n d c o n t r o l p l a n t s r e s p e c t i v e l y . T h e y may h a v e b e e n c o m p a r i n g p h o t o s y n t h e s i s r a t e s a t t w o v e r y d i f f e r e n t c h l o r o p h y l l c o n c e n t r a t i o n s w h e r e o n e ( o r b o t h ) o f t h e c h l o r o p h y l l c o n c e n t r a t i o n s m a y n o t h a v e b e e n l i m i t i n g p h o t o s y n t h e s i s . I f t h i s w a s t h e c a s e , p h o t o s y n t h e s i s r a t e s , o f c o n t r o l p l a n t s i n p a r t i c u l a r , w h e n e x p r e s s e d o n a p e r u n i t c h l o r o p h y l l b a s i s , may h a v e b e e n u n d e r e s t i m a t e d . 4 . 3 . 3 E f f e c t O f D u r a t i o n T h e e a r l i e s t m e a s u r e m e n t s o f C E R o f t h e f i r s t t r i f o l i a t e l e a v e s w e r e m a d e a f t e r a 6 d a y p e r i o d i n t h e t r e a t m e n t c h a m b e r s . A t t h a t t i m e , a r e d u c t i o n i n c a r b o n d i o x i d e e x c h a n g e c a p a c i t y w a s e v i d e n t ( f o r e x a m p l e , r e f e r t o T a b l e 4 . 4 i n s e c t i o n 4 . 3 . 4 ) T h e r e f o r e , a t l e a s t 6 d a y s o f h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n w e r e r e q u i r e d t o e l i c i t a r e d u c t i o n i n c a r b o n d i o x i d e e x c h a n g e c a p a c i t y . A l t h o u g h a r e d u c t i o n i n c a p a c i t y w a s e v i d e n t b e t w e e n 6 a n d 18 d a y s o f e n r i c h m e n t , t h e r e d u c e d n e t c a r b o n d i o x i d e e x c h a n g e 85 c a p a c i t y e x h i b i t e d by t h e f i r s t t r i f o l i a t e of e n r i c h e d p l a n t s was n o t p e r m a n e n t . I f CER was m e asured a f t e r 26 d a y s of t r e a t m e n t , a t a PI o f g r e a t e r t h a n 5 ( P l a s t o c h r o n Index was not an a d e q u a t e measure o f d e v e l o p m e n t a l s t a g e a f t e r t h i s p o i n t b e c a u s e o f e x t e n s i v e b r a n c h i n g and no f u r t h e r g r o w t h o f t h e main s t e m ) , t h e r e was no d i f f e r e n c e i n n e t c a r b o n d i o x i d e exchange c a p a c i t y between t h e f i r s t t r i f o l i a t e s of c o n t r o l and e n r i c h e d p l a n t s ( F i g . 4 . 4 ) . The r e s p o n s e o f CER t o C i was more l i n e a r t h a n a t t h e e a r l i e r s t a g e ( F i g . 4 . 1 ) , and t h e c a r b o n d i o x i d e c o m p e n s a t i o n p o i n t had i n c r e a s e d w i t h age i n b o t h t r e a t m e n t s . T hese r e s u l t s were s i m i l a r t o t h o s e o f H i c k l e n t o n and J o l l i f f e (1981b) who f o u n d no e f f e c t of e n r i c h m e n t on l e a v e s a t a PI of 10 w h i c h p r e v i o u s l y had shown an e f f e c t a t a PI of 5, a l b e i t i n a d i r e c t i o n o p p o s i t e t o t h a t r e p o r t e d h e r e . T h o s e a u t h o r s were u n a b l e t o d e t e r m i n e why t h e e f f e c t o f c a r b o n d i o x i d e d i s a p p e a r e d w i t h t i m e . H o f s t r a and H e s k e t h (1975) f o u n d t h a t e n r i c h e d p l a n t s - r e c o v e r e d f u l l c a r b o n d i o x i d e exchange c a p a c i t y a f t e r 3 d a y s , i f t r a n s f e r r e d t o ambient (340 u l l " 1 ) c a r b o n d i o x i d e c o n c e n t r a t i o n . T h e r e f o r e , t h e e f f e c t of h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n i n r e d u c i n g c a r b o n d i o x i d e exchange c a p a c i t y i s not p e rmanent, and may be a m e l i o r a t e d . 4.3.4 E f f e c t Of P h o t o s y n t h e t i c P h o t o n F l u x D e n s i t y One of t h e most w e l l e s t a b l i s h e d e f f e c t s of e l e v a t e d c a r b o n d i o x i d e c o n c e n t r a t i o n on p l a n t s i s t o i n c r e a s e r a t e s o f p h o t o s y n t h e s i s . The r e d u c e d n e t c a r b o n d i o x i d e exchange c a p a c i t y o f e n r i c h e d p l a n t s may be a f u n c t i o n o f t h e i r g r e a t e r CER d u r i n g g r o w t h . However, t h e CER d u r i n g g r o w t h i s a l s o a 86 i « CM 'E "o E 3. DC Ul o 2 0 0 4 0 0 6 0 0 8 0 0 C i, (ju I I"1) FIGURE 4.4 Response of net carbon d i o x i d e exchange ra te of the f i r s t t r i f o l i a t e of e n r i c h e d (1400 u l l ' 1 ) and c o n t r o l (340 u l l ' 1 ) p l a n t s (P l>5.0) to i n t e r c e l l u l a r space carbon d i o x i d e c o n c e n t r a t i o n . Measurement PPFD was 1000 umol m~2 s _ 1 . P l an t s were in the t reatment chambers fo r 26 days . The PPFD i n c i d e n t on the f i r s t t r i f o l i a t e d u r i n g the l a t t e r pa r t of t h i s p e r i o d was 100 umol m - 2 . - i C o n t r o l p l a n t s , open c i r c l e s ; e n r i c h e d p l a n t s , c l o s e d c i r c l e s . Measurements were ob t a i ned from 3 p l a n t s in each t r ea tment . Curves were f i t t e d to an asymptot i c equa t ion us i ng 8 and 9 va lues fo r c o n t r o l and e n r i c h e d t rea tments r e s p e c t i v e l y . 87 f u n c t i o n of PPFD. At low PPFD, the a b s o l u t e i n c r e a s e i n CER because of high carbon d i o x i d e c o n c e n t r a t i o n was much l e s s than at higher PPFD ( F i g . 4.2), although the percentage increase may be s i m i l a r . I f the reduced carbon d i o x i d e exchange c a p a c i t y was dependent on a higher CER i n e n r i c h e d p l a n t s , then under c o n d i t i o n s of PPFD l i m i t a t i o n of CER d u r i n g growth, where carbon d i o x i d e was l e s s e f f e c t i v e i n i n c r e a s i n g CER, there should a l s o be minimal d i f f e r e n c e i n net carbon d i o x i d e exchange c a p a c i t y between e n r i c h e d and c o n t r o l p l a n t s . Low l i g h t c o n d i t i o n s i n the canopy may i n f a c t e x p l a i n the disappearance through time of any d i f f e r e n c e i n net carbon d i o x i d e exchange c a p a c i t y between e n r i c h e d and c o n t r o l p l a n t s . P l a n t s at 26 days of treatment ( t o t a l of 37 days old) had a well-developed canopy, r e s u l t i n g i n e x t e n s i v e shading of lower leaves (<100 umol m"2 s" 1 PPFD). To t e s t the e f f e c t of l i g h t , i n t e n s i t y on the carbon d i o x i d e - i n d u c e d r e d u c t i o n of net carbon d i o x i d e exchange c a p a c i t y , p l a n t s were • t r a n s f e r r e d to treatment chambers a f t e r the f i r s t t r i f o l i a t e was f u l l y developed. At t h i s p o i n t some of the l e a f l e t s of the f i r s t t r i f o l i a t e were shaded with t r a c i n g paper, while the remainder were maintained at the higher PPFD. As seen in Table 4.4, there was minimal e f f e c t of carbon d i o x i d e enrichment on net carbon d i o x i d e exchange c a p a c i t y at 100 umol n r 2 s" 1 PPFD, even though comparable l e a f l e t s at 350 umol n r 2  s - i PPFD showed a 22% r e d u c t i o n . The a b s o l u t e net carbon d i o x i d e exchange r a t e s of low PPFD leaves were somewhat lower than those at high PPFD, and were probably due to a c c l i m a t i o n of those leaves to low PPFD. Such a c c l i m a t i o n i s t y p i c a l l y 88 TABLE 4 . 4 . The e f f e c t of shad ing du r i ng growth on the net carbon d i o x i d e exchange ra te of the f i r s t t r i f o l i a t e of e n r i c h e d (1400 u l 1~1) and c o n t r o l (340 u l 1 _ 1 ) beans grown at 26 C . 1 Treatment CER 2 (umol m~2 s " 1 ) %CER 3 C i f t (u l I ' 1) R e s p i r a t i o n 5 (umol m" 2 s " 1 ) 340 u l 1 " \ shaded 6 11.5 ±0.4 295 ±8 0.80 ±0.02 1 400 u l I' 1 , shaded 6 1 1 .6 ±0.8 ( + ) 1 302 ±3 0.80 ±0.02 340 u l I" 1 , unshaded 7 18.1 ±0.6 278 ±7 0.90 ±0.04 1 400 u l I" 1 , unshaded 7 14.1 ±0.6 22 289 ±3 1.30 ±0.02 Va lues are means of 3 l e a v e s , w i th s tandard e r r o r ; a l l measurements were taken a f t e r 6 days of t reatment CER measured at 1000 umol m" 2 s " 1 PPFD D i f f e r e n c e in CER between c o n t r o l and e n r i c h e d p l a n t s , measured at the same PPFD, and expressed as a percentage of the c o n t r o l v a l u e . C i r e f e r s to the CER measurements o n l y . R e s p i r a t i o n measured at 21% oxygen and 340 u l l " 1 carbon d i o x i d e c o n c e n t r a t i o n . 100 umol m" 2 s " 1 PPFD d u r i n g growth 350 umol nr 2 s " 1 PPFD d u r i n g growth 89 c h a r a c t e r i z e d by a g r e a t e r e f f i c i e n c y o f p h o t o s y n t h e s i s measured under low l i g h t c o n d i t i o n s , b u t w i t h r e d u c e d r a t e s under h i g h l i g h t (Boardman, 1977). When PPFD l i m i t e d CER d u r i n g g r o w t h , t h e e f f e c t of c a r b o n d i o x i d e e n r i c h m e n t was n e g l i g i b l e , s u g g e s t i n g t h a t t h e r e d u c t i o n i n n e t c a r b o n d i o x i d e exchange c a p a c i t y was r e l a t e d t o t h e e n h a n c e d CER o f e n r i c h e d p l a n t s . F u r t h e r m o r e , t h e d i s a p p e a r a n c e o f any e f f e c t of e n r i c h m e n t on t h e c a r b o n d i o x i d e exchange c a p a c i t y i n t h e f i r s t t r i f o l i a t e o f o l d e r l e a v e s was, a t l e a s t i n p a r t , e x p l a i n e d by s h a d i n g of t h e t r i f o l i a t e by t h e l e a v e s a b o v e . I t was a l s o p o s s i b l e t h a t s i n c e CER i s known t o d e c r e a s e w i t h age ( D a v i s and McCree, 1978), t h e lo w e r CER i n t h e o l d e r l e a v e s may a g a i n l i m i t any e f f e c t of e n r i c h m e n t . However, t h i s p o s s i b i l i t y was not i n v e s t i g a t e d . 4.3.5 Response To S i n k M a n i p u l a t i o n Reduced n e t c a r b o n d i o x i d e e xchange c a p a c i t y i n e n r i c h e d l e a v e s was n o t due t o d i f f e r e n c e s i n c h l o r o p h y l l c o n t e n t nor t o a l t e r e d r a t e s of p h o t o r e s p i r a t i o n , b o t h of w h i c h a r e i n t r i n s i c p r o p e r t i e s o f l e a v e s w h i c h i n f l u e n c e CER i n a d i r e c t and p r o p o r t i o n a l manner. Carbon d i o x i d e e n r i c h m e n t c o u l d i n f l u e n c e n e t c a r b o n d i o x i d e exchange c a p a c i t y by t h e somewhat more i n d i r e c t means o f a l t e r i n g t h e s o u r c e - s i n k r e l a t i o n s h i p s of t h e p l a n t , i n w h i c h c a s e , e n r i c h m e n t would n o t s i m p l y be a f f e c t i n g t h e l e a v e s a s i s o l a t e d u n i t s s e p a r a t e f r o m e a c h o t h e r , o r from t h e r e s t of t h e p l a n t . A s i n k i s d e f i n e d a s a p l a n t p a r t w h i c h u n d e r g o e s a n e t 90 i n p u t of c a r b o n from t h e r e s t o f t h e p l a n t ( Z e e v a a r t , 1979). C o n v e r s e l y , a s o u r c e i s a p l a n t p a r t showing a n e t e x p o r t of c a r b o n t o t h e r e m a i n d e r o f t h e p l a n t . S o u r c e or s i n k s t r e n g t h i s d e f i n e d as t h e p r o d u c t o f t h e s i z e and t h e a c t i v i t y o f t h e s o u r c e o r s i n k . The s o u r c e o r s i n k s t r e n g t h may change i n r e s p o n s e t o a change i n t h e demand f o r , or s u p p l y o f , a s s i m i l a t e s . T h e r e i s now a s u b s t a n t i a l body o f e v i d e n c e s u g g e s t i n g t h a t s i n k s t r e n g t h i n f l u e n c e s s o u r c e l e a f n et p h o t o s y t h e s i s r a t e s ( Z e e v a a r t , 1979; G i f f o r d and E v a n s , 1981). However, t h e e v i d e n c e f o r a mechanism by w h i c h t h e r e g u l a t i o n o c c u r s r e m a i n s c o n t r o v e r s i a l ( N e a l e s and I n c o l l , 1968). H i g h s t a r c h l e v e l s , w h i c h a r e r e l a t e d t o r e d u c e d p h o t o s y n t h e s i s ( N a f z i g e r and R o l l e r , 1976), have been p r o p o s e d as a means by w h i c h s i n k demand i n f l u e n c e s s o u r c e l e a f p h o t o s y n t h e s i s . S t a r c h s y n t h e s i s i n s o u r c e l e a v e s a p p e a r s t o be r e s p o n s i v e t o s i n k demand. S u c r o s e p h o s p h a t e s y n t h e t a s e , an enzyme i n v o l v e d i n s u c r o s e m e t a b o l i s m , may be an i m p o r t a n t c o n t r o l p o i n t (Huber and I s r e a l , 1982). F o r example, when s i n k demand f o r s u c r o s e was l i m i t e d by e x c i s i o n of t h e s o u r c e l e a f , s t a r c h a c c u m u l a t i o n i n c r e a s e d and s u c r o s e p h o s p h a t e s y n t h e t a s e a c t i v i t y f e l l i n t h e s o u r c e l e a f ; c o n v e r s e l y when s i n k demand was i n c r e a s e d by d e f o l i a t i o n of o t h e r s o u r c e l e a v e s , s t a r c h a c c u m u l a t i o n d e c r e a s e d w i t h a c o n c u r r e n t i n c r e a s e i n s u c r o s e p h o s p h a t e s y n t h e t a s e a c t i v i t y ( R u f t y and Huber, 1983). S t a r c h and s u c r o s e l e v e l s have been shown t o be i n v e r s e l y r e l a t e d and r e s p o n s i v e t o s i n k demand i n soybean ( T h o r n e and 91 R o l l e r , 1974), where a d i r e c t c o r r e l a t i o n was o b s e r v e d between s t a r c h c o n c e n t r a t i o n and m e s o p h y l l r e s i s t a n c e . I n t e r e s t i n g l y , a r e d u c e d CER a s s o c i a t e d w i t h h i g h s t a r c h c o n t e n t has been shown t o o c c u r i n w i d e l y d i v e r s e p h y s i o l o g i c a l s i t u a t i o n s , s u c h as a r e s u l t o f f l o o d i n g of t h e r o o t s of s u n f l o w e r (Wample and D a v i s , 1983), and i n n o n - t i l l e r i n g P a n g o l a a t c o o l n i g h t t e m p e r a t u r e s ( C h a t t e r t o n e t a l . , 1972). A z c o n - B i e t o (1983) a l s o f o u n d a d e c l i n e i n p h o t o s y n t h e s i s r a t e w h i c h . c o r r e l a t e d w i t h 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 wheat l e a v e s . E i t h e r i n c r e a s i n g t h e c a r b o n d i o x i d e c o n c e n t r a t i o n , o r r e d u c i n g t h e r a t e of t r a n s l o c a t i o n by c h i l l i n g t h e base of t h e s o u r c e l e a f , i n c r e a s e d t h e r a t e o f d e c l i n e i n p h o t o s y n t h e s i s when measured o v e r a 6 t o 8 hour p e r i o d . In v i e w of t h e a p p a r e n t r e l a t i o n s h i p between c a r b o h y d r a t e l e v e l and n e t c a r b o n d i o x i d e exchange c a p a c i t y , b o t h s t a r c h c o n c e n t r a t i o n and CER were d e t e r m i n e d i n e n r i c h e d and u n e n r i c h e d p l a n t s . E n r i c h m e n t t o 1400 u l l " 1 or 3000 u l l " 1 c a r b o n d i o x i d e r e s u l t e d i n a 21% and 18% d e c r e a s e i n n e t c a r b o n d i o x i d e exchange c a p a c i t y and a 4 . 4 - f o l d and 3 - f o l d i n c r e a s e i n s t a r c h c o n c e n t r a t i o n , r e s p e c t i v e l y , compared t o c o n t r o l p l a n t s ( T a b l e 4.5, p r e - s h a d e d v a l u e s ) . T h i s o b s e r v a t i o n a g r e e d w i t h p r e v i o u s s t u d i e s s h owing an i n c r e a s e i n s t a r c h c o n t e n t o f e n r i c h e d s o y b e a n s ( H o f s t r a and H e s k e t h , 1975; N a f z i g e r and R o l l e r , 1976) and c o t t o n (Mauney e t a l . , 1979) w h i c h c o r r e l a t e d w i t h r e d u c e d p h o t o s y n t h e s i s r a t e s . However, Fondy and G e i g e r (1982) f o u n d t h a t a l t h o u g h s t a r c h i n c r e a s e d from 20 t o 140 ug cm" 2 o v e r t h e c o u r s e of a 14 hour p h o t o p e r i o d , net c a r b o n d i o x i d e e xchange was 92 TABLE 4 . 5 . The e f f e c t of shading a l l but the f i r s t t r i f o l i a t e on s t a r c h conten t and net carbon d i o x i d e exchange ra te of the f i r s t t r i f o l i a t e of c o n t r o l (340 u l l " 1 ) and e n r i c h e d (1400 and 3000 u l l ' 1 ) p l a n t s . 1 Treatment [ S ta rch ] CER 1 %CER 2 (g m" 2 ) (umol m" 2 s " 1 ) Be fo re s h a d i n g 3 : 340 u l I" 1 1400 u l I'1 3000 u l I"1 7.26±0.68 31.92±3.88 21.60±2.21 31.1+1.3 24.6+0.5 25.6±1.6 A f t e r s e l e c t i v e s h a d i n g ' • 340 u l I"1 shaded 5 340 u l 1- 1 unshaded 6 1400 u l 1-1 shaded 1400 u l I" 1 unshaded 3000 u l I"1 shaded 3000 u l I" 1 unshaded 3.77±1.03 33.2±0.9 8.10±1.46 23 .7±1.5 26.88±1.97 24 .1±2.1 27.9612.20 19.310.3 28.5612.71 23 .511 .3 31.1213.01 20 .511 .3 29 20 13 CER measured at 1000 umol m" 2 s " 1 PPFD and 1000 u l l " 1 carbon d i o x i d e D i f f e r e n c e in CER between shaded and unshaded t rea tments w i t h i n each carbon d i o x i d e t r ea tmen t , expressed as a percentage of c o n t r o l v a l ues P re-shad ing va lues were taken a f t e r 8 days of carbon d i o x i d e t r ea tmen t ; v a l ues are the means of 8 p l a n t s , w i th s tandard e r r o r Measurements were made a f t e r 5 days of s e l e c t i v e s h a d i n g ; va lues a re the means of 4 p l a n t s , w i th s t anda rd e r r o r shaded l eaves ma in t a i ned at 90110 umol n r 2 s " 1 PPFD unshaded l eaves ma in t a ined at 350120 umol n r 2 s " 1 PPFD 93 not a l t e r e d . I t c a n be a r g u e d , however, t h a t s t a r c h r e d u c e s n e t c a r b o n d i o x i d e e xchange c a p a c i t y o n l y i f a c c u m u l a t i o n r e a c h e s some c r i t i c a l l e v e l , a c h i e v e d i n e n r i c h e d p l a n t s but not i n c o n t r o l p l a n t s i n my s t u d y . T h i s i s s u p p o r t e d by t h e f a c t t h a t c o t t o n showed a n e g a t i v e c o r r e l a t i o n between p h o t o s y n t h e s i s and s t a r c h c o n t e n t , but s o y b e a n , s u n f l o w e r , and sorghum, w h i c h d i d n o t a c c u m u l a t e as much s t a r c h , d i d not show any c o r r e l a t i o n (Mauney e t a l . , 1979). The r e d u c t i o n i n c a r b o n d i o x i d e e xchange c a p a c i t y i n e n r i c h e d p l a n t s i n my s t u d y , may have been a s s o c i a t e d w i t h t h e c a r b o h y d r a t e s t a t u s of t h e l e a v e s . The f a c t t h a t t h e d i f f e r e n c e i n c a p a c i t y o c c u r r e d a t h i g h , but not low PPFD, c o u p l e d w i t h t h e e s t a b l i s h e d i n f l u e n c e of s i n k demand on c a r b o h y d r a t e c o n t e n t and p h o t o s y n t h e s i s r a t e of s o u r c e l e a v e s , • would s u g g e s t t h a t t h e r e d u c t i o n i n n e t c a r b o n d i o x i d e e xchange c a p a c i t y of e n r i c h e d p l a n t s was i n f l u e n c e d by t h e r e l a t i v e r a t e s of c a r b o n a s s i m i l a t i o n and c a r b o n e x p o r t . The r a t e of t r a n s l o c a t i o n (Ho, 1977; P o t t e r , 1980), as w e l l as t h e e f f i c i e n c y o f t r a n s l o c a t i o n (Ho, 1977) have been shown t o i n c r e a s e i n e n r i c h e d p l a n t s . However, i f s i n k demand was i n f l u e n c i n g , and p e r h a p s l i m i t i n g , t h e r a t e of t r a n s l o c a t i o n f r o m e n r i c h e d s o u r c e l e a v e s i n my s t u d y , t h e g r e a t e r CER a t h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n s may have c o n t r i b u t e d t o r e d u c e d n e t c a r b o n d i o x i d e exchange c a p a c i t y . To t e s t t h e i n f l u e n c e o f s i n k demand on t h e c a r b o n d i o x i d e e x change c a p a c i t y of e n r i c h e d and c o n t r o l p l a n t s , p l a n t s were f i r s t grown f o r 9 d a y s i n one of t h r e e c a r b o n d i o x i d e 94 c o n c e n t r a t i o n s t o e s t a b l i s h d i f f e r e n c e s i n s t a r c h c o n t e n t and n e t c a r b o n d i o x i d e exchange c a p a c i t y i n t h e f i r s t t r i f o l i a t e l e a v e s ( T a b l e 4.5, b e f o r e s h a d i n g ) . A l l l e a v e s but t h e f i r s t t r i f o l i a t e were t h e n shaded w i t h opaque p a p e r t o i n c r e a s e s i n k demand on t h e u n s h a d e d l e a f ; o t h e r p l a n t s were l e f t u n s h a d e d . Low s i n k demand (unshaded p l a n t s ) r e s u l t e d i n d e c r e a s e d n e t c a r b o n d i o x i d e exchange c a p a c i t y r e l a t i v e t o h i g h s i n k demand i n a l l c a r b o n d i o x i d e t r e a t m e n t s , w i t h a r e d u c t i o n of 29, 20, and 13% i n 340, 1400, and 3000 u l l " 1 t r e a t e d p l a n t s r e s p e c t i v e l y ( T a b l e 4.5, a f t e r s e l e c t i v e s h a d i n g ) . C a r b o n d i o x i d e e n r i c h m e n t a l s o r e s u l t e d i n l o w e r c a r b o n d i o x i d e exchange c a p a c i t y . Those t r e n d s were a l s o f o u n d by C l o u g h e t a l . (1981) who m a n i p u l a t e d f r u i t number i n s o y b e a n s t o c r e a t e h i g h or low s i n k p l a n t s , a t two c a r b o n d i o x i d e c o n c e n t r a t i o n s . T h e i r e x p e r i m e n t d i f f e r e d f rom t h a t r e p o r t e d h e r e i n t h a t c a r b o n d i o x i d e and s i n k demand t r e a t m e n t s were i n i t i a t e d s i m u l t a n e o u s l y , and were not i n t e n d e d t o e v a l u a t e t h e r e s p o n s e of e n r i c h e d p l a n t s t o a change i n s i n k s t r e n g t h . F u r t h e r m o r e , t h e i r measurements of p h o t o s y n t h e s i s were b a s e d on t h e whole p l a n t . In my s t u d y , t h e r a t e of change i n net c a r b o n d i o x i d e e xchange c a p a c i t y o v e r t i m e i n u n s h a d e d p l a n t s was s i m i l a r i n a l l c a r b o n d i o x i d e t r e a t m e n t s ( d e c r e a s e s of 24, 22, and 20% i n 340, 1400, and 3000 u l 1~ 1 t r e a t m e n t s r e s p e c t i v e l y ) ( T a b l e 4.5), and t h e r e f o r e t h e r e s u l t s were n o t c o n f o u n d e d by a g e - r e l a t e d d i f f e r e n c e s i n n e t c a r b o n d i o x i d e exchange c a p a c i t y among t r e a t m e n t s . I n c r e a s i n g s i n k demand d e c r e a s e d t h e a g e - r e l a t e d d e c l i n e i n 95 c a r b o n d i o x i d e e xchange c a p a c i t y of t h e f i r s t t r i f o l i a t e , and i n t h i s r e s p e c t , e n r i c h e d p l a n t s d i d not r e s p o n d t o s h a d i n g as w e l l as c o n t r o l s . C o m p a r i n g c a r b o n d i o x i d e exchange c a p a c i t y w i t h i n s h a d e d ( h i g h demand) t r e a t m e n t s , t h e p e r c e n t r e d u c t i o n i n e n r i c h e d p l a n t s compared t o c o n t r o l s was even g r e a t e r t h a n t h a t b e f o r e s h a d i n g ( f o r example, i n c o m p a r i n g 3000 w i t h 340 u l l " 1 p l a n t s , t h e r e was an 18% r e d u c t i o n i n c a r b o n d i o x i d e e xchange c a p a c i t y b e f o r e s h a d i n g , but a 29% r e d u c t i o n a f t e r s h a d i n g ) . S i m i l a r r e s u l t s were o b t a i n e d when c o m p a r i n g t h e e f f e c t o f s h a d i n g on t h e c a p a c i t y w i t h i n c a r b o n d i o x i d e t r e a t m e n t s ; t h e r e was p r o g r e s s i v e l y l e s s d i f f e r e n c e i n c a r b o n d i o x i d e e x c h a n g e c a p a c i t y between shaded and u n s h a d e d p l a n t s w i t h i n c r e a s i n g c a r b o n d i o x i d e c o n c e n t r a t i o n t r e a t m e n t s ( T a b l e 4 . 5 ) . T h e r e f o r e , an i n c r e a s e i n s i n k demand d i d not a l l e v i a t e t h e r e d u c t i o n i n n e t c a r b o n d i o x i d e e xchange c a p a c i t y of e n r i c h e d compared t o c o n t r o l p l a n t s . S t a r c h c o n t e n t of t h e f i r s t t r i f o l i a t e was a l s o d e t e r m i n e d i n r e l a t i o n t o c a r b o n d i o x i d e e xchange c a p a c i t y . U nshaded c o n t r o l p l a n t s had v i r t u a l l y t h e same n e t c a r b o n d i o x i d e e x c h a n g e c a p a c i t y as s haded 3000 u l 1~ 1 p l a n t s , d e s p i t e a 3 - f o l d d i f f e r e n c e i n s t a r c h c o n t e n t , and a l s o showed a 24% r e d u c t i o n i n c a r b o n d i o x i d e e xchange c a p a c i t y compared t o i n i t i a l r a t e s , even t h o u g h s t a r c h c o n t e n t was u n a l t e r e d ( T a b l e 4 . 5 ) . T h e r e f o r e , c h a n g e s i n n e t c a r b o n d i o x i d e exchange c a p a c i t y b e c a u s e o f s h a d i n g ( s i n k demand) were i n d e p e n d e n t of s t a r c h c o n c e n t r a t i o n . T h i s o b s e r v a t i o n s u p p o r t s t h e work o f P o t t e r and B r e e n (1980) who f o u n d t h a t a l t h o u g h c o n t i n u o u s l i g h t f o r 52 h o u r s r e s u l t e d 96 i n s t a r c h a c c u m u l a t i o n t o 3.3 mg c m - 2 and r e d u c e d p h o t o s y n t h e t i c r a t e s i n r a p i d l y e x p a n d i n g s u n f l o w e r l e a v e s , o t h e r l e a v e s w h i c h were mature and s l o w l y e x p a n d i n g a c c u m u l a t e d l e s s s t a r c h but showed g r e a t e r p h o t o s y n t h e t i c r e d u c t i o n . Note t h a t b e c a u s e t h e l e a v e s were o f d i f f e r e n t a g e s , t h e i r s e n s i t i v i t y t o s t a r c h may have been d i f f e r e n t , as s u g g e s t e d by N a f z i g e r and R o l l e r ( 1 9 7 6 ) . However, as shown i n F i g u r e 4.5, when s t a r c h c o n t e n t was p l o t t e d v e r s u s CER f o r a l l t r e a t m e n t s , e n r i c h e d p l a n t s g e n e r a l l y had h i g h e r s t a r c h c o n t e n t and lo w e r n e t c a r b o n d i o x i d e exchange c a p a c i t y , a l t h o u g h t h e d a t a were q u i t e s c a t t e r e d . I t i s p o s s i b l e t h a t e n r i c h m e n t a f f e c t e d n e t c a r b o n d i o x i d e exchange c a p a c i t y t h r o u g h some a s p e c t o f s t a r c h a c c u m u l a t i o n w h i c h s e t a l o w e r maximum c a r b o n d i o x i d e exchange p o t e n t i a l . O t h e r f a c t o r s ( e g . s h a d i n g o r a g i n g ) may have t h e n m o d i f i e d t h e c a p a c i t y w i t h i n t h a t l i m i t , i n d e p e n d e n t l y of s t a r c h c o n c e n t r a t i o n . I t may be t h a t s t a r c h a c c u m u l a t i o n p e r se was n o t c a u s i n g t h e d e c l i n e i n n e t c a r b o n d i o x i d e exchange c a p a c i t y but was r a t h e r a r e f l e c t i o n of a complex m e t a b o l i c a d j u s t m e n t , some o t h e r a s p e c t o f w h i c h c o u l d have c a u s e d t h e r e d u c e d c a r b o n d i o x i d e e x c h a n g e c a p a c i t y . F o r example, T h o r n e and R o l l e r (1974) f o u n d t h a t a t h i g h s t a r c h l e v e l s , RuBP c a r b o x y l a s e a c t i v i t y was r e d u c e d . In a d d i t i o n , t h e r e i s c u r r e n t l y a g r e a t d e a l of i n t e r e s t i n t h e p o s s i b i l i t y t h a t r e g u l a t i o n of n e t p h o t o s y n t h e s i s r a t e may r e s i d e a t t h e ' l e v e l of c a r b o h y d r a t e t r a n s p o r t f r o m c h l o r o p l a s t s . T h i s p o s s i b i l i t y has been d i s c u s s e d w i t h p a r t i c u l a r r e f e r e n c e t o s i n k c o n t r o l o f s o u r c e l e a f p h o t o s y n t h e s i s r a t e s ( H e r o l d , 1980; Fondy and G e i g e r , 97 CM I O E a. tr o STA RCH g m -2 FIGURE 4.5 R e l a t i o n s h i p between net carbon d i o x i d e exchange r a te and s t a r c h content of the f i r s t t r i f o l i a t e of e n r i c h e d (1400 or 3000 u l l ' 1 ) and c o n t r o l (340 u l 1" 1) p l a n t s . The da ta are based on the measurements g iven in Tab le 4 . 5 . The curve was f i t t e d to a t h i r d - o r d e r po l ynomia l e q u a t i o n , u s i ng 40 v a l u e s . R 2 =0.46 98 1981). S u c r o s e i s t h e major m e t a b o l i t e t r a n s p o r t e d t h r o u g h t h e p l a n t , and t h e p r o p o s e d r e g u l a t i o n o f p h o t o s y n t h e s i s i n v o l v e s , a t t h e o u t s e t , s u c r o s e m e t a b o l i s m . S u c r o s e i s f i r s t s y n t h e s i z e d i n t h e c y t o p l a s m o f m e s o p h y l l c e l l s b e f o r e i t i s l o a d e d i n t o t h e phloem f o r e x p o r t . The s y n t h e s i s of s u c r o s e i n v o l v e s f i r s t t h e c o n v e r s i o n of t r i o s e p h o s p h a t e s , w h i c h a r e e x p o r t e d from t h e c h l o r o p l a s t , t o f r u c t o s e - 1 , 6 - b i s p h o s p h a t e . A f t e r d e p h o s p h o r y l a t i o n t o f r u c t o s e - 6 - p h o s p h a t e , and some c o n v e r s i o n t o U D P - g l u c o s e , t h e p r o d u c t s a r e t h e n c o n v e r t e d by s u c r o s e p h o s p h a t e s y n t h e t a s e t o s u c r o s e p h o s p h a t e , w h i c h i s t h e n c o n v e r t e d t o s u c r o s e by s u c r o s e p h o s p h a t e p h o s p h a t a s e . I f h i g h c y t o p l a s m i c s u c r o s e c o n c e n t r a t i o n s o c c u r r e d , f o r example, b e c a u s e of d e c r e a s e d l o a d i n g due t o low s i n k demand, t h e a c c u m u l a t i o n of s u c r o s e would l i m i t f u r t h e r s u c r o s e s y n t h e s i s by f e e d b a c k - i n h i b i t i o n of s u c r o s e p h o s p h a t e s y n t h e t a s e ( H e r o l d , 1980; Fondy and G e i g e r , 1981). A l t e r n a t i v e l y , h i g h s u c r o s e l e v e l s c o u l d 1-imit s u c r o s e p h o s p h a t e p h o s p h a t a s e a c t i v i t y ( H e r o l d , 1980). In e i t h e r c a s e , p h o s p h o r y l a t e d compounds would i n c r e a s e i n t h e c y t o p l a s m , and p h o s p h a t e ( P i ) c o n c e n t r a t i o n w ould d e c r e a s e . T h i s would f a v o u r r e t e n t i o n of t r i o s e p h o s p h a t e s i n t h e c h l o r o p l a s t s i n c e P i i s r e q u i r e d f o r exchange of t r i o s e p h o s p h a t e s a c r o s s t h e c h l o r o p l a s t i n n e r membrane. A D P - g l u c o s e p y r o p h o s p h o r y l a s e , an enzyme of s t a r c h s y n t h e s i s , i s s t i m u l a t e d by a h i g h t r i o s e p h o s p h a t e / P i r a t i o ( s e e Fondy and G e i g e r , 1981), and t r i o s e p h o s p h a t e s would t h u s be r e d i r e c t e d i n t o s t a r c h s y n t h e s i s . T r i o s e p h o s p h a t e s c o u l d c o n c e i v a b l y i n h i b i t p h o t o s y n t h e s i s 99 t h r o u g h a mass a c t i o n e f f e c t . A l t e r n a t i v e l y , low P i c o n c e n t r a t i o n s i n t h e c h l o r o p l a s t c o u l d r e d u c e n e t p h o t o s y n t h e s i s , p o s s i b l y t h r o u g h a r e d u c e d ATP/ADP r a t i o ( R o b i n s o n and W a l k e r , 1979) or t h r o u g h a d i r e c t e f f e c t on RuBP c a r b o x y l a s e a c t i v i t y ( H e l d t e t a l . , 1978). Low l e v e l s of P i i n l e a v e s have, i n f a c t , been c o r r e l a t e d w i t h r e d u c e d r a t e s o f n e t p h o t o s y n t h e s i s ( T e r r y and U l r i c h , 1973) and h i g h l e v e l s of l e a f s t a r c h ( J e n s e n , 1980). Low l e v e l s of P i would n i c e l y e x p l a i n a d e c r e a s e i n c a r b o n d i o x i d e e xchange c a p a c i t y w i t h i n c r e a s e d l e a 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 . As a c o r o l l o r y t o i n c r e a s i n g s i n k demand, an a d d i t i o n a l e x p e r i m e n t was d e s i g n e d t o i n v e s t i g a t e t h e e f f e c t o f a d e c r e a s e i n s i n k demand on t h e c a r b o n d i o x i d e - i n d u c e d r e d u c t i o n i n c a r b o n d i o x i d e e x c h a n g e c a p a c i t y . Demand f o r a s s i m i l a t e s was m a n i p u l a t e d by r e s t r i c t i n g p l a n t s i z e . P l a n t s were grown i n p o t s of two s i z e s , w i t h t h o s e i n t h e s m a l l e r p o t s g r o w i n g l e s s r a p i d l y b e c a u s e of r e s t r i c t e d r o o t d e v e l o p m e n t , and p r e s u m a b l y h a v i n g r e d u c e d s i n k demand. The d i f f e r e n c e i n n e t c a r b o n d i o x i d e e x c h a n g e c a p a c i t y between e n r i c h e d and c o n t r o l p l a n t s i n 3 i n c h p o t s , measured on an a r e a b a s i s a f t e r b o t h 11 and 18 d a y s , was s m a l l e r t h a n i n 5 i n c h p o t s ( T a b l e 4 . 6 ) . Hence, e n r i c h e d and c o n t r o l p l a n t s a p p r o a c h e d t h e same n e t c a r b o n d i o x i d e e x c h a n g e c a p a c i t y when g r o w t h was s e v e r e l y r e s t r i c t e d . The s i m i l a r i t y of exchange c a p a c i t y was a r e s u l t of e i t h e r t h e r e b e i n g no c a r b o n d i o x i d e e n r i c h m e n t e f f e c t , or b e c a u s e c o n t r o l p l a n t s were s u p p r e s s e d t o 100 TABLE 4 . 6 . E f f e c t of pot s i z e on the net carbon d i o x i d e exchange ra te of the f i r s t t r i f o l i a t e of e n r i c h e d (1400 u l l " 1 ) and c o n t r o l (340 u l l ' 1 ) p l a n t s . 1 Treatment CER 2 %CER 3 C h l o r o p h y l l • CER* %CER 5 PI (umol nr 2 (g m~2 ) (umol g " 1 s- 1 ) c h l . s " 1 ) 340 18.4±1.6 - 0.71010.025 2.59 - 4.5 u l I" 1 , 5" Pot 1400 12.111.1 44 0.71010.060 1.70 39 4.5 u l 1 " \ 5" Pot 340 11.710.6 - 0.38210.043 3.06 - 2.5 u l I" 1 , 3" Pot 1400 10.211.2 12 0.49010.041 2.09 32 2.5 u l 1 " \ 3" Pot 340 17.711.3 - 0.58210.018 3.05 - 4.5 u l I'1, 3" Pot 1400 15.311.1 14 0.58810.056 2.61 14 4.5 u l I'1, 3" Pot 1 Va lues are the means of 3 p l a n t s , w i th s tandard e r r o r ; a l l measurements were taken a f t e r 11 days of t r ea tment , except f o r the l a s t two rows, which were a f t e r 18 days 2 CER measured at 1000 umol n r 2 s _ 1 PPFD and 340 u l 1 _ 1 carbon d i o x i d e ; CER expressed on a u n i t a rea b a s i s 3 D i f f e r e n c e i n CER between c o n t r o l and e n r i c h e d t rea tments at the same pot s i z e and P l a s t o c h r o n Index, and on a per u n i t a rea b a s i s , expressed as a percentage of c o n t r o l v a l ue s 4 CER measured at 1000 umol n r 2 s _ 1 PPFD and 340 u l 1 _ 1 carbon d i o x i d e ; CER expressed on a u n i t c h l o r o p h y l l b a s i s s D i f f e r e n c e i n CER between c o n t r o l and e n r i c h e d t rea tments at the same pot s i z e and P l a s t o c h r o n Index, and on a per u n i t c h l o r o p h y l l b a s i s , exp ressed as a percentage of c o n t r o l v a l ues 101 t h e same e x t e n t a s e n r i c h e d p l a n t s . - J u d g i n g f r o m t h e s i m i l a r i t y of c a p a c i t y o f c o n t r o l p l a n t s i n 5 and 3 i n c h p o t s a t t h e same P l a s t o c h r o n Index, when e x p r e s s e d on a u n i t l e a f a r e a b a s i s , i t seems t h a t p o t s i z e d i d n o t s e r v e t o r e d u c e c a p a c i t y , a t l e a s t i n t h e f i r s t t r i f o l i a t e . At 11 d a y s , c h l o r o p h y l l l e v e l s of p l a n t s i n t h e s m a l l e r p o t s were r e d u c e d compared t o t h o s e i n l a r g e p o t s ( T a b l e 4 . 6 ) , and t h e n e t c a r b o n d i o x i d e e xchange c a p a c i t y p e r u n i t c h l o r o p h y l l was somewhat h i g h e r . R e f e r r i n g t o t h e argument p r e s e n t e d e a r l i e r , i t may be t h a t c o m p a r i n g two q u i t e d i f f e r e n t c h l o r o p h y l l l e v e l s , one o r more of w h i c h may be n o n - l i m i t i n g , g i v e s an u n d e r e s t i m a t e of CER, p a r t i c u l a r l y a t t h e h i g h e r c h l o r o p h y l l l e v e l ( c o n t r o l , 5 i n c h p o t ) . A l t e r n a t i v e l y , i t may be t h a t n e t c a r b o n d i o x i d e e xchange c a p a c i t y on a p e r u n i t c h l o r o p h y l l b a s i s was i n d e e d .somewhat i n c r e a s e d i n s m a l l p o t s p r e c i s e l y b e c a u s e of t h e d e c r e a s e i n c h l o r o p h y l l l e v e l . When p l a n t g r o w t h and s i n k demand were r e s t r i c t e d , h i g h e r c h l o r o p h y l l l e v e l s would p o s s i b l y r e d u c e n e t c a r b o n d i o x i d e exchange c a p a c i t y . A t low c h l o r o p h y l l l e v e l s , a b s o l u t e r a t e s of c a r b o n d i o x i d e exchange were c o r r e s p o n d i n g l y lower and so a r e d u c t i o n i n n e t c a r b o n d i o x i d e exchange c a p a c i t y was l e s s l i k e l y t o o c c u r . T h i s p o s s i b i l i t y was s u p p o r t e d by t h e f a c t t h a t e n r i c h e d and c o n t r o l p l a n t s a f t e r 18 d a y s i n s m a l l p o t s , and h a v i n g r e d u c e d c h l o r o p h y l l l e v e l s ( r e l a t i v e t o t h o s e i n l a r g e p o t s ) , showed a h i g h e r , but s i m i l a r , c a r b o n d i o x i d e e xchange c a p a c i t y when measured p e r u n i t l e a f a r e a , o r p e r u n i t c h l o r o p h y l l . Hence, i t seems t h a t r e d u c i n g s i n k demand by s i z e r e s t r i c t i o n 1 02 need n o t r e s u l t i n a r e d u c t i o n o f n e t c a r b o n d i o x i d e exchange c a p a c i t y i f c h l o r o p h y l l l e v e l s were r e d u c e d i n t h e p r o c e s s . 4.3.6 E f f e c t s On Dark R e s p i r a t i o n L e a v e s w h i c h showed r e d u c e d n e t c a r b o n d i o x i d e e xchange c a p a c i t y due t o e n r i c h m e n t a l s o showed g r e a t e r r e s p i r a t i o n r a t e s ( T a b l e 4.4, u n s h a d e d l e a v e s ) . D i f f e r e n c e s i n s t o m a t a l r e s i s t a n c e among t r e a t m e n t s were o b s e r v e d , but t h e measurements o f r e s p i r a t o r y c a r b o n e f f l u x were not c o m p l i c a t e d by d i f f e r e n c e s i n s t o m a t a l r e s i s t a n c e , u n l i k e measurements of p h o t o s y n t h e t i c c a r b o n f l u x . As s t o m a t a c l o s e d i n t h e d a r k , s t o m a t a l r e s i s t a n c e i n c r e a s e d . I n t e r c e l l u l a r c a r b o n d i o x i d e c o n c e n t r a t i o n c o r r e s p o n d i n g l y i n c r e a s e d b e c a u s e of r e s p i r a t o r y p r o d u c t i o n , w h i c h i n c r e a s e d t h e g r a d i e n t i n c o n c e n t r a t i o n f r o m t h e i n s i d e of t h e l e a f t o t h e o u t s i d e . An i n c r e a s e d d r i v i n g f o r c e t h e r e b y m a i n t a i n e d c o n s t a n t c a r b o n d i o x i d e e f f l u x i n s p i t e of an i n c r e a s e i n s t o m a t a l r e s i s t a n c e . As i l l u s t r a t e d i n F i g u r e 4 . 6 , . r e s p i r a t i o n r a t e was u n a f f e c t e d by s t o m a t a l c l o s u r e . C o n d i t i o n s of low PPFD, w h i c h d i d not e l i c i t a d e c r e a s e i n c a r b o n d i o x i d e exchange c a p a c i t y i n e n r i c h e d p l a n t s , a l s o d i d n o t c a u s e an i n c r e a s e i n r e s p i r a t i o n r a t e ( T a b l e 4.4, s h a d e d l e a v e s ) . T h i s e f f e c t o f low PPFD on r e s p i r a t i o n r a t e may have r e l a t e d t o a r e d u c e d CER o f b o t h e n r i c h e d and c o n t r o l p l a n t s . In my s t u d y , t h e CER of p l a n t s i n t h e i r r e s p e c t i v e t r e a t m e n t chambers was n o t d e t e r m i n e d . However, i t was a p p a r e n t f rom t h e u n i t l e a f r a t e d a t a ( C h a p t e r 3 ) , and f r o m F i g u r e 4.2 ( t h e CER of t h e f i r s t t r i f o l i a t e under g r o w t h c o n d i t i o n s can be a p p r o x i m a t e d 103 s s . o u t l e t d e w p o i n t o u t l e t C 0 2 k i * 9 1 8 2 7 3 6 4 5 T I M E , m i n FIGURE 4.6 Time-course of the response of l e a f water vapour and carbon d i o x i d e exchange to a l i g h t - d a r k t r a n s i t i o n . The l e a f was darkened at time zero . S o l i d l i n e s are chamber o u t l e t measurements and broken l i n e s are r e f e r e n c e i n l e t measurements. The a b s o l u t e magnitude of the d i f f e r e n c e between the i n l e t and o u t l e t i s unimportant here, s i n c e i t i s a f u n c t i o n of the degree of loop r e c i r c u l a t i o n i n the gas exchange measuring system. However, the r e l a t i v e changes in the d i f f e r e n c e between i n l e t and o u t l e t of the two gas exchanges should be noted. 1 04 a t a PPFD of 350 umol n r 2 s " 1 and a t a measurement c a r b o n d i o x i d e c o n c e n t r a t i o n of 340 u l l " 1 f o r c o n t r o l s and 1000 u l 1~ 1 f o r e n r i c h e d p l a n t s ) , t h a t e n r i c h e d p l a n t s had e n h a n c e d CER compared t o c o n t r o l s d u r i n g g r o w t h . A l i n e a r r e l a t i o n s h i p between n e t p h o t o s y n t h e s i s and t h e s u b s e q u e n t d a r k r e s p i r a t i o n r a t e has been r e p o r t e d ( P e n n i n g de V r i e s , 1972; Ludwig e t a l . , 1975). When p h o t o s y n t h e s i s was v a r i e d by c h a n g i n g e i t h e r l i g h t i n t e n s i t y or c a r b o n d i o x i d e c o n c e n t r a t i o n (Ludwig e t a l . , 1975), b o t h t r e a t m e n t s showed a l i n e a r p o s i t i v e r e l a t i o n s h i p between n e t p h o t o s y n t h e s i s r a t e and d a r k r e s p i r a t i o n r a t e , a l t h o u g h t h e r e s p o n s i v e n e s s t o c a r b o n d i o x i d e was somewhat l e s s . E n r i c h m e n t may have c a u s e d b o t h r e d u c e d n e t c a r b o n d i o x i d e e x change c a p a c i t y and e n h a n c e d r e s p i r a t i o n b e c a u s e of g r e a t e r c a r b o n d i o x i d e e xchange r a t e s . F u r t h e r , t h e g r e a t e r -c a r b o h y d r a t e c o n t e n t o f e n r i c h e d p l a n t s may have been r e l a t e d t o the. i n c r e a s e i n r e s p i r a t i o n . T h e r e i s some e v i d e n c e of a p o s i t i v e r e l a t i o n s h i p between carbohy-drate c o n t e n t and r e s p i r a t i o n r a t e . A z c o n - B i e t o and Osmond (1983) have f o u n d t h a t a h i g h c a r b o n d i o x i d e e f f l u x i n t h e d a r k , i m m e d i a t e l y f o l l o w i n g a p e r i o d of p h o t o s y n t h e s i s was c o r r e l a t e d w i t h a g r e a t e r c o n c e n t r a t i o n o f c a r b o h y d r a t e s i n wheat l e a v e s . 4.3.7 E f f e c t s On Stomata The s t o m a t a of b o t h e n r i c h e d and c o n t r o l p l a n t s c l o s e d i n r e s p o n s e t o i n c r e a s e d c a r b o n d i o x i d e c o n c e n t r a t i o n when measured a t 1000 umol m"2 s ~ 1 PPFD ( T a b l e 4 . 7 ) . However, s t o m a t a l r e s i s t a n c e o f e n r i c h e d p l a n t s was g e n e r a l l y l o w e r t h a n , o r e q u a l 105 TABLE 4 . 7 . S tomata l d i f f u s i v e r e s i s t a n c e of the p r imary l e a f and f i r s t t r i f o l i a t e of e n r i c h e d (1400 u l l ' 1 ) and c o n t r o l (340 u l l " 1 ) p l a n t s 1 . Treatment R s 2 at 340 u l 1' (s mm" 1) R s 3 at 1000 u l 1 (s mm" 1) Pr imary L e a f 340 u l 1- 1 1400 u l I" 1 51.6 ±5.4 38.4 ±3.2 F i r s t 340 12.4 ±0.7 24.3 ±2.9 T r i f o l i a t e 5 u l l ' 1 1400 9.6 ±0.7 22.3 ±6.3 u l I" 1 Va lues are the means of measurements of 4 l e a v e s , w i th s tandard e r r o r Rs was measured at an e x t e r n a l carbon d i o x i d e c o n c e n t r a t i o n of 340 u l 1~1 Rs was measured at an e x t e r n a l carbon d i o x i d e c o n c e n t r a t i o n of 1000 u l 1~1 Measured 6 days a f t e r t r a n s f e r to the t reatment chamber Measured 2 days a f t e r t r a n s f e r to the t reatment chamber, and from a d i f f e r e n t set of p l a n t s than the pr imary l e aves 106 t o , t h a t of c o n t r o l s when measured a t t h e same c a r b o n d i o x i d e c o n c e n t r a t i o n . The d a t a were q u i t e v a r i a b l e , and f u r t h e r work i s n e e d ed t o d e t e r m i n e t h e c i r c u m s t a n c e s of t h e somewhat r e d u c e d s t o m a t a l r e s i s t a n c e of l e a v e s of e n r i c h e d p l a n t s . The r e d u c t i o n of s t o m a t a l r e s i s t a n c e i n e n r i c h e d p l a n t s i m p l i e s t h a t s t o m a t a have a c c l i m a t e d t o h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n s , a t l e a s t u nder some c o n d i t i o n s . T h i s has n o t been p r e v i o u s l y r e p o r t e d . 4.3.8 E f f e c t On Tomato S e v e r a l a u t h o r s ( B i s h o p and W h i t t i n g h a m , 1968; Madsen, 1975) have f o u n d i n c r e a s e d p h o t o s y n t h e t i c r a t e s i n e n r i c h e d tomato p l a n t s compared t o c o n t r o l s , when measured a t t h e same e x t e r n a l c a r b o n d i o x i d e c o n c e n t r a t i o n . I n c r e a s e d p h o t o s y n t h e t i c r a t e s c o u l d , i n p a r t , have been due t o d i f f e r e n c e s i n s t o m a t a l a p e r t u r e between t r e a t m e n t s . H i c k l e n t o n and J o l l i f f e ( 1 9 8 0 b ) , however, showed t h a t e n r i c h e d t omato p l a n t s had i n c r e a s e d p h o t o s y n t h e t i c a b i l i t y e ven when measurements were b a s e d on c o m p a r a b l e i n t e r c e l l u l a r c a r b o n d i o x i d e c o n c e n t r a t i o n s . ' They a t t r i b u t e d t h e i n c r e a s e d c a p a c i t y t o a d e c r e a s e i n p h o t o r e s p i r a t i o n r a t e . I n t e r e s t i n g l y , t h e e f f e c t o c c u r e d o n l y i n e n r i c h e d p l a n t s a t 1000 u l l " 1 c a r b o n d i o x i d e and n o t i n t h o s e a t 5000 u l 1~ 1 c a r b o n d i o x i d e , and o n l y a t a p a r t i c u l a r age. The d a t a were i n c o n c l u s i v e as t o why t h e e f f e c t e v e n t u a l l y d i m i n i s h e d . I t i s p o s s i b l e t h a t s p e c i e s o r e n v i r o n m e n t a l d i f f e r e n c e s were r e s p o n s i b l e f o r t h e d i s c r e p a n c y between my d a t a and t h o s e of H i c k l e n t o n and J o l l i f f e ( 1 9 8 0 b ) . In an e f f o r t t o r e s o l v e t h e c o n t r a d i c t i o n , t h e same c u l t i v a r of tomato w h i c h was u s e d i n 1 07 t h e i r s t u d y was grown under c o n d i t i o n s a s s i m i l a r a s p o s s i b l e t o t h a t d e s c r i b e d i n t h e i r work; l i g h t i n t e n s i t y was 150 umol i r r 2  s-1 PPFD, t e m p e r a t u r e was 35 C, and p h o t o p e r i o d was 15 h o u r s . Measurements o f CER w h i c h were o b t a i n e d from l e a v e s a t t h e same node and a t t h e same d e v e l o p m e n t a l s t a g e as t h a t r e p o r t e d by H i c k l e n t o n and J o l l i f f e (1980b) a r e g i v e n i n T a b l e 4.8. A c o n c e n t r a t i o n of 1000 u l 1~ 1 c a r b o n d i o x i d e c a u s e d r e d u c e d n e t c a r b o n d i o x i d e e xchange c a p a c i t y , j u s t a s i n bean p l a n t s . T h e r e a r e , however, s e v e r a l p o i n t s of d i f f e r e n c e between my e x p e r i m e n t s and t h o s e of H i c k l e n t o n and J o l l i f f e ( 1 9 8 0 b ) . A l t h o u g h t h e g r o w t h chambers u s e d i n b o t h c a s e s were open and had c o n t i n u o u s f l o w of f r e s h a i r , my chambers had a t u r n o v e r t i m e of 1 m i n u t e , whereas t h e chambers u s e d by H i c k l e n t o n and J o l l i f f e had a 10 m i n u t e t u r n o v e r t i m e . T h i s would l e a d t o h i g h e r chamber h u m i d i t y and a 1.0-fold i n c r e a s e i n c o n c e n t r a t i o n of any v o l a t i l e s , s u c h as e t h y l e n e , p r o d u c e d by t h e p l a n t . E t h y l e n e i s a known p l a n t g r o w t h r e g u l a t o r and c o u l d c o n c e i v a b l y have a f f e c t e d CER. E t h y l e n e c o n c e n t r a t i o n i n my chambers was l e s s t h a n 0.2 u l l " 1 , but was n o t measured by H i c k l e n t o n and J o l l i f f e ( 1 9 8 0 b ) . 4.3.9 Summary Bush beans grown i n a t m o s p h e r e s e n r i c h e d w i t h c a r b o n d i o x i d e showed a r e d u c e d c a r b o n d i o x i d e exchange c a p a c i t y , t h u s s u p p o r t i n g t h e m a j o r i t y o f r e p o r t s d e a l i n g w i t h gas exchange i n e n r i c h e d p l a n t s of o t h e r s p e c i e s . The r e d u c t i o n i n c a p a c i t y was not a s s o c i a t e d w i t h c h a n g e s i n l e a f t h i c k n e s s , nor was i t due t o i n c r e a s e d s t o m a t a l r e s i s t a n c e , w h i c h has been p r e v i o u s l y 108 TABLE 4.8 Net carbon d i o x i d e exchange r a t e of c o n t r o l (340 u l l " 1 ) and e n r i c h e d (1000 u l l ' 1 ) tomato p l a n t s at a PI of 5 . 0 1 Treatment CER 2 at 340 u l I"1 (umol m - 2 %CER : CER" at 1000 u l r (umol m~2 s' %CER ! 340 u l I" 1 13.4 ±1.4 20.7 ±1.8 1000 u l I" 1 9.4 ±1.3 30 14.7 ±1.6 29 1 Va lues are the means of 3 p l a n t s , w i th s tandard e r r o r . - 1 PPFD and CER measured at 650 umol i r r 2 340 u l l " 1 carbon d i o x i d e D i f f e r e n c e in CER between c o n t r o l and e n r i c h e d p l a n t s measured at 340 u l l " 1 carbon d i o x i d e , and expressed as a percentage of c o n t r o l v a l ues CER measured at 650 umol nr 2 s~ 1 PPFD and 1000 u l 1~1 carbon d i o x i d e D i f f e r e n c e in CER between c o n t r o l and e n r i c h e d p l a n t s measured at 1000 u l l " 1 carbon d i o x i d e , and expressed as a percentage of c o n t r o l v a l ue s 109 a s s o c i a t e d w i t h t h e a p p a r e n t r e d u c t i o n i n c a p a c i t y (Imai and M u r a t a , 1 9 7 8 c ) . The r e d u c t i o n i n c a r b o n d i o x i d e exchange c a p a c i t y w i t h e n r i c h m e n t was a l s o not due t o r e d u c e d c h l o r o p h y l l l e v e l s . A r e d u c e d c h l o r o p h y l l c o n c e n t r a t i o n has been o b s e r v e d i n e n r i c h e d p l a n t s of o t h e r s p e c i e s , and c o u l d c o n c e i v a b l y have a c c o u n t e d f o r t h e d i f f e r e n c e i n c a r b o n d i o x i d e e xchange c a p a c i t y i n o t h e r s t u d i e s . E n r i c h e d beans showed t h e same d e g r e e of oxygen s e n s i t i v i t y as u n e n r i c h e d b e a n s , s u g g e s t i n g t h a t p h o t o r e s p i r a t i o n r a t e was u n a l t e r e d by e n r i c h m e n t and was t h e r e f o r e n o t a s s o c i a t e d w i t h t h e r e d u c e d c a r b o n d i o x i d e e xchange c a p a c i t y . T h i s c o n t r a s t s w i t h a d e c r e a s e i n oxygen s e n s i t i v i t y i n e n r i c h e d tomato p l a n t s w h i c h showed an i n c r e a s e d c a r b o n d i o x i d e exchange c a p a c i t y ( H i c k l e n t o n and J o l l i f f e , 1980b). • E n r i c h e d and u n e n r i c h e d l e a v e s m a i n t a i n e d a t a low PPFD, and hence a t low and s i m i l a r c a r b o n d i o x i d e e xchange r a t e s , d i d not show a d i f f e r e n c e i n c a r b o n d i o x i d e e xchange c a p a c i t y . T h i s o b s e r v a t i o n s u g g e s t s t h a t t h e r e d u c t i o n i n c a p a c i t y o f e n r i c h e d p l a n t s was a f u n c t i o n of t h e h i g h e r c a r b o n d i o x i d e exchange r a t e s a t t h e h i g h e r PPFD. T h i s r e l a t i o n s h i p has not been p r e v i o u s l y d e m o n s t r a t e d . I f c h l o r o p h y l l l e v e l s were r e d u c e d i n c o n t r o l and e n r i c h e d p l a n t s , t h e r e a g a i n was no d i f f e r e n c e i n c a r b o n d i o x i d e exchange c a p a c i t y . A s s u m i n g t h a t r e d u c e d c h l o r o p h y l l l e v e l s were l i m i t i n g CER i n b o t h c o n t r o l and e n r i c h e d p l a n t s , t h e e v i d e n c e would s u p p o r t t h e i d e a of a dependence of gas exchange c a p a c i t y on o v e r a l l CER. 110 As n o t e d i n p r e v i o u s s t u d i e s , e n r i c h e d p l a n t s showed i n c r e a s e d s t a r c h c o n t e n t i n t h e l e a v e s . However, t h e r e l a t i o n s h i p between s t a r c h c o n t e n t and c a r b o n d i o x i d e exchange c a p a c i t y , i n r e s p o n s e t o a change i n s i n k demand i n e n r i c h e d compared t o u n e n r i c h e d p l a n t s , has not been s t u d i e d p r e v i o u s l y . E n r i c h e d bean p l a n t s d i d n o t show t h e l a r g e c h a n g e s i n gas exchange c a p a c i t y f o u n d i n " u n e n r i c h e d p l a n t s when s i n k demand was a l t e r e d . The r e l a t i v e l y s m a l l c h a n g e s i n c a p a c i t y p o s s i b l y r e l a t e d t o a c o n s i s t e n t l y h i g h e r s t a r c h c o n t e n t i n e n r i c h e d p l a n t s . The h i g h e r s t a r c h l e v e l s may have r e d u c e d t h e upper l i m i t of c a r b o n d i o x i d e e xchange c a p a c i t y of e n r i c h e d p l a n t s . R e s p i r a t i o n r a t e i n c r e a s e d i n e n r i c h e d l e a v e s b ut o n l y i f t h e r e was a c o n c u r r e n t d e c r e a s e i n c a r b o n d i o x i d e e xchange c a p a c i t y . T h i s c o r r e l a t i o n i s a new and i n t e r e s t i n g f i n d i n g i n t h a t i t s u g g e s t s t h a t t h e two p r o c e s s e s were b e i n g i n f l u e n c e d by some common f a c t o r , p e r h a p s an i n c r e a s e d c a r b o h y d r a t e s t a t u s . 111 V. EFFECTS ON LEAF SENESCENCE 5. 1 INTRODUCTION D u r i n g t h e c o u r s e o f t h i s s t u d y , i t was o b s e r v e d t h a t s p e c i f i c l e a v e s of e n r i c h e d beans became p r e m a t u r e l y c h l o r o t i c . T h e r e have been o n l y a few o b s e r v a t i o n s of t h i s e f f e c t r e p o r t e d i n t h e l i t e r a t u r e ( W i t t w e r , 1967; H e s k e t h e t a l . , 1971; Chang, 1975). C a r b o n d i o x i d e - i n d u c e d s e n e s c e n c e c o r r e l a t e d w i t h a h i g h e r l e a f s t a r c h c o n t e n t i n c o t t o n and s o y b e a n , a l t h o u g h e n r i c h e d s u n f l o w e r , w h i c h a l s o had h i g h s t a r c h l e v e l s due t o e n r i c h m e n t , d i d not show a s i m i l a r r e l a t i o n s h i p ( H e s k e t h e t a l . , 1971). The s e n e s c e n c e was a l s o a s s o c i a t e d w i t h r e d u c e d p h o t o p h o s p h o r y l a t i o n and H i l l r e a c t i o n a c t i v i t y (Chang, 1975). Those o b s e r v a t i o n s c o n s t i t u t e t h e e x t e n t of knowledge a b o u t t h i s p o t e n t i a l l y i n t e r e s t i n g p h y s i o l o g i c a l p r o b l e m . I . t h e r e f o r e was i n t e r e s t e d i n e x a m i n i n g some of t h e e n v i r o n m e n t a l and p h y s i o l o g i c a l p a r a m e t e r s a s s o c i a t e d w i t h a c c e l e r a t e d c h l o r o s i s and s e n e s c e n c e of e n r i c h e d bean l e a v e s . The f o l l o w i n g q u e s t i o n s were a d d r e s s e d : 1. Which l e a v e s a r e i n d u c e d t o s e n e s c e by c a r b o n d i o x i d e e n r i c h m e n t , and how r a p i d i s t h e r e s p o n s e ? 2. How i s h i g h c a r b o n d i o x i d e - i n d u c e d s e n e s c e n c e m o d i f i e d by t h e e n v i r o n m e n t ? F o r example, how does t e m p e r a t u r e i n f l u e n c e t h e i n d u c t i o n or s u b s e q u e n t r a t e o f c a r b o n d i o x i d e - i n d u c e d s e n e s c e n c e ? S e c o n d l y , i s c a r b o n d i o x i d e - i n d u c e d s e n e s c e n c e i n f l u e n c e d by l i g h t i n t e n s i t y ? 3. S i n c e an i n c r e a s e i n s t a r c h c o n t e n t f i g u r e s so p r o m i n e n t l y 1 12 as a consequence of enrichment (see Chapter 4), i s i n c r e a s e d s t a r c h content r e l a t e d to the a c c e l e r a t e d senescence at high carbon d i o x i d e c o n c e n t r a t i o n s ? 4. Is high carbon d i o x i d e - i n d u c e d senescence r e l a t e d to changes in s o l u t e content of e n r i c h e d leaves? Conceivably, an i n c r e a s e in s o l u t e c o n c e n t r a t i o n may p a r a l l e l the i n c r e a s e i n s t a r c h , which c o u l d r e s u l t i n an o s m o t i c a l l y - i n d u c e d senescence. 5. Since carbon d i o x i d e causes stomatal c l o s u r e , i s the senescence caused by carbon d i o x i d e enrichment r e l a t e d to stomatal c l o s u r e ? Answers to these q u e s t i o n s w i l l c o n t r i b u t e s i g n i f i c a n t l y to an understanding of how carbon d i o x i d e enrichment i n f l u e n c e s l e a f senescence, and more g e n e r a l l y , how t h i s response r e l a t e s to other aspects of carbon d i o x i d e enrichment and p l a n t funct i o n . 1 1 3 5.2 MATERIALS AND METHODS P r e - t r e a t m e n t p l a n t c u l t u r e c o n d i t i o n s were as d e s c r i b e d p r e v i o u s l y ( C h a p t e r 3 ) . At 10 d a y s , when t h e p r i m a r y l e a v e s were f u l l y e x panded, 4 t o 8 p l a n t s were t r a n s f e r r e d t o e a c h t r e a t m e n t chamber and e x p o s e d t o e i t h e r 340±10U1 1" 1 ( c o n t r o l ) or 1400±20 u l l ' 1 ( e n r i c h e d ) c a r b o n d i o x i d e . D e p e n d i n g on t h e e x p e r i m e n t , g r o w t h t e m p e r a t u r e was e i t h e r 32 C day/25 C n i g h t (warm), o r 20 C day/15 C n i g h t ( c o o l ) . S u b s e q u e n t r e f e r e n c e t o th e g r o w t h t e m p e r a t u r e w i l l m e n t i o n t h e day t e m p e r a t u r e o n l y . A PPFD of 350 umol m~2 s " 1 was g e n e r a l l y u s e d , but i n some e x p e r i m e n t s , two l e v e l s of PPFD were u s e d . In t h i s i n s t a n c e , e a c h p l a n t was p o s i t i o n e d i n t h e chamber s u c h t h a t one p r i m a r y l e a f r e c e i v e d 350 umol m~2 s~'1 PPFD and t h e o t h e r 100 umol m"2  s - 1 PPFD. L i g h t i n t e n s i t y was a l s o m a n i p u l a t e d i n s i m i l a r -e x p e r i m e n t s by s h a d i n g one p r i m a r y l e a f w i t h a s i n g l e l a y e r of opaque p a p e r . P h o t o p e r i o d was 16 h o u r s u n l e s s o t h e r w i s e i n d i c a t e d . CER was m easured a t v a r i o u s t i m e s d u r i n g s e n e s c e n c e u s i n g a semi-open gas exchange s y s t e m ( s e e s e c t i o n 4 . 2 ) . In t i m e - c o u r s e s t u d i e s , e a c h l e a f c o u l d o n l y be u s e d once, b e c a u s e use o f t h e gas exchange c u v e t t e l e f t a r e s i d u a l r i n g of g r e a s e , d e p o s i t e d when t h e l e a f was s e a l e d i n t h e gas exchange chamber. Hence, t i m e - c o u r s e s t u d i e s of CER d u r i n g s e n e s c e n c e u s e d l e a v e s o f d i f f e r e n t p l a n t s a t e a c h s a m p l i n g d a t e . In a l l e x p e r i m e n t s , CER was d e t e r m i n e d a t 1000 umol m~2- s " 1 PPFD, and a t t h e p r e v a i l i n g g r o w t h t e m p e r a t u r e ; CER was b a s e d on an i n t e r c e l l u l a r c a r b o n d i o x i d e c o n c e n t r a t i o n o f 340 u l 1~ 1 1 1 4 ( i n t e r p o l a t e d f r o m c a r b o n d i o x i d e r e s p o n s e c u r v e s ) i n t h e t i m e -c o u r s e s t u d i e s , and on an e x t e r n a l c a r b o n d i o x i d e c o n c e n t r a t i o n o f 340 u l 1"' i n a l l o t h e r e x p e r i m e n t s . C h l o r o p h y l l d e t e r m i n a t i o n s were made a s p r e v i o u s l y d e s c r i b e d ( s e e s e c t i o n 4 . 2 ) . S p e c i f i c l e a f w e i g h t was d e t e r m i n e d by t a k i n g t h e oven d r y w e i g h t of a known a r e a of i n t e r v e i n a l l e a f t i s s u e . M easurements of s t o m a t a l r e s i s t a n c e were o b t a i n e d i n t h e t r e a t m e n t chambers and under g r o w t h c o n d i t i o n s . A d i f f u s i v e r e s i s t a n c e p o r o m e t e r was u s e d ( L i - C o r , model LI 6 0 ) . P l a s t o c h r o n Index was d e t e r m i n e d as i n S e c t i o n 4.2. In one e x p e r i m e n t , l e a v e s were e x c i s e d from t h e p l a n t a f t e r 10 d ays i n t h e p r e - t r e a t m e n t g r o w t h chamber, p l a c e d w i t h t h e i r p e t i o l e s i n s m a l l b e a k e r s of d i s t i l l e d w a t e r , and t r a n s f e r r e d t o t h e t r e a t m e n t chambers. Chamber t e m p e r a t u r e was m a i n t a i n e d a t 23 C day/17 C n i g h t , and l i g h t i n t e n s i t y was 370 umol m"2 s " 1 PPFD. C a r b o n d i o x i d e c o n c e n t r a t i o n was e i t h e r 340 or 3000 u l I" 1 . Water p o t e n t i a l s of l e a f d i s k s of c o n t r o l and e n r i c h e d p l a n t s were d e t e r m i n e d w i t h a Wescor dew p o i n t m i c r o v o l t m e t e r (model HR-33T, w i t h sample chamber C - 5 2 ) . The samples were t h e n f r o z e n on d r y i c e i n aluminum f o i l and s u b s e q u e n t l y m easured a g a i n w i t h t h e dew p o i n t m i c r o v o l t m e t e r t o d e t e r m i n e o s m o t i c p o t e n t i a l s . S p e c i f i c l e a f w e i g h t was a l s o d e t e r m i n e d u s i n g d i s k s o b t a i n e d f r o m t h e same l e a v e s . 115 5.3 RESULTS AND DISCUSSION As l e a v e s d e v e l o p , t h e y e x h i b i t a c h a r a c t e r i s t i c p a t t e r n i n p h o t o s y n t h e t i c c a p a c i t y . P h o t o s y n t h e s i s p e r u n i t l e a f a r e a i n c r e a s e s s t e a d i l y d u r i n g t h e t i m e of n e t i m p o r t a t i o n of a s s i m i l a t e s and r e a c h e s a maximum as t h e r a t e of i m p o r t r e a c h e s z e r o . At t h a t t i m e t h e l e a f i s a n e t e x p o r t e r ( G i a q u i n t a , 1978). P h o t o s y n t h e s i s r a t e t h e n d e c l i n e s i n t h e l a t t e r s t a g e s of l e a f d e v e l o p m e n t ( D a v i s and McCree, 1978). The d e c l i n e i n CER i n mature p r i m a r y l e a v e s i s e v i d e n t i n T a b l e 5.1. W i t h i n 17 d a y s o f t r a n s f e r t o c a r b o n d i o x i d e t r e a t m e n t chambers, CER d e c r e a s e d t o z e r o i n e n r i c h e d p l a n t s , i r r e s p e c t i v e of t h e g r o w t h t e m p e r a t u r e . The d e c l i n e i n CER of u n e n r i c h e d l e a v e s was much l e s s e v i d e n t . C h l o r o p h y l l l e v e l s a l s o d e c r e a s e d d r a m a t i c a l l y i n e n r i c h e d p r i m a r y l e a v e s a t b o t h t e m p e r a t u r e s ( T a b l e 5 . 1 ) . P r i m a r y l e a v e s of e n r i c h e d p l a n t s a t 20 C had l o s t v i r t u a l l y a l l c h l o r o p h y l l by 20 d a y s a f t e r t r a n s f e r ( F i g u r e 5 . 1 ) . Changes i n CER were f i r s t e v i d e n t i n p l a n t s a t 20 C ( F i g . 5.2) 6 d a y s a f t e r t r a n s f e r . 5.3.1 E f f e c t s Of T e m p e r a t u r e A c c e l e r a t e d s e n e s c e n c e may be due s i m p l y t o t h e n o r m a l advancement o f a g i n g o r i t may i n v o l v e d e t e r i o r a t i o n not a s s o c i a t e d w i t h n o r m a l a g i n g , f o r example, due t o e n v i r o n m e n t a l s t r e s s . P l a n t s a t t h e l o w e r t e m p e r a t u r e (20 C) d i d n o t grow as r a p i d l y as t h o s e a t t h e h i g h e r t e m p e r a t u r e (32 C ) , as shown by t h e i r l o w e r P l a s t o c h r o n Index ( T a b l e 5 . 1 ) . The i n i t i a t i o n o f s e n e s c e n c e , as e s t i m a t e d by t h e f i r s t s i g n s of c h l o r o p h y l l l o s s 116 TABLE 5 .1 . Net carbon d i o x i d e exchange ra te and c h l o r o p h y l l content of the pr imary l eaves be fo re and a f t e r 17 days of carbon d i o x i d e enr ichment at 32 or 20 C. 1 Treatment CER 2 C h l o r o p h y l l PI ( [C02]/T . ) (umol n r 2 s ^ H g irr 2 ) Be fore E n r i c h m e n t 3 : 340 u l l " 1 / 2 0 15.3±2.6 0.67±0.03 0.5 340 u l l " 1 / 3 2 15.9±1.8 0.67±0.03 0.5 A f t e r Enr i chment : 340 u l 1- 1 /20 10.5±1.3 0.6010.02 3.5 1 400 u l 1- 1 /20 O.OlO.O 0.2510.08 3.5 340 u l 1- V32 8.510.5 0.5410.04 >5.0 1400 u l 1- V32 O.OlO.O 0.13+0.07 >5.0 1 Va lues are the means of 3 measurements, w i th s tandard e r r o r 2 CER was measured at 1000 umol m" 2 s " 1 PPFD and an e x t e r n a l carbon d i o x i d e c o n c e n t r a t i o n s u f f i c i e n t to g i ve a C i of 340 u l l " 1 3 The same p l a n t s were used f o r a l l i n i t a l v a l u e s , hence on l y one i n i t i a l c h l o r o p h y l l va lue i s g i ven 117 FIGURE 5.1 P h o t o g r a p h of t h e p r i m a r y l e a v e s of e n r i c h e d (1400 u l l " 1 ) and c o n t r o l (340 u l l " 1 ) p l a n t s grown f o r 20 d a y s a t 20 C. 118 TIME AFTER TRANSFER (days) FIGURE 5.2 T i m e - c o u r s e of n e t c a r b o n d i o x i d e e x c h a n g e r a t e of p r i m a r y l e a v e s of e n r i c h e d (1400 u l 1~ 1 ) and c o n t r o l (340 u l l " 1 ) p l a n t s grown a t 20 C. CER was measured a t 1000 umol m"2 s ~ 1 PPFD and an i n t e r c e l l u l a r s p a c e c a r b o n d i o x i d e c o n c e n t r a t i o n of 340 u l l " 1 . E a c h v a l u e i s a measurement o f a d i f f e r e n t l e a f . E n r i c h e d p l a n t s , c l o s e d c i r c l e s ; c o n t r o l p l a n t s , open c i r c l e s . 119 o c c u r r e d somewhat s o o n e r i n p h y s i o l o g i c a l l y y o u n g e r l e a v e s a t c o o l t e m p e r a t u r e s t h a n i n p h y s i o l o g i c a l l y o l d e r l e a v e s a t warm t e m p e r a t u r e ( d a t a n o t p r e s e n t e d ) . F u r t h e r m o r e , b o t h e n r i c h e d and c o n t r o l p r i m a r y l e a v e s a t 32 C had a CER o f 13 umol n r 2 s " 1 a t day 8, s i m i l a r t o i n i t i a l r a t e s ( T a b l e 5 .1), but t h e CER of p r i m a r y l e a v e s of e n r i c h e d p l a n t s a t t h e c o o l t e m p e r a t u r e had d e c r e a s e d t o 5 umol n r 2 s ~ 1 by t h a t t i m e ( F i g . 5 . 2 ) . T h e r e f o r e , t h e i n d u c t i o n of s e n e s c e n c e was p r o b a b l y n o t due t o a c c e l e r a t e d a g i n g . T h i s c o n c l u s i o n r e q u i r e s t h a t t h e P l a s t o c h r o n Index be an e f f e c t i v e measure of p h y s i o l o g i c a l age. One must be c a r e f u l i n c o m p a r i n g r a t e s of s e n e s c e n c e a t two d i f f e r e n t t e m p e r a t u r e s . Once i n i t i a t e d , s e n e s c e n c e p r o c e e d e d a t a r a t e d e t e r m i n e d by t h e p r e v a i l i n g t e m p e r a t u r e , even t h o u g h t h e a c t u a l i n d u c t i o n of s e n e s c e n c e may have been e a r l i e r at- t h e c o o l e r t e m p e r a t u r e . A f t e r i n d u c t i o n , t h e r a t e of s e n e s c e n c e may have been s l o w e r b e c a u s e of a t e m p e r a t u r e - r e l a t e d s l o w i n g of m e t a b o l i s m . As a c a s e i n p o i n t , i f e n r i c h e d p l a n t s were t r a n s f e r r e d t o a h i g h e r t e m p e r a t u r e a t t h e o n s e t o f t h e f i r s t s i g n of c h l o r o s i s , f u r t h e r y e l l o w i n g p r o c e e d e d a t an a c c e l e r a t e d r a t e compared t o e n r i c h e d p l a n t s w h i c h were not t r a n s f e r r e d ( d a t a n o t shown). A l s o , c o n t r o l p l a n t s a t b o t h t e m p e r a t u r e s ( T a b l e 5.1) showed an e q u i v a l e n t d e c l i n e i n p h o t o s y n t h e s i s and c h l o r o p h y l l c o n t e n t , w h i c h may be m i s l e a d i n g f o r t h e r e a s o n s j u s t d i s c u s s e d . However, t h e r e d i d seem t o be a t e m p e r a t u r e - r e l a t e d e f f e c t on s e n e s c e n c e . T r i f o l i a t e s of e n r i c h e d p l a n t s a l s o showed c h l o r o s i s , b ut o n l y a t t h e c o o l t e m p e r a t u r e . The c h l o r o s i s 1 20 a p p e a r e d i n s m a l l p a t c h e s of i n t e r v e i n a l t i s s u e ( F i g . 5 . 3 ) , w h i c h e v e n t u a l l y p r o g r e s s e d t o c o m p l e t e y e l l o w i n g . Those l e a v e s d i d n o t expand t o n o r m a l s i z e , and t h e c h l o r o t i c l e s i o n s d e v e l o p e d when t h e t r i f o l i a t e s were s t i l l q u i t e s m a l l ( a b o u t 30% o f n o r m a l maximum l e a f s i z e ) . S e n e s c e n c e i s known t o be a c c e l e r a t e d by e x t r e m e s o f t e m p e r a t u r e (Thomas and S t o d d a r t , 1980). Two d a y s o f c h i l l i n g s t r e s s a t 10 C and a l i g h t i n t e n s i t y o f 215 W m-2 r e d u c e d p h o t o s y n t h e s i s t o z e r o i n Sorghum, a l t h o u g h c h l o r o p h y l l l e v e l s were not a f f e c t e d ( T a y l o r and Rowley, 1971). The e f f e c t s i n Sorghum were a t t r i b u t e d t o s o l a r i z a t i o n , w h i c h i s a l i g h t -d e p e n d e n t d e s t r u c t i o n of t h e p h o t o s y n t h e t i c a p p a r a t u s , o r as i n i t i a l l y d e f i n e d , a r e d u c e d a b i l i t y of c h l o r o p l a s t s t o f o r m s t a r c h (Holman, 1930). The t e m p e r a t u r e s t h a t a c c e l e r a t e d s e n e s c e n c e . i n my s t u d y were q u i t e m o d e r a t e by c o m p a r i s o n , and t h e PPFD was a maximum of o n l y 370 umol m~2 s " 1 (130 W n r 2 ) , q u i t e low compared t o t h a t g e n e r a l l y needed f o r s o l a r i z a t i o n . 5.3.2 E f f e c t s On S tomata S t o m a t a l c l o s u r e i s a s s o c i a t e d w i t h l e a f s e n e s c e n c e , a l t h o u g h t h e n a t u r e of t h e r e l a t i o n s h i p i s n o t c l e a r . S tomata a r e known t o c l o s e w i t h d e c r e a s i n g t e m p e r a t u r e , ( H o f s t r a and H e s k e t h , 1969) d e c r e a s i n g l i g h t i n t e n s i t y , or i n c r e a s i n g c a r b o n d i o x i d e c o n c e n t r a t i o n . I t was t h e r e f o r e c o n c e i v a b l e t h a t i n t h i s s t u d y , any one o r a l l of t h o s e p a r a m e t e r s may have a f f e c t e d s e n e s c e n c e t h r o u g h a change i n s t o m a t a l a p e r t u r e . S t o m a t a l r e s i s t a n c e was t h e r e f o r e measured i n s i t u , t h a t i s , i n l e a v e s i n t h e t r e a t m e n t chambers under t h e i r r e s p e c t i v e g r o w t h 121 FIGURE 5.3 P h o t o g r a p h of t h e c e n t r a l l e a f l e t of t h e f i r s t t r i f o l i a t e l e a f of e n r i c h e d (1400 u l 1~ 1) and c o n t r o l (340 u l 1~ 1) p l a n t s grown f o r 20 d a y s a t 20 C. 122 c o n d i t i o n s . F o r t h e f i r s t t h r e e d a y s of t r e a t m e n t w i t h two c a r b o n d i o x i d e c o n c e n t r a t i o n s and two l e v e l s of PPFD a t 20 C, t h e r e were no s i g n i f i c a n t d i f f e r e n c e s i n s t o m a t a l r e s i s t a n c e ( F i g . 5 . 4 ) . However, p l a n t s w h i c h s e n e s c e d t h e most r a p i d l y , t h o s e a t h i g h c a r b o n d i o x i d e (1400 u l l " 1 ) and h i g h l i g h t (370 umol m" 2 s " 1 ) , a l s o had t h e most r a p i d and e x t e n s i v e i n c r e a s e i n s t o m a t a l r e s i s t a n c e . By 17 d a y s , p l a n t s a t h i g h c a r b o n d i o x i d e and low PPFD a l s o began t o show s t o m a t a l c l o s u r e c o n c u r r e n t l y w i t h y e l l o w i n g . T e m p e r a t u r e a l s o had no i n i t i a l e f f e c t on s t o m a t a l r e s i s t a n c e . A f t e r 7 d a y s a t 32 C, s t o m a t a l r e s i s t a n c e was 0.91 ± 0.05 s mm"1 i n c o n t r o l s and 0.78 ±0.06 s mm"1 i n e n r i c h e d p l a n t s , v a l u e s s i m i l a r t o t h o s e a t 20 C ( F i g . 5 . 4 ) . The l a c k of i m m e d i a t e - t e m p e r a t u r e o r c a r b o n d i o x i d e e f f e c t s on s t o m a t a l r e s i s t a n c e was p r o b a b l y due t o a l i m i t a t i o n of s t o m a t a l o p e n i n g by t h e r e l a t i v e l y low PPFD. F u r t h e r m o r e , t h e two l e v e l s of PPFD use d were not s u f f i c i e n t l y d i f f e r e n t t o s i g n i f i c a n t l y a f f e c t s t o m a t a l a p e r t u r e ( F i g . 5 . 4 ) . I t has been s u g g e s t e d by Thimann and S a t l e r (1979) t h a t s t o m a t a l c l o s u r e i s not s i m p l y a r e s u l t of t h e s e n e s c e n c e p r o c e s s , but i s t h e c a u s e . Thimann (1980) s u p p o r t e d t h i s w i t h s e v e r a l l i n e s o f e v i d e n c e : (1) p e e l i n g of t h e e p i d e r m i s of t o b a c c o l e a f d i s c s g r e a t l y d e c r e a s e d t h e e f f e c t s o f k i n e t i n on s e n e s c e n c e , ( 2 ) c o a t i n g w i t h V a s e l i n e p r o m oted s e n e s c e n c e , (3) f u s i c o c c i n , w h i c h p romoted s t o m a t a l o p e n i n g i n t h e d a r k , d e l a y e d t h e o n s e t o f s e n e s c e n c e i n t h e d a r k , (4) p h e n y l m e r c u r i c 123 0 2 4 6 8 10 12 14 16 T I M E A F T E R T R A N S F E R ( d a y s ) FIGURE 5.4 T i m e - c o u r s e o f s t o m a t a l r e s i s t a n c e o f p r i m a r y l e a v e s a f t e r t r a n s f e r t o t r e a t m e n t chambers a t 20 C. Measurements were made i n t h e t r e a t m e n t c hambers. P r i m a r y l e a v e s were m a i n t a i n e d a t e i t h e r 370 ( s o l i d l i n e s ) or 100 ( b r o k e n l i n e s ) umol m~2 s " 1 PPFD, and 340 (open c i r c l e s ) o r 1400 ( c l o s e d c i r c l e s ) u l l " 1 c a r b o n d i o x i d e c o n c e n t r a t i o n . E a c h d a t a p o i n t i s t h e mean of measurements o f 4 t o 6 p l a n t s , w i t h s t a n d a r d e r r o r . 1 24 n i t r a t e , w h i c h promoted s t o m a t a l c l o s u r e i n t h e l i g h t , a l s o p r o m o ted s e n e s c e n c e i n t h e l i g h t , and (5) k i n e t i n was f o u n d t o c a u s e s t o m a t a l o p e n i n g i n t h e d a r k . L e a f gas exchange i s a l t e r e d b e c a u s e of s t o m a t a l c l o s u r e , and i t may be t h a t t h e c o n c e n t r a t i o n o f e t h y l e n e o r c a r b o n d i o x i d e i n t h e l e a f c o u l d be r e l a t e d t o t h e o n s e t of s e n e s c e n c e . The e v i d e n c e f o r and a g a i n s t a r o l e f o r e t h y l e n e i n s e n e s c e n c e has been p r e v i o u s l y d i s c u s s e d ( s e e C h a p t e r 2 ) . The p o s s i b i l i t y o f low c a r b o n d i o x i d e c o n c e n t r a t i o n s c a u s i n g s e n e s c e n c e i s s u p p o r t e d by r e c e n t work by S a t l e r and Thimann ( 1 9 8 3 a ) . They f o u n d t h a t o a t l e a v e s i n a i r minus c a r b o n d i o x i d e s e n e s c e d as r a p i d l y i n l i g h t as i n t h e d a r k . I b e l i e v e t h e r e a r e two p o s s i b l e e x p l a n a t i o n s f o r t h i s e f f e c t . F i r s t , i n t h e a b s e n c e of c a r b o n d i o x i d e , e l e c t r o n t r a n s p o r t i n t h e p h o t o r e a c t i o n s of p h o t o s y n t h e s i s would not - c o n t i n u e t o r e d u c e NADP, but would i n s t e a d r e d u c e oxygen t o h y d r o g e n p e r o x i d e i n t h e M e h l e r r e a c t i o n ( E l s t n e r , 1982). Over t h e l o n g term, t h i s c o u l d have damaging e f f e c t s on t h e p h o t o s y n t h e t i c a p p a r a t u s . S e c o n d l y , i f c a r b o n d i o x i d e d i s p l a c e s e t h y l e n e from i t s b i n d i n g s i t e s i n t h e l e a f , t h e a b s e n c e of c a r b o n d i o x i d e would c a u s e a g r e a t e r e f f e c t i v e c o n c e n t r a t i o n of e t h y l e n e i n t h e l e a f , w h i c h c o u l d i n f l u e n c e s e n e s c e n c e as p r e v i o u s l y d i s c u s s e d . Even i f low c a r b o n d i o x i d e c o n c e n t r a t i o n c a u s e s s e n e s c e n c e , t h e p o s s i b i l i t y of c a r b o n d i o x i d e c o n c e n t r a t i o n s b ecoming low enough t o c a u s e s e n e s c e n c e as a r e s u l t of s t o m a t a l c l o s u r e was not l i k e l y i n my s t u d y . S e n e s c e n c e was o b s e r v e d a t h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n s . Even when s t o m a t a c l o s e d , t h e 1 25 i n t e r c e l l u l a r c a r b o n d i o x i d e c o n c e n t r a t i o n would be h i g h e r t h a n a m b i e n t i n t h o s e t r e a t m e n t s . C l e a r l y , s t o m a t a l c l o s u r e , i f r e l a t e d t o s e n e s c e n c e , was n o t a c t i n g t h r o u g h a low c a r b o n d i o x i d e c o n c e n t r a t i o n . 5.3.3 E f f e c t s Of PPFD S i n c e c a r b o n d i o x i d e e n r i c h m e n t seemed t o i n f l u e n c e c a r b o n d i o x i d e e xchange c a p a c i t y t h r o u g h i t s e f f e c t on i n c r e a s i n g CER ( C h a p t e r 4 ) , s e n e s c e n c e may have been i n d u c e d f o r t h e same r e a s o n . In t h i s c o n t e x t , l i g h t a l s o c a u s e d c h l o r o p h y l l l o s s i n s u g a r c a n e (Edelman and S c h o o l a r , 1969), and i t may be s i g n i f i c a n t t h a t s u g a r c a n e i s a C4 s p e c i e s , and as s u c h was c a p a b l e o f v e r y h i g h p h o t o s y n t h e s i s r a t e s . I f s o , d e c r e a s i n g PPFD s h o u l d d e c r e a s e CER, and hence s e n e s c e n c e . F i g u r e 5.5 shows t h a t c a r b o n d i o x i d e e n r i c h m e n t had a g r e a t e r e f f e c t on l o s s of c a r b o n d i o x i d e exchange c a p a c i t y i n p r i m a r y l e a v e s m a i n t a i n e d a t 375 umol m~2 s " 1 t h a n i n t h o s e a t 90 umol n r 2 s ~ 1 . S i m i l a r l y , l o s s of c h l o r o p h y l l was more r a p i d a t t h e h i g h e r PPFD ( F i g . 5.6) At low PPFD, c a r b o n d i o x i d e e n r i c h m e n t had no c l e a r e f f e c t on c h l o r o p h y l l l e v e l s . I t s h o u l d be k e p t i n mind t h a t t h e l e v e l s of PPFD d i f f e r e d by o n l y 285 umol m"2 s " 1 , and d i f f e r e n c e s i n s e n e s c e n c e were s t i l l o b s e r v e d . The a b i l i t y o f l i g h t t o a c c e l e r a t e s e n e s c e n c e was not c o n s i s t e n t w i t h p r e v i o u s s t u d i e s showing t h a t l i g h t d e l a y e d s e n e s c e n c e compared t o d a r k t r e a t m e n t s i n o a t s (Thimann e t a l . , 1977) and P h a s e o l u s v u l g a r i s ( G o l d t h w a i t e and L a e t s c h , 1967). L i g h t was p o s t u l a t e d t o e x e r t t h i s e f f e c t t h r o u g h p h o t o s y n t h e s i s , and more s p e c i f i c a l l y i n t h e c a s e of o a t s , 126 CM J I I I I L TIME AFTER TRANSFER (days) FIGURE 5.5 Time-course of chlorophyll content and net carbon dioxide exchange rate of shaded and unshaded primary leaves of plants grown at 1400 ul l " 1 carbon dioxide and at 20 C. Shaded leaves (broken lines) were grown at 90 umol n r 2 s" 1 PPFD, and unshaded leaves ( s o l i d lines) were grown at 375 umol n r 2 s" 1 PPFD. Each data point i s a measurement of a d i f f e r e n t l e a f . 127 1400 340 1400 340 370 90 c o 2 P P F D FIGURE 5.6 Chlorophyll content of primary leaves grown at 340 or 1400 ul l ' 1 carbon dioxide, and 90 or 370 umol m~2 s" 1 photosynthetic photon flux density for 17 days. The growth temperature was 20 C. Each treatment i s the mean of 4 leaves, with standard error. 1 28 t h r o u g h c y c l i c p h o t o p h o s p h o r y l a t i o n . T h i s was b a s e d on t h e o b s e r v a t i o n t h a t 3 - ( 3 , 4 - d i c h l o r o p h e n y l ) - 1 , 1 - d i m e t h y l u r e a (DCMU) c o m p l e t e l y b l o c k e d s u g a r f o r m a t i o n i n p h o t o s y n t h e s i s , y e t d i d no t p r e v e n t t h e d e l a y of s e n e s c e n c e by l i g h t (Thimann e t a l . , 1977; Thimann, 1980). L i g h t may p o s s i b l y a l s o a c t t h r o u g h p h otochrome ( s e e Thomas and S t o d d a r t , 1980) a l t h o u g h i n P h a s e o l u s v u l g a r i s t h e r e s p o n s e t o l i g h t d i d n o t have t h e c h a r a c t e r i s t i c s - of a p h y t o c h r o m e r e s p o n s e ( G o l d t h w a i t e and L a e t s c h , 1967). E x p e r i m e n t s d e s i g n e d t o s t u d y t h e e f f e c t o f c a r b o n d i o x i d e on s e n e s c e n c e i n t h e d a r k compared t o i n t h e l i g h t were u n s u c c e s s f u l b e c a u s e of t h e t e c h n i c a l d i f f i c u l t i e s i n p l a c i n g e n t i r e p l a n t s i n a b s o l u t e d a r k n e s s w h i l e m a i n t a i n i n g a d e q u a t e v e n t i l a t i o n and c o n t r o l of c a r b o n d i o x i d e c o n c e n t r a t i o n . The c o n f l i c t i n g d a t a p e r t a i n i n g t o t h e r o l e o f l i g h t i n s e n e s c e n c e may be r e s o l v e d i f one assumes t h a t l i g h t i n f l u e n c e s two d i f f e r e n t p r o c e s s e s , d e p e n d i n g on i n t e n s i t y . Low l e v e l s o f PPFD may be r e q u i r e d t o m a i n t a i n a c e r t a i n d e g r e e of s t o m a t a l o p e n i n g , or l e v e l of e n e r g y c h a r g e i n m e s o p h y l l c e l l s t h r o u g h p h o t o s y n t h e s i s , i n o r d e r t o d e l a y s e n e s c e n c e . However, PPFD a t h i g h e r l e v e l s may have c o n t r i b u t e d t o s e n e s c e n c e t h r o u g h y e t o t h e r c o n s e q u e n c e s o f p h o t o s y n t h e t i c m e t a b o l i s m . In my s t u d y , l i g h t was p r o b a b l y not a c t i n g t h r o u g h a s o l a r i z a t i o n p r o c e s s , s i n c e t h e h i g h e s t l e v e l of PPFD was s t i l l r e l a t i v e l y low i n t h o s e e x p e r i m e n t s compared t o t h e l e v e l needed f o r s o l a r i z a t i o n . 1 29 5.3.4 E f f e c t s On L e a f Dry Weight T r e a t m e n t s w h i c h c a u s e d a g r e a t e r r a t e o f s e n e s c e n c e ( e g . 1400 u l l " 1 c a r b o n d i o x i d e / 3 7 0 umol m"2 s ~ 1 PPFD) a l s o c a u s e d a g r e a t e r SLW ( T a b l e 5 . 2 ) . S p e c i f i c l e a f w e i g h t was d e t e r m i n e d 6 d a y s a f t e r t h e i n i t i a t i o n of e a c h t r e a t m e n t , a t i m e when s e n e s c e n c e symptoms were not y e t a p p a r e n t . By s a m p l i n g a t t h i s t i m e , c h a n g e s i n l e a f d r y w e i g h t w h i c h were a r e s u l t o f s e n e s c e n c e were a v o i d e d . In s a m p l e s t a k e n a t 10 d a y s , when s e n e s c e n c e symptoms were e v i d e n t i n some t r e a t m e n t s , t h e t r e n d i n SLW was u n a l t e r e d ( d a t a not shown). As shown i n F i g u r e 5.7, a h i g h e r SLW c o r r e l a t e d w i t h a g r e a t e r s t a r c h c o n t e n t i n l e a v e s . The SLW o f young e x p a n d i n g f i r s t t r i f o l i a t e s o f e n r i c h e d p l a n t s a t c o o l t e m p e r a t u r e (60 ±1 g n r 2 ) was a l s o h i g h e r t h a n t h a t o f c o n t r o l s a l s o a t c o o l t e m p e r a t u r e (38 ±2 g n r 2 ) . A g a i n , t h e r e was a p o s i t i v e r e l a t i o n s h i p between l e a f d r y w e i g h t and s e n e s c e n c e . S i n c e t h o s e l e a v e s were p r e s u m a b l y s t i l l f u n c t i o n i n g as s i n k s , t h e h i g h e r s p e c i f i c l e a f w e i g h t s may r e s u l t , i n p a r t , from a g r e a t e r t r a n s p o r t of a s s i m i l a t e f r o m s o u r c e l e a v e s , w h i c h was t h e n b e i n g a c c u m u l a t e d , r a t h e r t h a n b e i n g u s e d i n l e a f a r e a e x p a n s i o n . 5.3.5 P r e l i m i n a r y E x p e r i m e n t s W i t h D e t a c h e d L e a v e s P r e v e n t i o n of a s s i m i l a t e movement o u t o f s o u r c e l e a v e s s h o u l d i n c r e a s e s p e c i f i c l e a f w e i g h t i f n e t a s s i m i l a t i o n c o n t i n u e s . F o r t h i s r e a s o n i t was of i n t e r e s t t o o b s e r v e how p r e v e n t i o n o f a s s i m i l a t e e x p o r t from i l l u m i n a t e d p r i m a r y l e a v e s would a f f e c t t h e r a t e of s e n e s c e n c e w i t h r e s p e c t t o c a r b o n 130 TABLE 5.2. S p e c i f i c l e a f weight and s t a r c h conten t of the pr imary l eaves of c o n t r o l (340 u l l " 1 ) and e n r i c h e d (1400 u l l " 1 ) p l a n t s at two temperatures and two l e v e l s of p h o t o s y n t h e t i c photon f l u x d e n s i t y . 1 340 u l I"1 1400 u l 1- 1 T PPFD SLW S t a r ch SLW S t a r ch (C) (umol m~2 (g m" 2 ) (g m" 2) (g m" 2) (g m" 2) s- 1 ) 32 340 33 ±2 1 .4 ±0.9 65 ±3 25.9 ±3.5 32 100 23 ±1 0.2 ±0.0 27 ±0 0.8 ±0.5 22 340 48 ±2 15.1 ±0.6 78 ±0 46.8 ±1.9 22 100 36 ±0 3.0 ±0.8 45 ±3 10.9 ±0.8 1 Va lues are the means of 3 measurements, w i th s tandard e r r o r 131 FIGURE 5.7 R e l a t i o n s h i p between s t a r c h content and s p e c i f i c l e a f weight of the pr imary l e a v e s . P l an t s were grown at e i t h e r 340 or 1400 u l l " 1 carbon d i o x i d e , and at e i t h e r 32 or 20 C. Pr imary l eaves were ma in ta ined at e i t h e r 90 or 370 umol n r 2 s~ 1 PPFD. Leaves were sampled a f t e r 6 days of t reatment The curve was c o m p u t e r - f i t t e d to a t h i r d -order po l ynomia l e q u a t i o n , u s i n g 13 data p o i n t s . R 2 =0.98 1 32 d i o x i d e c o n c e n t r a t i o n . I n o n e s e t o f e x p e r i m e n t s , t h e p e t i o l e s o f m a t u r e p r i m a r y l e a v e s w e r e h e a t - g i r d l e d t o k i l l t h e p h l o e m . G i r d l i n g c a u s e d a c c e l e r a t e d a b s c i s s i o n o f l e a v e s i n b o t h c a r b o n d i o x i d e t r e a t m e n t s a n d s o t h i s r e s u l t i n r e l a t i n g e x p o r t t o s e n e s c e n c e w a s i n c o n c l u s i v e . I n a s e c o n d e x p e r i m e n t , m a t u r e p r i m a r y l e a v e s w e r e d e t a c h e d f r o m t h e p l a n t a n d t r e a t e d w i t h e i t h e r 3 4 0 o r 3 0 0 0 u l l " 1 c a r b o n d i o x i d e a t 2 3 C . P r e l i m i n a r y r e s u l t s s h o w e d a m o r e r a p i d s e n e s c e n c e a n d g r e a t e r SLW i n t h e h i g h c a r b o n d i o x i d e l e a v e s ( d a t a n o t p r e s e n t e d ) . M o s t s t u d i e s o f s e n e s c e n c e e m p l o y d e t a c h e d l e a v e s o r l e a f d i s k s , b u t t h e s e n e s c e n c e - r e l a t e d b i o c h e m i s t r y o f e x c i s e d m a t e r i a l d i f f e r s f r o m t h a t o f a t t a c h e d l e a v e s ( L e w i n g t o n e t . a l . , 1 9 6 7 ) . S u c h d i f f e r e n c e s may b e d u e t o t h e . d i s r u p t i o n o f n o r m a l t r a n s p o r t p r o c e s s e s , t o a n d f r o m t h e l e a f , w h i c h a l t e r s h o r m o n a l a n d m e t a b o l i t e s t a t u s . T h i s i n f l u e n c e s b o t h t h e o n s e t o f s e n e s c e n c e a n d t h e e x p o r t o f a m i n o a c i d s a n d o t h e r p r o d u c t s d u r i n g s e n e s c e n c e ( S p e n c e r a n d T i t u s , 1 9 7 3 ) . N e v e r t h e l e s s , i t i s i n t e r e s t i n g t o n o t e t h a t c a r b o n d i o x i d e a c c e l e r a t e d s e n e s c e n c e i n b o t h a t t a c h e d a n d d e t a c h e d l e a v e s . A p p a r e n t l y , a n y c h a n g e s i n c u r r e d b y r e m o v a l f r o m t h e p l a n t d i d n o t u n d e r m i n e t h e r e l a t i o n s h i p b e t w e e n c a r b o n d i o x i d e a n d s e n e s c e n c e . C a r b o n d i o x i d e t h e r e f o r e h a d a f u n d a m e n t a l e f f e c t o n s e n e s c e n c e p e r s e , s e p a r a t e f r o m a n y p o s s i b l e e f f e c t o n t r a n s p o r t p r o c e s s e s w h i c h c o u l d i n d i r e c t l y i n f l u e n c e s e n e s c e n c e . T h e p o s s i b l e e f f e c t o f h i g h c a r b o n d i o x i d e o n t h e p r o d u c t i o n o r t r a n s p o r t o f g r o w t h r e g u l a t o r s f r o m o t h e r p l a n t 133 p a r t s i s a p p a r e n t l y not i n v o l v e d i n t h e c a r b o n d i o x i d e - i n d u c e d s e n e s c e n c e p r o c e s s . The p o s s i b i l i t y t h a t c a r b o n d i o x i d e i n d u c e d s e n e s c e n c e t h r o u g h a d e l e t e r i o u s a c c u m u l a t i o n of some p r o d u c t of p h o t o s y n t h e s i s i s s u p p o r t e d by o t h e r s t u d i e s r e l a t i n g c a r b o h y d r a t e l e v e l s t o s e n e s c e n c e . F o r example, l e a f d i s k s of X a n t h i u m f l o a t e d on 0.01M s u c r o s e s o l u t i o n s a t 3000 Lux l o s t 90% o f t h e i r c h l o r o p h y l l compared t o 60% l o s s i n c o n t r o l s f l o a t e d on water ( K h u d a i r i , 1970). The s u c r o s e e f f e c t was l i g h t and t e m p e r a t u r e d e p e n d e n t , and was n o t due t o an o s m o t i c e f f e c t of t h e s u c r o s e s o l u t i o n s , w h i c h had a s o l u t e p o t e n t i a l o f o n l y -3.0 k i l o p a s c a l s ( k P a ) . S i m i l a r l y , Moore e t a l . , (1974) f o u n d t h a t e x c i s e d S i n a p i s c o t y l e d o n s f l o a t e d on s u c r o s e s o l u t i o n s had g r e a t e r c h l o r o p h y l l l o s s , as w e l l as a g r e a t e r c a r b o h y d r a t e c o n t e n t , t h a n c o n t r o l s . S t a r c h c o n t e n t a l s o c o r r e l a t e s w i t h s e n e c e n c e . W i t t e n b a c h e t a l . , (1980) d e s c r i b e d an i n c r e a s e i n s t a r c h c o n t e n t c o i n c i d e n t w i t h t h e o n s e t of s e n e s c e n c e i n mature soybean l e a v e s , w h i c h was a s s o c i a t e d w i t h a d i s o r i e n t a t i o n of c h l o r o p l a s t l a m e l l a e p r e c e e d i n g a d e c l i n e i n p h o t o s y n t h e s i s , c h l o r o p h y l l c o n t e n t , and t o t a l l e a f n i t r o g e n c o n t e n t , and an i n c r e a s e i n p r o t e o l y t i c a c t i v i t y . S c h o o l a r and Edelman (1970) f o u n d t h a t i f s t a r c h s y n t h e s i s was p r e v e n t e d by a p p l i c a t i o n of i o d o a c e t a t e , t h e l i g h t d e p e n d e n t d e c r e a s e i n c h l o r o p h y l l c o n t e n t was s l o w e r . I t i s i n t e r e s t i n g t o s p e c u l a t e on how a c c u m u l a t e d c a r b o n may i n d u c e s e n e s c e n c e o f l e a v e s . W i t h e n h a n c e d p h o t o s y n t h e s i s and a c c u m u l a t i o n of f i x e d c a r b o n , one m i g h t e x p e c t a l o w e r 134 s o l u t e p o t e n t i a l . T h i s c o u l d i n d u c e s e n e s c e n c e i n a way s i m i l a r t o t h a t i n d u c e d by a w ater d e f i c i t . However', c a r b o n d i o x i d e -i n d u c e d s e n e s c e n c e d i d not seem t o be o s m o t i c i n n a t u r e , s i n c e t h e w a t e r p o t e n t i a l and s o l u t e p o t e n t i a l of l e a v e s sampled j u s t p r i o r t o t h e o n s e t of s e n e s c e n c e symptoms were not s i g n i f i c a n t l y d i f f e r e n t between c a r b o n d i o x i d e t r e a t m e n t s ( T a b l e 5 . 3 ) . A t h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n s , t h e c h l o r o p l a s t s p r e s u m a b l y assumed more of a s t o r a g e f u n c t i o n , w i t h p r o g r e s s i v e l y g r e a t e r a c c u m u l a t i o n s of s t a r c h . The d i s t o r t i o n of c h l o r o p l a s t s w h i c h r e s u l t s (Cave e t a l . , 1981; W u l f f and S t r a i n , 1982) c o u l d i n f l u e n c e t h e i n t e r c e p t i o n o f l i g h t , t h e d i f f u s i o n of c a r b o n d i o x i d e , o r c o u l d d i s r u p t t h e g r a n a . A l t e r n a t i v e l y , a m e t a b o l i c change c o u l d be i n v o l v e d . F o r example, Edelman and Hanson (1971) f o u n d t h a t s u c r o s e s u p p r e s s e s c h l o r o p l a s t d e v e l o p m e n t and c h l o r o p h y l l s y n t h e s i s i n a c a r r o t t i s s u e c u l t u r e w i t h low i n v e r t a s e a c t i v i t y . I t s h o u l d be k e p t i n mind, however, t h a t c a r b o n d i o x i d e may be i n f u e n c i n g s e n e s c e n c e by means o t h e r t h a n ( o r i n a d d i t i o n t o ) c a r b o h y d r a t e a c c u m u l a t i o n . C a r b o n d i o x i d e may c a u s e c h a n g e s i n t h e s e n s i t i v i t y t o g r o w t h r e g u l a t o r s , s u c h as e t h y l e n e , o r i n f l u e n c e t h e i r l e v e l s o r a c t i o n . P o s s i b l y c a r b o n d i o x i d e as b i c a r b o n a t e i n f l u e n c e s c e l l pH, t h e e x t e n t o f w h i c h would depend on t h e b u f f e r i n g c a p a c i t y o f t h e c e l l - ( s e e S m i t h and Raven, 1979) . 135 TABLE 5 .3 . Water and osmot ic p o t e n t i a l s , and s p e c i f i c l e a f l e a f weight of e n r i c h e d (1400 u l 1~1) and c o n t r o l (340 u l l " 1 ) p l a n t s grown f o r 7 days at 24 C . 1 Treatment Water p o t e n t i a l Osmotic p o t e n t i a l SLW (kPa) (kPa) (g m" 2) 340 u l I'1 -63 ±6 -100 ±17 59 ±4 1400 u l l " 1 -89 ±14 -121 ±13 1 13 ±6 1 Va lues are the means of 3 l e a v e s , w i th s t anda rd e r r o r . 1 36 5.3.6 Summary The i n d u c t i o n of l e a f s e n e s c e n c e by h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n s has not been p r e v i o u s l y c h a r a c t e r i z e d . C a r b o n d i o x i d e - i n d u c e d s e n e s c e n c e of p r i m a r y bean l e a v e s was p o s i t i v e l y c o r r e l a t e d w i t h l i g h t i n t e n s i t y ( P P FD). T r i f o l i a t e s a l s o showed a c c e l e r a t e d c h l o r o s i s , b ut o n l y a t c o o l t e m p e r a t u r e s ; s e n e s c e n c e of p r i m a r y l e a v e s may a l s o be i n f l u e n c e d by t e m p e r a t u r e but t h e d a t a were e q u i v o c a l on t h i s p o i n t . The f a c t t h a t h i g h e r PPFD c a u s e d a more r a p i d s e n e s c e n c e would s u g g e s t t h a t c a r b o n d i o x i d e - i n d u c e d s e n e s c e n c e was r e l a t e d t o t h e h i g h e r c a r b o n d i o x i d e exchange r a t e s of e n r i c h e d l e a v e s . F u r t h e r m o r e , t h o s e t r e a t m e n t s w h i c h c a u s e d t h e most r a p i d s e n e s c e n c e a l s o c a u s e d t h e g r e a t e s t SLW and l e a f s t a r c h c o n t e n t . 1 37 V I . GENERAL DISCUSSION By way of i n t r o d u c i n g a g e n e r a l d i s c u s s i o n of t h e f i n d i n g s of t h i s s t u d y , I w o u l d l i k e t o r e v i e w t h e major p o i n t s w h i c h were d e s c r i b e d . C o n t i n u o u s l o n g - t e r m c a r b o n d i o x i d e e n r i c h m e n t c a u s e d l a r g e i n c r e a s e s i n p l a n t d r y w e i g h t , w h i c h were r e f l e c t e d i n i n c r e a s e s i n g r o w t h r a t e compared t o u n e n r i c h e d p l a n t s . The i n c r e a s e s i n g r o w t h r a t e were d e p e n d e n t on i n c r e a s e s i n u n i t l e a f r a t e e a r l y i n d e v e l o p m e n t , and on i n c r e a s e s i n l e a f a r e a and t o a l e s s e r e x t e n t i n c r e a s e s i n u n i t l e a f r a t e , l a t e r i n d e v e l o p m e n t . The o b s e r v e d i n c r e a s e s i n u n i t l e a f r a t e o f whole p l a n t s were a l s o r e f l e c t e d i n i n c r e a s e s i n t h e net c a r b o n d i o x i d e exchange r a t e of i n d i v i d u a l l e a v e s o f t h o s e p l a n t s . In a d d i t i o n , t h e i n c r e a s e i n c a r b o n d i o x i d e exchange r a t e of e n r i c h e d l e a v e s c o r r e l a t e d w i t h a r e d u c e d c a r b o n d i o x i d e exchange c a p a c i t y and an i n c r e a s e d r e s p i r a t i o n r a t e i n t h o s e l e a v e s . An i n c r e a s e i n c a r b o n a c c u m u l a t i o n (measured, as an i n c r e a s e i n s p e c i f i c l e a f w e i g h t and an i n c r e a s e i n s t a r c h c o n t e n t ) , w h i c h r e s u l t e d , i n p a r t , f r o m t h e i n c r e a s e i n c a r b o n d i o x i d e e xchange r a t e of e n r i c h e d l e a v e s , c o r r e l a t e d w i t h a r e d u c t i o n i n c a r b o n d i o x i d e exchange c a p a c i t y of t h e f i r s t t r i f o l i a t e s , and a c c e l e r a t e d s e n e s c e n c e of t h e p r i m a r y l e a v e s . F u r t h e r m o r e , t h e c a r b o n d i o x i d e - i n d u c e d c h a n g e s i n r e l a t i v e g r o w t h r a t e and u n i t l e a f r a t e of whole p l a n t s , and t h e c h a n g e s i n c a r b o n d i o x i d e exchange c a p a c i t y , r e s p i r a t i o n r a t e , and s e n e s c e n c e of i n d i v i d u a l l e a v e s , were most e v i d e n t e a r l y i n p l a n t d e v e l o p m e n t . 138 In a number o f c a s e s , t h e m a g n i t u d e of a r e s p o n s e t o c a r b o n d i o x i d e e n r i c h m e n t c h a n g e d by i n c r e m e n t s w i t h an i n c r e a s i n g c a r b o n d i o x i d e c o n c e n t r a t i o n . The b e s t examples o f i n c r e m e n t a l change were t h e i n c r e a s e s i n a b s o l u t e g r o w t h r a t e , r e l a t i v e g r o w t h r a t e , and u n i t l e a f r a t e o f beans a t p r o g r e s s i v e l y g r e a t e r c a r b o n d i o x i d e c o n c e n t r a t i o n s ( C h a p t e r 3 ) . The i n c r e m e n t a l r e s p o n s e of t h o s e p a r a m e t e r s a p p e a r e d t o d e c r e a s e between 800 and 1200 u l l " 1 c a r b o n d i o x i d e , w h i c h s u g g e s t s an a p p r o a c h t o s a t u r a t i o n i n t h a t c o n c e n t r a t i o n r a n g e . C o m m e r c i a l g r e e n h o u s e s f r e q u e n t l y have e n r i c h e d a t m o s p h e r e s o f 1000 t o 1200 u l l " 1 c a r b o n d i o x i d e ( W i t t w e r and Robb, 1964). A c c o r d i n g t o my d a t a , a 1000 t o 1200 u l 1~ 1 c o n c e n t r a t i o n r a n g e would o p t i m i z e i n c r e a s e s i n g r o w t h w i t h a m i n i m a l c a r b o n d i o x i d e a d d i t i o n t o t h e g r e e n h o u s e a t m o s p h e r e . Bear i n mind t h a t e n v i r o n m e n t a l c o n d i t i o n s may m o d i f y t h e g r o w t h r e s p o n s e / c a r b o n d i o x i d e c o n c e n t r a t i o n r e l a t i o n s h i p . F o r example, b e c a u s e o f t h e s e v e r i t y of c a r b o n d i o x i d e - i n d u c e d s e n e s c e n c e a t c o o l t e m p e r a t u r e s ( C h a p t e r 5 ) , t h e growth r e s p o n s e a t 1000 t o 1200 u l l " 1 may be r e d u c e d when compared t o t h a t a t l o w e r c o n c e n t r a t i o n s . A c c e l e r a t e d l e a f s e n e s c e n c e i s an i m p o r t a t a n t c o n s i d e r a t i o n i n d e t e r m i n i n g optimum c a r b o n d i o x i d e c o n c e n t r a t i o n s f o r g r e e n h o u s e e n r i c h m e n t , p a r t i c u l a r l y a t c o o l t e m p e r a t u r e s . However, t h e d o s e - r e s p o n s e r e l a t i o n s h i p of c a r b o n d i o x i d e c o n c e n t r a t i o n and s e n e s c e n c e was not d e t e r m i n e d i n my s t u d y . The n e t c a r b o n d i o x i d e a s s i m i l a t i o n r a t e of l e a v e s 1 39 g e n e r a l l y i n c r e a s e s w i t h i n c r e a s i n g c a r b o n d i o x i d e c o n c e n t r a t i o n i f o t h e r e n v i r o n m e n t a l f a c t o r s a r e n o t l i m i t i n g ( G a a s t r a , 1963). The i m p o r t a n c e of c a r b o n d i o x i d e a s a d e t e r m i n a n t of net c a r b o n d i o x i d e a s s i m i l a t i o n r a t e c a n n o t be u n d e r e s t i m a t e d i n any s t u d y o f t h e e f f e c t s o f l o n g - t e r m c a r b o n d i o x i d e e n r i c h m e n t on p l a n t g r o w t h o r f u n c t i o n . The i n c r e a s e i n u n i t l e a f r a t e ( C h a p t e r 3) and CER ( C h a p t e r 4) of e n r i c h e d l e a v e s shows t h a t h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n r e s u l t e d i n an i n c r e a s e i n n e t c a r b o n d i o x i d e a s s i m i l a t i o n r a t e i n t h o s e p l a n t s . A b a s i c c o n c l u s i o n o f my s t u d y i s t h a t a h i g h e r n et c a r b o n d i o x i d e e x c h a n g e r a t e i n e n r i c h e d p l a n t s was r e s p o n s i b l e f o r many of t h e o b s e r v e d p h y s i o l o g i c a l e f f e c t s of e n r i c h m e n t . F o r example, an i n c r e a s e i n r e l a t i v e g r o w t h r a t e c o r r e l a t e d w i t h an i n c r e a s e i n u n i t l e a f r a t e ( C h a p t e r 3 ) . F u r t h e r m o r e , t h e h i g h e r CER o f e n r i c h e d l e a v e s c o r r e l a t e d w i t h a r e d u c t i o n i n c a r b o n d i o x i d e e x c h a n g e c a p a c i t y and an i n c r e a s e i n r e s p i r a t i o n r a t e compared t o c o n t r o l s ( C h a p t e r 4 ) . I f t h e d i f f e r e n c e i n CER between c o n t r o l and e n r i c h e d p l a n t s was r e d u c e d by s h a d i n g , o r by low c h l o r o p h y l l l e v e l s ( C h a p t e r 4 ) , t h e e f f e c t i v e n e s s of e n r i c h m e n t i n a l t e r i n g c a r b o n d i o x i d e exchange c a p a c i t y and d a r k r e s p i r a t i o n r a t e was d i m i n i s h e d . The e f f e c t of c a r b o n d i o x i d e e n r i c h m e n t on s e n e s c e n c e was a l s o d e p e n d e n t on t h e l e v e l of PPFD, once a g a i n showing a p r o b a b l e r e l a t i o n s h i p between e n h a n c e d CER and a c a r b o n d i o x i d e e n r i c h m e n t r e s p o n s e . S i m i l a r l y , a l i m i t a t i o n o f u n i t l e a f r a t e by low l i g h t i n t e n s i t y d u r i n g t h e l a t t e r s t a g e s of d e v e l o p m e n t p r o b a b l y 1 40 r e d u c e d t h e e f f e c t i v e n e s s of c a r b o n d i o x i d e e n r i c h m e n t i-n i n c r e a s i n g u n i t l e a f r a t e of whole p l a n t s ( C h a p t e r 3 ) . A t t e n u a t i o n o f l i g h t i n t e n s i t y t h r o u g h a s h o o t c a n o p y i s l o g a r i t h m i c i n n a t u r e , and hence t h e u n i t l e a f r a t e o f much of a w e l l - d e v e l o p e d s h o o t canopy may have been l i m i t e d by l i g h t i n t e n s i t y . In my s t u d y , t h e r e l a t i v e g r o w t h r a t e of b o t h e n r i c h e d and u n e n r i c h e d p l a n t s d e c r e a s e d w i t h m a t u r i t y , p r e s u m a b l y b e c a u s e an i n c r e a s i n g p r o p o r t i o n of t h e l e a v e s were f u n c t i o n i n g a t r e d u c e d l i g h t l e v e l s . At t h o s e i n t e n s i t i e s , t h e e f f e c t i v e n e s s of h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n i n i n c r e a s i n g u n i t l e a f r a t e was d i m i n i s h e d . T h i s i n t e r p r e t a t i o n i s s u p p o r t e d by d a t a on t h e r e s p o n s e of i n d i v i d u a l l e a v e s t o i n c r e a s i n g l i g h t i n t e n s i t y r e p o r t e d i n C h a p t e r 4. At low PPFD, t h e a b s o l u t e i n c r e a s e i n CER w i t h i n c r e a s e d c a r b o n d i o x i d e c o n c e n t r a t i o n was much l e s s t h a n t h e a b s o l u t e i n c r e a s e i n CER a t h i g h .PPFD. A l i g h t - c a r b o n d i o x i d e i n t e r a c t i o n has been o b s e r v e d p r e v i o u s l y , i n t h a t t h e e f f e c t of c a r b o n d i o x i d e was p r o g r e s s i v e l y g r e a t e r w i t h i n c r e a s i n g l i g h t i n t e n s i t y (Hopen and R i e s , 1962; F o r d and T h o r n e , 1967; Hughes and C o c k s h u l l , 1971). However, t h e p o s s i b i l i t y t h a t t h e s h a d i n g o f l e a v e s d u r i n g d e v e l o p m e n t was r e s p o n s i b l e f o r t h e d i m i n i s h i n g e f f e c t o f e n r i c h m e n t on r e l a t i v e g r o w t h r a t e has not been d i s c u s s e d i n t h e l i t e r a t u r e . My s t u d y t h e r e f o r e s u g g e s t s a number o f r e a s o n s why t h e g r o w t h of whole p l a n t s d i d not r e s p o n d t o e n r i c h m e n t as would be p r e d i c t e d f r o m s h o r t - t e r m c a r b o n d i o x i d e - r e s p o n s e c u r v e s of n e t p h o t o s y n t h e s i s r a t e . F i r s t , when CER was not l i m i t e d by l i g h t i n t e n s i t y , t h e n e t c a r b o n d i o x i d e exchange c a p a c i t y o f e n r i c h e d 141 l e a v e s was r e d u c e d compared t o t h e CER o f c o n t r o l l e a v e s . S e c o n d , when l i g h t i n t e n s i t y i n t h e canopy was low, t h e a b s o l u t e i n c r e a s e i n u n i t l e a f r a t e a t h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n was d i m i n i s h e d , t h e r e b y c o n t r i b u t i n g t o a l o w e r r e l a t i v e g r o w t h r a t e . T h i r d , a c c e l e r a t e d s e n e s c e n c e of l e a v e s of e n r i c h e d p l a n t s p r o b a b l y r e d u c e d t h e l e a f s u r f a c e a r e a a v a i l a b l e f o r n e t c a r b o n d i o x i d e e x c h a n g e . An o f t e n o v e r l o o k e d f a c t o r i n d e t e r m i n i n g t h e r e s p o n s e of u n i t l e a f r a t e t o c a r b o n d i o x i d e c o n c e n t r a t i o n i s t h e d e g r e e o f a i r t u r b u l e n c e w i t h i n t h e p l a n t c a n o p y . In a l l e x p e r i m e n t a l s i t u a t i o n s o u t l i n e d i n my s t u d y , chamber a i r was w e l l m i x e d . However, v e n t i l a t i o n c an be a c o n f o u n d i n g i n f l u e n c e when e x t r a p o l a t i n g t h e r e s p o n s e o f CER, o b t a i n e d f r o m gas exchange measurements i n w e l l - v e n t i l a t e d chambers or c u v e t t e s , t o a whole p l a n t or c r o p s i t u a t i o n . P r o f i l e s of c a r b o n d i o x i d e c o n c e n t r a t i o n w i t h i n an e n r i c h e d s h o o t canopy show g r a d i e n t s i n c o n c e n t r a t i o n w h i c h a r e d e p e n d e n t on t h e p o s i t i o n i n t h e canopy, and on p l a n t d e n s i t y (Duncan and B a r f i e l d , 1968). Hence, t h e c a r b o n d i o x i d e c o n c e n t r a t i o n a t t h e l e a f s u r f a c e may v a r y s u b s t a n t i a l l y w i t h l e a f p o s i t i o n or p l a n t d e n s i t y . As a r e s u l t , c a r b o n d i o x i d e r e s p o n s e c u r v e s g e n e r a t e d i n w e l l - v e n t i l a t e d measurement chambers may be q u i t e d i f f e r e n t t h a n t h o s e i n a f i e l d s i t u a t i o n . In a p o o r l y v e n t i l a t e d c r o p , I s u s p e c t t h a t u n i t l e a f r a t e would a p p r o a c h ' s a t u r a t i o n a t a h i g h e r c a r b o n d i o x i d e c o n c e n t r a t i o n t h a n one would e x p e c t f r o m l a b o r a t o r y m easurements. The e f f e c t i v e n e s s o f h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n i n 142 i n c r e a s i n g p l a n t g r o w t h , w h i c h i s c o n c u r r e n t l y l i m i t e d by o t h e r e n v i r o n m e n t a l f a c t o r s , s u c h a s l i g h t i n t e n s i t y ( C h a p t e r s 3 and 4 ) , has r e l e v a n c e a g r o n o m i c a l l y . Water and n u t r i e n t a v a i l a b i l i t y may a l s o l i m i t t h e g r o w t h p o t e n t i a l o f v e g e t a t i o n i n many a r e a s , but l i t t l e i n f o r m a t i o n i s a v a i l a b l e on t h e e f f e c t s of e n r i c h m e n t on g r o w t h when t h o s e c o n d i t i o n s a r e l i m i t i n g . In d e s c r i b i n g t h e e f f e c t o f c a r b o n d i o x i d e e n r i c h m e n t on g r o w t h w h i c h i s l i m i t e d by o t h e r f a c t o r s , t h e e f f e c t s of c a r b o n d i o x i d e on u n i t l e a f r a t e must be s e p a r a t e d from e f f e c t s on l e a f a r e a . F o r example, g r o w t h o f c o t t o n was i n c r e a s e d by c a r b o n d i o x i d e e n r i c h m e n t a t low n i t r a t e l e v e l s o n l y b e c a u s e o f an i n c r e a s e i n l e a f a r e a ; u n i t l e a f r a t e was n o t i n c r e a s e d by e n r i c h m e n t (Wong, 1979). C a r b o n d i o x i d e e n r i c h m e n t , a l s o a p p e a r s t o be of use i n i m p r o v i n g p l a n t g r o w t h under c o n d i t i o n s of low w a t e r a v a i l a b i l i t y . Growth o f wheat under w a t e r s t r e s s c o n d i t i o n s was b e t t e r i n e n r i c h e d t h a n u n e n r i c h e d p l a n t s ( G i f f o r d , 1979). When s t r e s s e d , e n r i c h e d p l a n t s showed g r e a t e r r e l a t i v e i n c r e a s e s i n g r o w t h t h a n u n e n r i c h e d p l a n t s , compared t o an u n s t r e s s s i t u a t i o n , a l t h o u g h t h e a b s o l u t e i n c r e a s e s i n growth were s m a l l e r . S i o n i t e t a l . (1980) f o u n d t h a t t h e b e t t e r g r o w t h o f e n r i c h e d compared t o u n e n r i c h e d wheat under water s t r e s s , a p p e a r e d t o be due t o b e t t e r t u r g o r m a i n t e n a n c e , as a r e s u l t o f l o w e r o s m o t i c p o t e n t i a l s i n e n r i c h e d p l a n t s . T h i s a l s o a l l o w e d b e t t e r s t o m a t a l o p e n i n g a t low w a t e r p o t e n t i a l s ( S i o n i t e t a l . , 1981b). 143 In summary, c a r b o n d i o x i d e e n r i c h m e n t a p p e a r s t o r e s u l t i n a g r o w t h i n c r e m e n t even when o t h e r e n v i r o n m e n t a l f a c t o r s may be l i m i t i n g . However, i t i s i m p o r t a n t t o r e c o g n i z e t h a t t h e g r e a t e s t a b s o l u t e g r o w t h i n c r e m e n t s w i l l r e s u l t when o t h e r e n v i r o n m e n t a l f a c t o r s do n o t c o n s t r a i n g r o w t h . I f g r o w t h i s r e s t r i c t e d by a s t r e s s , t h e r e l a t i v e i n c r e a s e i n g r o w t h due t o e n r i c h m e n t may be much l a r g e r t h a n t h e a b s o l u t e i n c r e a s e i n g r o w t h . T h i s i n c r e a s e i n g r o w t h p o t e n t i a l under r a t e - l i m i t i n g c o n d i t i o n s i s a r e s u l t o f t h e n a t u r e o f t h e i n v o l v e m e n t o f c a r b o n d i o x i d e i n p h o t o s y n t h e s i s . H i g h c a r b o n d i o x i d e c o n c e n t r a t i o n d e c r e a s e s t h e p e r c e n t a g e i n h i b i t i o n of t r u e p h o t o s y n t h e s i s c a u s e d by p h o t o r e s p i r a t i o n (Fock e t a l . , 1979), and t h e r e b y i n c r e a s e s t h e n e t p h o t o s y n t h e t i c r a t e , even when o t h e r e n v i r o n m e n t a l f a c t o r s may be l i m i t i n g . I t i s i n t e r e s t i n g t o n o t e t h a t t h e i n c r e m e n t s i n growth due t o e n r i c h m e n t may be tempered, not o n l y by o t h e r e v i r o n m e n t a l c o n s t r a i n t s , b u t a l s o by t h e e f f e c t of h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n s on s t o m a t a l a p e r t u r e . S t o m a t a l c l o s u r e a t h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n s would i n c r e a s e water use e f f i c i e n c y , b u t may c o n c u r r e n t l y l i m i t t h e g r o w t h r e s p o n s e . S t o m a t a l c l o s u r e i n r e s p o n s e t o c a r b o n d i o x i d e r e s u l t s i n one of two s i t u a t i o n s w i t h r e s p e c t t o n e t p h o t o s y n t h e s i s (van K e u l e n et. a l . , 1980). In t h e f i r s t i n s t a n c e , s t o m a t a l c l o s u r e i s s u f f i c i e n t t o o f f s e t an i n c r e a s e i n i n t e r c e l l u l a r c a r b o n d i o x i d e c o n c e n t r a t i o n , r e s u l t i n g i n no i n c r e a s e i n n e t p h o t o s y n t h e s i s , but a d e c r e a s e i n t r a n s p i r a t i o n . Hence, water use e f f i c i e n c y i n c r e a s e s . In t h i s c a s e , s t o m a t a l a p e r t u r e would a p p e a r t o 1 44 r e g u l a t e C i . The m a i n t e n a n c e of a c o n s t a n t C i has a l s o been o b s e r v e d when e x t r a c e l l u l a r c a r b o n d i o x i d e i s h e l d c o n s t a n t but t h e p h o t o s y n t h e t i c r a t e i s v a r i e d ; s t o m a t a l a p e r t u r e a d j u s t s w i t h c h a n g e s i n p h o t o s y n t h e s i s r a t e (Wong e t a l . , 1979). In t h e s e c o n d c a s e , s t o m a t a l a p e r t u r e d o e s not r e g u l a t e C i and an i n c r e a s e i n e x t r a c e l l u l a r c a r b o n d i o x i d e w i l l d e c r e a s e t r a n s p i r a t i o n r a t e , but i n c r e a s e n e t p h o t o s y n t h e s i s r a t e . A g a i n w ater use e f f i c i e n c y i n c r e a s e s , but b o t h t r a n s p i r a t i o n r a t e and n e t a s s i m i l a t i o n r a t e have c h a n g e d . In my s t u d y , beans showed s t o m a t a l c l o s u r e ( i . e . i n c r e a s e d s t o m a t a l r e s i s t a n c e ) i n r e s p o n s e t o h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n s , when measured a t l i g h t i n t e n s i t i e s a p p r o a c h i n g s a t u r a t i o n (1000 umol n r 2 s ~ 1 PPFD). However, C i a p p e a r e d not t o be r e g u l a t e d by s t o m a t a l a p e r t u r e , and i n c r e a s e s i n C i r e s u l t e d i n a g r e a t e r CER. A l s o , s t o m a t a l • r e s i s t a n c e m e asured i n p l a n t s u nder g rowth c o n d i t i o n s (350 umol i r r 2 s " 1 PPFD) was n o t a f f e c t e d by c a r b o n d i o x i d e c o n c e n t r a t i o n ( C h a p t e r 5 ) . T h e r e a l s o a p p e a r e d t o be some a c c l i m a t i o n of s t o m a t a t o c o n t i n u o u s c a r b o n d i o x i d e e n r i c h m e n t ( C h a p t e r 4 ) , w h i c h would s e r v e t o m a i n t a i n somewhat g r e a t e r s t o m a t a l o p e n i n g a t h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n s . T h e r e f o r e , s t o m a t a l c l o s u r e i n beans was not an i m p o r t a n t f a c t o r i n l i m i t i n g t h e g r o w t h r e s p o n s e t o e n r i c h m e n t . As p r e v i o u s l y e n r i c h m e n t e f f e c t s a r e l a t i v e l y h i g h e r o u t l i n e d , many of t h e c a r b o n d i o x i d e d e s c r i b e d i n t h i s s t u d y were a s s o c i a t e d w i t h CER i n e n r i c h e d t h a n c o n t r o l p l a n t s . 1 45 One p o s s i b l e way a higher CER c o u l d i n f l u e n c e carbon d i o x i d e exchange c a p a c i t y , r e s p i r a t i o n r a t e , or senescence of en r i c h e d p l a n t s , i s through an i n c r e a s e i n the carbon a s s i m i l a t i o n / c a r b o n export r a t i o of source l e a v e s , r e s u l t i n g i n an i n c r e a s e i n the l e a f carbohydrate s t a t u s . A high SLW and s t a r c h content c o r r e l a t e d with a l l those responses to enr ichment. The r e l a t i o n s h i p between s t a r c h content and p h o t o s y n t h e t i c c a p a c i t y has been debated e x t e n s i v e l y i n the l i t e r a t u r e . A negative r e l a t i o n s h i p between s t a r c h content and p h o t o s y n t h e t i c c a p a c i t y was observed by N a f z i g e r and R o l l e r (1976) with s h o r t -term (16 hour) carbon d i o x i d e enrichment of soybean. Mauney e_t a l . (1979) and H o f s t r a and Hesketh (1975) have observed the same c o r r e l a t i o n i n long-term enrichment s t u d i e s . In f a c t , the negative c o r r e l a t i o n between s t a r c h content and p h o t o s y n t h e t i c c a p a c i t y i s evident i n a l l long-term carbon d i o x i d e enrichment s t u d i e s to date. The r e l a t i o n s h i p between s t a r c h content and p h o t o s y n t h e t i c c a p a c i t y has a l s o been p o s t u l a t e d as an i n t e g r a t o r of sink and source a c t i v i t y i n p l a n t s . I t has been suggested that sink demand c o n t r o l s source l e a f p h o t o s y n t h e s i s through an e f f e c t on s t a r c h content of source l e a v e s . For example, i f sink s t r e n g t h decreases r e l a t i v e to source s t r e n g t h , s t a r c h would presumably accumulate i n source l e a v e s , and i n h i b i t p h o t o s y n t h e s i s i n those l e a v e s . Although the r e l a t i o n s h i p has been documented i n many s t u d i e s (Herold, 1980), others do not f i n d a c o r r e l a t i o n , p a r t i c u l a r l y i n short-term s t u d i e s (Geiger, 1976; P o t t e r and 1 46 B r e e n , 1980; Fondy and G e i g e r , 1982). I t has been s u g g e s t e d (Mauney e t a l . , 1979) t h a t where no a p p a r e n t r e l a t i o n s h i p between s t a r c h c o n t e n t and p h o t o s y n t h e t i c c a p a c i t y i s a p p a r e n t , t h e s t a r c h l e v e l s were not h i g h enough. However, i n a s t u d y o f s o y b e a n s e x p o s e d t o c o n t i n u o u s l i g h t f o r 54 h o u r s , t h e r e was no d e c l i n e i n p h o t o s y n t h e s i s , d e s p i t e a 14-f o l d i n c r e a s e i n s t a r c h c o n t e n t , w h i c h s u g g e s t s t h a t t h e r e was no r e l a t i o n s h i p ( P o t t e r and B r e e n , 1980). D a t a o f C h a p t e r 4 may be r e l e v a n t i n t h i s d i s c u s s i o n . T h e r e a p p e a r s t o be a n e g a t i v e r e l a t i o n s h i p between s t a r c h c o n t e n t and t h e maximum c a r b o n d i o x i d e e xchange c a p a c i t y . However, c h a n g e s i n c a r b o n d i o x i d e e x change c a p a c i t y w i t h i n t h e l i m i t s d e t e r m i n e d by s t a r c h c o n t e n t were e v i d e n t l y i n d e p e n d e n t of s t a r c h l e v e l s . W i t h t h i s d i s c u s s i o n i n mind, I would l i k e t o r e v i e w s e v e r a l e f f e c t s of c a r b o n d i o x i d e e n r i c h m e n t on l e a f p h y s i o l o g y w h i c h were o b s e r v e d i n t h i s s t u d y . F i r s t , c a r b o n d i o x i d e e n r i c h m e n t was l e s s e f f e c t i v e i n d e c r e a s i n g t h e c a r b o n d i o x i d e exchange c a p a c i t y o f l e a v e s i f c h l o r o p h y l l c o n t e n t of t h e l e a v e s was low, o r t h e l e v e l of PPFD was l i m i t i n g . T h i s r e d u c e d t h e CER of e n r i c h e d , as w e l l as c o n t r o l p l a n t s , but a l s o d e c r e a s e d t h e d i f f e r e n c e i n CER between t h o s e t r e a t m e n t s . The r e l a t i o n s h i p between p h o t o s y n t h e t i c r a t e and p h o t o s y n t h e t i c c a p a c i t y i n e n r i c h e d p l a n t s has not r e c e i v e d much a t t e n t i o n i n t h e l i t e r a t u r e . At low CER, a more f a v o r a b l e l e a f c a r b o h y d r a t e b a l a n c e would p r e s u m a b l y be a c h e i v e d , and h i g h c a r b o n d i o x i d e would t h e r e f o r e be l e s s e f f e c t i v e i n r e d u c i n g c a r b o n d i o x i d e exchange c a p a c i t y . 147 In t h i s c o n t e x t , I c o n s i d e r t h e r e l a t i o n s h i p between c h l o r o p h y l l c o n t e n t and c a r b o n d i o x i d e exchange c a p a c i t y t o be p a r t i c u l a r l y n o t e w o r t h y . When g r o w t h , and hence s i n k demand, was s e v e r e l y r e s t r i c t e d i n p l a n t s g r o w i n g i n s m a l l c o n t a i n e r s ( C h a p t e r 4 ) , t h e lo w e r c h l o r o p h y l l l e v e l s f o u n d i n s o u r c e l e a v e s would l i k e l y r e d u c e c a r b o n a c c u m u l a t i o n i n t h o s e l e a v e s . A r e d u c t i o n i n c a r b o n d i o x i d e e xchange c a p a c i t y , a t l e a s t p e r u n i t c h l o r o p h y l l , whould t h e r e f o r e be l e s s l i k e l y t o o c c u r . To t h e b e s t of my knowledge, t h i s r e l a t i o n s h i p has n o t been d i s c u s s e d i n t h e l i t e r a t u r e . An i n c r e a s e i n s i n k demand d i d n o t a l l e v i a t e t h e d i f f e r e n c e i n c a r b o n d i o x i d e exchange c a p a c i t y among e n r i c h e d and c o n t r o l t r e a t m e n t s , s u g g e s t i n g t h a t t h e l e a f c a r b o n b a l a n c e was n o t s u f f i c i e n t l y a f f e c t e d , p e r h a p s b e c a u s e of a c o n s t a n t s u p p l y of new a s s i m i l a t e . In f a c t , e n r i c h e d p l a n t s d i d not r e s p o n d as r e a d i l y as c o n t r o l s t o a change i n s i n k demand i n terms of an i n c r e a s e d c a r b o n d i o x i d e e x c h a n g e c a p a c i t y o r r e d u c e d s t a r c h c o n t e n t . E n r i c h e d p l a n t s were, t h e r e f o r e , n ot as f l e x i b l e i n a d j u s t i n g s o u r c e l e a f c a r b o n d i o x i d e exchange c a p a c i t y t o a new demand s i t u a t i o n . T h i s has not been o b s e r v e d b e f o r e . R e p r o d u c t i v e g r o w t h , a c t i n g t h r o u g h an i n c r e a s e d s i n k demand, i s known t o i n c r e a s e CER i n s o u r c e l e a v e s ( G i f f o r d and E v a n s , 1981). A l t h o u g h r e p r o d u c t i v e g r o w t h may c o n t r i b u t e t o a l o w e r SLW i n e n r i c h e d p l a n t s ( s e e C h a p t e r 3 ) , i t i s n o t known whether t h e o n s e t of r e p r o d u c t i v e g r o w t h i n f l u e n c e s t h e t h e c a r b o n d i o x i d e exchange c a p a c i t y of t h o s e p l a n t s . C a r b o n d i o x i d e - i n d u c e d s e n e s c e n c e was a l s o a s s o c i a t e d w i t h 1 48 h i g h c a r b o n r e t e n t i o n i n t h e l e a v e s ; SLW and s t a r c h c o n t e n t were e x t r e m e l y h i g h i n t h o s e l e a v e s w h i c h were t h e most r a p i d l y s e n e s c e n t . I n c r e a s i n g SLW, e i t h e r by l o w e r i n g t h e t e m p e r a t u r e or by i n c r e a s i n g net a s s i m i l a t i o n a t a h i g h e r l i g h t i n t e n s i t y , r e s u l t e d i n a more r a p i d s e n e s c e n c e o f e n r i c h e d p l a n t s . Here a g a i n , as i n t h e r e l a t i o n s h i p between s t a r c h c o n t e n t and c a r b o n d i o x i d e e xchange c a p a c i t y , one must be c a u t i o u s i n s u g g e s t i n g t h a t a h i g h s t a r c h c o n t e n t c a u s e s s e n e s c e n c e . Even s o , I f e e l i t i s p h y s i o l o g i c a l l y s i g n i f i c a n t t h a t t h e c o r r e l a t i o n o c c u r r e d i r r e s p e c t i v e of t h e manner i n w h i c h s e n e s c e n c e was i n d u c e d . C a r b o n d i o x i d e was v e r y e f f e c t i v e i n i n d u c i n g s e n e s c e n c e i n p r i m a r y l e a v e s , w h i c h p r o b a b l y r e l a t e d t o t h e f a c t t h a t t h o s e l e a v e s a t t a i n e d t h e h i g h e s t SLW ( C h a p t e r 3 ) . C a r b o n d i o x i d e i n d u c e d s e n e s c e n c e a t 1400 u l l " 1 , w h i c h i s u n u s u a l l y low when compared t o t h e h i g h c o n c e n t r a t i o n s ( >10000 u l 1~ 1) r e q u i r e d i n o t h e r d e v e l o p m e n t a l p r o c e s s e s , s u c h as i n t e r a c t i o n w i t h e t h y l e n e ( A b e l e s , 1973) or i n f l o w e r i n d u c t i o n i n P h a r b i t i s n i l under a n o n - i n d u c t i v e l o n g - d a y p h o t o p e r i o d ( H i c k l e n t o n and J o l l i f f e , 1980a). T h i s l a r g e d i f f e r e n c e i n c o n c e n t r a t i o n s u g g e s t s t h a t t h e mode of a c t i o n of c a r b o n d i o x i d e i n t h e i n d u c t i o n of s e n e s c e n c e may be u n i q u e , compared t o i t s r o l e i n o t h e r d e v e l o p m e n t a l p r o c e s s e s . An i n c r e a s e i n SLW may be an i n d i c a t i o n t h a t t h e p l a n t was u n a b l e t o use c a r b o n a t a r a t e s u f f i c i e n t t o keep pace w i t h i t s a s s i m i l a t i o n . C o n s e q u e n t l y , s e n e s c e n c e of t h o s e l e a v e s was a means of r e d u c i n g t o t a l l e a f a r e a and t h e r e b y a c h i e v i n g a more e q u i t a b l e b a l a n c e of s u p p l y and demand. Hence, s e n e s c e n c e i n t h i s c a s e was not as much a 1 49 f u n c t i o n of age as i t was of an a l t e r e d c a r b o n b a l a n c e , a r e l a t i o n s h i p not w i d e l y r e c o g n i z e d i n t h e l i t e r a t u r e . E v i d e n t l y , t h o s e f a c t o r s w h i c h . i n d u c e d s e n e s c e n c e were a l s o r e s p o n s i b l e f o r a s h i f t i n l e a f s t a r c h m e t a b o l i s m , e i t h e r b e c a u s e o f a change i n c a r b o n p a r t i t i o n i n g , o r s i m p l y b e c a u s e of g r e a t e r l e a f c a r b o n a c c u m u l a t i o n under t h o s e c o n d i t i o n s ( h e r e , a c c u m u l a t i o n / e x p o r t r e m a i n s t h e same). In t h i s r e s p e c t , s o u r c e / s i n k r e l a t i o n s h i p s may be q u i t e i m p o r t a n t i n d e t e r m i n i n g t h e i n d u c t i o n or s u b s e q u e n t r a t e of s e n e s c e n c e . U s i n g a t e c h n i q u e of s u r g i c a l l y m a n i p u l a t i n g t r a n s p o r t from t h e f l a g l e a f of wheat, L a z a n e t a l . , (1983) showed t h a t r e s t r i c t e d e x p o r t of c a r b o n and n i t r o g e n p r o m o t e d s e n e s c e n c e . The o n s e t of s e n e s c e n c e was a s s o c i a t e d w i t h t h e a t t a i n m e n t of a t h r e s h o l d l e v e l o f e t h a n o l - s o l u b l e c a r b o h y d r a t e . S i m i l a r l y , t h e e f f e c t o f t e m p e r a t u r e on s o u r c e / s i n k r e l a t i o n s h i p s may be i m p o r t a n t i n c a r b o n d i o x i d e - i n d u c e d s e n e s c e n c e . P l a n t g r o w t h was much s l o w e r a t 20 C t h a n a t 32 C, and t h i s was p r e s u m a b l y a s s o c i a t e d w i t h a l o w e r demand f o r a s s i m i l a t e s by d e v e l o p i n g s i n k s , and h e n c e , r e d u c e d t r a n s l o c a t i o n r a t e s . The r e d u c e d demand may have r e s u l t e d i n a g r e a t e r c a r b o h y d r a t e c o n t e n t of e n r i c h e d p r i m a r y l e a v e s ( s e e T a b l e 5 . 2 ) , and c o n t r i b u t e d t o an e a r l i e r o n s e t o f s e n e s c e n c e and a more e x t e n s i v e s e n e s c e n c e , t h a n i n e n r i c h e d p l a n t s a t a h i g h e r t e m p e r a t u r e . T h e r e i s e v i d e n c e t h a t t h e t r a n s l o c a t i o n / p h o t o s y n t h e s i s r a t i o i n c r e a s e s w i t h t h e measurement t e m p e r a t u r e , from 10 t o 30 C (bean) and from 10 t o 40 C ( s o y b e a n ) , i n p l a n t s p r e v i o u s l y grown a t 20 C ( M a r o w i t c h e t 150 a l . , 1982). Hence, a l o w e r t r a n s l o c a t i o n / p h o t o s y n t h e s i s r a t i o a t c o o l e r t e m p e r a t u r e s may have c o n t r i b u t e d t o an a c c u m u l a t i o n of c a r b o h y d r a t e s i n s o u r c e l e a v e s i n my s t u d y . A l t e r n a t i v e l y , a change i n t e m p e r a t u r e may have i n f l u e n c e d c a r b o h y d r a t e s t a t u s i n d e p e n d e n t l y o f an e f f e c t on p l a n t s o u r c e / s i n k r e l a t i o n s h i p s . The m a j o r i t y of s t u d i e s w h i c h e x p l o r e s o u r c e - s i n k r e l a t i o n s h i p s and s e n e s c e n c e do so s p e c i f i c a l l y i n r e l a t i o n t o m o n o c a r p i c s e n e s c e n c e . Seed r e m o v a l i n legume s p e c i e s g e n e r a l l y d e l a y s s e n e s c e n c e . However, c a r b o h y d r a t e l e v e l s do not seem t o be i n v o l v e d s i n c e pod r e m o v a l d e l a y e d c h l o r o s i s of s o y b e a n , d e s p i t e l a r g e i n c r e a s e s i n l e a f s t a r c h and s o l u b l e c a r b o h y d r a t e c o n t e n t (Mondal e t a l . , 1978). S i m i l a r l y , m a l e - s t e r i l e s o y b e a n s s e t 85% fewer pods t h a n n o n - s t e r i l e s o y b e a n s , an e f f e c t w h i c h was a s s o c i a t e d w i t h d e l a y e d s e n e s c e n c e and a h i g h e r n i t r o g e n c o n t e n t ( W i l s o n e t a l , . 1978). Y e t m a l e - s t e r i l e s o y b e a n s a c c u m u l a t e d more c a r b o h y d r a t e s i n l e a v e s . Seed r e m o v a l i n some non-legumes, on t h e o t h e r hand, a c c e l e r a t e s s e n e s c e n c e . - E a r r e m o v a l r e s u l t e d i n a c c e l e r a t e d s e n e s c e n c e and 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 l e a v e s of c o r n ( A l l i s o n and Weinmann, 1970; C h r i s t e n s e n e t a l . , 1981), and b a r l e y (Mandahar and G a r g , 1975). Removal of f l o w e r s i n p e p p e r a c c e l e r a t e d t h e a g e - r e l a t e d l o s s of RuBP c a r b o x y l a s e a c t i v i t y ( H a l l and B r a d y , 1977). In an e f f o r t t o r e s o l v e t h o s e c o n f l i c t i n g d a t a , Thomas and S t o d d a r t (1980) p r o p o s e d t h a t s i n k demand f o r n i t r o g e n was r e d u c e d when s e e d s o f legumes ( w h i c h have a r e l a t i v e l y h i g h N:C r a t i o ) were removed, t h e r e b y d e l a y i n g s e n e s c e n c e t h r o u g h a 151 g r e a t e r r e t e n t i o n of n i t r o g e n i n t h e l e a v e s . C o n v e r s e l y , r e m o v a l of s i n k s w i t h a h i g h demand f o r f i x e d c a r b o n but o n l y a l i m i t e d demand f o r m o b i l e n u t r i e n t s ( f o r example i n non-l e g u m e s ) , c a u s e s c a r b o h y d r a t e a c c u m u l a t i o n and e v e n t u a l s e n e s c e n c e . T h i s view i s b a s e d on t h e n u t r i e n t d i v e r s i o n h y p o t h e s i s of s e n e s c e n c e , and as s u c h , f i t s n i c e l y w i t h t h e s e l f - d e s t r u c t c o n c e p t of S i n c l a i r and de Wit ( 1 9 7 5 ) , who have s u g g e s t e d t h a t t h e n i t r o g e n r e q u i r e m e n t of d e v e l o p i n g s e e d s of legumes i s so g r e a t t h a t i t r e s u l t s i n e x t e n s i v e n i t r o g e n l o s s f r o m l e a v e s , and s u b s e q u e n t s e n e s c e n c e o f t h o s e l e a v e s . In my s t u d y , c a r b o n d i o x i d e - i n d u c e d s e n e s c e n c e of P h a s e o l u s was u n r e l a t e d t o n u t r i e n t r e m o v a l . F i r s t , s e n e s c e n c e o c c u r r e d W e l l b e f o r e pod d e v e l o p m e n t , so ' t h e o n l y o r g a n s r e q u i r i n g m o b i l e n u t r i e n t s were new l e a v e s and r o o t s , b o t h of w h i c h p r e s u m a b l y -had r e l a t i v e l y low. demands. Second, d e t a c h e d l e a v e s , w h i c h c o u l d not t r a n s p o r t m o b i l e n u t r i e n t s ( o t h e r t h a n l e a k a g e from t h e p e t i o l e ) , showed t h e same t r e n d s as a t t a c h e d l e a v e s ( C h a p t e r 5 ) . I f c a r b o n d i o x i d e - i n d u c e d s e n e s c e n c e i s u n r e l a t e d t o m o n o c a r p i c d e a t h , one c o u l d s u g g e s t t h a t i t may be a c t i n g as a s t r e s s . A h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n i s not u s u a l l y c o n s i d e r e d an e n v i r o n m e n t a l s t r e s s . On t h e c o n t r a r y , h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n s a r e g e n e r a l l y a s s o c i a t e d w i t h e n h a n c e d p l a n t g r o w t h ; d e l e t e r i o u s a f f e c t s o f h i g h c o n c e n t r a t i o n s have r a r e l y been c o n s i d e r e d or even o b s e r v e d . I n s t a n c e s of s t r e s s , i n w h i c h d e f i c i e n c y of r e s o u r c e s c a u s e s e n e s c e n c e a r e w e l l known. F o r example, m i n e r a l , l i g h t , o r 152 w a t e r d e f i c i e n c i e s a l l can i n d u c e s e n e s c e n c e . C o n v e r s e l y , e x c e s s i v e c o n d i t i o n s of t e m p e r a t u r e o r l i g h t ( s o l a r i z a t i o n ) may a l s o l e a d t o p h o t o s y n t h e t i c d y s f u n t i o n and e v e n t u a l d e a t h . In t h i s r e s p e c t , c a r b o n d i o x i d e - i n d u c e d s e n e s c e n c e i s u n l i k e s o l a r i z a t i o n . In an e a r l y s t u d y o f s o l a r i z a t i o n , Holman (1930) f o u n d t h a t beans w h i c h had l a r g e s t a r c h d e p o s i t s i n t h e l e a v e s were l e s s p r o n e t o s o l a r i z a t i o n t h a n t h o s e w i t h l e s s s t a r c h . F u r t h e r , t h e t e n d a n c y t o w a r d s o l a r i z a t i o n was o f f s e t by c a r b o n d i o x i d e e n r i c h m e n t . I t i s now a p p a r e n t t h a t e x c e s s i v e c o n c e n t r a t i o n s o f c a r b o n d i o x i d e w i l l a l s o i n d u c e s e n e s c e n c e , t h e r e l a t i o n s h i p s of w h i c h a r e u n i q u e compared t o o t h e r e n v i r o n m e n t a l e x t r e m e s . F u r t h e r c a r b o n d i o x i d e e n r i c h m e n t s t u d i e s a r e needed t o f o l l o w up s e v e r a l p o i n t s a r i s i n g f r o m t h e p r e s e n t work. B e t t e r i n f o r m a t i o n i s needed on t h e r e l a t i v e r a t e s of t r a n s l o c a t i o n and c a r b o n s t o r a g e i n l e a v e s , a s i n f l u e n c e d by e n r i c h m e n t . T h i s has b e a r i n g on t h e e f f e c t on c a r b o n . d i o x i d e e n r i c h m e n t on d r y m a t t e r p a r t i t i o n i n g i n p l a n t s . F u t u r e work s h o u l d a l s o i n c l u d e s t u d i e s of t h e growth r e s p o n s e o f e n r i c h e d p l a n t s when g r o w t h i s r e s t r i c t e d by o t h e r e n v i r o n m e n t a l f a c t o r s . My work s u g g e s t s t h a t t h e growth r e s p o n s e of e n r i c h e d p l a n t s was r e d u c e d d u r i n g t h e l a t t e r s t a g e s of d e v e l o p m e n t by a c o m b i n a t i o n o f l i g h t l i m i t a t i o n a s w e l l as by a r e d u c e d c a r b o n d i o x i d e e xchange c a p a c i t y of l e a v e s . F u t u r e work m i g h t i n c l u d e s t u d i e s of g r o w t h and c a r b o n p a r t i t i o n i n g i n e n r i c h e d p l a n t s when t e m p e r a t u r e l i m i t s g r o w t h . A s t u d y of t h e 1 53 g r o w t h r e s p o n s e of e n r i c h e d p l a n t s i n r e l a t i o n t o t e m p e r a t u r e i s p a r t i c u l a r l y r e l e v a n t i n view of t h e e f f e c t o f c a r b o n d i o x i d e on s e n e s c e n c e a t c o o l t e m p e r a t u r e s . The o b s e r v a t i o n s r e c o r d e d i n t h i s s t u d y a r e r e l e v a n t i n w i d e l y d i v e r s e a r e a s o f p h y s i o l g i c a l i n v e s t i g a t i o n . B e c a u s e of t h e a p p l i c a b i l i t y of c a r b o n d i o x i d e e n r i c h m e n t t o s u c h a r e a s of s t u d y as p l a n t s e n e s c e n c e and s o u r c e - s i n k r e l a t i o n s h i p s , i t i s c o n c e i v a b l e t h a t f u t u r e work may i n c o r p o r a t e c a r b o n d i o x i d e e n r i c h m e n t as a t o o l of i n v e s t i g a t i o n o f t h o s e p r o c e s s e s . T h r e e s u c h a r e a s of i n v e s t i g a t i o n come t o mind. C a r b o n d i o x i d e e n r i c h m e n t e f f e c t i v e l y c h a n g e s t h e s o u r c e s t r e n g t h of l e a v e s , and t h i s may be u s e f u l i n d e f i n i n g how n e t p h o t o s y n t h e s i s r a t e i s c o n t r o l l e d by i n t e g r a t i o n of p l a n t s o u r c e - s i n k a c t i v i t y . S e c o n d , t h e i n c r e a s e s i n d a r k r e s p i r a t i o n r a t e i n e n r i c h e d p l a n t s may be a f u n c t i o n o f t h e h i g h e r CER o f t h o s e p l a n t s . Hence, c a r b o n d i o x i d e e n r i c h m e n t s t u d i e s may be u s e f u l i n e x p l o r i n g t h e n a t u r e of t h e r e l a t i o n s h i p between n e t p h o t o s y n t h e s i s r a t e and r e s p i r a t i o n r a t e . T h i r d , c a r b o n d i o x i d e - i n d u c e d s e n e s c e n c e of l e a v e s seems t o be r e l a t e d t o c a r b o h y d r a t e m e t a b o l i s m of t h o s e l e a v e s . T h i s may have a p p l i c a b i l i t y i n more g e n e r a l s t u d i e s of s e n e s c e n c e . F o r example, i t w o u l d be w o r t h w h i l e t o d e t e r m i n e t h e i n f l u e n c e o f g r o w t h r e g u l a t o r s on t h e c a r b o h y d r a t e m e t a b o l i s m of l e a v e s i n d u c e d t o s e n e s c e a t h i g h c a r b o n d i o x i d e c o n c e n t r a t i o n s . I t i s c l e a r t h a t an u n d e r s t a n d i n g o f c a r b o n d i o x i d e e n r i c h m e n t e f f e c t s of p l a n t s must encompass d e v e l o p m e n t a l , as w e l l as p h o t o s y n t h e t i c r e s p o n s e s . Much more work i s r e q u i r e d t o 1 54 determine the i n t e r - r e l a t i o n s h i p s between those two broad s e t s of responses to carbon d i o x i d e enrichment. 1 55 V I I . CONCLUSIONS C o n t i n u o u s l o n g - t e r m c a r b o n d i o x i d e e n r i c h m e n t o f bean p l a n t s i n c r e a s e d p l a n t g r o w t h r a t e . The p r i m a r y i n f l u e n c e of c a r b o n d i o x i d e e n r i c h m e n t on r e l a t i v e g r o w t h r a t e o c c u r r e d e a r l i e r i n d e v e l o p m e n t t h a n d i d t h e major i n f l u e n c e on a b s o l u t e g r o w t h r a t e . The i n c r e a s e i n r e l a t i v e g r o w t h r a t e o f e n r i c h e d p l a n t s e a r l y i n d e v e l o p m e n t was p a r a l l e l e d by an i n c r e a s e i n u n i t l e a f r a t e a t t h a t t i m e . T h e r e f o r e , t h e i n c r e a s e i n r e l a t i v e g r o w t h r a t e was p r o b a b l y a r e f l e c t i o n of an i n c r e a s e d u n i t l e a f r a t e . S i m i l a r l y , t h e majo r i n c r e a s e i n c a r b o n d i o x i d e e x c h a n g e r a t e of i n d i v i d u a l l e a v e s of e n r i c h e d p l a n t s o c c u r r e d e a r l y i n l e a f d e v e l o p m e n t . A c c o m p a n y i n g t h e i n c r e a s e i n u n i t l e a f r a t e or c a r b o n d i o x i d e e x c h a n g e r a t e were a d e c r e a s e i n c a r b o n d i o x i d e e xchange c a p a c i t y and an i n c r e a s e i n r e s p i r a t i o n r a t e o f t r i f o l i a t e l e a v e s , and a c c e l e r a t e d s e n e s c e n c e o f p r i m a r y l e a v e s . I t was t h e r e f o r e c o n c l u d e d t h a t t h e e f f e c t s o f c a r b o n d i o x i d e e n r i c h m e n t on c a r b o n d i o x i d e e xchange c a p a c i t y , r e s p i r a t i o n r a t e , and l e a f s e n e s c e n c e r e s u l t e d f r o m t h e l a r g e i n c r e a s e s i n u n i t l e a f r a t e or c a r b o n d i o x i d e e xchange r a t e w h i c h o c c u r r e d e a r l y i n p l a n t o r l e a f d e v e l o p m e n t r e s p e c t i v e l y . C a r b o n d i o x i d e e n r i c h m e n t c a u s e d l a r g e i n c r e a s e s i n l e a f s t a r c h c o n t e n t w h i c h a c c o m p a n i e d t h e r e d u c t i o n i n c a r b o n d i o x i d e e x change c a p a c i t y of e n r i c h e d l e a v e s . A c o n s i s t e n t l y h i g h e r l e a f s t a r c h c o n t e n t may e x p l a i n why t h e c a r b o n d i o x i d e exchange c a p a c i t y o f e n r i c h e d s o u r c e l e a v e s d i d n o t r e s p o n d a s s t r o n g l y as u n e n r i c h e d l e a v e s t o an i n c r e a s e i n s i n k demand. The i n c r e a s e i n l e a f s t a r c h c o n t e n t was even more p r o n o u n c e d i n 1 56 e n r i c h e d primary leaves which showed a c c e l e r a t e d senescence. T h e r e f o r e , the e f f e c t of an i n c r e a s e d carbon d i o x i d e exchange rate on carbon d i o x i d e exchange c a p a c i t y , r e s p i r a t i o n r a t e , and senescence in- e n r i c h e d p l a n t s may have been through a g r e a t e r l e a f carbohydrate s t a t u s . Changes i n the growth temperature or l i g h t i n t e n s i t y i n f l u e n c e d l e a f carbohydrate s t a t u s and the rate of primary l e a f senescence. L i g h t i n t e n s i t y a l s o m o d i f i e d the a f f e c t of carbon d i o x i d e enrichment on l e a f carbon d i o x i d e exchange c a p a c i t y of t r i f o l i a t e s . T h e r e f o r e , i t was concluded that the e f f e c t i v e n e s s of carbon d i o x i d e enrichment i n e l i c i t i n g those p l a n t responses was i n f l u e n c e d by the environment, p o s s i b l y through an e f f e c t on l e a f carbohydrate s t a t u s . 157 BIBLIOGRAPHY 1. ABELES, F.B. 1973. E t h y l e n e i n p l a n t b i o l o g y . Academic P r e s s , New Y o r k . 2. AHARONI, N. and M. LIEBERMAN. 1979. E t h y l e n e a s a r e g u l a t o r of s e n e s c e n c e i n t o b a c c o l e a f d i s c s . P l a n t P h y s i o l . 64:801-804. 3. ALLISON, J.C.S. and H. WEINMANN. 1970. E f f e c t o f a b s e n c e of d e v e l o p i n g g r a i n on c a r b o h y d r a t e c o n t e n t and s e n e s c e n c e of maize l e a v e s . P l a n t P h y s i o l . 46:435-436. 4. AOKI, M. and K. YABUKI. 1977. S t u d i e s on t h e c a r b o n d i o x i d e e n r i c h m e n t f o r p l a n t g r o w t h , V I I . Changes i n d r y m a t t e r p r o d u c t i o n and p h o t o s y n t h e t i c r a t e o f cucumber d u r i n g c a r b o n d i o x i d e e n r i c h m e n t . A g r . Met. 18:475-485. 5. ARTECA, R.N. and B.W. POOVAIAH. 1982. Changes i n p h o s p h o e n o l p y r u v a t e c a r b o x y l a s e and r i b u l o s e 1, 5-b i s p h o s p h a t e c a r b o x y l a s e i n Solanum t u b e r o s u m L. a s a f f e c t e d by r o o t zone a p p l i c a t i o n s of c a r b o n d i o x i d e . H o r t S c i e n c e 17:396-398 6. AZCON-BIETO, J . 1983. 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 c a r b o h y d r a t e s i n wheat l e a v e s . P l a n t P h y s i o l . 73:681-686. 7. AZCON-BIETO, J . and C B . OSMOND. 1983. R e l a t i o n s h i p between p h o t o s y n t h e s i s and r e s p i r a t i o n . The e f f e c t of c a r b o h y d r a t e s t a t u s on t h e r a t e of C02 p r o d u c t i o n by r e s p i r a t i o n i n d a r k e n e d and i l l u m i n a t e d wheat l e a v e s . P l a n t P h y s i o l . 71:574-581. 8. BACH, W. 1980. The c a r b o n d i o x i d e p r o b l e m . An i n t e r d i s c i p l i n a r y s u r v e y . ( I n t r o d u c t i o n ) . E x p e r i e n t i a 36:767-768 9. BARTON, R. 1966. F i n e s t r u c t u r e of m e s o p h y l l c e l l s i n s e n e s c i n g l e a v e s of p h a s e o l u s . P l a n t a 71:314-325. 10. BASSI, P.K., E.B.TREGUNNA, and P.A. J O L L I F F E . 1976. C a r b o n d i o x i d e e xchange and phy t o c h r o m e c o n t r o l of f l o w e r i n g i n X a n t h i u m p e n n s y I v a n i c u m . Can. J . B o t . 54:2881-2887. 11. BASSI, P.K. and M.S. SPENCER. 1982. E f f e c t of c a r b o n d i o x i d e and l i g h t on e t h y l e n e p r o d u c t i o n i n i n t a c t s u n f l o w e r p l a n t s . P l a n t P h y s i o l . 69:1222-1225. 12. BATT, T. and H.W. WOOLHOUSE. 1975. C h a n g i n g a c t i v i t i e s d u r i n g s e n e s c e n c e and s i t e s of s y n t h e s i s o f p h o t o s y n t h e t i c enzymes i n l e a v e s of t h e l a b i a t e , P e r i l l a f r u t e s c e n s ( L . ) 1 58 B r i t t . J . E x p t . B o t . 26:569-579. 13. BISHOP, P.M. and C P . WH IT T INGHAM. 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 c a r b o n d i o x i d e e n r i c h e d a t m o s p h e r e . P h o t o s y n t h e t i c a 2:31-38. 14. BOARDMAN, N.K. 1977. C o m p a r a t i v e p h o t o s y n t h e s i s of sun and shade p l a n t s . Ann. Rev. P l a n t P h y s i o l . 28:355-377. 15. BOLAS B.D. and R. MELVILLE. 1935. The e f f e c t on t h e tomato p l a n t o f c a r b o n d i o x i d e p r o d u c e d by c o m b u s t i o n . Ann. A p p l . B i o l . 22:1-15. 16. 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 o f c a r b o n d i o x i d e i n t h e a i r on t h e p h o t o s y n t h e t i c p r o c e s s of l e a v e s and on t h e mode of g r o w t h of p l a n t s . P r o c . R o y a l S o c . 70:397-413. 17. BRUINSMA, J . 1963. The q u a n t i t a t i v e a n a l y s i s of c h l o r o p h y l l s a and b i n p l a n t e x t r a c t s . Photochem. and P h o t o b i o l . 2:241-249. 18. CALVERT, A. and G. SLACK. 1976. E f f e c t o f c a r b o n d i o x i d e e n r i c h m e n t on g r o w t h , d e v e l o p m e n t and y i e l d of g l a s s h o u s e t o m a t o e s . 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 of e n r i c h m e n t . J . H o r t . S c i . 51:401-409. 19. CANVIN, D.T. 1979. P h o t o r e s p i r a t i o n : C o m p a r i s o n between C 3 a n d C4 p l a n t s . . I_n : M. G i b b s and E. L a t z k o , e d s . , P h o t o s y n t h e s i s I I . P h o t o s y n t h e t i c Carbon. M e t a b o l i s m and R e l a t e d P r o c e s s e s . E n c y c l o p e d i a of P l a n t P h y s i o l o g y , New S e r i e s V o l . 6. S p r i n g e r - V e r l a g . B e r l i n . pp. 368-396. 20. CAVE, G., L.C.TOLLEY, and B.R. STRAIN. 1981. E f f e c t o f c a r b o n d i o x i d e e n r i c h m e n t on c h l o r o p h y l l c o n t e n t , s t a r c h c o n t e n t and s t a r c h g r a i n s t r u c t u r e i n T r i f o l i u m s u b t e r r a n e u m l e a v e s . P l a n t P h y s i o l . 51:171-174. 21. CHANG, C.W. 1975. C a r b o n d i o x i d e and s e n e s c e n c e i n c o t t o n p l a n t s . P l a n t P h y s i o l . 55:515-519. 22. CHATTERTON, N . J . , G.E. CARLSON, W.E. HUNGERFORD, and D.R. L E E . 1972. E f f e c t o f t i l l e r i n g and c o o l n i g h t s on p h o t o s y n t h e s i s and c h l o r o p l a s t s t a r c h i n P a n g o l a . C r o p S c i . 12:206-208. 23. CHRISTENSEN, L . E . , F.E. BELOW, and R.H. HAGEMAN. 1981. The e f f e c t s of e a r r e m o v a l on s e n e s c e n c e and m e t a b o l i s m o f m a i z e . P l a n t P h y s i o l . 68:1180-1185. 24. CLOUGH, J.M., M.M. PEET, and P . J . KRAMER. 1981. E f f e c t s o f h i g h a t m o s p h e r i c C02 and s i n k s i z e on r a t e s o f p h o t o s y n t h e s i s o f a soybean c u l t i v a r . P l a n t P h y s i o l . 67:1007-1010. 159 25. DAVIS, S.D. and K . J . McCREE. 1978. P h o t o s y n t h e t i c r a t e and d i f f u s i o n c o n d u c t a n c e a s a f u n c t i o n of age i n l e a v e s of bean p l a n t s . C r o p S c i . 18:280-282. 26. DAVIS, T.D. and J.R. POTTER. 1983. H i g h c a r b o n d i o x i d e a p p l i e d t o c u t t i n g s : e f f e c t s on r o o t i n g and s u b s e q u e n t g r o w t h i n o r n a m e n t a l s p e c i e s . H o r t S c i e n c e 18:194-196. 27. DHAWAN,K.R., P.K BASSI, and M.S. SPENCER. 1981. E f f e c t s of c a r b o n d i o x i d e on e t h y l e n e p r o d u c t i o n and a c t i o n i n i n t a c t s u n f l o w e r p l a n t s . P l a n t P h y s i o l . 68:831-834. 28. DUNCAN, W.G. and B . J . BARFIELD. 1968. P r e d i c t i n g e f f e c t s of c a r b o n d i o x i d e e n r i c h m e n t w i t h s i m u l a t i o n models and a d i g i t a l c o m p u t e r . I_n : C o n t r o l l e d A t m o s p h e r e s f o r P l a n t G rowth. Amer. S oc. of A g r i c u l t u r a l E n g i n e e r s , pp. 246. 29. EDELMAN, J . and A . I . SCHOOLAR. 1969. L i g h t a s a major f a c t o r i n c h l o r o p h y l l d e s t r u c t i o n i n s u g a r cane l e a f t i s s u e . Z. P f l a n z e n p h y s i o l . 60:470-471. 30. EDELMAN, J . and A.D. HANSON. 1971. S u c r o s e s u p p r e s s i o n o f c h l o r o p h y l l s y n t h e s i s i n c a r r o t c a l l u s c u l t u r e s . 1 P l a n t a 98:150-156. 31. EHLERINGER, J . and 0. BJORKMAN. 1977. Quantum y i e l d s f o r C02 u p t a k e i n C3 and C4 p l a n t s . Dependence on t e m p e r a t u r e , C02, and 02 c o n c e n t r a t i o n . P l a n t P h y s i o l . 59:86-90. 32. EHRET, D.L. and P.A. J O L L I F F E . 1983. A semi - o p e n s y s t e m f o r measurements of p l a n t gas exchange r a t e s . P h o t o s y n t h e t i c a ( i n p r e s s ) . 33. ELSTNER, E . F . 1982. Oxygen a c t i v a t i o n and oxygen t o x i c i t y . Ann. Rev. P l a n t P h y s i o l . 33:73-96. 34. ERICKSON, R.O. and F . J . MICHELINI. 1957. The p l a s t o c h r o n i n d e x . Amer. J . B o t . 44:297-304. 35. 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 c a r b o n 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 Hordeum v u l g a r e 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 o f c a r b o n d i o x i d e and o x y g e n . Ann. B o t . 37:1035-1039. 36. FINN, G.A. and W. A. BRUN. 1982. E f f e c t of a t m o s p h e r i c C02 e n r i c h m e n t on gr o w t h , n o n s t r u c t u r a l c a r b o h y d r a t e c o n t e n t , and r o o t n o d u l e a c t i v i t y i n s o y b e a n . P l a n t P h y s i o l . 69:327-331. 37. FOCK, H., K. KLUG, and D.T. CANVIN. 1979. E f f e c t of 160 c a r b o n d i o x i d e and te-mperature on p h o t o s y n t h e t i c C02 u p t a k e and p h o t o r e s p i r a t o r y C02 e v o l u t i o n i n s u n f l o w e r l e a v e s . P l a n t a 145:219-223. 38. FONDY, B.R. and D.R. GEIGER. 1981. R e g u l a t i o n o f e x p o r t by i n t e g r a t i o n of s i n k and s o u r c e a c t i v i t y . What's New i n P l a n t P h y s i o l o g y 12:33-36. 39. FONDY, B.R. and D.R. GEIGER. 1982. D i u r n a l p a t t e r n o f t r a n s l o c a t i o n and c a r b o h y d r a t e m e t a b o l i s m i n s o u r c e l e a v e s of B e t a v u l g a r i s L. P l a n t P h y s i o l . 70:671-676. 40. FORD, M.A. and G.N. THORNE. 1967. E f f e c t of C02 c o n c e n t r a t i o n on g r o wth of s u g a r - b e e t . b a r l e y , k a l e and m a i z e . Ann. B o t . 31:628-643. 41. FRIEDRICH, J.W. and R.C. HUFFAKER. 1980. P h o t o s y n t h e s i s , l e a f r e s i s t a n c e s , and r i b u l o s e 1, 5-b i s p h o s p h a t e c a r b o x y l a s e d e g r a d a t i o n i n s e n e s c i n g b a r l e y l e a v e s . P l a n t P h y s i o l . 65:1103-1107. 42. 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 o f c a r b o n 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. '43. GAASTRA, P. 1963. C l i m a t i c c o n t r o l o f p h o t o s y n t h e s i s and r e s p i r a t i o n . I_n : L.T. E v a n s , e d ., E n v i r o n m e n t a l C o n t r o l o f P l a n t Growth. Academic P r e s s . New Y o r k . pp. 113-140. 44. GEIGER, D.R. 1976. E f f e c t s of t r a n s l o c a t i o n and a s s i m i l a t e demand on p h o t o s y n t h e s i s . Can. J . B o t . 54:2337-2345. 45. GIAQUINTA, R. 1978. S o u r c e and s i n k l e a f m e t a b o l i s m i n r e l a t i o n t o phloem t r a n s l o c a t i o n . P l a n t P h y s i o l . 61:380-385. 46. GIFFORD, R.M. 1977. Growth p a t t e r n , c a r b o n d i o x i d e e x change and d r y w e i g h t d i s t r i b u t i o n i n wheat g r o w i n g under d i f f e r i n g p h o t o s y n t h e t i c e n v i r o n m e n t s . A u s t . J . P l a n t P h y s i o l . 4:99-110 47. GIFFORD, R.M. 1979. Growth and y i e l d of c a r b o n d i o x i d e -e n r i c h e d wheat under w a t e r - l i m i t e d c o n d i t i o n s . A u s t . J . P l a n t P h y s i o l . 6:367-378. 48. GIFFORD, R.M. and L.T. EVANS. 1981. P h o t o s y n t h e s i s , c a r b o n p a r t i t i o n i n g , and y i e l d . Ann. Rev. P l a n t P h y s i o l . 32:485-509. 49. GOLDTHWAITE, J . J . and W.M. LAETSCH. 1967. R e g u l a t i o n o f s e n e s c e n c e i n bean l e a f d i s c s by l i g h t and c h e m i c a l g r o w t h r e g u l a t o r s . P l a n t P h y s i o l . 42:1757-1762. 161 50. GRODZINSKI, B., I . BOESEL, and R.F. HORTON. 1983. L i g h t s t i m u l a t i o n of e t h y l e n e r e l e a s e from l e a v e s of Gomphrena g l o b o s a L. P l a n t P h y s i o l . 71:588-593. 51. GUSTAFSON, S.W. and P . J . BREEN. 1982. E f f e c t s of C02 e n r i c h m e n t on p h o t o s y n t h e s i s , d r y m a t t e r a c c u m u l a t i o n and y i e l d of beans, P h a s e o l u s v u l g a r i s L. H o r t S c i e n c e 17 ( s u p p l . ) a b s t r a c t 43. 52. HALES, S. 1727. V e g e t a b l e s t a t i c k s . R e p u b l i s h e d by A m e r i c a n E l s e v i e r , ( 1 9 6 9 ) . New Y o r k . 53. HALL, A . J . and C . J . BRADY. 1977. A s s i m i l a t e s o u r c e -s i n k r e l a t i o n s h i p s i n C a p s i c u m annuum L. I I . E f f e c t s o f f r u i t i n g and d e f l o r a t i o n on t h e p h o t o s y n t h e t i c c a p a c i t y and s e n e s c e n c e of t h e l e a v e s . A u s t . J . P l a n t P h y s i o l . 4:771-783. 54. HANSEN, J . , D. JOHNSON, A. L A C I S , S. LEBEDEFF, P. L E E , D. RIND, and G. RUSSELL. 1981. C l i m a t e i m p a c t of i n c r e a s i n g a t m o s p h e r i c c a r b o n d i o x i d e . S c i e n c e 213:957-966. 55. HARDMAN, L . L . and W.A. BRUN. 1971. E f f e c t of a t m o s p h e r i c c a r b o n d i o x i d e e n r i c h m e n t a t d i f f e r e n t d e v e l o p m e n t a l s t a g e s on g r o w t h and y i e l d components o f s o y b e a n s . C r o p S c i e n c e . 11:886-888. 56. HARDWICK, K., M. WOOD and H.W. WOOLHOUSE. 1968. P h o t o s y n t h e s i s and r e s p i r a t i o n i n r e l a t i o n t o l e a f age i n P e r i l l a F r u t e s c e n s ( L . ) B r i t t . New P h y t o l . 67:79-86. 57. HARDY, R.W.F. and U.D.HAVELKA. 1977. P o s s i b l e r o u t e s t o i n c r e a s e t h e c o n v e r s i o n of s o l a r e n e r g y t o f o o d and f e e d by g r a i n legumes and c e r e a l g r a i n s ( c r o p p r o d u c t i o n ) : c a r b o n d i o x i d e and n i t r o g e n f i x a t i o n , f o l i a r f e r t i l i z a t i o n , and a s s i m i l a t e p a r t i t i o n i n g . I_n : A. M i t s u i , S. M i y a c h i , A. San P i e t r o and S. Tamura, e d s . , B i o l o g i c a l S o l a r E n e r g y C o n v e r s i o n . A c ademic P r e s s , New Y o r k . pp. 299-322. 58. HELDT, H.W., C . J . CHON, and G.H. LORIMER. 1978. P h o s p h a t e r e q u i r e m e n t f o r t h e l i g h t a c t i v a t i o n of r i b u l o s e 1, 5 b i s p h o s p h a t e c a r b o x y l a s e i n i n t a c t s p i n a c h c h l o r o p l a s t s . FEBS L e t t e r s 92:234-240. 59. HEROLD, A. 1980. R e g u l a t i o n o f p h o t o s y n t h e s i s by s i n k a c t i v i t y - t h e m i s s i n g l i n k . New P h y t o l . 86:131-144. 60. HESKETH, J.D., K.C. FRY, G. GUINN,and J.R. MAUNEY. 1971. E x p e r i m e n t a l a s p e c t s o f g r o w t h m o d e l l i n g : p o t e n t i a l c a r b o h y d r a t e r e q u i r e m e n t of c o t t o n b o l l s . I_n : T r e e Growth Dynamics and M o d e l i n g . Duke U n i v e r s i t y , pp. 123-1 62 127. 61. HESKETH, J.D., W.L. OGREN, M.E. HAGEMAN, and D.B. PETERS. 1981. C o r r e l a t i o n s among l e a f c a r b o n d i o x i d e exchange r a t e s , a r e a s and enzyme a c t i v i t i e s among soybean c u l t i v a r s . P h o t o s y n t h e s i s R e s e a r c h 2:21-30. 62. HESKETH, J.D., E.M. LARSON, A . J . GORDON, and D.B. PETERS. 1983. I n t e r n a l f a c t o r s i n f l u e n c i n g p h o t o s y n t h e s i s and r e s p i r a t i o n . lr\ : J . E . D a l e and F . L . M i l t h o r p e , e d s . , The Growth and F u n c t i o n i n g of L e a v e s . Cambridge U n i v . P r e s s . C a m b r i d g e . pp. 381-405. 63. HICKLENTON, P.R. and P.A. J O L L I F F E . 1978. E f f e c t s of g r e e n h o u s e C02 e n r i c h m e n t on t h e y i e l d and p h o t o s y n t h e t i c p h y s i o l o g y of tomato p l a n t s . Can. J . P l a n t S c i . . 58:801-817 64. HICKLENTON, P.R. and P. A. J O L L I F F E . 1980a. C a r b o n d i o x i d e and f l o w e r i n g i n P h a r b i t i s n i l C h o i s y . P l a n t P h y s i o l . 66:13-17. 65. HICKLENTON, P.R. and P.A. J O L L I F F E . 1980b. A l t e r a t i o n s i n t h e p h y s i o l o g y o f C02 e xchange i n tomato p l a n t s grown i n C 0 2 - e n r i c h e d a t m o s p h e r e s . Can. J . B o t . 58:2181-2189. 66. HO, L.C. 1977. E f f e c t s of C02 e n r i c h m e n t on t h e r a t e s o f p h o t o s y n t h e s i s and t r a n s l o c a t i o n of tomato l e a v e s . Ann. A p p l . B i o l . 87:191-200. 67. HOFSTRA, G. and J.D. HESKETH. 1969. The e f f e c t o f t e m p e r a t u r e on s t o m a t a l a p e r t u r e i n d i f f e r e n t s p e c i e s . Can. J . B o t . 47:1307-1310.. 68. HOFSTRA, G. and J.D. HESKETH. 1975. The e f f e c t s of t e m p e r a t u r e and C02 e n r i c h m e n t on p h o t o s y n t h e s i s i n s o y b e a n . I_n : R. M a r c e l l e , e d . , E n v i r o n m e n t a l 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 . D r . W. J u n k . The Hague. pp.71-81. 69. HOLMAN, R. 1930. On s o l a r i z a t i o n of l e a v e s . U n i v e r s i t y of C a l i f o r n i a P u b l i c a t i o n s i n B o t a n y . 16:139-151. 70. HOPEN, H.J. and S.K. RIES. 1962. The m u t u a l l y c o m p e n s a t i n g e f f e c t o f c a r b o n 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 t h e g r o w t h of Cumumis S a t i v u s L. P r o c e e d i n g s o f t h e A m e r i c a n S o c i e t y f o r H o r t i c u l t u r a l Sc i e n c e 81 : 358. 71. HUBER, S.C. and D.W. ISRAEL. 1982. B i o c h e m i c a l b a s i s f o r p a r t i t i o n i n g of p h o t o s y n t h e t i c a l l y f i x e d c a r b o n between s t a r c h and s u c r o s e i n soybean ( G l y c i n e max M e r r . ) l e a v e s . P l a n t P h y s i o l . 69:691-696. 1 63 72. HUGHES, A.P. and P.R. FREEMAN. 1967. Growth a n a l y s i s u s i n g f r e q u e n t s m a l l h a r v e s t s . J . A p p l . E c o l . 4:553-560. 73. HUGHES, A.P. and K.E. COCKSHULL. 1971. The v a r i a t i o n i n r e s p o n s e t o l i g h t i n t e n s i t y and c a r b o n 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 o f Chrysanthemum  m o r i f o l i u m grown i n c o n t r o l l e d e n v i r o n m e n t s a t two t i m e s o f y e a r . Ann. B o t . 35:933-45 . 74. HURD, R.G. 1968. E f f e c t s of C02- e n r i c h m e n t on t h e gro w t h o f young tomato p l a n t s i n low l i g h t . Ann. B o t . 32:531-542. 75. HUXLEY, P.A. 1971. L e a f volume: a s i m p l e method f o r measurement and some n o t e s on i t s use i n s t u d i e s o f l e a f g r o w t h . J . A p p l . E c o l . 8:147-153. 76. IMAI, K. and Y. MURATA. 1976. E f f e c t o f c a r b o n d i o x i d e c o n c e n t r a t i o n on g r o w t h and d r y m a t t e r p r o d u c t i o n o f c r o p p l a n t s . 1. E f f e c t s on l e a f a r e a , d r y m a t t e r , t i l l e r i n g , d r y m a t t e r d i s t r i b u t i o n r a t i o , and t r a n s p i r a t i o n . P r o c . C r o p S c i . S o c. J a p a n . 45:598-606. 77. IMAI, K. and Y. MURATA. 1977. E f f e c t o f c a r b o n d i o x i d e c o n c e n t r a t i o n on g r o w t h and d r y m a t t e r p r o d u c t i o n o f c r o p p l a n t s . I I . S p e c i f i c and v a r i e t a l d i f f e r e n c e s i n t h e r e s p o n s e of d r y m a t t e r p r o d u c t i o n . J a p a n J o u r . C r o p S c i . 46:291-297. 78. IMAI, K. and Y. MURATA. 1978a. E f f e c t o f c a r b o n d i o x i d e c o n c e n t r a t i o n on growth and d r y m a t t e r p r o d u c t i o n o f c r o p p l a n t s . I I I . R e l a t i o n s h i p between C02 c o n c e n t r a t i o n and n i t r o g e n n u t r i t i o n i n some C3- and C4-s p e c i e s . J a p a n J o u r . C r o p S c i . 47:118-123. 79. IMAI, K. and Y. MURATA, 1978b. E f f e c t o f c a r b o n d i o x i d e c o n c e n t r a t i o n on g r o w t h and d r y m a t t e r p r o d u c t i o n o f c r o p p l a n t s . IV. A f t e r - e f f e c t s of c a r b o n d i o x i d e - t r e a t m e n t s on t h e a p p a r e n t p h o t o s y n t h e s i s , d a r k r e s p i r a t i o n and d r y m a t t e r p r o d u c t i o n J a p a n . J o u r . C r o p S c i . 48:328-335. 80. IMAI, K. and Y. MURATA. 1978c. E f f e c t o f c a r b o n d i o x i d e c o n c e n t r a t i o n on growth and d r y m a t t e r p r o d u c t i o n of c r o p p l a n t s . V. A n a l y s i s of a f t e r - e f f e c t of c a r b o n d i o x i d e - t r e a t m e n t on a p p a r e n t p h o t o s y n t h e s i s . J a p a n . J o u r . C r o p S c i . 47:587-595. 81. JENSEN, R.G. 1980. B i o c h e m i s t r y o f t h e c h l o r o p l a s t . Iri : T h e ' B i o c h e m i s t r y of P l a n t s , V o l . 1:7. 82. J O L L I F F E , P.A. and E.B. TREGUNNA. 1968. E f f e c t of t e m p e r a t u r e , c a r b o n d i o x i d e c o n c e n t r a t i o n , and l i g h t 164 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. 83. RANNANGARA, C.G. and H.W. WOOLHOUSE. 1968. Changes i n t h e enzyme a c t i v i t y of s o l u b l e p r o t e i n f r a c t i o n s i n t h e c o u r s e of f o l i a r s e n e s c e n c e i n P e r i l l a F r u t e s c e n s L. B r i t t . New P h y t o l . 67:533-542. 84. van REULEN, H., H.H. van LAAR, W. LOUWERSE, and J . G o u d r i a a n . 1980. P h y s i o l o g i c a l a s p e c t s of i n c r e a s e d c a r b o n d i o x i d e c o n c e n t r a t i o n . E x p e r i e n t i a 36:786-792. 85. RHANNA, R., R. WAGNER, W. JUNGE, and GOVINDJEE. 1980. E f f e c t s of C 0 2 - d e p l e t i o n on p r o t o n u p t a k e and r e l e a s e i n t h y l a k o i d membranes. FEBS L e t t e r s . 121:222-224. 86. RHUDAIRI, A.R. 1970. C h l o r o p h y l l d e g r a d a t i o n by l i g h t i n l e a f d i s c s i n t h e p r e s e n c e of s u g a r . P h y s i o l . P l a n t . 23:613-622. 87. RRIEDEMANN, P.E., R . J . SWARD, and W.J.S. DOWNTON. 1976. V i n e r e s p o n s e t o c a r b o n d i o x i d e e n r i c h m e n t d u r i n g h e a t t h e r a p y . A u s t . J . P l a n t . P h y s i o l . 3:605-618. 88. RRIZER, D.T., R.H. ZIMMERMAN, H.H. RLUETER, and W.A. BAILEY. 1971. Growth of c r a b a p p l e s e e d l i n g s i n c o n t r o l l e d e n v i r o n m e n t s : e f f e c t of c a r b o n d i o x i d e l e v e l , and t i m e and d u r a t i o n of carbon- d i o x i d e t r e a t m e n t . J . • Amer. S o c . H o r t . S c i . 96:285-288. 89. LAZAN,H.B., E.W.R. BARLOW, and C . J . BRADY. 1983. The s i g n i f i c a n c e o f v a s c u l a r c o n n e c t i o n i n r e g u l a t i n g s e n e s c e n c e o f t h e d e t a c h e d f l a g l e a f o f wheat. J . E x p t . B o t . 34:726-736. 90. LEWINGTON, R . J . , M. TALBOT, and E.W. SIMON. 1967. The y e l l o w i n g of a t t a c h e d and d e t a c h e d cucumber c o t y l e d o n s . J . E x p t . B o t . 18:526-534. 91. LORIMER, G.H., M.R. BADGER, and H.W. HELDT. 1978. The a c t i v a t i o n of r i b u l o s e 1, 5 ~ b i s p h o s p h a t e c a r b o x y l a s e / o x y g e n a s e . I_n : H.W.J. S i e g e l m a n and G. H i n d , e d s . , P h o t o s y n t h e t i c C a r b o n A s s i m i l a t i o n . Plenum P r e s s . New Y o r k . pp. 283-306. 92. LUDLOW, M.M. 1975. E f f e c t of water s t r e s s on t h e d e c l i n e of l e a f n e t p h o t o s y n t h e s i s w i t h age. I_n : R. M a r c e l l e , e d . , E n v i r o n m e n t a l 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. J u n k . The Hague. pp. 123-134. 93. LUDWIG, L . J . , D.A. CHARLES-EDWARDS and A.C. WITHERS. 1975. Tomato l e a f p h o t o s y n t h e s i s and r e s p i r a t i o n i n v a r i o u s l i g h t and c a r b o n d i o x i d e e n v i r o n m e n t s . I_n : R. M a r c e l l e , e d . , E n v i r o n m e n t a l and B i o l o g i c a l C o n t r o l o f 1 65 P h o t o s y n t h e s i s . D r . W. Junk. The Hague. pp. 29-36. 94. MACDOWALL, F.D.H. 1972. Growth k i n e t i c s o f M a r q u i s wheat. I I . C a r b o n d i o x i d e d e p e n d e n c e . Can. J . B o t . . 50:883-889. 95. MADSEN, E. 1968. E f f e c t - of C 0 2 - c o n c e n t r a t i o n on t h e a c c u m u l a t i o n of s t a r c h and s u g a r i n tomato l e a v e s . P h y s i o l . P l a n t . 21:168-175. 96. MADSEN, E. 1973a. The e f f e c t o f C 0 2 - c o n c e n t r a t i o n on d e v e l o p m e n t and d r y m a t t e r p r o d u c t i o n i n young tomato p l a n t s . A c t a . A g r i c u l t u r a e S c a n d i n a v i c a 23:235-240. 97. MADSEN, E. 1973b. E f f e c t of C 0 2 - c o n c e n t r a t i o n on t h e m o r p h o l o g i c a l , h i s t o l o g i c a l and c y t o l o g i c a l c h a n g e s i n tomato p l a n t s . A c t a A g r i c u l t u r a e S c a n d i n a v i c a 23:241-246. 98. MADSEN, E. 1974. E f f e c t of C02 c o n c e n t r a t i o n on growth and f r u i t p r o d u c t i o n of tomato p l a n t s . . A c t a A g r i c u l t u r a e S c a n d i n a v i c a 24:242-246. 99. MADSEN, E. 1975. E f f e c t of C 0 2 - e n r i c h m e n t on g r o w t h , d e v e l o p m e n t , f r u i t p r o d u c t i o n and f r u i t q u a l i t y o f tomato, from a p h y s i o l o g i c a l v i e w p o i n t . I_n : P. C h o v a r d and N. de B i l d e r l i n g , e d s . , P h y t o t r o n i c s 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 R e s e a r c h . B o r d a s . pp. 319-329. 319-329. 100. MANDAHAR, C.L. and I.D. GARG. 1975. E f f e c t of e a r r e m o v a l on s u g a r s and c h l o r o p h y l l s o f b a r l e y l e a v e s . P h o t o s y n t h e t i c a . 9:407-409. 101. MAROWITCH, J . , C. RICHTER, and J . HODDINOTT. 1982. The i n f l u e n c e of t e m p e r a t u r e on t r a n s l o c a t i o n and p h o t o s y n t h e s i s i n beans and s o y b e a n . C a n a d i a n B o t a n i c a l A s s o c i a t i o n - C a n a d i a n S o c i e t y o f P l a n t P h y s i o l o g i s t s , J o i n t M e e t i n g . A b s t a c t 59. 102. MATILE, PH. and F. WINKENBACH. 1971. F u n c t i o n of l y s o s o m e s and l y s o s o m a l enzymes i n t h e s e n e s c i n g c o r o l l a of t h e m o r n i n g g l o r y Ipomoea p u r p u r e a . J . E x p t . B o t . 22:759-771. 103. MAUNEY, J.R., K.E. FRY, and G GUINN. 1978. R e l a t i o n s h i p of p h o t o s y n t h e t i c r a t e t o growth and f r u i t i n g o f c o t t o n , s o y b e a n , sorghum, and s u n f l o w e r . C r o p S c i . 18:259-263. 104. MAUNEY, J.R., G. GUINN, K.E. FRY, and J.D. HESKETH. 1979. C o r r e l a t i o n o f p h o t o s y n t h e t i c c a r b o n d i o x i d e u p t a k e and 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 c o t t o n , s o y b e a n , s u n f l o w e r and sorghum. P h o t o s y n t h e t i c a 13:260. 105. MONDAL, M.H., W.A. BRUN, and M.L. BRENNER. 1978. E f f e c t s of s i n k r e m o v a l on p h o t o s y n t h e s i s and s e n e s c e n c e 166 in leaves of soybean Glycine max L. plants. Plant Physiol. 61:394-397. 106. MOORE, K.G., A. ILLSLEY, and P.H. LOVELL. 1974. Effects of sucrose on pet i o l a r carbohydrate accumulation and photosynthesis in excised Sinapis cotyledons. J . Expt. Bot. 25:887-898. 107. NAFZIGER, E.D. and H.R. ROLLER. 1976. Influence of leaf starch concentration on C02 assimilation in soybean. Plant Physiol. 57:560-563. 108. NEALES, T.F. and L.D. INCOLL. 1968. The control of leaf photosynthesis rate by the le v e l of assimilate concentration in the le a f : a review of the hypothesis. The Botanical Review 34:107-125. 109. NEALES, T.F. and A.O. NICHOLLS. 1978. Growth responses of young wheat plants to a range of ambient C02 l e v e l s . Aust. J. Plant Physiol. 5:45-59. 110. NOODEN, L.D. and A.C. LEOPOLD. 1978. Phytohormones and the endogenous regulation of senescence and abscission. In : D.S. Letham, P.B. Goodwin, and T.J.V. Higgins, eds., Phytohormones and Related Compounds-A Comprehensive Treatise, Volume I I . Phytohormones and The Development of Higher Plants. Elsevier/North-Holland Biomedical Press. New York. pp. 329-361. 111. OSBORNE, D. 1978. Ethylene. In : D.S. Letham, P.B. Goodwin, and T.J.V. Higgins, eds., Phytohormones and Related Compounds-A e Comprehensive Treatise Vol. I. Elsevier/North-Holla-nd Biomedical Press. New York. pp. . 265-294. 112. PATTERSON, D.T. and E.P. FLINT. 1980. Potential effects of global atmospheric C02 enrichment on the growth and competitiveness of C3 and C4 weed and crop plants. Weed S c i . 28:71-75. 113. PATTERSON, D.T. and E.P. FLINT. 1982. Interacting effects of C02 and nutrient concentration. Weed S c i . 30:389-394. 114. PENNING de VRIES, F.W.T. 1972. Respiration and growth. In : A.R. Rees, R.E. Cockshull, D.W. Hand, and R.G. Hurd, eds., Crop Processes in Controlled Environments. Academic Press. London and New York. pp 327-347. 115. PEREZ-TREJO, M.S., H.W. JANES, and C. Frenkel. 1981. Mobilization of respiratory metabolism in potato tubers by carbon dioxide. . Plant Physiol. 67:514-517. 116. PHILLIPS, D.A.-, R.D. NEWELL, S.A. HASSELL, and C E . 167 FELLING. 1976. The e f f e c t of C02 e n r i c h m e n t on r o o t n o d u l e d e v e l o p m e n t and s y m b i o t i c N2 r e d u c t i o n i n p i s u m  s a t ivum 1. Amer. J . B o t . 63:356-362. 117. POTTER, J.R. 1980. P a r t i t i o n i n g o f p h o t o s y n t h a t e between s t a r c h and e x p o r t e d m a t e r i a l i n s u n f l o w e r l e a v e s e x p o s e d t o 2000 u l / 1 C02. P l a n t P h y s i o l . ( s u p p l . ) a b s t r a c t 402. 118. POTTER, J.R. and P . J . BREEN. 1980. M a i n t a i n e n c e o f h i g h p h o t o s y n t h e t i c r a t e s d u r i n g t h e a c c u m u l a t i o n o f h i g h l e a f s t a r c h l e v e l s i n s u n f l o w e r and s o y b e a n . P l a n t P h y s i o l . 66:528-531. 119. RABINOWITCH, E. and GOVINDJEE. 1969. I_n : P h o t o s y n t h e s i s . John W i l e y and Sons, I n c . New Y o r k . 120. RAPER, JR. C D . , and F. PEEDIN. 1978. P h o t o s y n t h e t i c r a t e d u r i n g s t e a d y - s t a t e growth as i n f l u e n c e d by c a r b o n -d i o x i d e c o n c e n t r a t i o n . B o t . Gaz. 139:147-149. 121. RASCHKE, K. 1975. S t o m a t a l A c t i o n . Ann. Rev. P l a n t P h y s i o l . 26:309-340. 122. REINSCH, C H . 1971.. S m o o t h i n g by s p l i n e f u n c t i o n s . I I . Num. Math. 16:451-454. 123. RHODES, M.J.C. 1980. R e s p i r a t i o n and s e n e s c e n c e o f p l a n t o r g a n s . I_n : P.K. Stumpf and E.E. Conn, e d s . , The B i o c h e m i s t r y o f P l a n t s , . Volume I I . M e t a b o l i s m and R e s p i r a t i o n . Academic P r e s s . New Y o r k . pp. 419-462. 124. ROBINSON, S.P. and D.A. WALKER. 1979. The c o n t r o l o f 3- p h o s p h o g l y c e r a t e r e d u c t i o n i n i s o l a t e d c h l o r o p l a s t s by t h e c o n c e n t r a t i o n s of ATP, ADP, and 3 - p h o s p h o g l y c e r a t e . B i o c h i m i c a e t B i o p h y s i c a A c t a . 545:528-536. 125. RUFTY, JR. T.W., C D . RAPER JR., and W.A. JACKSON. 1981. N i t r o g e n a s s i m i l a t i o n , r o o t g r o w t h and whole p l a n t r e s p o n s e s o f soybean t o r o o t t e m p e r a t u r e , and t o c a r b o n d i o x i d e and l i g h t i n t h e a e r i a l e n v i r o n m e n t . New P h y t o l . 88:607-619. 126. RUFTY, JR. T.W. and S.C. HUBER. 1983. Changes i n s t a r c h f o r m a t i o n and a c t i v i t i e s of s u c r o s e p h o s p h a t e s y n t h a s e and c y t o p l a s m i c f r u c t o s e - 1 , 6 - b i s p h o s p h a t a s e i n r e s p o n s e t o s o u r c e - s i n k a l t e r a t i o n s . P l a n t P h y s i o l . 72:474-480. 127. SATLER, S.O. and K.V. THIMANN. 1983a. M e t a b o l i s m of o a t l e a v e s d u r i n g s e n e s c e n c e . P l a n t P h y s i o l . 71:67-70. 128. SATLER, S.O. and K.V. THIMANN. 1983b. R e l a t i o n between r e s p i r a t i o n and s e n e s c e n c e i n o a t l e a v e s . P l a n t P h y s i o l . 72:540-546. 168 129. SCHOOLAR, A . I . and J.EDELMAN. 1970. P r o d u c t i o n and s e c r e t i o n o f s u c r o s e by s u g a r - c a n e l e a f t i s s u e . J . E x p t . B o t . 130. SINCLAIR, T.R. and C T . DEWIT. 1975. P h o t o s y n t h a t e and n i t r o g e n r e q u i r e m e n t s f o r s e e d p r o d u c t i o n by v a r i o u s c r o p s . S c i e n c e . 189:565-567. 131. SIONIT, N., H.HELLMERS, and B.R. STRAIN .1980. Growth and y i e l d of wheat under c a r b o n d i o x i d e e n r i c h m e n t and water s t r e s s . C r o p S c i . 20:687-690. 132. SIONIT, N., D.A. MORTENSEN, B.R. STRAIN, and H. HELLMERS. 1981a. Growth r e s p o n s e of wheat t o C02 e n r i c h m e n t and d i f f e r e n t l e v e l s of m i n e r a l n u t r i t i o n . A g r o n . J . 73:1023-1027. 133. SIONIT, N., B.R. STRAIN, H. HELLMERS, AND P . J . KRAMER. 1981b. E f f e c t s of a t m o s p h e r i c c a r b o n d i o x i d e c o n c e n t r a t i o n and water s t r e s s on water r e l a t i o n s of wheat. B o t . Gaz. 142:191-196. 134. SMALL, T. and H.L. WHITE. 1930. C a r b o n d i o x i d e i n r e l a t i o n t o g r e e n h o u s e c r o p s . IV. The e f f e c t on t o m a t o e s of an e n r i c h e d a t m o s p h e r e m a i n t a i n e d by means of a s t o v e . •Ann. A p p l . B i o l . . 17:81-89. 135. SMITH, F.A. and J'.A. RAVEN. 1979. I n t r a c e l l u l a r pH and i t s r e g u l a t i o n . Ann. Rev. P l a n t P h y s i o l . 30:289-311. 136. SPENCER, P.W. and J . S . TITUS. 1973. A p p l e l e a f s e n e s c e n c e : l e a f d i s c compared t o a t t a c h e d l e a f . P l a n t P h y s i o l . 51:89-92. 137. STEMLER, A. and GOVINDJEE. 1973. B i c a r b o n a t e i o n as a c r i t i c a l f a c t o r i n p h o t o s y n t h e t i c oxygen e v o l u t i o n . P l a n t P h y s i o l . 52:119-123. 138. TAYLOR, A.O. and A.S. CRAIG. 1971. P l a n t s under c l i m a t i c s t e s s . I I . Low t e m p e r a t u r e , h i g h l i g h t e f f e c t s on c h l o r o p l a s t u l t r a s t r u c t u r e . P l a n t P h y s i o l . 47: 719-725. 139. TAYLOR, A.O. and J.A. ROWLEY. 1971. P l a n t s under c l i m a t i c s t r e s s . I . Low t e m p e r a t u r e , h i g h l i g h t e f f e c t s on p h o t o s y n t h e s i s . P l a n t P h y s i o l . 47:713-718. 140. TERRY, N. and A. ULRICH. 1973. E f f e c t s o f p h o s p h o r u s d e f i c i e n c y on t h e p h o t o s y n t h e s i s and r e s p i r a t i o n of l e a v e s of s u g a r b e e t . P l a n t P h y s i o l . 51:43-47. 141. TETLEY, R.M. and K.V. THIMANN. 1974. The m e t a b o l i s m o f o a t l e a v e s d u r i n g s e n e s c e n c e . I . R e s p i r a t i o n , 169 c a r b o h y d r a t e m e t a b o l i s m , and t h e a c t i o n - of c y t o k i n i n s . P l a n t P h y s i o l . 54: 294-303. 142. THIMANN, R.M. TETLEY, and B.M. KRIVAK. 1977. M e t a b o l i s m o f o a t l e a v e s d u r i n g s e n e s c e n c e . P l a n t P h y s i o l . 59:448-454. 143. THIMANN, K.V. and S.O. SATLER. 1979. R e l a t i o n between l e a f s e n e s c e n c e and s t o m a t a l c l o s u r e : s e n e s c e n c e i n l i g h t . P r o c . N a t l . A c a d . S c i . 76:2295-2298. 144. THIMANN, K.V. 1980. The s e n e s c e n c e of l e a v e s . In. : K.V. Thimann, e d . , S e n e s c e n c e i n P l a n t s . CRC. P r e s s . Boca B a t o n , F l a . pp. 87-109. 145. THOMAS, H. and J . L . STODDART. 1975. S e p a r a t i o n o f c h l o r o p h y l l d e g r a d a t i o n from o t h e r s e n e s c e n c e p r o c e s s e s i n l e a v e s of a mutant g e n o t y p e of meadow f e s c u e ( F e s t u c a  p r a t e n s i s L . ) . P l a n t P h y s i o l . 56:438-441. 146. THOMAS, H. and J . L . STODDART. 1980. L e a f s e n e s c e n c e . Ann. Rev. P l a n t P h y s i o l . 31:83-111. 147. THORNE, J.H. and RH.R. ROLLER. 1974. I n f l u e n c e of a s s i m i l a t e demand on p h o t o s y n t h e s i s , d i f f u s i v e r e s i s t a n c e s , ' t r a n s l o c a t i o n , and c a r b o h y d r a t e l e v e l s of so y b e a n l e a v e s . P l a n t P h y s i o l . 54:201-207. 148. TOGNONI, F., A.H. HALEVY, and S.H. WITTWER. 1967.. Growth o f bean and tomato p l a n t s a s e f f e c t e d by r o o t a b s o r b e d g r o w th s u b s t a n c e s and a t m o s p h e r i c c a r b o n d i o x i d e . P l a n t a 72:43-52 149. WAGGONER, P.E., D.M. MOSS, and J.D. HESKETH. 1963. R a d i a t i o n i n t h e p l a n t e n v i r o n m e n t and p h o t o s y n t h e s i s . A g r o n . J . 55:36-38. 150. WALCZAK, T. and H. GABRYS. 1981. The C02 e f f e c t on 1 i g h t - i n d u c e d c h l o r o p l a s t t r a n s l o c a t i o n s i n h i g h e r p l a n t l e a v e s . Z. P f l a n z e n p h y s i o l . 101 .: 367-375. 151. WAMPLE, R. L. and R. W. DAVIS. 1983. E f f e c t o f f l o o d i n g on s t a r c h a c c u m u l a t i o n i n c h l o r o p l a s t s of s u n f l o w e r ( H e l i a n t h u s annus L.) P l a n t P h y s i o l . 73:195-1 9 8 . 152. WHITE, H.L. 1930. C a r b o n d i o x i d e i n r e l a t i o n t o g l a s s h o u s e c r o p s . V. An a n a l y s i s o f t h e r e s p o n s e o f t h e tomato c r o p t o an a t m o s p h e r e e n r i c h e d w i t h c a r b o n d i o x i d e . Ann. A p p l . B i o l . 17:755-766. 153. WILSON, R.F., J . W. BURTON, J . A. BUCK, AND C. A. BRIM. 1978. S t u d i e s on g e n e t i c m a l e - s t e r i l e s o y b e a n s . 1. D i s t r i b u t i o n of p l a n t c a r b o h y d r a t e and n i t r o g e n - d u r i n g 1 70 d e v e l o p m e n t . P l a n t P h y s i o l . 61:838-841. 154. WITTENBACH, V.A., R.C.ACKERSON, R.T. GlAQUINTA, and R.R. HEBERT. 1980. Changes i n p h o t o s y n t h e s i s , r i b u l o s e b i s p h o s p h a t e c a r b o x y l a s e , p r o t e o l y t i c a c t i v i t y , and u l t r a s t r u c t u r e of soybean l e a v e s d u r i n g s e n e s c e n c e . C r o p S c i . 20:225-230. 155. WITTWER, S.H. and W. ROBB. 1964. C a r b o n d i o x i d e e n r i c h m e n t o f g r e e n h o u s e a t m o s p h e r e s f o r f o o d c r o p p r o d u c t i o n . E c o n . B o t . 18:34-56. 156. WITTWER, S.H. 1967. C a r b o n d i o x i d e and i t s r o l e i n p l a n t g r o w t h . P r o c e e d i n g s of t h e XVII I n t e r n a t i o n a l H o r t i c u l t u r a l C o n g r e s s 111:311-322. 157. WITTWER, S.H. 1983. R i s i n g a t m o s p h e r i c c a r b o n d i o x i d e and c r o p p r o d u c t i v i t y . H o r t S c i e n c e 18:667-673. 158. WONG, S.C. 1979. E l e v a t e d a t m o s p h e r i c p a r t i a l p r e s s u r e of c a r b o n d i o x i d e and p l a n t g r o w t h . O e c o l o g i a 44:68-74. 159. WONG, S . C , I.R. COWAN, and C D . FARQUHAR. 1979. S t o m a t a l c o n d u c t a n c e c o r r e l a t e s w i t h p h o t o s y n t h e t i c c a p a c i t y . N a t u r e 282:424-426. 160. WRIGHT, S.T.C. and R.W.P. HIRON. 1969. (+)- A b s c i s i c a c i d , t h e growth i n h i b i t o r i n d u c e d i n d e t a c h e d wheat l e a v e s f o l l o w i n g a p e r i o d of w i l t i n g . N a t u r e (London) 224:719-720. 161. WULFF, R.D. and B.R. STRAIN. 1982. E f f e c t s of C02 e n r i c h m e n t on growth and p h o t o s y n t h e s i s i n Desmodium  p a n i c u l a t u m . Can. J . B o t . 60:1084-1.091 162. WYSE, R. 1980. Growth o f s u g a r b e e t s e e d l i n g s i n v a r i o u s a t m o s p h e r e s of oxygen and c a r b o n d i o x i d e . C r o p S c i . 20:456-458. 163. ZEEVAART, J.A.D. 1979. R e g u l a t i o n of a s s i m i l a t e p a r t i t i o n i n g . I_n V I I . P a r t i t i o n i n g of A s s i m i l a t e s . Summary R e p o r t s of a Workshop h e l d a t M i c h i g a n S t a t e U n i v e r s i t y , E a s t L a n s i n g , M i c h i g a n . 171 APPENDIX A - PRIMARY DATA OF TRIAL 2 The p r i m a r y d a t a of t r i a l s 1 and 2 were n o t p o o l e d b e c a u s e of s i g n i f i c a n t b l o c k e r r o r ( s e e T a b l e 3 . 1 ) . Hence, t h e p r i m a r y d a t a f o r b o t h t r i a l s were a n a l y s e d s e p a r a t e l y . The p r i m a r y d a t a of l e a f a r e a o v e r t i m e ( F i g . A1) and p l a n t d r y w e i g h t o v e r t i m e ( F i g . A2) a r e g i v e n f o r t h e 4 c a r b o n d i o x i d e t r e a t m e n t s of t r i a l 2. As i n t r i a l 1, t h e l e a f a r e a and p l a n t d r y w e i g h t of e n r i c h e d p l a n t s i n c r e a s e d i n c o m p a r i s o n t o c o n t r o l s , w i t h t h e c h a n g e s i n p l a n t d r y w e i g h t b e i n g more p r o n o u n c e d t h a n t h e c h a n g e s i n l e a f a r e a . G e n e r a l l y , t h e t r e n d s i n l e a f a r e a and p l a n t d r y w e i g h t o v e r t i m e among c a r b o n d i o x i d e t r e a t m e n t s were s i m i l a r t o t h e c h a n g e s i n t h o s e p a r a m e t e r s i n t r i a l 1, a l t h o u g h t h e d a t a were more v a r i a b l e i n t r i a l 2. 172 0.24 -P L A N T AGE, d a y s FIGURE A1 Leaf area over time of plants grown at 340, 500, 1200, or 3000 ul l " 1 carbon dioxide. The data are from T r i a l 2. Each value is the mean of 4 plants, with standard deviation. Curves are cubic spline functions f i t ted to data for each carbon dioxide treatment 173 10 20 30 40 50 P L A N T A G E , days FIGURE A2 The dry weight over time of p l a n t s grown at 340, 500, 1200, or 3000 u l l " 1 carbon d i o x i d e . The data are from T r i a l 2. Each va lue i s the mean of 4 p l a n t s , w i th s tandard d e v i a t i o n . Curves are cub i c s p l i n e f u n c t i o n s f i t t e d to data fo r each carbon d i o x i d e t r ea tment . 

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