Open Collections

UBC Theses and Dissertations

UBC Theses Logo

UBC Theses and Dissertations

Insect growth inhibitors from asteraceous plant extracts Salloum, Gregory Stewart 1987

Your browser doesn't seem to have a PDF viewer, please download the PDF to view this item.

Item Metadata

Download

Media
831-UBC_1987_A6_7 S24.pdf [ 5.13MB ]
Metadata
JSON: 831-1.0097049.json
JSON-LD: 831-1.0097049-ld.json
RDF/XML (Pretty): 831-1.0097049-rdf.xml
RDF/JSON: 831-1.0097049-rdf.json
Turtle: 831-1.0097049-turtle.txt
N-Triples: 831-1.0097049-rdf-ntriples.txt
Original Record: 831-1.0097049-source.json
Full Text
831-1.0097049-fulltext.txt
Citation
831-1.0097049.ris

Full Text

INSECT GROWTH INHIBITORS FROM ASTERACEOUS PLANT EXTRACTS by GREGORY STEWART SALLOUM B.Sc,  Macdonald C o l l e g e o f M c G i l l U n i v e r s i t y , 1983  A THESIS SUBMITTED IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF MASTER OF SCIENCE  in THE FACULTY OF GRADUATE STUDIES (Department o f P l a n t  Science)  We a c c e p t t h i s t h e s i s as conforming to the r e q u i r e d  standard  THE UNIVERSITY OF BRITISH COLUMBIA A p r i l 1987 ^Gregory Stewart Salloum  In presenting this thesis in partial fulfilment of the requirements for an advanced degree at the University of British Columbia, I agree that the Library shall make it freely available for reference and study. I further agree that permission for extensive copying of this thesis for scholarly purposes may be granted by the head of my department or by his or her representatives.  It is understood that copying or  publication of this thesis for financial gain shall not be allowed without my written permission.  Department of  Plant  Science  The University of British Columbia 1956 Main Mall Vancouver, Canada V6T 1Y3 Date  April  28,  1987  Gregory Stewart Salloum A p r i l 1987 Department o f P l a n t S c i e n c e  INSECT GROWTH INHIBITORS FROM ASTERACEOUS PLANT EXTRACTS  ABSTRACT  P e t r o l and e t h a n o l i c e x t r a c t s o f s i x a s t e r a c e o u s weeds were added t o artificial  d i e t and screened f o r i n h i b i t i o n o f l a r v a l growth on v a r i e g a t e d  cutworm, Peridroma s a u c i a Artemisia  (Hbn.).  P e t r o l and e t h a n o l i c e x t r a c t s o f  t r i d e n t a t a and Chamomilla suaveolens and e t h a n o l i c e x t r a c t s o f  Chrysothamnus nauseosus and Centaurea d i f f u s a were h i g h l y i n h i b i t o r y a t f i v e times the n a t u r a l l y o c c u r r i n g c o n c e n t r a t i o n s .  The two C. suaveolens  e x t r a c t s and the e t h a n o l e x t r a c t o f _A. t r i d e n t a t a were a c t i v e a t t h e natural concentration 80% o f t h i s l e v e l .  (100%) and were f u r t h e r examined a t 20, 40, 60, and  I n h i b i t i o n o f l a r v a l growth was d i r e c t l y r e l a t e d t o  concentration  f o r each o f t h e t h r e e e x t r a c t s t e s t e d .  EC^Q'S ( e f f e c t i v e  concentration  t o i n h i b i t growth by 50% r e l a t i v e t o c o n t r o l s ) f o r the t h r e e  e x t r a c t s were 36-42% o f the n a t u r a l l y o c c u r r i n g l e v e l i n the p l a n t s . N u t r i t i o n a l i n d i c e s were c a l c u l a t e d f o r second i n s t a r P_. s a u c i a f e e d i n g on the a c t i v e e t h a n o l i c A^. t r i d e n t a t a e x t r a c t and the p e t r o l e x t r a c t from C. s u a v e o l e n s .  The r e l a t i v e growth r a t e (RGR) o f P_. s a u c i a  l a r v a e f e d the e t h a n o l i c e x t r a c t o f _A. t r i d e n t a t a i n a r t i f i c i a l  d i e t was  s i g n i f i c a n t l y lower than t h a t i n l a r v a e f e d d i e t w i t h t h e p e t r o l e x t r a c t o f C.  s u a v e o l e n s and l a r v a e on c o n t r o l d i e t .  D i e t a r y u t i l i z a t i o n was  s i g n i f i c a n t l y lower f o r l a r v a e f e d the _A. t r i d e n t a t a e x t r a c t .  R e s u l t s of a f i e l d  trial  i n d i c a t e d t h a t a s i n g l e treatment of _A.  t r i d e n t a t a e x t r a c t at the e q u i v a l e n t  of 0.2  g/ml  s i g n i f i c a n t l y b e t t e r than the c a r r i e r s o l v e n t water as measured by a v i s u a l damage e s t i m a t e .  could  p r o t e c t cabbage  (30% aq e t h a n o l ) or  distilled  An i n s e c t i c i d e s t a n d a r d ,  TM deltamethrin  (17.9  ug/1  w i t h 0.4%  Superspred  ), suppressed pest damage  s i g n i f i c a n t l y b e t t e r than the /A. t r i d e n t a t a - e x t r a c t treatment. A r e s i d u a l o v i p o s i t i o n deterrency field  results.  t o P i e r i s rapae was  Caged experiments i n the l a b o r a t o r y  o v i p o s i t i o n deterrency  found i n the  confirmed the  of the A^. t r i d e n t a t a e x t r a c t a t 0.2  g/ml.  O f f s p r i n g o f f i e l d - c o l l e c t e d P_. s a u c i a l a r v a e grew 2 . 5 - f o l d than l a r v a e from the l a b o r a t o r y c o l o n y .  However, d i e t with the  t r i d e n t a t a e x t r a c t i n h i b i t e d both f i e l d - c o l l e c t e d and  contact  laboratory  heavier _A. reared  s a u c i a l a r v a e e q u a l l y when compared to t h e i r r e s p e c t i v e c o n t r o l s f e d untreated  diet.  In summary, these r e s u l t s i n d i c a t e the p o t e n t i a l b e n e f i t of  using  s p e c i f i c unrefined  p l a n t e x t r a c t s f o r growth i n h i b i t o r s and o v i p o s i t i o n  deterrents  insect pests.  against  The  c o n t r i b u t i o n of i n d i v i d u a l  p h y t o c h e m i c a l s i n the _A. t r i d e n t a t a e t h a n o l i c e x t r a c t to growth i n h i b i t i o n or o v i p o s i t i o n d e t e r r e n c y  i s currently  iii  speculative.  TABLE OF CONTENTS  Page  I.  INTRODUCTION A.  II.  III.  Objectives  1 of Thesis  2  LITERATURE REVIEW  4  A.  Chemical B a s i s f o r P l a n t R e s i s t a n c e  to Insects  B.  Insect Bioassays of Phytochemicals  C.  Behavioral  D.  P h y t o c h e m i c a l s and P h y s i o l o g i c a l S t r e s s  13  E.  P l a n t E x t r a c t s i n Crop P r o t e c t i o n  15  Response t o P l a n t Defense Chemicals  5 8 10  MATERIALS AND METHODS  19  A.  Plant Extracts  19  B.  Biological  19  1.  I n i t i a l Screening  2.  B i o a s s a y o f Unextracted and E x t r a c t e d 21  Second S c r e e n i n g  21  Dose-Response B i o a s s a y s  21  1.  Dose-Response B i o a s s a y on P_. s a u c i a Neonates  21  2.  Dose-Response B i o a s s a y u s i n g t h e A l f a l f a  3. D.  19  Plant Material 3. C.  Screenings  Looper, Autographa c a l i f o r n i c a Speyer  22  Sensitivity  22  o f Older P_. s a u c i a Larvae  F i n a l Determination o f Extract f o r F i e l d T r i a l 1.  2.  D e t a i l e d Growth A n a l y s i s o f P. s a u c i a Larvae F e e d i n g on t r i d e n t a t a and C. s u a v e o l e n s Extracts Formulation of A c t i v e E x t r a c t s f o r F o l i a r A p p l i c a t i o n and S t a b i l i t y o f Crude E x t r a c t s  iv  22  22 23  E.  F.  G.  H.  F i e l d T r i a l o f t h e _A. t r i d e n t a t a e x t r a c t on Cabbage  23  L a b o r a t o r y E v a l u a t i o n o f the O v i p o s i t i o n Deterrence of _A. t r i d e n t a t a E x t r a c t on Caged P. rapae  25  Growth Comparison o f L a b o r a t o r y Reared and Wild C o l o n i e s o f P_. s a u c i a Larvae on Standard D i e t and D i e t s w i t h A d d i t i o n o f an E t h a n o l i c A_. t r i d e n t a t a Crude E x t r a c t  26  Phytochemical I n v e s t i g a t i o n  26  1.  2.  Chromatographic S e p a r a t i o n _A. t r i d e n t a t a E x t r a c t  of the E t h a n o l i c 26  Chromatography o f S e s q u i t e r p e n e Lactones P r e v i o u s l y I s o l a t e d from A^ t r i d e n t a t a  IV.  V.  RESULTS  29  A.  Laboratory Screenings  29  B.  D e t a i l e d Growth A n a l y s i s o f P_. s a u c i a Larvae on the C. s u a v e o l e n s and t h e k_. t r i d e n t a t a E x t r a c t s F i e l d T r i a l o f t h e _A. t r i d e n t a t a E x t r a c t on  42  C.  Cabbage  46  D.  O v i p o s i t i o n o f P_. rapae;  E.  Q u a l i t y o f Cabbage Heads from t h e F i e l d T r i a l  54  F.  Comparison o f W i l d Versus L a b o r a t o r y Reared  54  G.  Preliminary  VII. VIII.  L a b o r a t o r y Experiment  P_. s a u c i a  Phytochemical I n v e s t i g a t i o n  54  57  DISCUSSION A.  VI.  28  68  Screening Asteraceous E x t r a c t s f o r Insect Inhibitors  Growth 68  B.  P h y t o c h e m i c a l s and I n s e c t Growth I n h i b i t o r s  72  C.  F i e l d T r i a l s of Plant E x t r a c t s  76  CONCLUSIONS  84  BIBLIOGRAPHY  86  APPENDICES  99  v  LIST OF TABLES  Page Table I .  Table I I .  P l a n t s p e c i e s , components e x t r a c t e d , h a r v e s t l o c a t i o n s i n B r i t i s h Columbia and dates of m a t e r i a l bioassayed i n i n i t i a l s c r e e n i n g  20  E f f e c t s of weed e x t r a c t s i n c o r p o r a t e d i n t o a r t i f i c i a l d i e t on growth and s u r v i v a l of neonate P. s a u c i a i n an i n i t i a l s c r e e n i n g bioassay  30  T a b l e I I I . Growth and s u r v i v a l of P_. s a u c i a neonate l a r v a e fed on a r t i f i c i a l d i e t s w i t h u n e x t r a c t e d p l a n t powder or on the e x t r a c t e d marcs T a b l e IV.  T a b l e V.  Table VI.  31  E f f e c t s of s e l e c t e d weed e x t r a c t s i n c o r p o r a t e d i n t o a r t i f i c a l d i e t at natural concentrations on the weight and s u r v i v a l o f neonate P_. s a u c i a fed f o r 11 days  33  E f f e c t s of d i e t a r y _A. t r i d e n t a t a e x t r a c t (EtOH) and C_. s u a v e o l e n s e x t r a c t ( p e t r o l ) on 2nd i n s t a r P_. s a u c i a d i g e s t i b i l i t y o f food (AD), r e l a t i v e growth r a t e (RGR), r e l a t i v e consumption r a t e (RCR), and g r o s s (ECI) and net (ECD) d i e t a r y u t i l i z a t i o n s  A3  Mean growth and percent s u r v i v a l of neonate P_. s a u c i a l a r v a e f e d a r t i f i c i a l d i e t s i n c o r p o r a t i n g f r e s h and e i g h t month o l d e t h a n o l i c e x t r a c t s , hot water, room temp water and 20% aq e t h a n o l f o r m u l a t i o n s o f A^. t r i d e n t a t a and C_. s u a v e o l e n s f o r 16 days  45  T a b l e V I I . Mean v i s u a l q u a l i t y e s t i m a t e s of cabbage t r e a t e d w i t h an _A. t r i d e n t a t a e t h a n o l i c e x t r a c t , d e l t a m e t h r i n , 30% aq e t h a n o l , and water, recorded 25 days p o s t - t r e a t m e n t  55  T a b l e VIII.Mean P_. s a u c i a l a r v a l weight of a F^ f i e l d c o l l e c t e d p o p u l a t i o n compared t o the l a b o r a t o r y c o l o n y f e d the s t a n d a r d a r t i f i c i a l d i e t and d i e t c o n t a i n i n g a 50% _A. t r i d e n t a t a e t h a n o l i c e x t r a c t (dwt/dwt) f o r 8 days  56  T a b l e IX.  T a b l e X.  Mean l a r v a l weight of neonate P_. s a u c i a f e d a r t i f i c i a l d i e t admixed w i t h c h r o m a t o g r a p h i c a l l y separated f r a c t i o n s o f an A^. t r i d e n t a t a e t h a n o l i c e x t r a c t compared t o the o r i g i n a l e x t r a c t a t e c o l o g i c a l c o n c e n t r a t i o n s and the standard d i e t  58  P h y t o c h e m i c a l c o n s t i t u e n t s p r e v i o u s l y i s o l a t e d from Artemisia tridentata  74  LIST OF FIGURES  Page F i g u r e 1.  Percent growth (o) r e l a t i v e t o c o n t r o l growth and percent t o t a l m o r t a l i t y (•) o f P_. s a u c i a neonates f e d e t h a n o l i c e x t r a c t s from A) _A. t r i d e n t a t a , and B) C. suaveolens and a p e t r o l e x t r a c t from C) C. suaveolens admixed t o a r t i f i c i a l d i e t s (n=25 l a r v a e per concentration )  36  Percent growth (o) r e l a t i v e t o c o n t r o l growth and percent t o t a l m o r t a l i t y (•) o f s i x d a y - o l d £. saucia l a r v a e f e d e t h a n o l i c e x t r a c t s from A) A_. t r i d e n t a t a , and B) C. s u a v e o l e n s and a p e t r o l e x t r a c t from C) C. suaveolens admixed t o a r t i f i c i a l d i e t s (n=25 l a r v a e per c o n c e n t r a t i o n )  38  Percent growth (o) r e l a t i v e t o c o n t r o l growth and percent t o t a l m o r t a l i t y (•) o f neonate _A. c a l i f o r n i c a fed e t h a n o l i c e x t r a c t s from A) _A. t r i d e n t a t a , and B) C. suaveolens, and a p e t r o l e x t r a c t from C) C_. suaveolens admixed t o a r t i f i c i a l d i e t s (n=25 l a r v a e per c o n c e n t r a t i o n )  40  Percentage change i n cabbage l o o p e r e q u i v a l e n t s (CLE) a f t e r f i e l d s p r a y i n g cabbage w i t h a ) 30% aq e t h a n o l i c s o l u t i o n o f A. t r i d e n t a t a (0.2 g/ml), b) 30% aq e t h a n o l , c ) d e l t a m e t h r i n 2.5 EC (17 u g / l a . i . ) w i t h 0.1% Superspred or d) d i s t i l l e d water, J u l y 24, 1985.  48  Percent change i n imported cabbageworm, JP. rapae l a r v a l p o p u l a t i o n s b e f o r e and a f t e r f i e l d s p r a y i n g cabbage w i t h a ) 30% aq e t h a n o l i c s o l u t i o n o f A^. tridentata (0.2 g/ml), b) 30% aq e t h a n o l , c ) d e l t a m e t h r i n 2.5 EC (17 p g / l a . i . ) w i t h 0.1% Superspred or d) d i s t i l l e d water, J u l y 24, 1985.  50  Percent change i n imported cabbageworm, P_. rapae, o v i p o s i t i o n surveyed b e f o r e and a f t e r f i e l d s p r a y i n g cabbage with a) 30% aq e t h a n o l i c s o l u t i o n o f _A. tridentata (0.2 g/ml), b) 30% aq e t h a n o l , c ) d e l t a m e t h r i n 2.5 EC (17 p g / l a . i . ) w i t h 0.1% Superspred or d) d i s t i l l e d water, J u l y 24, 1985.  52  t  F i g u r e 2.  t  F i g u r e 3.  F i g u r e 4.  F i g u r e 5.  F i g u r e 6.  vii  F i g u r e 7.  T h i n - l a y e r chromatograph of f r a c t i o n s 2 (fr#2) and 4 (fr#4) from the s e p a r a t i o n of a crude e t h a n o l i c A . t r i d e n t a t a e x t r a c t chromatographed with phytochemicals from A r t e m i s i a spp. The pure s e s q u i t e r p e n e l a c t o n e s compared to the A^. t r i d e n t a t a f r a c t i o n s were: Dehydroleucodin ( d h l ) , d i h y d r o s a n t a m a r i n ( d h s ) , a r b u s c u l i n A (abA), a r b u s c u l i n C (abC), m a t r i c a r i n (mat), d e a c e t o x y m a t r i c a r i n (dom), d e a c e t y l m a t r i c a r i n (dam), and dehydroreynosin ( d h r ) . Nonf l u o r e s c e n t c o l o u r s o c c u r r e d a f t e r d e v e l o p i n g the p l a t e w i t h a v a n i l l i n reagent and the arrows i n d i c a t e a c o l o u r s h i f t a f t e r 24 h. TLC developed w i t h petroleum ether:CHCl2:Et20Ac (2:2:1) i n a non-saturated tank.  60  T h i n - l a y e r chromatograph of f r a c t i o n s 2 (fr#2) and 4 (fr#4) from the s e p a r a t i o n of a crude e t h a n o l i c _A. t r i d e n t a t a e x t r a c t chromatographed with phytochemicals from A r t e m i s i a spp. The pure s e s q u i t e r p e n e l a c t o n e s compared to the _A. t r i d e n t a t a f r a c t i o n s were: d e h y d r o l e u c o d i n ( d h l ) , d i h y d r o s a n t a m a r i n ( d h s ) , a r b u s c u l i n A (abA), t a t r i d i n A ( t t A ) , m a t r i c a r i n (mat), d e a c e t o x y m a t r i c a r i n (dom), and d e a c e t y l m a t r i c a r i n (dam). N o n - f l u o r e s c e n t c o l o u r s o c c u r r e d a f t e r d e v e l o p i n g the p l a t e with a v a n i l l i n reagent and the arrows i n d i c a t e a c o l o u r s h i f t a f t e r 24 h. TLC developed w i t h CHC13:acetone (6:1) i n a s a t u r a t e d tank.  62  T h i n - l a y e r chromatograph of f r a c t i o n s 2 (fr#2) and 4 ( f r # 4 ) from the s e p a r a t i o n of a crude e t h a n o l i c A^. t r i d e n t a t a e x t r a c t chromatographed w i t h phytochemicals from A r t e m i s i a spp. The pure s e s q u i t e r p e n e l a c t o n e s compared t o the A_. t r i d e n t a t a f r a c t i o n s were: Dehydroleucodin ( d h l ) , a r b u s c u l i n A (abA), a r b u s c u l i n B (abB), a r b u s c u l i n C (abC), t a t r i d i n A ( t t A ) , m a t r i c a r i n (mat), d e a c e t o x y m a t r i c a r i n (dom), d e a c e t y l m a t r i c a r i n (dam), and d e h y d r o r e y n o s i n ( d h r ) . N o n - f l u o r e s c e n t c o l o u r s o c c u r r e d a f t e r d e v e l o p i n g the p l a t e w i t h a v a n i l l i n reagent and the arrows i n d i c a t e a c o l o u r s h i f t a f t e r 24 h. TLC developed w i t h CHC1 :acetone (6:1) i n a s a t u r a t e d tank.  64  F i g u r e 10. T h i n - l a y e r chromatograph of f r a c t i o n s 2 (fr#2) and 4 (fr#4) from the s e p a r a t i o n of a crude e t h a n o l i c A^. t r i d e n t a t a e x t r a c t chromatographed w i t h phytochemicals from A r t e m i s i a spp. The pure s e s q u i t e r p e n e l a c t o n e s compared t o the _A. t r i d e n t a t a f r a c t i o n s were: a r b u s c u l i n A (abA), a r b u s c u l i n B (abB), t a t r i d i n A ( t t A ) , m a t r i c a r i n (mat), d e a c e t o x y m a t r i c a r i n (dom), and d e a c e t y l m a t r i c a r i n (dam). N o n - f l u o r e s c e n t c o l o u r s o c c u r r e d a f t e r d e v e l o p i n g the p l a t e w i t h a v a n i l l i n reagent and the arrows i n d i c a t e a c o l o u r s h i f t a f t e r 24 h. TLC developed w i t h petroleum ether:CHClo:Et 0Ac (2:2:1) i n a non-saturated tank  66  F i g u r e 8.  F i g u r e 9.  3  9  viii  ACKNOWLEDGEMENTS  I s i n c e r e l y thank my and  Dr. Murray B. Isman, f o r h i s guidance  c o n t i n u a l s u p p o r t , throughout t h i s p r o j e c t , and  development. C.  supervisor,  I earnestly  thank my  Runeckles f o r h i s a d v i c e  i n my  personal  t h e s i s committee, s p e c i f i c a l l y , Dr.  V.  i n the phytochemical i n v e s t i g a t i o n , Dr. G.  H.  N. Towers f o r h i s guidance on p o t e n t i a l p l a n t s p e c i e s t o e x t r a c t , and R.  S. Vernon f o r h i s a s s i s t a n c e d e s i g n i n g  the f i e l d  experiment.  I thank Susan Schwab f o r e x c e l l e n t t e c h n i c a l a s s i s t a n c e , Rick for sesquiterpene  l a c t o n e s , Gordon Ayer f o r P.  Dr.  s a u c i a eggs, Roy  Kelsey  Cranston rpw  f o r C_. d i f f u s a p l a n t s , R. and  S. Vernon f o r p r o v i d i n g cabbage seed and  G. H. N. Towers f o r use  p a r t by an NSERC o p e r a t i n g  of h i s l a b o r a t o r y . grant  (A2729) t o M.  I a l s o acknowledge J . C a r l s o n , Johnson and  D.  T h i s work was B.  Decis  supported i n  Isman.  Champagne, L. G i l k e s o n , C. Guppy, S.  the graduate s t u d e n t s i n the Department of P l a n t S c i e n c e  their assistance.  I extend a s p e c i a l thanks t o J i l l  c o n t i n u a l support during  for  E. Evans f o r p r o v i d i n g  the arduous c o m p l e t i o n of t h i s manuscript.  ix  ,  1  I.  Present l i b e r a l use  day  crop p r o t e c t i o n s t r a t e g i e s o f t e n r e l y h e a v i l y on  of broad-spectrum i n s e c t i c i d e s .  a r e beginning 1980).  Introduction  t o reduce the use  the  Novel pest management  of t h e s e n o n - s e l e c t i v e  biocides  (Huffaker  Improvements to crop p r o t e c t i o n methods are n e c c e s i t a t e d by  e n v i r o n m e n t a l , and insecticides  P u b l i c concern r e g a r d i n g  Basol  1980,  Metcalf  1980).  i n s e c t i c i d e s i s o f t e n the r e s u l t  chemophobia or the f e a r t h a t widespread use human h e a l t h .  The  of  of c h e m i c a l s w i l l damage the  f e a r i s j u s t i f i a b l e , f o r example, where  t o x i c i n s e c t i c i d e s have been misused c a u s i n g i n c l u d i n g groundwater c o n t a m i n a t i o n ( Z i t t e r  e n v i r o n m e n t a l damage, 1984).  A g r i c u l t u r a l agencies  could better inform l e g i t i m a t e i n t e r e s t s ( i . e . ,  the p u b l i c ) of the  and  Isman 1986)  b e n e f i t s of i n s e c t i c i d e use  provide The  o p t i o n s and  health,  economic concerns a s s o c i a t e d w i t h the use of s y n t h e t i c  (Luck e t a l . 1977,  environment and  tactics  (Czerwinski  a l t e r n a t i v e s t o the use  and  and  risks  be a b l e  to  of b i o c i d e s .  c o s t of i n s e c t i c i d e usage i s i n c r e a s i n g f o r s e v e r a l r e a s o n s .  Where i n s e c t i c i d e s are used i n t e n s i v e l y , i n s e c t s o f t e n develop r e s i s t a n c e to i n s e c t i c i d e s (Luck e t a l . 1977).  Almost i n v a r i a b l y , broad-spectrum  i n s e c t i c i d e s i n c r e a s e the development of r e s i s t a n t pest genotypes. i n s e c t i c i d e s are sprayed, major or minor pest s p e c i e s may problem i f t h e i r n a t u r a l enemies are e l i m i n a t e d . resurge  use h i g h e r  change c o n t r o l c h e m i c a l s . those no l o n g e r e f f e c t i v e .  When p e s t  populations other  doses, i n c r e a s e the frequency of s p r a y i n g , New  i n s e c t i c i d e s are thus needed to  I n s e c t s t h a t have a q u i r e d  e x i s t i n g i n s e c t i c i d e s are o f t e n p r e d i s p o s e d new  become a s e r i o u s  or develop r e s i s t a n c e t o an i n s e c t i c i d e , farmers and  a p p l i c a t o r s , may  When  i n s e c t i c i d e s (Devonshire and  or  replace  r e s i s t a n c e to  to develop c r o s s - r e s i s t a n c e to  Moore 1982).  Cross-resistance  has  2  decreased the average l i f e s p a n o f a new (Metcalf  1980).  m i l l i o n ) and  A d d i t i o n a l economic problems, such as c o s t  development time (8-10  1985), suggest t h a t i n n o v a t i v e p e s t s and  y e a r s ) f o r new  approaches a r e  products  years  (about  $25  (Kinoshita  needed t o c o n t r o l i n s e c t  manage e x t a n t i n s e c t i c i d e s ( C r o f t 1982).  Natural  p r o d u c t s from p l a n t s a r e  p r o t e c t i o n agents and as v e g e t a b l e o i l s , Greek and  i n s e c t i c i d e t o about two  p o t e n t i a l s o u r c e s of u s e f u l c r o p  novel pest c o n t r o l s t r a t e g i e s .  are known t o have been used f o r crop p r o t e c t i o n i n e a r l y  Roman a g r i c u l t u r e (Smith and  e x t r a c t s have been i n v e s t i g a t e d A l i Khan 1984) diseases.  and  P l a n t p r o d u c t s , such  fungal  (Kuc  Secoy 1975).  More r e c e n t l y ,  f o r c o n t r o l o f both v i r a l and  S h a i n 1977,  (Verma and  El-Shazly  However, e x t e n s i v e produced few  (Mclndoo and  Sievers  screenings for acutely  1924,  Abid  e t a l . 1981)  Numerous p l a n t e x t r a c t s have a l s o been examined f o r t h e i r  i n s e c t i c i d a l properties  plant  crop acute  Jacobson 1958).  t o x i c p h y t o c h e m i c a l s have  c o m m e r c i a l l y e x p l o i t e d b o t a n i c a l i n s e c t i c i d e s (Jacobson  and  Crosby 1971). A.  Objectives  The  of  Thesis  primary o b j e c t i v e of t h i s i n v e s t i g a t i o n was  to s c r e e n crude  e x t r a c t s from a s t e r a c e o u s weeds as p o t e n t i a l i n s e c t c o n t r o l a g e n t s .  Plants  i n the A s t e r a c e a e were s e l e c t e d because of t h e i r r i c h n e s s i n secondary m e t a b o l i t e s (Herout 1970). the v a r i e g a t e d i n s e c t pest  The  i n s e c t used i n the  cutworm, Peridroma s a u c i a  extract screening  (Hbn.), because i t i s a  of many c r o p s i n Canada ( B e i r n e  1971), and  is  c o n s i s t e n t l y and  a v a i l a b l e i n l a r g e q u a n t i t i e s i n Dr. M.  laboratory.  p l a n t s chosen ( T a b l e  current  The  economic v a l u e .  p o l a r and  The  non-polar s o l v e n t s  was  serious  reared  B.  Isman's  I ) were a l l weedy s p e c i e s w i t h  d r i e d p l a n t powders were e x t r a c t e d  no  w i t h both  to a s s e s s b i o l o g i c a l a c t i v i t y of f r a c t i o n s  3 containing  d i f f e r e n t phytochemical m i x t u r e s .  growth i n h i b i t i n g e x t r a c t s  showed the r e l a t i v e e f f e c t i v e n e s s of  e x t r a c t s as l a r v a l growth i n h i b i t o r s and i n h i b i t o r y response.  Dose-response of the most  the nature of the  further studied  determination of n u t r i t i o n a l i n d i c e s f o r larvae feeding admixed w i t h The  next o b j e c t i v e was  to prepare a f o r m u l a t i o n  extract's  e f f i c a c y against  cabbage i n s e c t  insured  the s u r v i v a l of the cabbage p l a n t s f o r the f i e l d  Two  laboratory  One  was  term p h y t o t o x i c  field  of the most a c t i v e  A short  t e s t on cabbage p r i o r t o the f i e l d  the f i e l d  observation  the cabbage t r e a t e d with A r t e m i s i a  b u t t e r f l i e s i n the l a b o r a t o r y .  The  e t h a n o l i c _A. t r i d e n t a t a e x t r a c t on from the final  laboratory  field  t r i d e n t a t a e x t r a c t using  caged  other compared the e f f e c t s of f i e l d - c o l l e c t e d VC  growth i n h i b i t i o n was  the  larvae r e l a t i v e to  colony.  s e c t i o n of t h i s t h e s i s i n v o l v e d an examination of  i n conjunction  trial.  of lowered P i e r i s rapae egg-counts  p h y t o c h e m i s t r y i n the most g r o w t h - i n h i b i t i n g separation  test  trial.  experiments were performed subsequent to the  to confirm  The  by  on a r t i f i c i a l d i e t  pests.  larvae  and  extracts.  e x t r a c t t o a s s e s s the  in  growth  To d i s t i n g i s h between g r o s s b e h a v i o r a l  p h y s i o l o g i c a l e f f e c t s , growth i n h i b i t i o n was  the  extract.  the  Chromatographic  with an i n s e c t b i o a s s a y i n v e s t i g a t e d whether  a t t r i b u t a b l e t o one  or s e v e r a l groups of compounds.  Chromatographic comparisons of pure compounds i s o l a t e d from c l o s e l y r e l a t e d p l a n t s w i t h a c t i v e f r a c t i o n s allowed t e n t a t i v e i d e n t i f i c a t i o n of c h e m i c a l c l a s s of compounds r e s p o n s i b l e  f o r the  growth  the  inhibition.  4 II.  L i t e r a t u r e Review  The l i t e r a t u r e on the i n t e r a c t i o n s between p l a n t s and t h e i r herbivores i s large.  insect  T h i s i n c l u d e s numerous reviews and books on the  b r e e d i n g o f p l a n t s r e s i s t a n t or t o l e r a n t t o i n s e c t a t t a c k ( P a i n t e r 1951, Maxwell and J e n n i n g s 1980, Hedin  1983), i d e n t i f i c a t i o n o f p l a n t  c h a r a c t e r i s t i c s c o n f e r r i n g i n s e c t r e s i s t a n c e ( T h o r s t e i n s o n 1960, Chapman 1974,  Rosenthal and Janzen  1979, D e t h i e r 1980, Hedin  1983), and the  p h y s i o l o g i c a l f a c t o r s t h a t a l l o w i n s e c t s t o p e r c e i v e and m e t a b o l i z e phytochemicals  ( F r a e n k e l 1959, B r a t t s t e n  1979, B e l l and Carde  a d d i t i o n , many c l a s s e s o f secondary compounds t h a t i n f l u e n c e  1984).  In  insect  behavior and p h y s i o l o g y have been examined (Chapman 1974, Rosenthal and Janzen  1979, Hedin 1983, Whitehead and Bower  1983).  S e v e r a l s t u d i e s have c o n s i d e r e d s u b l e t h a l ways o f m a n i p u l a t i n g i n s e c t p o p u l a t i o n s w i t h p h y t o c h e m i c a l s , i n c l u d i n g i n h i b i t i o n o f f e e d i n g (Jermy e t al.  1981), growth ( M c M i l l i a n e t a l . 1969) and o v i p o s i t i o n ( M i t c h e l l and  Heath 1985). Whittaker  These c h e m i c a l s a r e o f t e n termed  (1970) a s c h e m i c a l s t h a t mediate  ' a l l e l o c h e m i c s ' , d e f i n e d by  non-nutritional  interspecific  interactions. Many p h y t o c h e m i c a l s have been shown t o reduce i n s e c t growth, and p r o l o n g the l i f e c y c l e  (Chapman 1974, Reese and Beck 1976).  i n s e c t ' s l i f e c y c l e has been shown t o i n c r e a s e exposure (Wesloh  e t a l . 1983) and other e n v i r o n m e n t a l hazards  Extending an  to predators  (Courtney  1986).  Phytophagous i n s e c t s t h a t develop more s l o w l y than normal a r e more l i k e l y to  d i e o f d i s e a s e (Courtney 1981) and the e a r l y onset of w i n t e r (Chew  1975).  Natural m o r t a l i t y of f i r s t  i n s t a r O s t r i n i a n u b i l a l i s L. i s about  90%  (Beck 1960).  Introduction  of new, and enhancement of e x i s t i n g  mortality f a c t o r s at t h i s vulnerable i t s population  stage may play an important r o l e i n  b i o l o g y and thus i t s e f f e c t i n a g r i c u l t u r a l systems.  O v i p o s i t i o n i n t e r f e r e n c e i s another s u b l e t h a l mode through which a l l e l o c h e m i c a l s a i d plants i n escaping  insect herbivory.  Both h o s t and  non-host p l a n t e x t r a c t s have been shown t o d e t e r o v i p o s i t i o n o f s e v e r a l insect species  ( T i n g l e and M i t c h e l l 1984, Renwick and Radke 1985).  authors have r e c o g n i z e d  the importance o f u s i n g p h y t o c h e m i c a l  of i n s e c t b e h a v i o r t o p r o t e c t crops from h e r b i v o r y 1974,  Jermy 1983).  Many  disruptions  (Munakata 1970, Chapman  P l a n t a l l e l o c h e m i c a l s t h a t have s u b l e t h a l e f f e c t s on  i n s e c t p e s t s a r e p o t e n t i a l l y u s e f u l f o r a g r i c u l t u r e as v a l u a b l e c r i t e r i a or as a p p l i e d p r o t e c t a n t s , humans, e n v i r o n m e n t a l l y  provided  breeding  t h a t they a r e n o n - t o x i c t o  sound, economical t o produce and c o n v i e n i e n t t o  apply. A.  Chemical B a s i s  for Plant Resistance  to Insects  Some i n s e c t s feed on s e v e r a l d i f f e r e n t p l a n t f a m i l i e s whereas r e l y on a r e s t r i c t e d number o f p l a n t s o r even a s i n g l e host (Thorsteinson by i n s e c t s .  1960). Several  others  species  In a d d i t i o n , some p l a n t s p e c i e s a r e r a r e l y  attacked  f a c t o r s i n f l u e n c e t h e p a t t e r n of i n s e c t a t t a c k  available plant species.  on the  P l a n t a r c h i t e c t u r e , i n c l u d i n g s i z e , shape, and  c o l o r , as w e l l as p l a n t h a b i t a t and d i s t r i b u t i o n , a r e i m p o r t a n t considerations  i n host  s e l e c t i o n by phytophagous i n s e c t s .  chemical content of the plant i s o f t e n considered determining host-plant  specificity  (Haniotakis  However, t h e  t h e most i m p o r t a n t f a c t o r  and V o y a d j o g l o u 1978,  Hardman and E l l i s 1978, Rosenthal and Janzen 1979). Many c l a s s e s o f phytochemicals have been shown t o p l a y an important role i n insect-plant interactions.  Sugars, amino a c i d s and p r o t e i n s a r e  6 important f o r i n s e c t growth and stimulants  (Bernays and  development and  Simpson 1982,  Dethier  can play a r o l e as  1973).  These  feeding  ubiquitous  c o n s t i t u e n t s of p l a n t s are commonly known as primary m e t a b o l i t e s .  Their  broad d i s t r i b u t i o n s i n the p l a n t kingdom make them u n l i k e l y c a n d i d a t e s f o r i n s e c t host  s p e c i f i c i t y or p l a n t d e f e n s i v e  c h e m i s t r y (Bernays and  Chapman  1978). The  other  metabolites, not  major group of p h y t o c h e m i c a l s a r e c a l l e d  e i t h e r because they are  according The  These important p h y t o c h e m i c a l s a r e u s u a l l y  1973,  abiotic factors.  be i n f l u e n c e d by  The  differ  Duffey 1982), p r e v i o u s  the i n s e c t s p e c i e s examined ( E i s n e r  route  of e n t r y and  Isman and  Duffey  a t o x i c r e a c t i o n at a higher  The  i n s e c t s i n the same order  dose.  Nepetalactone, a  (Eisner  or no e f f e c t  1964).  d i s t r i b u t i o n of secondary c h e m i c a l s i s f r e q u e n t l y l i m i t e d to  p l a n t taxon, w h i l e o c c u r r i n g s p o r a d i c a l l y i n s y s t e m a t i c a l l y groups.  (1982)  e l i c i t a growth i n h i b i t o r y reponse a t  monoterpenoid, i s known t o r e p e l some i n s e c t s but have l i t t l e on other  Glucosinolates  Isman  the simultaneous o c c u r r e n c e of  Chapman 1978).  have shown t h a t a phytochemical may and  Chew 1980,  1964,  exposure to a l l e l o c h e m i c a l s (Jermy e t a l . 1982),  p h y t o c h e m i c a l s (Bernays and  concentration  depending  insect-allelochemical  Chew 1980), i n s e c t growth s t a g e (Reese 1979,  as w e l l as c o n c e n t r a t i o n ,  one  classified  e f f e c t s of a s i n g l e a l l e l o c h e m i c a l on i n s e c t s may  i n t e r a c t i o n may  other  1974,  t o t h e i r s t r u c t u r e or b i o s y n t h e t i c pathways.  on a v a r i e t y of b i o t i c and  Dethier  are  Numerous secondary  are known to mediate i n s e c t - p l a n t i n t e r a c t i o n s (Chapman  Hedin 1983).  and  p o o r l y understood or because they  i n v o l v e d i n primary metabolism, or both.  metabolites  "secondary"  or mustard o i l g l y c o s i d e s p r o v i d e  o c c u r r i n g o f t e n i n the B r a s s i c a c e a e  and  o c c u r r i n g o c c a s i o n a l l y i n the u n r e l a t e d  other  one  unrelated  a good example,  C a p p a r a l e s f a m i l i e s , but  plant family Caricaceae  (Bjorkman  7 1976).  S e s q u i t e r p e n e l a c t o n e s are another example, o c c u r r i n g widely i n  A s t e r a c e a e and al.  l e s s frequently  i n the Apiaceae and  the  Magnoliaceae (Heywood et  1977). S p e c i f i c a l l e l o c h e m i c a l s ( e . g . , o v i p o s i t i o n cues) can  have a  b e n e f i c i a l e f f e c t on oligophagous i n s e c t s ( i . e . , i n s e c t s having a r e s t r i c t e d range of food p l a n t s of r e l a t e d p l a n t o r d e r s  or even of a s i n g l e  genus), w h i l e at the same time c o n t r i b u t i n g t o the p l a n t ' s c h e m i c a l defence a g a i n s t more g e n e r a l Brassicaceae  herbivores.  A l l y l g l u c o s i n o l a t e , present  i n many  p l a n t s , i s innocuous to the growth of the c r u c i f e r  specialist  P i e r i s rapae, but i n h i b i t s the growth of the polyphagous Spodoptera eridania  , and  i s a c u t e l y t o x i c when fed t o the Apiaceae s p e c i a l i s t P a p i l i o  polyxenes ( B l a u e t a l . 1978). may  Even compounds p r e s e n t  cause a n t i b i o s i s (sensu P a i n t e r  of o l i g o p h a g o u s i n s e c t s . C u c u r b i t a c e a e deter tredecimnotata,  or reduce the f i t n e s s and  t r i t e r p e n o i d c u c u r b i t a c i n s of  vigor  f e e d i n g by the c u c u r b i t - s p e c i a l i s t E p i l a c h n a  ( C a r r o l l and  beetle,  Hoffman 1980).  i n s e c t s r e l y e n t i r e l y on s p e c i f i c chemical cues f o r  or o v i p o s i t i o n .  plants  the  but a c t as a t t r a c t a n t s to the s t r i p e d cucumber  Acalymma v i t t a t a Very few  The  1951)  i n t h e i r host  feeding  Most oligophagous i n s e c t s do not appear t o r e l y on  the  presence or absence of a s i n g l e compound f o r host a c c e p t a b i l i t y , but t h e i r chemosensory response to the t o t a l p h y t o c h e m i c a l mixture 1973).  Studies  of food consumption by c r u c i f e r o u s f l e a  P h y l l o t r e t a spp.,  on  (Dethier  beetles,  show t h a t the amount consumed i s u s u a l l y dependent on  b a l a n c e of s t i m u l a n t  and  deterrent  chemicals (Nielsen  the  1978).  Although the e f f e c t s of phytochemicals have o f t e n been i n v e s t i g a t e d i n d i v i d u a l l y , an i n s e c t ' s sensory p e r c e p t i o n  of t h e i r n a t u r a l h a b i t a t must  i n c l u d e the complexity of chemical m i x t u r e s i n non-host p l a n t s .  Many  8 p l a n t s have compounds t h a t , i n combination with o t h e r phytochemicals, increase  i n s e c t a n t i b i o s i s more so than a s i n g l e compound alone (Adams and  Bernays 1978, B.  Insect  Kubo e t a l . 1984,  b i o a s s a y s have been used t o d e t e c t  agents i n p l a n t e x t r a c t s . including mortality  utilization  egg  (Isman and  B l a i s i n g e r 1978)  B i o a s s a y s may  and  Proksch 1985), r e p r o d u c t i v e  or  and  p o t e n t i a l (Robert  'no-choice' experiments.  by e i t h e r an  B i o a s s a y s may 'all-or-none'  on which t o feed i n s e c t has  be designed so t h a t  and  growth i n h i b i t i o n may  p l a n t powders or e x t r a c t s i n a r t i f i c i a l i n s e c t s (Hsiao and  or a graded type of and  Fraenkel  weighed.  1968).  be examined by  d i e t s and  feeding  After feeding,  Bernays (1983) and  the  incorporating  the  d i e t s to t e s t  surviving larvae  p r o c e s s of e x t r a c t i n g a l l e l o c h e m i c a l s from p l a n t  incorporating  them i n t o a r t i f i c a l  d i e t s could  p o t e n t i a l l y i n c r e a s i n g or d e c r e a s i n g Chapman 1978).  An  alter  mask the  tissue  and  allelochemicals,  their effectiveness  a r t i f i c i a l d i e t may  are  Smith (1978) d i s c u s s c r i t e r i a f o r  c h o o s i n g between d i f f e r e n t b i o a s s a y methods. The  the  ' a l l or none' response.  L a r v a l feeding  counted and  or  a single  L a r v a l growth, f o r example, i s u s u a l l y a graded measure i s an  occur  Choice experiments o f f e r  or more s u b s t r a t e s  or o v i p o s i t i o n a l s u b s t r a t e .  response.  and  e t a l . 1982).  In the so c a l l e d no-choice experiments the  i n s e c t s are evaluated  oviposition  of b e h a v i o r a l l y a c t i v e a l l e l o c h e m i c a l s may  the t e s t i n s e c t a s e l e c t i o n of two  mortality  measure a v a r i e t y of f a c t o r s ,  consumption ( B e n t l e y  form of c h o i c e  control  counts (Jacobson e t a l . 1978), growth, d i e t a r y  Laboratory evaluation  oviposit.  phytophagous pest  (Freedman e t a l . 1979), f e e d i n g  p u n c t u r e s , l a r v a l and  feeding  Neal 1985).  B i o a s s a y s of Phytochemicals  Several  i n the  Berenbaum and  (Bernays  presence of  and  feeding  9 deterrents increased  (Bernays and feeding  Chapman 1977).  The  masking may  s t i m u l a t i o n , absence of o t h e r f i t n e s s - r e d u c i n g compounds  or a b e t t e r n u t r i e n t source than h o s t p l a n t s . artificial  However, i n s e c t s fed  d i e t s provide a good r e l a t i v e measure of l a r v a l growth  i n h i b i t i o n using extracted  phytochemicals.  a l l e v i a t e s problems i n h e r e n t  i n the use  E x p e r i m e n t a l e r r o r may  The  use  of a r t i f i c i a l  of l i v e p l a n t m a t e r i a l s  d i r e c t comparisons with other c h e m i c a l l y  the  be a r e s u l t of  defined  i n c r e a s e w i t h the use  food  diets  and  allows  sources.  of l i v e p l a n t m a t e r i a l  potential for large allelochemical differences  (Risch  1985),  due  to  and  p h y s i c a l d i f f e r e n c e s l i k e l e a f toughness (Reese 1983). Although many compounds i n d i f f e r e n t c h e m i c a l c l a s s e s have been shown to i n h i b i t i n s e c t growth and  increase  development time, b i o a s s a y s do  o f t e n d i s t i n g i s h between g r o s s l y d i f f e r e n t modes of a c t i o n . c o u n t s , f o r example, may they do not  be e x c e l l e n t f o r d e t e c t i n g  d i s t i n g i s h between b e h a v i o r a l  between treatments.  Behavioral  and  feeding  Fecal  not  pellet  i n h i b i t o r s but  physiological differences  reasons f o r reduced f e c a l p e l l e t counts i n  a treatment i n c l u d e reduced p h a g o s t i m u l a t i o n and  feeding  deterrency.  P o s s i b l e p h y s i o l o g i c a l causes f o r reduced f e c a l p e l l e t c o u n t s r e l a t e to t o x i c i t y , i n c l u d i n g reduced food  u t i l i z a t i o n and  i n h i b i t i o n of metabolism.  Cockroaches are known to i n c r e a s e consumption to compensate f o r d i l u t e d with a non-nutritive, non-toxic c e l l u l o s e f i l l e r In a d d i t i o n , R i s c h  (1985) has  shown t h a t f e e d i n g  depending upon whether l e a f d i s k s or whole l e a v e s  food  ( B i g n e l l 1978).  p r e f e r e n c e s can are used i n  change,  feeding  bioassays. Unless behavioral  and  p h y s i o l o g i c a l e f f e c t s can  be s e p a r a t e d , the mode  of a c t i o n at even a s u p e r f i c i a l l e v e l remains s p e c u l a t i v e . e f f i c i e n c y s t u d i e s are used to d i s t i n g i s h between b e h a v i o r a l p h y s i o l o g i c a l components of i n s e c t growth i n h i b i t i o n  Dietary and  (Reese 1979,  Isman  and  10 Duffey  1982, Isman and Rodriguez 1984) as w e l l as d e t e r m i n i n g  of an i n s e c t ' s food source Kogan and Cope 1974).  (Soo Hoo and F r a e n k e l  the adequacy  1966, Waldbauer 1968,  I n d i c e s o f d i e t a r y u t i l i z a t i o n a r e c a l c u l a t e d from  measurements o f food consumption, weight g a i n , and e x c r e t a p r o d u c t i o n . growth r a t e may then be separated and  dietary u t i l i z a t i o n .  bioassay C.  i n t o i t s c o n s t i t u e n t s , consumption r a t e  Some 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 w i t h  this  method have r e c e n t l y been examined by Schmidt and Reese (1986).  B e h a v i o r a l Response t o P l a n t Defence Chemicals  Feeding  and o v i p o s i t i o n i n phytophagous i n s e c t s a r e r e g u l a t e d by  s e v e r a l f a c t o r s , such a s chemostimulants ( H s i a o 1969) and d e t e r r e n t s and  The  (Shurr  Holdaway 1970, Renwick and Radke 1985) i n host and non-host p l a n t s  (Jermy 1966, Bernays 1983, Thibout  and Auger 1983).  The term 'non-  p r e f e r e n c e ' has o f t e n been used t o d e s c r i b e a b e h a v i o r a l non-event but i s l e s s than i d e a l due t o i t s a n t h r o p o c e n t r i c s p e c i f y a substance t h a t when c o n t a c t e d  bias.  prevents  a c t i v i t y , i n c l u d i n g f e e d i n g or o v i p o s i t i o n (sensu  'Deterrent'  i s used t o  or i n t e r r u p t s b e h a v i o r a l Schoonhoven 1982).  A  ' s t i m u l a n t ' f o r t h e purpose o f t h i s d i s c u s s i o n i s the antonym of d e t e r r e n t and  i s a substance t h a t , when p h y s i c a l l y c o n t a c t e d ,  incites a positive  b e h a v i o r a l response such as f e e d i n g or o v i p o s i t i o n . Two d i f f e r e n t n e u r a l events may produce t h e same b e h a v i o r a l r e s p o n s e . A d e t e r r e n t may a c t d i r e c t l y on a chemoreceptor (Schoonhoven 1982) or by masking the e f f e c t of a chemostimulant ( M i t c h e l l and S u t c l i f f e  1984).  Chemoreceptors i n silkworm, Bombyx mori, l a r v a e c o n t a i n s p e c i a l i s t t h a t respond d i r e c t l y t o both s t i m u l a n t s and d e t e r r e n t s  (Ishikawa  cells 1966).  Sparteine, a phytoalkaloid feeding i n h i b i t o r , i s responsible f o r inhibiting  the response of t h e s u g a r - s e n s i t i v e c e l l .  A l a c k o f response  from the r e c e p t o r d e t e c t i n g b e h a v i o r a l s t i m u l a n t s w i l l produce t h e same  11  response as a d e t e r r e n t and  ( M i t c h e l l and S u t c l i f f e  1984).  Both t h e c e n t r a l  the p e r i p h e r a l nervous system have i m p o r t a n t r o l e s i n the f e e d i n g  behavior  of herbivorous  i n s e c t s (Dethier  1980).  Many i n s e c t s examined do n o t have s p e c i f i c r e c e p t o r s f o r i n d i v i d u a l chemicals.  Dethier  feeding deterrents  (1973, 1980) h y p o t h e s i z e s i n v o l v e s the p r o c e s s i n g  several receptors.  Herbivore  that neuro-reception  of  of information c e n t r a l l y  g e n e r a l i s t s and s p e c i a l i s t s may have a  s i m i l i a r c a p a c i t y t o d e t e c t c h e m i c a l s but t h e i n f o r m a t i o n may be differently stimulants  ( D e t h i e r 1980).  from  processed  P h y t o c h e m i c a l s may thus s e r v e as f e e d i n g  t o some phytophagous i n s e c t s w h i l e i n h i b i t i n g f e e d i n g and growth  i n non-adapted s p e c i e s . P l a n t r e s i s t a n c e t o i n s e c t s may o f t e n be t r a c e d t o p h y t o c h e m i c a l defenses.  Host-plant  c h e m i c a l s may a i d p l a n t b r e e d e r s by f o c u s i n g  a t t e n t i o n on f a c t o r s c o n t r i b u t i n g t o a r t h r o p o d t h a t may p r e v e n t arthropod  aphid  e t a l . 1985).  L e p t i n o t a r s a decemlineata, are deterred i n Solanum chacoense (Sinden  O v i p o s i t i o n deterrents, present 1960,  e t a l . 1975, de P o n t i  crops 1977).  from b a r l e y , i s r e s p o n s i b l e f o r r e s i s t a n c e t o t h e  S c h i z a p h i s graminum (Zuniga  present  Allelochemicals  a t t a c k have been i d e n t i f i e d i n s e v e r a l  i n c l u d i n g tomatoes and cucumbers ( P a t t e r s o n Gramine, an a l k a l o i d  resistance.  their  H s i a o and F r a e n k e l  Colorado potato  beetles,  from f e e d i n g by g l y c o a l k a l o i d s  e t a l . 1986). i n many p l a n t s (Gupta and T h o r s t e i n s o n  1968a), can occur  c h e m i c a l s t h a t a r e r e l e a s e d upon f e e d i n g .  as c u t i c u l a r components or as Field  t e s t s with  the melonworm,  D i a p h a n i a h y a l i n a t a , and the pickleworm, D i a p h a n i a n i t i d a l a s , showed t h a t the p r i n c i p l e mechanism of r e s i s t a n c e i n two v a r i e t i e s o f b u t t e r n u t C u r c u r b i t a moschata, was o v i p o s i t i o n d e t e r r e n c y c o r n b o r e r , 0_. n u b i l a l i s ,  ( E l s e y 1985).  squash,  European  females a v o i d o v i p o s i t i n g i n f i e l d s where damage  to c o r n r e l e a s e s host v o l a t i l e s  (Shurr  and Holdaway 1970).  12 O v i p o s i t i o n and plants.  Spraying  mating and 1985)  and  f e e d i n g d e t e r r e n t s a r e o f t e n present  p l a n t e x t r a c t s as i n s e c t c o n t r o l agents has  o v i p o s i t i o n of oligophagous (Robert polyphagous i n s e c t s ( B u r n e t t and  sesquiterpene  i n non-host  and  B l a i s i n g e r 1978,  Jones 1978).  l a c t o n e , g l a u c o l i d e A, present  inhibited  The  i n Vernonia  non-host  spp.  d e t e r s f e e d i n g by l a r v a e of s e v e r a l s p e c i e s of polyphagous  Dover  (Asteraceae),  lepidopterans  ( B u r n e t t e t a l . 1974). P h y t o c h e m i c a l s may occurring mixtures. non-toxic  show b e h a v i o r a l d e t e r r e n c y  Woodhead and  only as n a t u r a l l y  Bernays (1977) have shown t h a t s e v e r a l  p h e n o l i c compounds a t n a t u r a l c o n c e n t r a t i o n s  deterrency  o n l y when combined.  i s o l a t e d , they  Even where dominant compounds have been  seldom account f o r host  i n s e c t s (Berenbaum 1985).  produce f e e d i n g  d i s c r i m i n a t i o n even i n o l i g o p h a g o u s  Although g l u c o s i n u l a t e s a r e known to s t i m u l a t e  some c r u c i f e r f e e d i n g c a t e r p i l l a r s , the t o t a l response can be a t t r i b u t e d t o more than one  group of phytochemicals (Gupta and T h o r s t e i n s o n  Furthermore, N i e l s e n (1978a) s t a t e s t h a t the a c c e p t a b i l i t y of c r u c i f e r host p l a n t s to f l e a b e e t l e s , P h y l l o t r e t a spp.,  their  c o u l d not  accounted f o r s o l e l y on the b a s i s o f g l u c o s i n o l a t e c o n t e n t , due  1960).  be  but was  likely  to a composite of a l l e l o c h e m i c a l s . P l a n t s commonly c o n t a i n f e e d i n g d e t e r r e n t s  Isman and  Duffey  1982).  I n s e c t s , however, do not always a v o i d  because of t o x i c phytochemicals. harmless p l a n t s and plants.  (Woodhead and  c o n v e r s e l y may  I n s e c t s may  Bernays  1977,  plants  be d e t e r r e d from f e e d i n g  on  be i n t o x i c a t e d by consuming poisonous  C e r t a i n tomato c u l t i v a r s t h a t c o n t a i n the t o x i c g l y c o a l k a l o i d ,  tomatine, a r e consumed with impunity a n t i d o t a l e f f e c t of f o l i a r s t e r o l s  by H e l i o t h i s zea l a r v a e because of  (Campbell and  Duffey  1981).  With  c a u t e r i z e d chemoreceptors, tobacco hornworm l a r v a e , Manduca s e x t a , consume n o n - t o x i c  p l a n t s p r e v i o u s l y avoided  the  (de Boer e t a l . 1977).  readily  13 P h y t o c h e m i c a l s , such as f e e d i n g allow  deterrents,  t h a t p r o t e c t a p l a n t and  s u r v i v a l o f s u s c e p t i b l e i n s e c t genotypes may p r o t e c t p l a n t s  longer  than potent chemicals that q u i c k l y s e l e c t f o r r e s i s t a n t i n s e c t genotypes. Gould (1986), i n h i s s i m u l a t i o n model t o p r e d i c t the d u r a b i l i t y of wheat germplasm r e s i s t a n t t o the H e s s i a n f l y , M a y e t i o l a  destructor, indicated  t h a t t h e most d u r a b l e r e s i s t a n c e would occur i f t o t a l l y c u l t i v a r s are planted  with a r e s i s t a n t c u l t i v a r .  susceptible  T h i s suggests t h a t  plant  r e s i s t a n c e w i l l be more durable when i n s e c t s a r e not under severe s e l e c t i o n pressure,  such as would occur when a monoculture o f a r e s i s t a n t c u l t i v a r i s  planted.  P l a n t e x t r a c t s used i n t h e f i e l d may mimic t h e p h y t o c h e m i c a l  p r o f i l e o f a mixed c r o p p i n g D.  system.  Phytochemicals and P h y s i o l o g i c a l S t r e s s  P l a n t defense c h e m i c a l s cause many adverse p h y s i o l o g i c a l e f f e c t s on i n s e c t s , f o r example reduced d i g e s t i o n , s u p p r e s s i o n  o f microsomal enzymes,  d i s r u p t i o n o f endosymbiotic organisms, i n t e r f e r e n c e with hormonal processes,  reduction of reproductive  Immature i n s e c t s a r e c o n f r o n t e d  c a p a c i t y and death. w i t h c h e m i c a l and p h y s i c a l p l a n t  d e f e n s e s when o b t a i n i n g v i t a l n u t r i e n t s .  Dietary nitrogen  and water, a r e  o f t e n t h e most important f a c t o r s l i m i t i n g l a r v a l growth (Mattson 1980, S c r i b e r and Slansky  1981).  Many a d u l t female i n s e c t s o v i p o s i t on or near p o t e n t i a l food The  plants.  g r a v i d female must be g e n e t i c a l l y 'wired' t o d i s c r i m i n a t e among  p o t e n t i a l l a r v a l food p l a n t s and t h e l a r v a e must be a b l e t o d e t e c t and avoid  i n g e s t i o n of t o x i n s .  In f a c t , neonate l e p i d o p t e r a n  larvae are highly  s u s c e p t i b l e t o a l l e l o c h e m i c a l s (Reese 1979) p o s s i b l y because they have l e s s a c t i v e or fewer d e t o x i f i c a t i v e enzymes, a t h i n p e r i t r o p h i c gut membrane or l a c k endosymbiotic organisms f o r x e n o b i o t i c d e t o x i f i c a t i o n .  The  ubiquitous  m i x e d - f u n c t i o n oxidase (MFO) system i s the major  d e t o x i f i c a t i o n system i n i n s e c t s (Ahmad 1986, Dauterman and Hodgson 1978). Adaptation  t o an a l l e l o c h e m i c a l may r e s u l t from an i n c r e a s e d  e f f i c i e n c y of  the n o n s p e c i f i c MFO system. Polyphagous i n s e c t s , such as some l e p i d o p t e r a n  l a r v a e , are p o t e n t i a l l y  exposed t o a broad range o f s u b l e t h a l p l a n t t o x i n s and may be b e t t e r to detoxify ubiquitous  able  f i t n e s s - r e d u c i n g a l l e l o c h e m i c a l s than o l i g o p h a g o u s  i n s e c t s , as the former possess higher  gut MFO l e v e l s ( K r i e g e r e t a l . 1971).  S p e c i f i c a l l e l o c h e m i c a l s , however, may be b e t t e r d e a l t w i t h by s p e c i a l i s t i n s e c t s f e e d i n g on t h e i r host p l a n t s  ( B l a u e t a l . 1978).  How does t h e metabolism o f x e n o b i o t i c s insecticides?  Insect resistance often i n v o l v e s increased  a c t i v i t y and when t h i s occurs,  Spider  enzymatic  cross-resistance to chemically  compounds i s q u i t e common (Agosin 1982).  a f f e c t insect resistance to  and P e r r y  mites bred f o r t o l e r a n c e  unrelated  1974, Devonshire and Moore  t o an i n s e c t r e s i s t a n t cucumber  v a r i e t y were, i n t e r e s t i n g l y , c r o s s - r e s i s t a n t t o s e v e r a l i n s e c t i c i d e s and a v a r i e t y of unrelated investigated  p l a n t s , but t h e mechanism o f r e s i s t a n c e was n o t  (Gould e t a l . 1982).  S p e c i a l i s t s and g e n e r a l i s t s may s e q u e s t e r a wide v a r i e t y o f t o x i c phytochemicals. causing  Sequestration  damage t o t h e i n s e c t .  may e f f e c t i v e l y prevent t h e x e n o b i o t i c The monarch b u t t e r f l y , Danaus  which s e q u e s t e r s c a r d i a c g l y c o s i d e s  from i t s milkweed h o s t ,  from  plexippus, i s an e x c e l l e n t  example (Roeske e t a l . 1976). N i c o t i n e i s h i g h l y t o x i c t o many i n s e c t s but some i n s e c t p e s t s o f tobacco a v o i d 1964,  t o x i c i t y by e f f i c i e n t metabolism and e x c r e t i o n ( S e l f e t a l .  B r a t t s t e n 1979).  L-Canavanine i s an abundant n o n - p r o t e i n  amino a c i d  found i n seeds o f the C e n t r a l American legume, D i o c l e a megacarpa, and i s h i g h l y t o x i c t o most i n s e c t s .  However the b r u c h i d ,  Caryedes b r a s i l i e n s i s ,  15 not  only f e e d s on J). megacarpa seeds but uses the a r g i n i n e a n a l o g , L -  canavanine, i n p r o t e i n p r o d u c t i o n ( R o s e n t h a l e t a l . 1982). One must be c a u t i o u s when drawing a n a l o g i e s between the a r t h r o p o d response t o i n s e c t i c i d e s , and t h e i r response t o p h y t o c h e m i c a l m i x t u r e s . I n s e c t s r e s i s t a n t t o a s i n g l e p y r e t h r o i d can develope s t r o n g  cross-  r e s i s t a n c e t o o t h e r p y r e t h r o i d s ( P r i e s t e r and Georghiou 1980) and o t h e r c l a s s e s o f i n s e c t i c i d e s ( F u n a k i and Motoyama 1986).  In c o n t r a s t ,  d e t o x i f i c a t i o n of p h y t o c h e m i c a l m i x t u r e s i n p l a n t s has r e c e i v e d attention.  However, Gould e t a l . (1982) have shown t h a t  several  scant  organophosphorous  r e s i s t a n t m i t e s a r e as s e n s i t i v e t o a t o x i c h o s t p l a n t as a r e s u s c e p t i b l e strains.  These r e s u l t s suggest t h a t t h e d e t o x i f i c a t i o n mechanism o f  a l l e l o c h e m i c a l m i x t u r e s i s d i f f e r e n t from organophosphorous and t h a t o t h e r mechanisms may E.  insecticides  be i n v o l v e d .  P l a n t E x t r a c t s i n Crop P r o t e c t i o n  P l a n t p r o d u c t s have been used t o c o n t r o l i n s e c t s s i n c e man cultivating plants.  first  began  An e x t r a c t from the f l o w e r s o f Chrysanthemum  c i n e r a r i a e f o l i u m , c a l l e d pyrethrum, i s perhaps the most w i d e l y used i n s e c t i c i d a l plant product. 1916  ( M a l l i s 1982).  Pyrethrum was  Rotenone,  first  s o l d i n North America i n  from D e r r i s spp. and Lonchocarpus spp., and  n i c o t i n e , from N i c o t i a n a r u s t i c a , a r e o t h e r c o m m e r c i a l l y a v a i l a b l e i n s e c t i c i d a l plant constituents.  Other b o t a n i c a l i n s e c t i c i d e s a r e used  mainly where they a r e i n d i g e n o u s , i n c l u d i n g Ryania s p e c i o s a , tung seed ( A l e u r i t e s spp.) and s a b a d i l l a from Schoenocaulon spp.(Jacobson and Crosby 1971). R e c e n t l y , crude e x t r a c t s and i s o l a t e d  p h y t o c h e m i c a l s from the neem  t r e e , A z a d i r a c h t a i n d i c a , have been i n v e s t i g a t e d as i n s e c t c o n t r o l a g e n t s . A z a d i r a c h t i n , a limonoid i s o l a t e d  from neem, c o m p l e t e l y i n h i b i t e d  feeding  16 of the migratory  l o c u s t , S c h i s t o c e r c a g r e g a r i a , a t l e v e l s as low as 1  2 ng/cm  on l e a f d i s k s (Kubo and N a k a n i s h i  P r o t e c t i o n Agency of the U n i t e d formulation  (Larson  1977).  The E n v i r o n m e n t a l  S t a t e s has r e g i s t e r e d a  patented  1985) o f neem o i l f o r use on non-food c r o p s , and  r e g i s t r a t i o n on food crops  i s pending (Jacobson 1986).  Both pyrethrum and neem o i l c o n t a i n more than one i n s e c t i c i d a l compound.  The pyrethrum a c t i v i t y i s d e r i v e d  from s i x p y r e t h r i n e s t e r s  ( E l l i o t and Janes 1973) and the i n s e c t i c i d a l e f f e c t s o f neem o i l a r e a r e s u l t o f s e v e r a l t e t r a n o r t r i t e r p e n o i d s (Jacobson 1986). B o t a n i c a l s may p r o v i d e agents.  entomologists  Mixtures of defensive  with  novel c r o p  c h e m i c a l s have evolved  evidence suggests p l a n t s m i t i g a t e damage by having p h y t o c h e m i c a l s (Berenbaum 1985).  protection  i n p l a n t s and some  combinations o f  Adams and Bernays (1978) showed t h a t  f o u r t e e n phytochemicals i n n a t u r a l l y o c c u r r i n g c o n c e n t r a t i o n s produce a measurable e f f e c t on L o c u s t a alone  but d e t e r r e d  evolved  m i g r a t o r i a f e e d i n g when  f e e d i n g when presented  as a mixture.  presented  I f plants,  over m i l l i o n s o f y e a r s , use m i x t u r e s o f chemicals t o defend  herbivory  perhaps we can a l s o use t h i s  'novel'  d e t e r r e n t s ) may p r o v i d e from h e r b i v o r y  (Bernays 1983).  having against  strategy.  N a t u r a l l y o c c u r r i n g i n s e c t growth i n h i b i t o r s  crops  d i d not  (e.g.,  feeding  e f f e c t i v e t o o l s f o r crop management by p r o t e c t i n g  while a v o i d i n g d e s t r u c t i o n of b e n e f i c i a l i n s e c t s  Recent s t u d i e s suggest t h a t s y n t h e t i c i n s e c t i c i d e s c o n f e r  p a r t o f t h e i r b e n e f i t due t o s u b l e t h a l e f f e c t s .  Aldicarb at sublethal  doses r e d u c e s t h e a b i l i t y t o f l y and probe, as w e l l as the f e c u n d i t y , o f potato  aphids,  Macrosiphum euphorbiae ( B o i t e a u e t a l . 1985).  f l y i n g and probing v i r a l diseases.  a l s o decreased t h e a b i l i t y o f t h i s a p h i d  Reduced to transmit  The carbamate, methomyl, i n h i b i t s the growth and  development of f a l l armyworm, Spodoptera f r u g i p e r d a , l a r v a e a t s u b l e t h a l  17 concentrations at  ( J a v i d and  A l l 1984).  Some of the p y r e t h r o i d s show promise  s u b l e t h a l doses because they d e t e r i n s e c t f e e d i n g and  development (Dobrin and  Hammond 1985,  Kumar and  inhibit  Chapman 1984).  P h y s i o l o g i c a l or b e h a v i o r a l s t r e s s impeding o p t i m a l l a r v a l reduces i n s e c t s ' r e s i s t a n c e to d i s e a s e  (Boucias  t h e i r s u s c e p t i b i l i t y to n a t u r a l enemies.  increase l a r v a l  increases  deterrents infection  T r i c h o p l u s i a n i l a r v a e i n the e a r l y i n s t a r s are more s u s c e p t i b l e t o  i n f e c t i o n by the entomopathogenic fungus S p i c a r i a r i l e y i 1975).  and  For example, f e e d i n g  used i n c o n j u n c t i o n with i n s e c t pathogens may rates.  e t a l . 1984)  growth  I f a d e t e r r e n t can m a i n t a i n  (Ignoffo et a l .  an i n s e c t i n an e a r l y i n s t a r  through  growth i n h i b i t i o n , then o t h e r m o r t a l i t y f a c t o r s can p l a y a g r e a t e r r o l e i n population regulation. s u b l e t h a l e f f e c t s may populations  The  studies provide  have an important  (Reese and  F i e l d use  Laboratory  evidence  that chronic  but d e f e r r e d impact on i n s e c t  Beck 1976).  of f e e d i n g d e t e r r e n t s has  been l i m i t e d t o a few  compounds.  best example of a f e e d i n g d e t e r r e n t t e s t e d on a l a r g e s c a l e i s the  s y n t h e t i c compound, 4 ' - d i m e t h y l t r i a z e n o - a c e t a n i l i d e ( c i t e d i n Bernays 1983).  T h i s product  was  an e f f e c t i v e f e e d i n g d e t e r r e n t i n f i e l d  against s e v e r a l herbivorous  i n s e c t s i n c l u d i n g , the cabbage l o o p e r , T. n i ,  the c o t t o n leafworm, Alabama a r g i l l a c e a . and grandis.  However, no c o n t r o l was  g o s s y p i e l l a . and  t h e c o d l i n g moth,  These r e s u l t s emphasize t h a t f e e d i n g and o v i p o s i t i o n  deterrents are often species The  the b o l l w e e v i l , Anthonomus  observed f o r s e v e r a l o t h e r p e s t i n s e c t s ,  such as the pink bollworm, P e c t i n o p h o r a C y d i a pomonella.  tests  use of f e e d i n g and  specific. oviposition deterrents integrate well  contemporary i n t e g r a t e d p e s t management (IPM).  IPM  r e q u i r e s the  of p e s t s to determine when b i o c i d a l agents a r e needed.  The  with monitoring  a p p l i c a t i o n of  d e t e r r e n t s c o u l d be p r o p h y l a c t i c or a p p l i e d when pest p o p u l a t i o n s  or  crop  damage r e a c h an economic t h r e s h o l d . deterrents  Advantages of a p p l y i n g  phytochemical  i n c l u d e s e l e c t i v e pest c o n t r o l and minimal e n v i r o n m e n t a l  disturbances. Many host and non-host phytochemicals a r e known t o i n h i b i t oviposition. reducing  Laboratory  growth and  r e p o r t s on t h e i s o l a t i o n of i n s e c t f i t n e s s -  p h y t o c h e m i c a l s a r e numerous (e.g., T r i a l and Dimond 1979, D e l l e  Monache e t a l . 1984).  Most o f t h e s t u d i e s a r e not d i r e c t l y concerned  with  the a p p l i c a t i o n o f the c h e m i c a l s i n pest c o n t r o l s i t u a t i o n s , but d e a l  with  e c o l o g i c a l or p h y s i o l o g i c a l considerations. in  Most phytochemicals examined  the l a b o r a t o r y have not been s t u d i e d i n f i e l d  u l t i m a t e aim o f d e v e l o p i n g experimental  field  t e s t s and r a r e l y w i t h t h e  a u s e f u l a g r i c u l t u r a l product.  The l a c k o f  data on the use of s u b l e t h a l phytochemicals undermines  the many l a b o r a t o r y s t u d i e s on the s u b j e c t .  19  III.  A.  Plant The  MATERIAL AND  METHODS  extracts p l a n t s were a l l c o l l e c t e d from southern B r i t i s h Columbia, a i r TM  d r i e d and  f i n e l y ground i n a Wiley  l o c a t i o n of h a r v e s t , plant material  (200  (petroleum ether,  and  harvest  g) was  mill.  Plant species, parts  dates a r e l i s t e d  i n Table I .  t h o r o u g h l y mixed with 1 l i t e r  The  s l u r r y was  t h e n the e x t r a c t s were reduced under vacuum t o 10-60 respective B.  Powdered  of e i t h e r p e t r o l  b o i l i n g range 30-60°C) or 95% aq e t h a n o l  soaked f o r 24 h a t room temp (21°C).  extracted,  (EtOH) and  f i l t e r e d and  rinsed,  ml depending on  their  viscosities.  B i o l o g i c a l Screenings 1. I n i t i a l  Screening  E x t r a c t s 5 - f o l d of those n a t u r a l l y o c c u r r i n g , c a l c u l a t e d as the weight o f p l a n t powder e x t r a c t e d  to the dry weight o f a r t i f i c i a l  (dwt/dwt), were admixed w i t h the dry p o r t i o n of the a r t i f i c i a l ( B i o s e r v I n c . , Frenchtown, NJ, removed i n a fume hood.  no.  9682) and  diet  t r e a t e d w i t h the c a r r i e r s o l v e n t s a l o n e ( p e t r o l and  EtOH).  2 neonate P_. s a u c i a l a r v a e from a l a b o r a t o r y  were p l a c e d  g (wwt)  was  similarly  Upon  hatching, on about 2  a l i q u o t s o f d i e t i n 30 ml p l a s t i c cups a t room temperature.  r e a r i n g cups were p l a c e d  l a r v a l growth was  diet.  U s i n g l i v e l a r v a l weights (n=30),  measured as a percentage of the c o n t r o l s a f t e r 14  l a r v a l gravimetric  The  i n p l a s t i c boxes w i t h moistened paper t o w e l s t o  p r e v e n t d e s i c c a t i o n of l a r v a e and  The  colony  diet  diet  the c a r r i e r s o l v e n t  C o n t r o l s c o n s i s t e d of a r t i f i c i a l  dry  d a t a was  a n a l y s i s i n each experiment.  1 ° 8 ^ Q transformed p r i o r t o  days.  statistical  20 Table  I.  P l a n t s p e c i e s , components e x t r a c t e d , h a r v e s t  Columbia and dates of m a t e r i a l b i o a s s a y e d  Plant Species  Artemisia tridentata Centaurea d i f f u s a Chrysotharanus nauseosus Chamomilla suaveolens S e n e c i o iacobaea Tragopon dubius  stms=stems, l v s = l e a v e s ,  Components Extracted  stms, stms, stms, whole stms, stms,  lvs, f ls lvs, fls lvs plant lvs lvs, fls  fls=flowers  in initial  Location  Summerland Kamloops Keromeos Vancouver Abbottsford Hedley  locations i n British  screening  Harvest date  10--83 10--83 05--84 05--84 05--84 05--84  21 2.  B i o a s s a y of Unextracted and The  Extracted  Plant  Material  p l a n t r e s i d u e r e m a i n i n g from the i n i t i a l e x t r a c t i o n , h e r e a f t e r  r e f e r r e d t o as the  'marc', and  t h e i r r e s p e c t i v e unextracted  were assayed f o r b i o l o g i c a l a c t i v i t y a g a i n s t  P_. s a u c i a neonates to  determine the e f f i c i e n c y of the e x t r a c t i o n p r o c e s s . the a r t i f i c i a l  d i e t c o n s i s t e d of B i o s e r v  p o r t i o n of marc or p l a n t powder (1:1 one  no.  w/w).  p l a n t powders  The  9682 with an  dry i n g r e d i e n t s equivalent  C o n t r o l d i e t was  p a r t powdered c e l l u l o s e ( a l p h a c e l ) t o one  prepared  part a r t i f i c i a l  using  diet.  An  a d d i t i o n a l treatment c o n s i s t e d of the c o n t r o l d i e t without c e l l u l o s e . treatment was  the p r e v i o u s 3.  Second The  for  The  the same as  most i n h i b i t o r y e x t r a c t s to P_. s a u c i a l a r v a l growth were s e l e c t e d Artificial  d i e t s were f r e s h l y prepared  (dwt/dwt) of the p l a n t e x t r a c t s .  t r e a t e d w i t h the c a r r i e r s o l v e n t .  using  C o n t r o l d i e t s were  P_. s a u c i a neonates were i n d i v i d u a l l y  on c a . 1 g of d i e t (n=25) and  a l l o w e d t o feed f o r 11 days and  A l l s u r v i v i n g l a r v a e were p l a c e d  allowed t o c o n t i n u e f e e d i n g  on c o n t r o l d i e t on day  11  then and  to determine the p e r s i s t e n c e of growth  i n h i b i t o r y e f f e c t s through p u p a t i o n and C.  was  Screening  natural concentrations  weighed.  e x p e r i m e n t a l design  experiment.  a further bioassay.  placed  This  used to determine the e f f e c t of d i l u t i n g the c o n t r o l d i e t  w i t h c e l l u l o s e on l a r v a l growth. in  of  emergence.  Dose-Response B i o a s s a y s 1.  Dose-Response B i o a s s a y on P_. s a u c i a Neonates Four c o n c e n t r a t i o n s  (20, 40,  60,  and  the e t h a n o l i c e x t r a c t of _A. t r i d e n t a t a and were assayed as above w i t h e t h a n o l  and  80% of n a t u r a l cone, dwt/dwt) of both e x t r a c t s of C_j_ suaveolens  petroleum ether  solvent  A f t e r 15 days P_. s a u c i a l a r v a e were counted, weighed, and  controls.  then allowed  to  22 feed on  the c o n t r o l d i e t u n t i l p u p a t i o n .  were recorded inhibiting  f o r each t r e a t m e n t .  l a r v a l growth by  p r o b i t a n a l y s i s (Finney 2.  50%  EC^QS ( e f f e c t i v e  relative  to c o n t r o l s ) were c a l c u l a t e d  was  species.  Field  on a r t i f i c i a l  performed to determine i f the  s a u c i a was  same as  3.  described  d i e t ( B i o s e r v no.  determine how  for  9682).  The  bioassay  r e s u l t i n g F^  and  neonates  the  Larvae  b i o l o g i c a l a c t i v i t y of the  plant extracts  another dose-response experiment was  with older c a t e r p i l l a r s .  Neonate P_. s a u c i a  standard  to  data a n a l y s i s were  i n f l u e n c e d by l a r v a l age,  the  other  i n the neonate P_. s a u c i a dose-response experiment.  S e n s i t i v i t y of O l d e r P_. s a u c i a To  a l s o evident  c o l l e c t e d A., c a l i f o r n i c a l a r v a e were r e a r e d  were used f o r t h i s experiment (n=25); the the  using  Speyer  b i o l o g i c a l e f f e c t of e x t r a c t s on P.  maturity  concentrations  the A l f a l f a Looper, Autographa  Another dose-response b i o a s s a y  noctuid  emergence  1971).  Dose-Response B i o a s s a y u s i n g  californica  Rates of p u p a t i o n and  control diet.  The  initiated  l a r v a e were f e d f o r s i x days on  r e s u l t i n g second i n s t a r l a r v a e ( c a .  mg)were then t r a n s f e r r e d t o the treatment d i e t s (n=25). d a t a a n a l y s i s were as p r e v i o u s l y d e s c r i b e d  was  The  7  bioassay  i n the neonate P_. s a u c i a  and  dose-  response experiment. D.  F i n a l Determination of E x t r a c t f o r F i e l d 1.  and  D e t a i l e d Growth A n a l y s i s of  Trial  s a u c i a Larvae F e e d i n g on  A. t r i d e n t a t a  C_. s u a v e o l e n s E x t r a c t s To  d i s t i n g u i s h between b e h a v i o r a l  and  physiological contibutions  l a r v a l growth i n h i b i t i o n , a d e t a i l e d growth a n a l y s i s was second i n s t a r P_. s a u c i a •  L a r v a e (10.9  t h e i r natural concentrations  (100%  ± 1.5  dwt/dwt).  mg, An  n=15)  initiated  to  on  were f e d d i e t s a t  EtOH e x t r a c t of  k_.  23 t r i d e n t a t a and standard  a p e t r o l e x t r a c t of C_. s u a v e o l e n s were compared with  d i e t treated with p e t r o l .  The  duration  the  of the experiment was  48  h, a l t h o u g h l a r v a e were weighed a t 24 h as w e l l as 48 h to determine r e l a t i v e f e e d i n g and  growth r a t e s over the  o t h e r w i s e i n d i c a t e d , a l l measurements are i n d i c e s were c a l c u l a t e d as d e s c r i b e d 2.  by  two  24 h p e r i o d s .  Except where  based on dry w e i g h t s .  S c r i b e r and  Slansky  (1981).  F o r m u l a t i o n of A c t i v e E x t r a c t s f o r F o l i a r A p p l i c a t i o n and  of Crude  Growth  Stability  Extracts  B i o l o g i c a l a c t i v i t y o f two  aqueous e x t r a c t s and  a 20% aq EtOH e x t r a c t  were compared t o the o r i g i n a l EtOH e x t r a c t (8 months o l d ) and  a freshly  prepared EtOH e x t r a c t from the o r i g i n a l p l a n t m a t e r i a l f o r both _A. t r i d e n t a t a and  C_. s u a v e o l e n s .  The  two  aqueous e x t r a c t s were prepared  adding 10 g of ground p l a n t powder t o 40 ml of d i s t i l l e d water. these s l u r r i e s was  brought t o a r o l l i n g  room temp f o r 24 h r s b e f o r e  being  b o i l and  filtered.  The  One  by of  then both were kept a t 20% aq EtOH e x t r a c t s were  prepared f o l l o w i n g the same procedure as the room temp water e x t r a c t s . aqueous e x t r a c t s were then l y o p h i l i z e d t o reduce t h e i r volume. l a r v a e f e d on a r t i f i c i a l  d i e t s t r e a t e d w i t h water or EtOH.  s a u c i a l a r v a e (n=25) were used f o r t h i s b i o a s s a y E.  F i e l d T r i a l of the A. An  experiment was  designed t o t e s t the  pyrethroid i n s e c t i c i d e , deltamethrin.  b l o c k s and  27,  1985.  above.  f i e l d e f f i c a c y of the  _A.  (30% aq EtOH), water and  Cabbage ( c v . E a r l y M a r v e l )  P l o t s of cabbage were assigned  treatments were randomized w i t h i n each b l o c k .  to four  the was  complete  Plots consisted  2 of a row  of seven cabbage p l a n t s  equivalent  row  P.  t r i d e n t a t a E x t r a c t on Cabbage  t r i d e n t a t a e x t r a c t r e l a t i v e to i t s c a r r i e r  seeded on May  Control  Neonate  as d e s c r i b e d  The  (11.7  o f unsprayed cabbage.  m ) and  were separated  by  an  In a d d i t i o n , guard p l a n t s were  24 s i t u a t e d a t both ends of each p l o t .  Spray a p p l i c a t i o n s were made with a  hand-held t r i g g e r s p r a y e r and the n o z z l e was c a l i b r a t e d to d e l i v e r an e q u i v a l e n t volume (14 ml) of s o l u t i o n t o each p l a n t .  The n o z z l e was  c a l i b r a t e d by measuring 10 p u l l s o f the t r i g g e r i n t o a graduated as a f i n e m i s t .  Four t r i g g e r p u l l s were executed  cylinder  from d i r e c t l y over the  p l a n t and t h r e e p u l l s f o r coverage t o t h e s i d e s and t h r e e p u l l s f o r the lower s i d e o f t h e l e a v e s . aq EtOH. of  This c a r r i e r  a spreader  g/ml.  The A. t r i d e n t a t a e x t r a c t was formulated  g i v e s an even f o l i a r coverage without  or s t i c k e r .  i n 30%  the a d d i t i o n  The r e s u l t i n g s o l u t i o n was the e q u i v a l e n t o f 0.2  The 3 o t h e r treatments  c o n s i s t e d o f the c a r r i e r s o l v e n t , 30% aq  EtOH, d i s t i l l e d water, and t h e s t a n d a r d p e s t c o n t r o l agent, d e l t a m e t h r i n TM (Decis  2.5 EC, Hoechst) a t 17.9 ug a i . / l p l u s 1 ml/1 o f t h e s p r e a d e r TM TM TM s t i c k e r , Superspred (Decis and Superspred were p r o v i d e d c o u r t e s y o f Dr. Robert  S. Vernon, A g r i c u l t u r e Canada, Vancouver, B.C). Spray  a p p l i c a t i o n s were made t o a l l p l o t s on J u l y 24, 1985 a t 6:30 am. w i t h the cabbage a t post-heading experimental  (59 days from s e e d ) .  p l a n t s were surveyed  The above ground p a r t s o f a l l  2 days b e f o r e s p r a y i n g t o e s t a b l i s h t h e  b a s e l i n e i n s e c t p o p u l a t i o n s i n each of the experimental trial  was i n i t i a t e d , a l l e x p e r i m e n t a l  plots.  p l a n t s were monitored  Once t h e  1, 6, 9, and 25  days a f t e r s p r a y i n g t o a s s e s s t h e e f f e c t s on the major i n s e c t p e s t s o f cabbage.  The pre-count  survey and t h e f i r s t  t h r e e post-treatment  counts  were n o n - d e s t r u c t i v e v i s u a l o b s e r v a t i o n s o f both s i d e s o f a l l non-head leaves.  The f i n a l i n s e c t count a t 25 days post-treatment  sampling  o f the above ground cabbage.  l o o p e r ( C L ) , T_. ni^, imported  The p e s t s monitored  was a d e s t r u c t i v e were t h e cabbage  cabbageworm (ICW), P_. rapae, and diamondback  moth l a r v a e (DBM), P l u t e l l a x y l o s t e l l a L. s e p a r a t e l y where numbers warranted  Cabbage p e s t s were a n a l y z e d  and t o g e t h e r as cabbage l o o p e r  25 e q u i v a l e n t s (CLE)  :  Data from the f i e l d  1 CLE = 1 CL = 1.5 trial  was  ICW  = 20 DBM  analyzed i n a completely  design with repeated o b s e r v a t i o n s over time.  The  'day' e f f e c t was  Then the sum  randomized b l o c k  'treatment'  freedom were p a r t i t i o n e d w i t h i n d i v i d u a l treatment The  ( S h e l t o n e t a l . 1982).  contrast  p a r t i t i o n e d w i t h p o l y n o m i a l expansion  of squares  from the  'treatment  X day'  X day'  coefficients.  interaction  was Significant  i n t e r a c t i o n s e s t a b l i s h e d d i f f e r e n c e s i n v a r i a t i o n among  p e s t p o p u l a t i o n l e v e l s w i t h the f o u r spray treatments survey  of  comparisons.  p a r t i t i o n e d f o r the l i n e a r , q u a d r a t i c and r e s i d u a l v a r i a t i o n . 'treatment  degrees  d u r i n g the  five  dates.  A q u a l i t y e s t i m a t e was  o b t a i n e d by r a t i n g each of the cabbage heads on  the l a s t day o f the experiment  (25 days p o s t - s p r a y ) .  on the amount o f v i s i b l e damage present i n and  The r a t i n g was  on each head a f t e r  based  the  wrapper l e a v e s were removed (4=marketable cabbage, no e x t e r i o r damage; 3=sauerkraut  grade, e x t e r i o r damage o n l y , no h o l e s ; 2=garden grade,  h o l e i n t o head; l=unmarketable, An a n a l y s i s of v a r i a n c e was  more than one h o l e ; 0=no head r e m a i n i n g ) .  performed  on the v i s u a l damage e s t i m a t e s  means were compared by p a r t i t i o n i n g the treatment F.  sum  of  L a b o r a t o r y E v a l u a t i o n of the O v i p o s i t i o n Deterrence  E x t r a c t on Caged P. A field  one  and  squares. of A.  tridentata  rapae  c o l l e c t e d colony of P_. rapae l a r v a e was  reared to pupation  on  cabbage ( c v . E a r l y M a r v e l ) .  E i g h t a d u l t s of each sex were p l a c e d i n a 50 X  50 X 50 cm screened cage and  f e d a s o l u t i o n of 10% s u c r o s e .  on a l a b o r a t o r y bench and  l a t e summer l i g h t  from the south and west  supplemented w i t h a 100 watt lamp f o r 16 h each day. six  cabbage l e a v e s i n 50 ml f l a s k s f i l l e d  The cage  was  was  On the f o l l o w i n g day  with water were o f f e r e d t o the  b u t t e r f l i e s i n a c h o i c e o v i p o s i t i o n experiment.  Two  l e a v e s from each o f  26  the f o l l o w i n g treatments were i n c l u d e d :  A_. t r i d e n t a t a e x t r a c t a t  equivalent  distilled  of 0.2  g/ml,  30%  aq EtOH, and  were hand-painted onto the l e a v e s . i n a c i r c l e i n a random o r d e r . experiment was G.  repeated.  The  l e a v e s and  The  solutions  the f l a s k s were arranged  Eggs were counted a f t e r 48 h and  Results  the  from the 2 r e p l i c a t e s were pooled.  Growth Comparison o f L a b o r a t o r y Reared and  Larvae on Standard D i e t and  water.  the  Wild  Colonies  of P.  D i e t s w i t h A d d i t i o n of an E t h a n o l i c  saucia  A.  t r i d e n t a t a Crude E x t r a c t A chronic  feeding  bioassay  n a t u r a l P_. s a u c i a p o p u l a t i o n population  on standard  was  used t o e s t a b l i s h the response of a  i n comparison with the £ . s a u c i a  a r t i f i c i a l d i e t and  EtOH-A. t r i d e n t a t a e x t r a c t .  Neonate  s a u c i a and  P_. s a u c i a  two  laboratory reared  groups (n=30).  e i t h e r the standard  One  d i e t s w i t h the a d d i t i o n of  l a r v a e from f i e l d (>20  group from each colony  allowed t o feed  into  on  c o n t r o l d i e t or d i e t admixed with a e t h a n o l i c _A.  the neonates were each p l a c e d of d i e t .  was  the  c o l l e c t e d £.  g e n e r a t i o n s ) were d i v i d e d  t r i d e n t a t a e x t r a c t (50% n a t . cone, dwt/dwt).  mg  laboratory  As i n p r e v i o u s  experiments  i n i n d i v i d u a l feeding containers with ca.  A f t e r 8 days the l a r v a e were counted and  weights were log^Q t r a n s f o r m e d and  analyzed  by a two  weighed. way  The  500  larval  a n a l y s i s of  variance. H.  Phytochemical I n v e s t i g a t i o n 1.  Chromatographic S e p a r a t i o n  of the E t h a n o l i c _A. t r i d e n t a t a E x t r a c t  C e n t r i f u g a l t h i n l a y e r chromatography was  used to s e p a r a t e the a c t i v e  component(s) o f the crude e t h a n o l i c e x t r a c t of A^. t r i d e n t a t a . of e x t r a c t e q u i v a l e n t 10 cm  silica  to e i g h t grams of p l a n t powder was  g e l column (60-200 mesh) with EtOH and  reduced e x t r a c t was  loaded  onto a Chromatatron  TM  An a l i q u o t  e l u t e d on a 1.5  reduced to 3 ml.  p l a t e (2 mm)  and  x  The  developed  27 with s u c c e s s i v e l y more p o l a r s o l v e n t s  (hexane, hexanetCHCl^ ( 1 : 1 ) t  C H C l : a c e t o n e ( 6 : 1 ) , CHC1 , EtOH, and  MeOH) to give ten 10-ml  3  5-ml  3  f r a c t i o n s and  eluates  followed  fractions.  2 50-ml f r a c t i o n s .  The  first  by 44 l a r g e l y acetone and  The  eighty-nine  chromatography (TLC)  5 f r a c t i o n s were hexane  CHCl^ e l u a t e s and  f r a c t i o n s were s p o t t e d  plates containing  f r a c t i o n s , 77  on s i l i c a  40 EtOH-MeOH gel thin layer  a f l u o r e s c e n t i n d i c a t o r (Baker-  TM flex  , IB2-F).  sesquiterpene  The  p l a t e s were developed u s i n g a standard  lactones  (Pieman et a l . 1980)  p l a t e s were observed under s h o r t - and  according  observations  containing  were m a i n l y CHCl^ e l u a t e s 5 contained  EtOH and  f u r t h e r separated  hexane:CHC1 spotted  3  on TLC  4 (36-60).  4.  major f r a c t i o n nos.  as i t contained  included  2,3  and  4  major f r a c t i o n  TLC 3 was  3  was  s p o t s common to eluted  with  CHCl^ g i v i n g 60 f r a c t i o n s t h a t were analyzed  as above.  The  60 f r a c t i o n s  3 were d i v i d e d between major f r a c t i o n nos.  2 (1-35)  r e s u l t i n g 4 major f r a c t i o n s were b i o a s s a y e d w i t h  neonated P_. s a u c i a l a r v a e (n=20) u s i n g an 80% major f r a c t i o n .  1  Major f r a c t i o n no.  Major f r a c t i o n no.  p l a t e s , developed and  The  o f : major f r a c t i o n no.  MeOH f r a c t i o n s 49-89. TM  2 and  then  then reduced i n volume  ( f r a c t i o n s 6-13,14-26,27-48) and  (2:1,1:1,1:2,1:6) and  of major f r a c t i o n no. and  ( f r a c t i o n s 1-5),  on the Chromatatron  both major f r a c t i o n s nos.  l i g h t and  were pooled i n t o 5 major f r a c t i o n s  Pooled f r a c t i o n s c o n s i s t e d  hexane e l u a t e s  TLC  Fractions containing similar  t o t h e i r p h y t o c h e m i c a l c o n s t i t u e n t s and  under vacuum.  no.  w i t h CHCl^-acetone ( 6 : 1 ) .  l o n g - wave u l t r a v i o l e t  sprayed w i t h a v a n i l l i n spray r e a g e n t . compounds based on TLC  method f o r  cone (dwt/dwt) of the  extract  A p o s i t i v e c o n t r o l c o n s i s t i n g of the o r i g i n a l e x t r a c t  as w e l l as a s o l v e n t  (CHC1-) c o n t r o l .  was  28  2.  Chromatography of S e s q u i t e r p e n e Lactones P r e v i o u s l y I s o l a t e d  from  tridentata To determine the c h e m i c a l n a t u r e o f the h i g h l y a c t i v e _A. t r i d e n t a t a e x t r a c t , pure s e s q u i t e r p e n e l a c t o n e s t a n d a r d s were chromatographed w i t h the most a c t i v e major f r a c t i o n s from the p r e v i o u s experiment.  Ten  s e s q u i t e r p e n e l a c t o n e s were o b t a i n e d from Dr. R i c h a r d G. K e l s e y , Dept. o f Chemistry, U n i v e r s i t y of Montana t h a t had p r e v i o u s l y been i s o l a t e d  from _A.  t r i d e n t a t a and i t s c l o s e l y r e l a t e d s u b s p e c i e s ( K e l s e y and S h a f i z a d e h Dehydroleucodin, d i h y d r o s a n t a m a r i n , a r b u s c u l i n A, a r b u s c u l i n B, C,  1979).  arbusculin  ( t a t r i d i n A - purity questionable), matricarin, deacetoxymatricarin,  d e a c e t y l m a t r i c a r i n , and d e h y d r o r e y n o s i n were chromatographed w i t h the major f r a c t i o n s o b t a i n e d i n the p r e v i o u s experiment tridentata extract. CHCl :acetone 3  and the o r i g i n a l e t h a n o l i c  The two s o l v e n t systems used c o n s i s t e d of  (6:1) (Pieman e t a l . 1980)  ( 2 : 2 : l ) ( G r e i s s m a n and G r i f f i n  1971).  and  petroleum  ether:CHCl :Et 0Ac  P l a t e s were developed  3  2  i n either  s a t u r a t e d or n o n - s a t u r a t e d TLC tanks and examined under l o n g - and  short-  wave u l t r a v i o l e t l i g h t , and t r e a t e d w i t h the v a n i l l i n c o l o u r reagent as d e s c r i b e d above.  A^  29  IV.  A.  Laboratory  RESULTS  Screenings  T a b l e I I shows the growth i n h i b i t o r y e f f e c t s on P_. s a u c i a l a r v a e of crude p l a n t e x t r a c t s added a t 5 times the n a t u r a l c o n c e n t r a t i o n to a r t i f i c i a l d i e t .  S i x of the  (sensu P a i n t e r 1951)  t o proceed t o the next s c r e e n i n g .  concentrations  were h i g h , o n l y  12 e x t r a c t s e x h i b i t e d s u f f i c i e n t a n t i b i o s i s  the  Since  i n h i b i t e d growth advanced to the second s c r e e n i n g .  survived  on the d i e t s w i t h e t h a n o l i c and  C.  No  p e t r o l e x t r a c t s of  d i f f u s a were s i m i l a r l y a c t i v e .  e x t r a c t s i g n i f i c a n t l y d i f f e r e n t from the c o n t r o l s was ethanol  e x t r a c t , but a t the h i g h c o n c e n t r a t i o n  considered The  r e s u l t s of the u n e x t r a c t e d  shown i n T a b l e I I I .  The  The  p l a n t powder and  unextracted l e a s t one  larvae  the only  bioassay  other  it  was  screening. marc b i o a s s a y  are  e f f i c i e n c y of the e x t r a c t i o n p r o c e s s i n removing  growth i n h i b i t o r y p h y t o c h e m i c a l s i s determined by t e s t i n g the plant residue  or  the j>. jacobaea  used i n the  to be s u f f i c i e n t l y a c t i v e f o r f u r t h e r  extract  tridentata  suaveolens; the e t h a n o l i c e x t r a c t from C_. nauseosus and  e t h a n o l i c e x t r a c t from C.  not  the  s i x treatments t h a t c o m p l e t e l y  severely  and  (dwt/dwt)  f o r l a r v a l growth i n h i b i t i o n .  p l a n t powder s e v e r e l y  In cases where d i e t w i t h  the  i n h i b i t e d P_. s a u c i a l a r v a l growth, a t  of the r e s p e c t i v e marcs was  i n h i b i t o r y agents removed.  extracted  shown to have had  the  growth  30 Table  II.  E f f e c t s o f weed e x t r a c t s ^ i n c o r p o r a t e d i n t o a r t i f i c i a l  growth and s u r v i v a l o f neonate P_. s a u c i a i n an i n i t i a l  PLANT SPECIES  A.  tridentata  C. d i f f u s a C. nauseosus C. s u a v e o l e n s S.  iacobaea  GROWTH i(% OF CONTROL) Petrol  Ethanol  screening  SURVIVORSHIP (% ) Petrol  0  0  8c  63  3  80ab  0c  67  0  0c  0c  0  0  3  106a  105a  18bc  90  37  T. d u b i u s  107a  72ab  80  80  control  100a  77  73  Extract concentrations 2  100a  bioassay  2  Ethanol  0c  Oc  d i e t on  were f i v e times the n a t u r a l cone.(dwt/dwt).  N=30  3 Treatments f o l l o w e d by the same l e t t e r a r e not s i g n i f i c a n t l y d i f f e r e n t (Tukey's s t u d e n t i z e d range (HSD) t e s t , p=0.05).  Table I I I . artificial  Growth and s u r v i v a l o f P_. s a u c i a neonate l a r v a e f e d on d i e t s with unextracted  PLANT  POWDER  SPECIES  GROWTH  2  A. t r i d e n t a t a C. d i f f u s a C.  nauseosa  C.  og  ETHANOL MARC  SURVIVAL  4  47cdef  p l a n t powder o r on the e x t r a c t e d  0 87  3  GROWTH SURVIVAL  121abc 34defg  PETROL MARC GROWTH SURVIVAL  50  19fg  53  77  31efg  70  34bcde  50  149ab  40  163ab  90  suaveolens  18g  30  114abcd  77  190ab  80  S.  iacobaea  15g  30  98abcd  73  145ab  77  T.  dubius  96abcd  67  43  277a  73  147ab  Standard d i e t  272a  Control  100abed 80  (with c e l l u l o s e )  Plant material incorporated  marcs  70  with a r t i f i c i a l  d i e t (1:1  dwt/dwt).  i  Taken a s t h e percentage o f l a r v a l growth o f the c o n t r o l treatment w i t h c e l l u l o s e f i l l e r s i m u l a t i n g the p l a n t m a t e r i a l . 'Percentage o f t o t a l l a r v a l s u r v i v o r s  (N=30).  Treatments f o l l o w e d by the same l e t t e r are not s i g n i f i c a n t l y d i f f e r e n t (Tukey*s s t u d e n t i z e d range (HSD) test;p=0.05).  32 S p e c i f i c a l l y , the d i e t c o n t a i n i n g  the u n e x t r a c t e d  jacobaea powders produced s i g n i f i c a n t l y s m a l l e r than both of t h e i r marcs.  C_. s u a v e o l e n s and £5.  l a r v a e and higher  Larvae f e d the marc d i e t s from these p l a n t s grew  as w e l l , or b e t t e r than c o n t r o l l a r v a e .  A n t i b i o s i s was s i m i l i a r l y  among t h e JP. s a u c i a l a r v a e f e d the d i e t s w i t h u n e x t r a c t e d powder, i n which no s u r v i v o r s were observed. l a r v a e f e d t h e _A. t r i d e n t a t a e t h a n o l from t h e c o n t r o l s .  mortality  high  k_. t r i d e n t a t a  Furthermore t h e weights o f  marc d i e t d i d not d i f f e r  significantly  The e x t r a c t s from _A. t r i d e n t a t a and C. s u a v e o l e n s t h a t  produced these marcs were a l s o shown t o be the most a c t i v e P_. s a u c i a  larval  growth i n h i b i t o r s ( T a b l e I V ) . These r e s u l t s demonstrate t h a t i n n e a r l y every c a s e where P_. s a u c i a growth i n h i b i t o r s were p r e s e n t unextracted extracting  i n the  p l a n t powders they were removed by one or both of t h e solvents.  P_. s a u c i a l a r v a e f e d d i e t without c e l l u l o s e were, on the average, over two and a h a l f times h e a v i e r dwt).  than those f e d d i e t c o n t a i n i n g  c e l l u l o s e (50%  The d i e t a r y a d d i t i o n o f p l a n t powders or marcs, however, d i d not  always reduce l a r v a l growth. r e s u l t e d i n heavier standard The  Larvae f e d seven o f the treatment d i e t s  l a r v a e on average than the c o n t r o l l a r v a e f e d the  d i e t with c e l l u l o s e . r e s u l t s o f the second s c r e e n i n g  concentrations experiment.  experiment a t n a t u r a l  a r e comparable to those o f t h e p l a n t powder and marc  Even a t the reduced c o n c e n t r a t i o n  (100% dwt/dwt) the d i e t  c o n t a i n i n g t h e A. t r i d e n t a t a e t h a n o l i c e x t r a c t r e s u l t e d i n 100% P_. s a u c i a l a r v a l m o r t a l i t y (Table I V ) .  T a b l e IV. E f f e c t s o f s e l e c t e d weed e x t r a c t s i n c o r p o r a t e d diet at natural concentrations  into a r t i f i c a l  on t h e weight and s u r v i v a l o f neonate P_.  s a u c i a f e d f o r 11 days  PLANT  EXTRACT  SPECIES  LARVAL WEIGHT  SURVIVORSHIP  (% OF CONTROL)  (% OF TOTAL)  63ab  51  A. t r i d e n t a t a  PETROL  A. t r i d e n t a t a  ETHANOL  Od  C. d i f f u s a  ETHANOL  44ab  92  C. nauseosus  ETHANOL  31bc  96  C. suaveolens  PETROL  9d  36  C. s u a v e o l e n s  ETHANOL  control  N=25 neonate P. s a u c i a  12cd 100a  2  0  8 96  larvae  Treatments f o l l o w e d by the same l e t t e r a r e not s i g n i f i c a n t l y (p=0.05) u s i n g Tukey's s t u d e n t i z e d range (HSD) t e s t .  different  34 In a d d i t i o n , l a r v a e  f e d d i e t s w i t h both C. suaveolens e x t r a c t s  s i g n i f i c a n t l y l e s s and had reduced control diet. experiment  s u r v i v o r s h i p compared t o l a r v a e  grew f e d on  These r e s u l t s agree w i t h the p l a n t powder and marc  showing t h a t d i e t s i n c o r p o r a t i n g  p l a n t powders o f A_. t r i d e n t a t a  and C_. s u a v e o l e n s a r e t h e most b i o l o g i c a l l y a c t i v e towards J?. s a u c i a larvae. Larvae s u r v i v i n g t h e second s c r e e n i n g  were t r a n s f e r r e d t o c o n t r o l d i e t  to examine l a t e n t e f f e c t s o f n e o n a t a l growth i n h i b i t i o n on l a t e r  larval  growth and development.  Between 50 and 100% o f t h e s u r v i v i n g JP. s a u c i a  l a r v a e emerged a s a d u l t s  from a l l t r e a t m e n t s .  do not appear  These t r e a t m e n t s ,  t o cause any obvious p h y s i o l o g i c a l damage which p e r s i s t s  through t h e pupal t o t h e a d u l t  stage.  F i g . 1 shows t h e r e s u l t s o f t h e dose-response and  therefore,  experiment  using  ethanol  p e t r o l e x t r a c t s o f C_. suaveolens and t h e _A. t r i d e n t a t a e t h a n o l i c  e x t r a c t a t four concentrations. related  I?, s a u c i a l a r v a l weight was i n v e r s e l y  t o the p l a n t e x t r a c t c o n c e n t r a t i o n  f o r a l l 3 crude e x t r a c t s .  The  r e s u l t i n g EC^Q'S were 36, 39, and 42% f o r t h e C_. suaveolens EtOH- and p e t r o l e x t r a c t , and _A. t r i d e n t a t a E t O H - e x t r a c t , r e s p e c t i v e l y . r e s u l t s were o b t a i n e d when t h i s experiment  was r e p l i c a t e d .  Similar  EC^^'s f o r the  p e t r o l e x t r a c t o f C_. suaveolens and t h e EtOH e x t r a c t o f _A. t r i d e n t a t a were 37 and 35%, r e s p e c t i v e l y . S u r v i v a l o f P_. s a u c i a l a r v a e was m a i n l y c o n c e n t r a t i o n - d e p e n d e n t f o r each e x t r a c t .  Survivorship  showed a l i n e a r response  e t h a n o l i c and C. s u a v e o l e n s p e t r o l e x t r a c t s  f o r t h e A^. t r i d e n t a t a  2 (with r  v a l u e s o f 0.84 and  0.87, r s p e c t i v e l y ) , b u t s u r v i v o r s h i p on t h e C_. suaveolens e t h a n o l i c  extract  2 d i d n o t c o r r e l a t e w e l l w i t h the l i n e a r e q u a t i o n ( r = 0.52) ( F i g . P o o l e d r e s u l t s from two dose-response  experiments  suggest t h a t  1).  low d i e t a r y  35 concentrations  of the p l a n t e x t r a c t s  e f f e c t on s u r v i v a l .  The  highest  (0-40% n a t u r a l c o n e ) , had  dietary concentration  (80%)  little  usually  r e s u l t e d i n g r e a t l y reduced s u r v i v o r s h i p .  F i g . 1 shows m o r t a l i t y i s  largely uneffected  increased  f o r the t h r e e  u n t i l the c o n c e n t r a t i o n  both s u r v i v a l and evident  and  60%  extracts.  A n t i b i o s i s increased with concentration  was  between 40%  as shown by the r e s u l t s of  growth i n h i b i t i o n ( F i g . 1 ) .  Dose-dependant a n t i b i o s i s  i n i t i a l l y as l a r v a l growth i n h i b i t i o n and  both i n c r e a s e d  growth i n h i b i t i o n and  mortality.  b i o a c t i v e p l a n t e x t r a c t s on P_. s a u c i a l a r v a e was l a t e r experiment measuring consumption and utilization.  As i n the p r e v i o u s  The  at higher  doses as  mode of a c t i o n of  examined f u r t h e r i n a  growth r a t e s a l o n g  with d i e t a r y  experiment, s u r v i v i n g l a r v a e p l a c e d  c o n t r o l d i e t , allowed to pupate and  the  on  emerge, showed no o b v i o u s p e r s i s t e n t  physiological effects. Age-dependent e f f e c t s were a l s o examined i n a dose-response experiment. the  Six-day o l d , second i n s t a r l a r v a e appeared more t o l e r a n t t o  p l a n t e x t r a c t s than neonates when t e s t e d a t the same  For example, even a t an e x t r a c t c o n c e n t r a t i o n consistently growth was  the m o r t a l i t y  15% or l e s s r e l a t i v e t o the c o n t r o l s ( F i g . 2 ) .  33,  52 and  s u a v e o l e n s EtOH- and The  o f 80%  concentrations.  b i o l o g i c a l a c t i v i t y of these e x t r a c t s i s not  saucia larvae.  Larvae of another polyphagous n o c t u i d ,  A_. c a l i f o r n i c a , were a l s o t e s t e d i n a c h r o n i c experiment ( F i g . 3 ) .  Furthermore,  72% of c o n t r o l s f o r the A_. t r i d e n t a t a EtOH and p e t r o l e x t r a c t s , r e s p e c t i v e l y , at t h i s  The  feeding  was  the  C_.  concentration.  restricted  t o JP.  the a l f a l f a  looper,  dose-response  ECL^'s of A_. c a l i f o r n i c a neonates were 10-20%  36 Figure  1.  mortality tridentata suaveolens  Percent growth (o) r e l a t i v e t o c o n t r o l growth and (•)  of £ .  , and  s a u c i a neonates f e d e t h a n o l i c e x t r a c t s  B) C.  s u a v e o l e n s , and  percent  from A)  a p e t r o l e x t r a c t from C)  _A.  C.  admixed t o a r t i f i c i a l d i e t s (n=25 l a r v a e per c o n c e n t r a t i o n  each e x t r a c t ) .  E r r o r bars on the growth p o i n t s are the standard  total  with  deviation.  37  (-HAinviaoiAj %  H - ) H 1 M 0 H 9  %  38 Figure  2.  mortality A) A.  Percent growth (o) r e l a t i v e t o c o n t r o l growth and (•)  of s i x d a y - o l d  t r i d e n t a t a , and  suaveolens  P.  saucia  larvae  B) C_. s u a v e o l e n s , and  percent  fed e t h a n o l i c e x t r a c t s  a p e t r o l e x t r a c t from C)  admixed to a r t i f i c i a l d i e t s (n=25 l a r v a e per c o n c e n t r a t i o n  each e x t r a c t ) .  E r r o r bars on the  growth p o i n t s are the s t a n d a r d  total from C. with  deviation.  % GROWTH  % MORTALITY 6£  M  (-•-)  40 Figure  3.  mortality tridentata suaveolens  Percent growth (o) r e l a t i v e to c o n t r o l growth and (•)  of neonate A_. c a l i f o r n i c a  , and  B) C_. s u a v e o l e n s , and  percent  fed e t h a n o l i c e x t r a c t s a petrol extract  from A)  from C)  E r r o r bars on the  growth p o i n t s are the s t a n d a r d  A^  C.  admixed to a r t i f i c i a l d i e t s (n=25 l a r v a e per c o n c e n t r a t i o n  each e x t r a c t ) .  total  with  deviation.  20 4 0 60 80 0  20 4 0 60 80  % Natural Cone, (dwt)  42 ( n a t u r a l cone.) f o r a l l three e x t r a c t s .  These E C ^ Q ' S are about h a l f  observed E C ^ Q ' S f o r P_. s a u c i a l a r v a e and  no A_. c a l i f o r n i c a l a r v a e  the  60 or 80%  concentrations  f o r any  B.  D e t a i l e d Growth A n a l y s i s of P.  of the  the  survived  extracts.  s a u c i a L a r v a e on the C. suaveolens  and  A. t r i d e n t a t a E x t r a c t s .  The  r e s u l t s of d e t a i l e d growth a n a l y s i s of second i n s t a r P.  d i e t s c o n t a i n i n g the A. t r i d e n t a t a EtOH e x t r a c t and p e t r o l e x t r a c t , are shown i n T a b l e V. f o r the t h r e e  l a r v a l cohorts  growth r a t e (RGR)  d i d not  The  p e t r o l e x t r a c t was  The  70%  RGR  fed  the C_. suaveolens  approximate d i g e s t i b i l i t y  differ significantly.  The  (AD)  relative  i s a product of r e l a t i v e consumption r a t e (RCR)  dietary u t i l i z a t i o n .  and  f o r l a r v a e f e d d i e t s with the C_. suaveolens  of the c o n t r o l - d i e t f e d l a r v a e .  t h i s growth i n h i b i t i o n appears t o be a s s o c i a t e d i n d i c a t e d by the s i g n i f i c a n t l y not  saucia,  lower RCR,  The  majority  with behavioral  of  f a c t o r s as  whereas d i e t a r y u t i l i z a t i o n s  d i f f e r s i g n i f i c a n t l y from the c o n t r o l s .  The  do  P_. s a u c i a l a r v a e f e d d i e t s  w i t h the A_. t r i d e n t a t a EtOH e x t r a c t s produced even lower growth r a t e s than l a r v a e f e d the C.  suaveolens e x t r a c t .  however, appears l a r g e l y due the extremely low  net  T h i s s e v e r e growth i n h i b i t i o n ,  to p h y s i o l o g i c a l e f f e c t s . T h i s i s i n d i c a t e d  (ECI) and  g r o s s (ECD)  the consumption r a t e remained about 60%  d i e t a r y u t i l i z a t i o n even w h i l e  of the c o n t r o l .  Separate c o n s i d e r a t i o n of the RGRs over the 48 h experiment r e v e a l i n t e r e s t i n g phenomenon. e x t r a c t s during  the  first  The  RGR  24 h p e r i o d was  of the c o n t r o l .  an  f o r l a r v a e f e d the C_^ suaveolens p e t r o l severely  retarded  r e l a t i v e to  c o n t r o l s , but i n the second 24 h p e r i o d the growth r a t e a c c e l e r a t e d e q u a l the RGR  by  to  the  A3 T a b l e V. E f f e c t s of d i e t a r y _A. t r i d e n t a t a e x t r a c t (EtOH) and C_. suaveolens extract  ( p e t r o l ) on second i n s t a r P_. s a u c i a d i g e s t i b i l i t y o f food (AD),  r e l a t i v e growth r a t e (RGR), r e l a t i v e consumption r a t e (RCR), and gross ( E C I ) ^ and net (ECD)''" d i e t a r y u t i l i z a t i o n s .  Nutritional Index  C. suaveolens  n= AD (mg  D i e t a r y Supplement A. t r i d e n t a t a  13  IA ±SD 59.6 ± 15.2 dwt/mg dwt-day x 100) 2  RGR ±SD O.AA ± 0.09 b (mg dwt/mg dwt-day) RCR ±SD 2.2 ± (mg dwt/mg dwt-day)  Control  4  0.6ab  ECI ±SD 20.A + 6.3 a (mg dwt/mg dwt-day x 100) ECD ±SD AA.9 ± A8.2 ab (mg dwt/mg dwt-day x 100)  58.2  + 33.8  0.03  + 0.15  15 51.8  c  1.6 + 1.2 b  -3.6  + 2A.6 b  0.8 + 59.5  ECI=efficiency of conversion of ingested ECD=efficiency of conversion of digested  b  ±  lA.6nsd  3  0.58 ± 0.10 a  2.7 ± 0.A a  22.0  ± 5.1 a  52.9  ± AA.6 a  food food  SD = standard d e v i a t i o n nsd = not s i g n i f i c a n t l y  different  Means i n a row f o l l o w e d by the same l e t t e r a r e not s i g n i f i c a n t l y (Tukey's s t u d e n t i z e d range (HSD) t e s t , p=0.05).  different  AA The s t a b i l i t y of crude e x t r a c t s i s important f o r r e l i a b l e of l a b o r a t o r y  reproduction  experiments and f o r i n s e c t c o n t r o l s i n the f i e l d .  The  growth  i n h i b i t o r y response of P_. s a u c i a t o e x t r a c t s kept a t A°C f o r e i g h t months was  numerically  though not s i g n i f i c a n t l y d i f f e r e n t (p<0.05) from f r e s h l y  prepared e x t r a c t s ( T a b l e V I ) .  However, i n both t r e a t m e n t s o f the f r e s h as  compared t o the s t o r e d e x t r a c t s , l a r v a l m o r t a l i t y was (orthogonal  comparisons; p<0.05).  significantly  reduced  F r e s h l y prepared e x t r a c t s were  subsequently used i n f u r t h e r experiments. The r e s u l t s of the f o r m u l a t i o n  t r i a l s o f crude A_. t r i d e n t a t a and  suaveolens e x t r a c t s a r e shown i n T a b l e V I .  There was an almost  C.  complete  l o s s i n a c t i v i t y of the C_. suaveolens e x t r a c t s when d i l u t e d w i t h water. The c o n s i s t e n t growth i n h i b i t o r y a c t i v i t i e s of the A_. t r i d e n t a t a e x t r a c t s f o r m u l a t e d i n 20% aq EtOH and the added  p h y s i o l o g i c a l component of  i n h i b i t i o n determined i t s s e l e c t i o n f o r the f i e l d t a r and suspended  p a r t i c u l a t e s i n the water  p r e s e n t problems i n the l a b o r a t o r y w i t h the a r t i f i c i a l  diet.  F i e l d spraying,  residues.  The  undissolved  f o r m u l a t e d e x t r a c t s d i d not  b i o a s s a y because  medium, thus a 30% aq EtOH s o l u t i o n was extract  trial.  growth  the e x t r a c t was  admixed  however, r e q u i r e d a more s o l u b l e  used t o d i s s o l v e most of the  45 T a b l e VI.  Mean growth and percent s u r v i v a l of neonate P_. s a u c i a l a r v a e f e d  artificial  d i e t s i n c o r p o r a t i n g f r e s h and e i g h t month o l d e t h a n o l i c  e x t r a c t s , hot water, room temp water and 20% aq e t h a n o l f o r m u l a t i o n s of A_. t r i d e n t a t a and C_. suaveolens f o r 16 days  Treatment  % Survival  Mean weight ± SD  A. t r i d e n t a t a 95% e t h a n o l fresh  24  2.6 +  2.5fg  aged  68  13.8 +  6.6def  96  34.9 + 25.5cd  100  41.9 + 23.3bc  water Hot water 20% aq EtOH  +  40  8.7  fresh  12  1.1 +  aged  52  5.8  water  100  98.2  + 45.7a  Hot water  100  121.5  ± 62.4a  20% aq EtOH  100  79.4  96  101.7  2  5.3ef  C. suaveolens 95% e t h a n o l  Control SD=standard  +  0.6g 5.3fg  + 26.6ab + 35.6a  deviation  i  Means f o l l o w e d by the same l e t t e r are not s i g n i f i c a n t l y d i f f e r e n t (Tukey's s t u d e n t i z e range (HSD) t e s t , p=0.05). E x t r a c t s were kept a t 4°C f o r e i g h t months.  46 C.  F i e l d T r i a l of the A. t r i d e n t a t a E x t r a c t on Cabbage The r e s u l t s of the f i e l d  6.  spraying  are i l l u s t r a t e d  i n F i g u r e s 4, 5, and  S i g n i f i c a n t d i f f e r e n c e s between treatments were observed i n the  aggregate number of cabbage p e s t s computed as CLEs ( F i g . 4, Appendix I ) . When treatments were compared f o r the e n t i r e d u r a t i o n o f the experiment, the cabbage t r e a t e d w i t h _A. t r i d e n t a t a e x t r a c t had s i g n i f i c a n t l y fewer CLEs compared w i t h the c a r r i e r s o l v e n t the water sprayed cabbage.  (30% aq EtOH) treatment and  T h e r e was no s i g n i f i c a n t d i f f e r e n c e i n CLEs  between the 30% aq EtOH and t h e water sprayed cabbage. i n s e c t i c i d e treatment of d e l t a m e t h r i n of the l e p i d o p t e r a n  cabbage p e s t s .  t h a t t h e r e was a g e n e r a l  (p<0.05)  In a d d i t i o n , the  was shown t o g i v e e x c e l l e n t c o n t r o l  The s i g n i f i c a n t l i n e a r e f f e c t i n d i c a t e s  i n c r e a s e i n the number o f pest  cabbage over the d u r a t i o n of the experiment.  l a r v a e on the  The s i g n i f i c a n t r e s i d u a l i n  the p o l y n o m i a l a n a l y s i s shows t h a t the f i r s t - o r d e r p o l y n o m i a l d i d not account f o r a l l the v a r i a t i o n i n the experiment.  The  'treatment X day'  i n t e r a c t i o n was s i g n i f i c a n t l y d i f f e r e n t (p<0.001) and the s e p a r a t i o n  of the  sum of squares showed t h a t the l i n e a r i n t e r a c t i o n o f t h e d e l t a m e t h r i n ,  in  c o n t r a s t w i t h the r e s t of the t r e a t m e n t s , accounts f o r most (62%) of the variation.  The s i g n i f i c a n t q u a d r a t i c  improvement due t o f i t t i n g  component measures the a d d i t i o n a l  the second-order p o l y n o m i a l .  the v a r i a t i o n i n the CLE of the d e l t a m e t h r i n  T h i s shows t h a t  treatment c o n t r a s t e d  against  the r e s t o f the treatments does not c l o s e l y f o l l o w the l i n e a r r e l a t i o n s h i p . Separate c o n s i d e r a t i o n s  o f t h e key cabbage p e s t s r e v e a l h i g h l y  s i g n i f i c a n t d i f f e r e n c e s (p<0.001) i n the l a r v a l counts of ICW between the A^. t r i d e n t a t a treatment and the 30% aq EtOH and water t r e a t e d cabbage p l o t s (Fig.  5, Appendix I I ) .  About 35% o f t h e v a r i a t i o n i n ICW  due t o t h i s s i g n i f i c a n t d i f f e r e n c e .  l a r v a l counts i s  The p l o t s t r e a t e d w i t h  deltamethrin  47 had s i g n i f i c a n t l y plots.  (p<0.001) fewer ICW  l a r v a e than a l l the other treatment  The d e l t a m e t h r i n c o n t r a s t , however, accounted  variation.  ICW  f o r 62% o f the t o t a l  l a r v a l p o p u l a t i o n counts were not s i g n i f i c a n t l y  between the water t r e a t e d p l a n t s and the 30% EtOH t r e a t e d Separation of  'treatment X day' i n t e r a c t i o n sum  different  plots.  of squares f o r the  d e l t a m e t h r i n v e r s u s the o t h e r t r e a t m e n t s i n Appendix I I a g a i n shows s i g n i f i c a n t l i n e a r and q u a d r a t i c v a r i a t i o n . One  of the most encouraging r e s u l t s comes from the a n a l y s i s of the  egg c o u n t s .  Low  numbers o f ICW  eggs on the A.  suggest an o v i p o s i t i o n d e t e r r i n g e f f e c t sprayed cabbage had s i g n i f i c a n t l y  t r i d e n t a t a sprayed cabbage  (Fig. 6).  The A^  (p<0.001) fewer ICW  EtOH and water sprayed p l a n t s (Appendix  tridentata  eggs than the 30% aq  I I I ) . Interestingly,  plants  exposed to the d e l t a m e t h r i n - s p r e a d e r treatment had s i g n i f i c a n t l y h i g h e r ICW  ICW  egg counts than a l l o t h e r t r e a t m e n t s .  (p<0.001)  Reduced numbers o f  ICW  eggs i n the _A. t r i d e n t a t a t r e a t e d p l o t s r e l a t i v e t o p l o t s t r e a t e d w i t h i t s c a r r i e r suggest one reason f o r the c o n t i n u e d s u p p r e s s i o n of ICW t h e A. t r i d e n t a t a t r e a t e d p l o t s ( F i g . 5 ) .  larvae i n  48 F i g u r e 4. Percentage change i n cabbage l o o p e r e q u i v a l e n t s  (CLE) a f t e r  field  s p r a y i n g cabbage with a) 30% aq e t h a n o l i c s o l u t i o n of _A. t r i d e n t a t a  (0.2  g/ml), b) 30% aq e t h a n o l , c ) d e l t a m e t h r i n  0.1%  Superspred or d) d i s t i l l e d water, J u l y 24,  2.5 EC (17 ug/1 a . i . ) w i t h 1985.  '  1  1  6  9  Days from spray  25  F i g u r e 5. Percent  change i n imported  p o p u l a t i o n s before and a f t e r f i e l d e t h a n o l i c s o l u t i o n of  tridentata  cabbageworm, P_. rapae  s p r a y i n g cabbage with a ) 30% aq (0.2 g/ml), b) 30% aq e t h a n o l , c )  d e l t a m e t h r i n 2.5 EC (17 u g / l a . i . ) w i t h 0.1% Superspred water, J u l y 24, 1985.  larval  or d) d i s t i l l e d  P i e r i s larvae (% change) to  O  o  TS  o  O  ^  o  O  O  o  O  CO  o  F i g u r e 6. surveyed  Percent change i n imported b e f o r e and a f t e r f i e l d  s o l u t i o n of k_. t r i d e n t a t a  cabbageworm, P_. rapae,  oviposition  s p r a y i n g cabbage with a) 30% aq e t h a n o l i c  (0.2 g/ml),  b) 30% aq e t h a n o l , c) d e l t a m e t h r i n TM  2.5 24,  EC (17 p g / l a . i . ) with 0.1% 1985.  Superspred  or d) d i s t i l l e d water, J u l y  54 D.  O v i p o s i t i o n of P. rapae: L a b o r a t o r y  Experiment  The o v i p o s i t i o n d e t e r r i n g e f f e c t o f the k_. t r i d e n t a t a e x t r a c t i n the f i e l d was of  confirmed i n a c o n t r o l l e d l a b o r a t o r y experiment.  over 105 eggs were l a i d  were l a i d  i n the two  While a t o t a l  e x p e r i m e n t a l t r i a l s o n l y two  on the cabbage l e a v e s p a i n t e d w i t h the A^. t r i d e n t a t a  eggs  extract.  Leaves w i t h e t h a n o l and water s o l u t i o n s r e c e i v e d almost a l l o f the eggs, and 45 r e s p e c t i v e l y .  Female J?. rapae were observed a l i g h t i n g on  58  the  e x t r a c t sprayed cabbage l e a v e s but without o v i p o s i t i n g e i t h e r i n the l a b o r a t o r y or i n the f i e l d E.  experiments.  Q u a l i t y of Cabbage Heads from t h e F i e l d  Trial  The v i s u a l q u a l i t y e s t i m a t e o f the f i e l d sprayed cabbage ( T a b l e V I I ) showed t h a t the s i n g l e s p r a y i n g o f the _A. t r i d e n t a t a e x t r a c t cabbage of s i g n i f i c a n t l y water sprayed p l a n t s .  (p<0.05) h i g h e r q u a l i t y than the 30% EtOH and  The cabbage sprayed w i t h d e l t a m e t h r i n , however,  r e c e i v e d the h i g h e s t v i s u a l q u a l i t y e s t i m a t e , which was h i g h e r (p<0.001) than a l l the o t h e r t r e a t m e n t s . (p=0.7) was F.  Comparison of Wild Versus L a b o r a t o r y Reared of the growth response o f two  significantly  No s i g n i f i c a n t  d e t e c t e d between the EtOH and water sprayed  A comparison  produced  difference  treatments.  P. s a u c i a L a r v a e s e p a r a t e p o p u l a t i o n s o f I?.  s a u c i a f e d d i e t s w i t h and w i t h o u t a growth i n h i b i t i n g e x t r a c t a r e shown i n Table V I I I .  The F^ JP. s a u c i a l a r v a e from the f i e l d - c o l l e c t e d p o p u l a t i o n  grew s i g n i f i c a n t l y  f a s t e r than l a r v a e from the l a b o r a t o r y c o l o n y .  f e e d i n g on the s t a n d a r d d i e t f o r 8 days t h e P. s a u c i a from the p o p u l a t i o n had grown an average larvae.  of 137 mg  v e r s u s 57 mg  After  field  f o r the l a b - r e a r e d  The P_. s a u c i a l a r v a e f e d d i e t w i t h k_. t r i d e n t a t a e x t r a c t grew, as  55 T a b l e V I I . Mean v i s u a l q u a l i t y e s t i m a t e s of cabbage t r e a t e d with an A^ t r i d e n t a t a e t h a n o l i c e x t r a c t , d e l t a m e t h r i n , 30% aq e t h a n o l , and water, r e c o r d e d 25 days  post-treatment.  Treatment  Concentration  Mean e s t i m a t e ±SD^ 2  A., t r i d e n t a t a (30% aq EtOH)  0.2 g-eq  1.6 ± 0.7b  Deltamethrin  17.9 pg/1  2.5 ± 0.7a  Ethanol d i s t i l l e d water  30% aq  1.3 ± 0.8c 1.3 ± 0.6c  ^"Visual q u a l i t y e s t i m a t e s ± s t a n d a r d d e v i a t i o n ; based on a s c a l e from 0-4. The s c a l e i s an e s t i m a t e o f market q u a l i t y i e . 0=no head r e m a i n i n g , l=unmarketable more than one h o l e i n t h e head, 2=garden grade, one h o l e i n t o head, 3=sauerdraut grade, e x t e r i o r damage o n l y , no h o l e s , 4=marketable cabbage, no e x t e r i o r o r i n t e r i o r damage. 2 Means f o l l o w e d by the same l e t t e r a r e not s i g n i f i c a n t l y d i f f e r e n t means s e p a r a t e d by o r t h o g o n a l c o n t r a s t s ) .  (p>0.05,  56  Table V I I I . Mean P_. s a u c i a l a r v a l weight of a  field  collect  compared t o t h e l a b o r a t o r y colony f e d the standard a r t i f i c i a l  population d i e t and d i e t  c o n t a i n i n g a 50% _A. t r i d e n t a t a e t h a n o l i c e x t r a c t (dwt/dwt) f o r 8 days.  %  D i e t Treatment Source o f l a r v a e Standard  Survival (n=30)  Mean weight ±SD  %R(T  diet  Lab c o l o n y F i e l d colony  90 100  56.9 136.6  ± 27.9 ± 43.6  90 80_  13.5 34.8  ± 8.7 ± 14.7  3 _A. t r i d e n t a t a  diet  Lab c o l o n y F i e l d colony  23.7 25.5  standard d e v i a t i o n 2 % of r e s p e c t i v e c o n t r o l  3 50% (dwt/dwt) c o n c e n t r a t i o n  Two-way oafn aV la y iance Source r is ai tsi o o nf v a rDF Model 3 between p o p u l a t i o n s 1 between d i e t s 1 1 popul. * d i e t s 104 Error  F  SS 16.9 8.3 7.5 0.2 22.9  a  25.6 37.8 34.0 0.8  Probability 0.0001 0.0001 0.0001 0.3833  Sum o f squares o f l a r v a l growth a r e a d j u s t e d f o r m o r t a l i t y ,  expected, s i g n i f i c a n t l y However, growth of the  l e s s than the l a r v a e fed the s t a n d a r d f i e l d P_. s a u c i a l a r v a e was  diet.  4 - f o l d more than the  lab-  r e a r e d P_. s a u c i a on e x t r a c t - t r e a t e d d i e t . I n t e r e s t i n g l y , t h e r e was o r i g i n and  no  s i g n i f i c a n t i n t e r a c t i o n between l a r v a l  response t o d i e t a r y _A. t r i d e n t a t a e x t r a c t .  p r o p o r t i o n a l growth i n h i b i t i o n (75%) between the two  l a r v a l populations,  was  not  Preliminary  The was  significantly different  for laboratory  separated  experiments.  e t h a n o l i c e x t r a c t from A^. t r i d e n t a t a  pooled i n t o f o u r major groups with g e n e r a l l y d i s t i n c t  a final  the  Phytochemical I n v e s t i g a t i o n  chromatographically  profiles:  words,  thus i n d i c a t i n g t h a t percentage of  c o n t r o l l a r v a l growth i s a v a l i d e x p r e s s i o n  G.  In other  a non-polar hexane f r a c t i o n , two  group of EtOH and MeOH e l u a t e s .  chemical  groups from CHCl^ e l u a t e s ,  T a b l e IX shows the growth  and  and  m o r t a l i t y of P_. s a u c i a f e d d i e t s i n c o r p o r a t i n g these e l u t i o n s .  The  CHClg f r a c t i o n s accounted f o r most of the growth i n h i b i t i o n and  mortality  i n J?. s a u c i a l a r v a e .  There was  no  two  s i g n i f i c a n t d i f f e r e n c e among the  other  f r a c t i o n s or the c o n t r o l . The  d i r e c t chromatography of s e s q u i t e r p e n e  l a c t o n e s w i t h the  b i o l o g i c a l l y a c t i v e f r a c t i o n s i s shown i n F i g s . 7,8,9, and b i o l o g i c a l a c t i v i t y of the two f r a c t i o n s of /U  t r i d e n t a t a c o u l d not  the t e n pure s e s q u i t e r p e n e v a l u e s and  chromatographically  10.  separated  two  The  high  major  be c o n s i s t e n t l y c o r r e l a t e d w i t h any  lactones a v a i l a b l e .  c o l o u r r e a c t i o n s corresponded i n one  no unambiguous matches i n both s o l v e n t  Although some o f solvent  of  the  system, t h e r e were  systems or tank arrangements.  T a b l e IX. Mean l a r v a l weight o f neonate P. s a u c i a f e d a r t i f i c i a l admixed w i t h c h r o m a t o g r a p h i c a l l y s e p a r a t e d f r a c t i o n s ethanolic  extract  compared  t o the o r i g i n a l extract  c o n c e n t r a t i o n s and the s t a n d a r d  TREATMENT  A.  diet  o f an _A. t r i d e n t a t a  at ecological  diet^.  % SURVIVAL  % LARVAL WEIGHT  (n=20)  (of  #1-Hexane  95  63.5a  #2-CHCl  3  40  2.8b  #A-CHC1  3  30  1.3b  95  74.2a  0  0.0c  95  100.0a  control)  tridentata  ELUANT  #5-MeOH,EtOH Original  extract  standard  diet  The s t a n d a r d d i e t was t r e a t e d  with  2  petrol  L a r v a l growth f o l l o w e d by t h e same l e t t e r a r e not s i g n i f i c a n t l y d i f f e r e n t , Tukey's s t u d e n t i z e d range (HSD) t e s t (p=0.05).  59 Major f r a c t i o n  no.  2 i s a h i g h l y complex phytochemical  mixture  c o n t a i n i n g seven major c o n s t i t u e n t s , of which f i v e g i v e a p o s i t i v e c o l o u r r e a c t i o n w i t h the v a n i l l i n r e a g e n t . and  long-wave u l t r a v i o l e t  vanillin  u s i n g c o l o u r r e a c t i o n s w i t h the  reagent.  Major f r a c t i o n major TLC s p o t s . ultraviolet  no.  no. 4 appears as a c h e m i c a l l y s i m p l e r mixture of  Two  l i g h t and  v a n i l l i n reagent. fraction  l i g h t and  Ten o t h e r s p o t s were v i s i b l e i n s h o r t -  4.  five  of the s p o t s f l u o r e s c e d blue with long-wave the o t h e r 3 s p o t s gave p o s i t i v e c o l o u r r e a c t i o n s w i t h  S i x o t h e r minor c o n s t i t u e n t s were a l s o d e t e c t e d i n major  60 F i g u r e 7.  T h i n - l a y e r chromatograph of f r a c t i o n s 2 (fr#2) and 4 (fr#4) from  the s e p a r a t i o n of a crude e t h a n o l i c _A. t r i d e n t a t a e x t r a c t chromatographed w i t h phytochemicals  from A r t e m i s i a spp.  The  compared t o the J\. t r i d e n t a t a f r a c t i o n s were: dihydrosantamarin  pure s e s q u i t e r p e n e l a c t o n e s Dehydroleucodin  (dhl),  ( d h s ) , a r b u s c u l i n A (abA), a r b u s c u l i n C (abC), m a t r i c a r i n  (mat), d e a c e t o x y m a t r i c a r i n (dom), d e a c e t y l m a t r i c a r i n (dam), and dehydroreynosin  (dhr).  Non-fluorescent colours occurred a f t e r developing  the p l a t e w i t h a v a n i l l i n reagent and a f t e r 24 h. non-saturated  the arrows i n d i c a t e a c o l o u r s h i f t  TLC developed w i t h petroleum e t h e r : C H C l : E t 0 A c 3  tank.  2  (2:2:1) i n a  0.9  COLOR LEGEND  0.8  Bk Br B f  o  0.7 0.6 R.  o  P**  0.1 0  P PB Pp *  OPp**  p-  0  C L op**  Br  O** p  ON 9P?-P  O  B r  *  o P*Br**  o  P»Br**  ~ p p  O Pp«»Bk  O  P*I  O  Pp  O  fY-Bk  OBr  frfil  *  o **  B Y  0.3 0.2  N  Y  0.5 0.4  Bk*  black brown blue f l u o r e s c e s longwave UV navy b l u e pink pale blue purple quenches longwave UV quenches shortwave UV yellow  f #4 r  O abC  abA  dhl  dom  mat  dhs  dhr  dam  P*>Bk**  62 F i g u r e 8.  T h i n - l a y e r chromatograph of f r a c t i o n s 2 (fr#2) and  A (fr#A) from  the s e p a r a t i o n of a crude e t h a n o l i c _A. t r i d e n t a t a e x t r a c t chromatographed w i t h phytochemicals from A r t e m i s i a spp.  The  compared t o the A^. t r i d e n t a t a f r a c t i o n s were: d i h y d r o s a n t a m a r i n ( d h s ) , a r b u s c u l i n A (abA), (mat), d e a c e t o x y m a t r i c a r i n fluoresent colours occurred r e a g e n t and  (dom), and  pure s e s q u i t e r p e n e dehydroleucodin  (dhl),  tatridin A (ttA), matricarin  d e a c e t y l m a t r i c a r i n (dam).  a f t e r developing  the p l a t e w i t h a  the arrows i n d i c a t e a c o l o u r s h i f t a f t e r 2A h.  w i t h CHC13:acetone (6:1)  lactones  i n a saturated  tank.  Non-  vanillin  TLC  developed  CD P B *  e  fb  ©  2  o  .  **  i*  v  PB N Br P fb  fb  O  fb  o  p  O  £j  B  r  Br  O~p p  -  pale blue navy blue brown pink fluoresces blue, longwave UV Bk - black Pp - purple Y - yellow * - quenches longwave UV quenches shortshort** ~ quenches "" wave UV  fb  O  O  COLOR LEGEND  OP-  Brj»  oPp-Brl  '  p  0 Br  0 Bk —  J  fr//2  0 r B  1—  fr/M  1  1  1  1  dhl  dom  mat  abA  I dhs  , dam  u_ ttA  64 F i g u r e 9.  T h i n - l a y e r chromatograph of f r a c t i o n s 2 (fr#2) and 4 (fr#4) from  the s e p a r a t i o n of a crude e t h a n o l i c with phytochemicals  t r i d e n t a t a e x t r a c t chromatographed  from A r t e m i s i a spp.  The pure s e s q u i t e r p e n e l a c t o n e s  compared t o the A_. t r i d e n t a t a f r a c t i o n s were:  Dehydroleucodin  (dhl),  a r b u s c u l i n A (abA), a r b u s c u l i n B (abB), a r b u s c u l i n C (abC), t a t r i d i n A ( t t A ) , m a t r i c a r i n (mat), d e a c e t o x y m a t r i c a r i n (dom), d e a c e t y l m a t r i c a r i n (dam), and  dehydroreynosin  (dhr).  Non-fluorescent colours occurred a f t e r  d e v e l o p i n g the p l a t e w i t h a v a n i l l i n reagent and the arrows i n d i c a t e a c o l o u r s h i f t a f t e r 24 h. saturated  tank.  TLC developed  w i t h CHCl^:acetone  (6:1) i n a  P-Br **  o o  COLOR LEGEND Bk black Br brown f fluoresceslong-UV PB pale blue B blue 0 orange Y yellow N navy b l u e P pink purple PP * quenches l o n g - U V quenches s h o r t - U V  P-Br**  Pp-Br  o  Pp-Bk  O P . Br*.  o —I  fr//2  1  f //4 r  I abB  -JL_ abC  -J dhl  u. dom  I mat  i abA  l dhr  1 dam  Pp-Bk  i ttA  66 F i g u r e 10.  T h i n - l a y e r chromatograph of f r a c t i o n s 2 (fr#2) and 4 (fr#4)  from the s e p a r a t i o n of a crude e t h a n o l i c  tridentata  extract  chromatographed w i t h p h y t o c h e m i c a l s from A r t e m i s i a spp.  The  pure  s e s q u i t e r p e n e l a c t o n e s compared t o the _A. t r i d e n t a t a f r a c t i o n s were: a r b u s c u l i n A (abA), a r b u s c u l i n B (abB), t a t r i d i n A ( t t A ) , m a t r i c a r i n d e a c e t o x y m a t r i c a r i n (dom), and d e a c e t y l m a t r i c a r i n (dam).  (mat),  Non-fluorescent  c o l o u r s o c c u r r e d a f t e r d e v e l o p i n g the p l a t e with a v a n i l l i n reagent and arrows  i n d i c a t e a c o l o u r s h i f t a f t e r 24 h.  TLC developed w i t h petroleum  ether:CHClo:Et 0Ac (2:2:1) i n a n o n - s a t u r a t e d 9  tank.  the  COLOR LEGEND Bk Br B PB f N P PP Y *  **  black brown blue pale blue f l u o r e s c e s longwave UV l i g h t navy b l u e pink purple yellow quenches longwave UV l i g h t quenches longwave UV l i g h t  O'Br DP*PP  C^Pp-Br  QP-  **  O fr//2  fr//A  abB  abA  dom  mat  dam  P-Br** *  O  ^  ttA  p  P*Bk  68 V.  A. Screening  DISCUSSION  A s t e r a c e o u s E x t r a c t s f o r Insect Growth I n h i b i t o r s  P l a n t e x t r a c t s from many f a m i l i e s have been screened as i n s e c t c o n t r o l agents.  Many l a r g e s c r e e n i n g s  for botanical i n s e c t i c i d e s occurred  the advent of s y n t h e t i c i n s e c t i c i d e s (Jacobson and p u b l i c i n t e r e s t i n a l t e r n a t i v e s to c o n v e n t i o n a l the s c i e n t i f i c e f f o r t t o f i n d e n v i r o n m e n t a l l y ( A b i v a r d i & Benz 1984, c o n t r o l s has  Bernays 1983).  o f t e n focused  p r i o r to  Crosby 1971). Recent  i n s e c t i c i d e s has  supported  sound i n s e c t c o n t r o l s  Screening  on acute t o x i c i t y but  for botanical insect the past  ten y e a r s have  seen a renewed i n t e r e s t i n m a t e r i a l s w i t h more s u b t l e a c t i o n s , such growth r e g u l a t o r s , l a r v a l growth i n h i b i t o r s , and  as  oviposition deterrents.  E x t r a c t s from c e r t a i n a s t e r a c e o u s p l a n t s a r e r e p o r t e d  to i n f l i c t  v a r i e t y of d e l e t e r i o u s e f f e c t s on i n s e c t s ; some e x t r a c t s from  a  the  Asteraceae have been shown to a c t as i n s e c t r e p e l l e n t s (Hwang e t a l . 1985), feeding  i n h i b i t o r s (Isman and  o v i p o s i t i o n deterrents  Rodriguez 1984,  (Lundgren 1975,  Burnett  insecticides  (Jacobson and  a c t i v i t y and  c h r o n i c t o x i c i t y of e t h a n o l i c and  i n the Asteraceae (Table  Crosby 1971).  The  Nawrot e t a l . and  1982)  Jones 1978)  and  contact  i n s e c t growth i n h i b i t o r y p e t r o l e x t r a c t s of s i x weeds  I ) have a l s o been a s s e s s e d .  Many f a c t o r s determine an i n s e c t ' s response t o p l a n t a l l e l o c h e m i c a l s , such as i n s e c t s p e c i e s , stage of development, c o n c e n t r a t i o n a l l e l o c h e m i c a l , and  the c h e m i c a l c o n t e x t  presented to the i n s e c t . i n h i b i t o r s has age  of  the  i n which the a l l e l o c h e m i c a l i s  L e p i d o p t e r a n l a r v a l s e n s i t i v i t y t o growth  been shown p r e v i o u s l y t o be i n v e r s e l y c o r r e l a t e d w i t h  (Reese 1983,  Isman and  Duffey 1982).  These r e s u l t s f o l l o w t h i s  i n t h a t younger P_. s a u c i a l a r v a e were more s e n s i t i v e t o e f f e c t s of phytochemical growth i n h i b i t o r s than o l d e r l a r v a e  (Fig 1 & 2).  larval pattern  the  Neonate  P.  s a u c i a l a r v a e grew l e s s , and s u f f e r e d g r e a t e r larvae fed a r t i f i c i a l  diet containing  e x t r a c t s ( F i g 1 and 2 ) .  m o r t a l i t y t h a n , second i n s t a r  the same l e v e l s o f a s t e r a c e o u s  Younger l e p i d o p t e r a n  l a r v a e may possess fewer  endosymbiotic microorganisms needed f o r d e t o x i f i c a t i o n (Jones e t a l . 1981), or lower l e v e l s o f c o n s t i t u a t i v e d e t o x i f y i n g enzymes (Ahmad 1986). Insect feeding  growth i n h i b i t i o n c o u l d r e s u l t from b e h a v i o r a l  deterrency),  s u p p r e s s i o n ) o r both.  Schroeder (1976) has shown t h a t d e c r e a s e s i n l a r v a l  food a v e r s i o n  nutrient u t i l i z a t i o n .  (e.g.,  p h y s i o l o g i c a l f a c t o r s ( e . g . , microsomal enzyme  food u t i l i z a t i o n c a n be induced by food behavioral  factors  deprivation.  S t a r v a t i o n or  r e s u l t i n g i n a lower RCR may a l s o  decrease  T a b l e V shows t h a t when JP. s a u c i a l a r v a e were f e d  the k_. t r i d e n t a t a e x t r a c t , even though the RCR was 60% o f t h e c o n t r o l s , the ECI  o f those l a r v a e was o n l y  likely  5% o f t h e c o n t r o l s ( T a b l e V ) .  Thus i t i s  t h a t the s e v e r e l y reduced food u t i l i z a t i o n o f t h e A^. t r i d e n t a t a f e d  l a r v a e was due t o p h y s i o l o g i c a l f a c t o r s r a t h e r than a lower consumption rate. L a r v a l growth on C_^ s u a v e o l e n s - p e t r o l  diet i s i n i t i a l l y  shown by a RGR o f 51% o f t h a t o f t h e c o n t r o l s f o r t h e f i r s t 48 hr n u t r i t i o n a l experiment. l a r v a e recovered  i n h i b i t e d as 24 h r s o f t h e  In the second h a l f o f t h e experiment these  from t h e p r i o r i n h i b i t i o n and a t t a i n e d an RGR equal t o the  control fed larvae.  In c o n t r a s t , t h e RGR f o r P_. s a u c i a l a r v a e f e d an _A.  t r i d e n t a t a - e t h a n o l i c d i e t was s i g n i f i c a n t l y lower (Tukey's  studentized  range (HSD) t e s t ; p=0.05) than the c o n t r o l s f o r both 24 h r p e r i o d s and remained e s s e n t i a l l y t h e same a t 27 and 29% o f c o n t r o l f e d l a r v a l respectively. behavioral  RGR,  T h i s i m p l i e s t h a t growth i n h i b i t o r s t h a t f u n c t i o n m a i n l y as  feeding  deterrents  can be r e a d i l y overcome by i n s e c t s , whereas  growth i n h i b i t o r s t h a t decrease n u t r i e n t u t i l i z a t i o n may p r o t e c t b e t t e r because o f t h e i r more p e r s i s t e n t a c t i v i t y .  plants  The  ability  of the e x t r a c t i o n p r o c e s s t o remove i n s e c t growth  i n h i b i t o r s i s an important step i n an e f f i c i e n t  screening  process.  Table  I I I shows t h a t s o l v e n t e x t r a c t i o n removed i n s e c t growth i n h i b i t o r s i n n e a r l y every case where i n h i b i t o r s were p r e s e n t powders.  plant  I n c r e a s e s i n l a r v a l growth on the m a r c - d i e t s i n d i c a t e t h a t  e x t r a c t i o n p r o c e s s was the p l a n t The  i n the unextracted  the  e f f i c i e n t i n removing i n s e c t growth i n h i b i t o r s from  material. d i l u t i o n of i n s e c t d i e t w i t h a n o n - n u t r i t i v e ,  substance has  been shown t o i n c r e a s e  i n s e c t s (Dadd 1970).  food  non-deterrent  consumption i n a number of  Cockroaches have been shown to i n c r e a s e  response t o d i e t a r y d i l u t i o n s of c e l l u l o s e ( B i g n e l l 1978).  feeding  In  in  contrast,  growth i s reduced i n P_. s a u c i a l a r v a e by d i e t a r y a d d i t i o n s of c e l l u l o s e (Table I I I ) .  T h i s may  phagostimulation, imply i n c r e a s e d  r e s u l t from a d e c r e a s e i n a v a i l a b l e n u t r i t i o n ,  or b o t h .  Increased  f i t n e s s but may  consumption does not  l e a d t o decreased d i e t a r y  u l t i m a t e l y r e s u l t i n g i n growth r e d u c t i o n .  Results  P_. s a u c i a l a r v a e f e d many of the d i e t s ( e . g . , CJ. nauseosa and  the p e t r o l m a r c - d i e t from C.  i n T a b l e I I I show t h a t  suaveolens and  growth i n h i b i t i o n and  been r e p o r t e d deterrency, spectrum.  i n asteraceous p l a n t s .  The  species  control diet. a c t i v i t y has  growth i n h i b i t i o n and  previously feeding  s p e c i f i c r a t h e r than broad  Nawrot e t a l . (1982) screened 23 e x t r a c t s from a s t e r a c e o u s  plants against  three c o l e o p t e r a n  e x t r a c t s possessed s t r o n g consistency  Artificial  possess a d d i t i o n a l n u t r i e n t s or  feeding deterrent  however, appears t o be  T_. d u b i u s ) grew  control diet.  phagostimulants l a c k i n g i n the c e l l u l o s e c o n t a i n i n g Insect  utilization,  the e t h a n o l i c marc-diet from  more than d i d l a r v a e f e d the c e l l u l o s e c o n t a i n i n g d i e t s i n c l u d i n g p l a n t m a t e r i a l may  necessarily  feeding  p e s t s of s t o r e d p r o d u c t s and deterrency.  found t h a t  I n t e r e s t i n g l y , t h e r e was  between the e x t r a c t s ' a c t i v i t y amongst the i n s e c t  species  7 no  tested.  These a u t h o r s l a t e r confirmed t h a t s e s q u i t e r p e n e  part responsible  f o r the f e e d i n g deterrency  (Nawrot e t a l . 1984,  Harmatha and  e x t r a c t s examined, C.  of the a s t e r a c e o u s e x t r a c t s  Nawrot 1984).  suaveolens and  T a b l e IV shows t h a t of  A_. t r i d e n t a t a had  i n h i b i t o r y a c t i v i t y on P_. s a u c i a l a r v a l growth. t r i d e n t a t a was  the  A. c a l i f o r n i c a l a r v a e  the RGR  and  of the P.  ( F i g . 3) and  An e t h a n o l i c e x t r a c t of  feeding i n h i b i t o r s .  V i l l a n i and  i n v e s t i g a t e d crude e x t r a c t s from twelve Asteraceae and Artemisia  d r a c u n c u l u s and  Santolina virens, deterred  wireworm, Melanotus communis. A s t e r a c e a e (and moth, C y d i a  14 other  Suomi and  lowered  V).  S e v e r a l i n v e s t i g a t o r s have chosen e x t r a c t s of A r t e m i s i a potent i n s e c t growth and  associates  species  Gould  feeding  by  two,  corn  (1986) examined to l a r v a l  deterrency  in  as  (1985)  found t h a t  plants) f o r feeding deterrency  pomonella. They found the s t r o n g e s t  eleven  codling  the  A s t e r a c e a e e x t r a c t s from A r t e m i s i a a b s i n t h i u m, Chrysothamnus nauseosus Tanacetum v u l g a r e .  Yang (1983) has  reported  t h a t two  the buds of A_. c a p i l l a r i s were f e e d i n g d e t e r r e n t s cabbageworm, P i e r i s rapae . reported  A.  performance  because i t s i g n i f i c a n t l y  saucia larvae (Table  the  strongest  chosen f o r f i e l d e v a l u a t i o n because of s u p e r i o r  against  ECI  l a c t o n e s were i n  phenlylalkynes  f o r the  and  from  imported  In c h o i c e experiments Jermy e t a l . (1981)  t h a t Colorado p o t a t o b e e t l e ( L e p t i n o t a r s a d e c e m l i n e a t a ) l a r v a e  were i n h i b i t e d from f e e d i n g on l e a f d i s k s coated w i t h an e t h a n o l i c from _A. t r i d e n t a t a .  In my  study I have shown t h a t an e t h a n o l i c e x t r a c t  A. t r i d e n t a t a s t r o n g l y i n h i b i t s l a r v a l growth of two  lepidopteran  A_. c a l i f o r n i c a and  The  P. s a u c i a , i n f e e d i n g b i o a s s a y s .  i n d i c a t e t h a t e x t r a c t s of A r t e m i s i a phytophagous i n s e c t  pests.  extract of  larvae,  above r e s u l t s  s p e c i e s have broad spectrum a c t i v i t y  on  72 B.  Phytochemicals and I n s e c t Growth I n h i b i t i o n Research on i n s e c t growth i n h i b i t o r s and  f e e d i n g d e t e r r e n t s has o f t e n  focused on the i s o l a t i o n of s p e c i f i c p h y t o c h e m i c a l s . have emphasized  Many i n v e s t i g a t o r s  i n d i v i d u a l compounds and s i n g l e c l a s s e s of  as the key t o i n s e c t - p l a n t i n t e r a c t i o n s . i n s e c t s are always  exposed  phytochemicals  In n a t u r e however, phytophagous  t o complex m i x t u r e s o f p h y t o c h e m i c a l s .  C o n s i d e r i n g the w i t h i n p l a n t d i v e r s i t y o f c h e m i c a l s , i n t e r a c t i o n s among phytochemicals may (Berenbaum  be a common d e t e r m i n i n g f a c t o r i n i n s e c t / p l a n t  relations  1985).  P l a n t defense s t r a t e g i e s u s i n g c h e m i c a l m i x t u r e s p r o b a b l y occur more f r e q u e n t l y than d e f e n s i v e s t r a t e g i e s u s i n g s i n g l e a l l e l o c h e m i c a l s . p l a n t s c o n t a i n more than one d e f e n s i v e p h y t o c h e m i c a l (Berenbaum Harborne 1982),  Most  1985,  but, few s t u d i e s have examined the growth i n h i b i t o r y  a c t i v i t i e s among c o - o c c u r r i n g p h y t o c h e m i c a l s .  In the l i m i t e d number o f  cases where c o - o c c u r r i n g c h e m i c a l s have been examined, the underscore the importance o f phytochemical i n t e r a c t i o n s .  results Adams and  Bernays  (1978) examined the e f f e c t s of f o u r t e e n s i m p l e p h e n o l i c c h e m i c a l s from Sorghum b i c o l o r f e d t o L o c u s t a m i g r a t o r i a a t n a t u r a l l y concentrations.  These phytochemicals produced  d e t e r r e n c y o n l y when combined.  a measurable  feeding  When f e e d i n g d e t e r r e n t s from u n r e l a t e d  c h e m i c a l groups were combined i n b i n a r y combinations tomatine) d e t e r r e n t e f f e c t s were o f t e n a d d i t i v e Phytochemicals presented as a mixture may e f f e c t on i n s e c t s .  occurring  (e.g., s i n i g r i n  (Adams and Bernays  1978).  have a g r e a t e r than a d d i t i v e  Berenbaum and Neal (1985) r e p o r t the  synergistic  e f f e c t s of the methylene d i o x y p h e n y l compound, m y r i s t i c i n and the c o o c c u r r i n g furanocoumarin, concentrations.  xanthotoxin, at n a t u r a l l y  I n s e c t growth may  and  occurring  be reduced more e f f e c t i v e l y by a  73 chemical  mixture c a u s i n g d i f f e r e n t b e h a v i o r a l and  than a s i n g l e d e t e r r e n t  physiological activity  chemical.  Polyphagous p e s t s a r e g e n e r a l l y more r e s i s t a n t to growth i n h i b i t o r s than i n s e c t s w i t h a narrow host range (Bernays 1983) evidence present  and  thus may  of a broader spectrum of growth i n h i b i t o r y a c t i v i t y .  In  provide the  t h e s i s , the growth of the h i g h l y polyphagous P_. s a u c i a l a r v a e  was  s i g n i f i c a n t l y reduced r e l a t i v e t o the c o n t r o l s by e t h a n o l i c e x t r a c t s from f i v e of the s i x p l a n t s i n v e s t i g a t e d but o n l y two  of s i x p e t r o l e x t r a c t s  t e s t e d a t f i v e times the n a t u r a l c o n c e n t r a t i o n .  This i n d i c a t e s that  the  growth i n h i b i t o r s i n the p l a n t s chosen ( T a b l e I ) c o n t a i n m o s t l y p o l a r compounds.  The  g r e a t e r p r o p o r t i o n of the e t h a n o l e x t r a c t s e x h i b i t i n g  potent a c t i v i t y support  the r e s u l t s of Freedman e t a l . (1979).  A_. t r i d e n t a t a i s known t o c o n t a i n many p h y t o c h e m i c a l s ( T a b l e X) some a r e r e p o r t e d to have i n s e c t growth and K e l s e y and  Shafizadeh  feeding i n h i b i t o r y  activity.  (1979) have i s o l a t e d s e v e r a l s e s q u i t e r p e n e  from A. t r i d e n t a t a and  i t s subspecies.  of t h e s e , d e a c e t y l m a t r i c a r i n .  and  lactones  Jermy e t a l . (1981) bioassayed  They r e p o r t e d good f e e d i n g  a c t i v i t y a g a i n s t l a r v a l C o l o r a d o potato  one  deterrent  b e e t l e but noted t h a t  significant  f e e d i n g d e t e r r e n t a c t i v i t y remained i n the e x t r a c t even a f t e r removal of deacetylmatricarin.  Wisdom e t a l . (1983) t e s t e d f i v e  l a c t o n e s a g a i n s t H_. zea and t r i d e n t a t a , dehydroleucodin, Sesquiterpene i n s e c t growth and sesquiterpene  found t h a t o n l y a g u a i a n o l i d e  from _A.  s i g n i f i c a n t l y reduced growth.  l a c t o n e s from o t h e r feeding.  sesquiterpene  Isman and  p l a n t s have been shown t o a f f e c t Rodriguez (1983) found t h a t s e v e r a l  l a c t o n e s e x t r a c t e d from Parthenium s p e c i e s  i n h i b i t e d l a r v a l growth of H_. z e a .  Burnett  and  (Asteraceae)  co-workers (1974) r e p o r t e d  t h a t of s i x l e p i d o p t e r a n l a r v a l s p e c i e s examined, f o u r were d e t e r r e d f e e d i n g on Vernonia  spp.  (Asteraceae)  containing sesquiterpene  from  lactones.  TABLE IX:  Phytochemical  c o n s t i t u e n t s p r e v i o u s l y i s o l a t e d from A r t e m i s i a  tridentata  Monoterpenes camphor 1,8-cineole delta-3-carene s a n t o l i n y l ester alpha-pinene camphene  Sesquiterpene lactones' matricarin t a t r i d i n A, B, C deacetoxymatricarin deacetylmatricarin ridentin d e t a t i n A, B dehydroleucodin a r b u s c u l i n A, B, C  1  Coumarins esculin umbelliferone cichoriin isoscopoletin scopoletin scoparon esculetin artelin  Flavonoids q u e r c e t a g e t i n 3,6-dimethyl e t h e r quercetagetin 3,6,7-trimethyl ether kaempferol 3 , 6 , 7 - t r i m e t h y l e t h e r luteolin luteolin-7-0-glucoside 6-methoxy l u t e o l i n axillarin  Buttkus e t a l . (1977) essential oils  The l i s t e d monoterpenes comprise  2  Seaman (1982)  3  Brown e t a l . (1975)., Murray e t a l . (1982) p h e n o l i c f r a c t i o n o f an A^. t_. spp. vaseyana  4  Rodriguez  e t a l . (1972)  80% o f the  These compounds a r e 80% of the extract.  However, Jones e t a l . (1979) r e p o r t e d yellow  t h a t cabbage l o o p e r , T. nd^,  and  w o o l l y b e a r , Spilosoma v i r g i n i c a , were not i n h i b i t e d from f e e d i n g  diet containing  the s e s q u i t e r p e n e  lactones, glaucolide-A.  r e s u l t s i n d i c a t e that s e v e r a l sesquiterpene i n h i b i t o r y and sesquiterpene  feeding  deterrent  l a c t o n e s are  a c t i v i t y against a l l insect species In the p r e s e n t  above  l a c t o n e s have i n s e c t growth  properties.  e f f e c t i v e and  The  However, not a l l  those t h a t are do not  show  tested.  study f o u r f r a c t i o n s of a c h r o m a t o g r a p h i c a l l y  e t h a n o l i c e x t r a c t of _A. t r i d e n t a t a were assayed, and  two  separated  fractions  accounted f o r n e a r l y a l l o f the growth i n h i b i t o r y a c t i v i t y of the e x t r a c t (Table  IX).  T h i n l a y e r chromatographic s e p a r a t i o n s  showed that s e v e r a l major s p o t s t h a t these were s e s q u i t e r p e n e Camphor and  1,8-cineole,  reacted  lactones  1 , 8 - c i n e o l e has  to a v a n i l l i n r e a g e n t ,  suggesting  (Pieman e t a l . 1980).  been shown t o r e p e l the American c o c k r o a c h ,  major f r a c t i o n s i n t h i s t h e s i s r e v e a l e d t h a t may  (TLC)  s e v e r a l spots  be monoterpenes (Croteau  and  that  Ronald  i n the most growth i n h i b i t o r y f r a c t i o n s ( F i g s . 7-10).  coumarins r e p o r t e d feeding  feeding deterrency  from A^. t r i d e n t a t a and  of C o l o r a d o p o t a t o b e e t l e l a r v a e .  shown t o i n h i b i t l a r v a l growth and  7-10  and  Periplaneta  T h i n l a y e r chromatographic  Jermy e t a l . (1981) examined the  in  insects.  t o be a mosquito r e p e l l e n t (Hwang e t a l . 1985),  quenched u l t r a v i o l e t l i g h t 1983)  8-11)  major monoterpenes i n the e s s e n t i a l o i l o f _A.  americana ( S c r i v e n and Meloan 1984). study of the two  initial  (Figs.  t r i d e n t a t a have p r e v i o u s l y been shown to be h i g h l y a c t i v e a g a i n s t Camphor i s r e p o r t e d  on  spots  several  found t h a t none of t h e s e reduced Coumarin has,  however, been  development as w e l l as a d u l t  the c o t t o n leafworm, Spodoptera l i t t o r a l i s s e v e r a l TLC  of  showed a weak blue  (Mansour 1982).  fluorescence  f r a c t i o n s of A^. t r i d e n t a t a t h a t c o u l d be coumarins.  i n the  Isman and  fertility In  Figures  active Rodriguez  76 (1983) r e p o r t e d that q u e r c e t a g e t i n 3,7-dimethy e t h e r , a f l a v o n o i d  from  a l a r v a l growth i n h i b i t o r o f l\_. zea  guayule (Parthenium argentatum) was  and  J3. exigua whereas a c l o s e l y r e l a t e d 6-hydroxykaempferol 3 , 6 , 7 - t r i m e t h y l e t h e r was i n A.  stimulatory  to both i n s e c t s p e c i e s .  t r i d e n t a t a have not The  been i n v e s t i g a t e d  a e r i a l p a r t s o f A^. t r i d e n t a t a are  b i o l o g i c a l l y a c t i v e compounds.  The  o t h e r p h e n o l i c compounds  for insect  activity.  known t o p o s s e s s a wide range of  Indigenous peoples of B r i t i s h Columbia used  A^. t r i d e n t a t a f o l i a g e as a d i s i n f e c t a n t , i n s e c t r e p e l l e n t and deodorant when h a n d l i n g c o r p s e s (Turner 1979). 1967)  and  non-volatile  (Ramirez 1969)  possess a n t i b a c t e r i a l a c t i v i t y . been r e p o r t e d from v o l a t i l e and  non-volatile  a  V o l a t i l e (Nagy and  components of the  In a d d i t i o n ,  as  leaves are  Tengerdy known t o  allelopathic activity  has  l e a f f r a c t i o n s (Groves  and  Anderson 1981). Seasonal (Kelsey e t a l . 1982) 1971)  v a r i a t i o n s i n the  terpenoid  and  c o n t e n t of _A. t r i d e n t a t a have been  r e p o r t e d ; u n t i l the a c t i v e i n g r e d i e n t s occurring  compounds, one  i n h i b i t o r y and K e l s e y and  are  known and  bioassayed with  must be c a u t i o u s i n i n t e r p r e t i n g i n s e c t  o v i p o s i t i o n deterrence  the  growth  biological activity  be a r e s u l t of synergism between the  of  v o l a t i l e essential  o t h e r secondary compounds l i k e s e s q u i t e r p e n e l a c t o n e s  phenolics.  co-  activity.  co-workers (1983) suggest t h a t  A_. t r i d e n t a t a may o i l s and  i n t r a s p e c i f i c (Shafizadeh et a l .  and  Although t h i s h y p o t h e s i s i s i n t r i g u i n g i t i s n o n e t h e l e s s  speculative.  C.  F i e l d T r i a l s of P l a n t The  requires and  Extracts  proper s e l e c t i o n of b o t a n i c a l s the  evaluation  greenhouse s t u d i e s  of e x t r a c t s have not  as  i n the  field-active control  agents  target area.  laboratory  always p r e d i c t e d  the  Since  e f f e c t s i n the  field  77 ( O b r y c k i and Tauber 1984,  Haverty and  Robertson 1982), f i e l d  e s s e n t i a l part of a complete s c r e e n i n g p r o c e d u r e .  save f o r work on neem e x t r a c t s .  F i g u r e s 4-6  an  While l a b o r a t o r y  s c r e e n i n g s of crude p l a n t e x t r a c t s f o r growth i n h i b i t o r s and d e t e r r e n t s are not uncommon, r e p o r t s of f i e l d  s t u d i e s are  trials  feeding  on e x t r a c t s i s s c a r c e ,  show the r e s u l t s of a  field  t r i a l on cabbage u s i n g an e t h a n o l i c e x t r a c t of _A. t r i d e n t a t a s e l e c t e d i n the l a b o r a t o r y . The  s u i t a b l e bioassay  f o r both the p e s t and  intended a p p l i c a t i o n  should be c a r e f u l l y chosen when s c r e e n i n g p l a n t e x t r a c t s a g a i n s t insects.  For example, the a c r i d i d , L o c u s t a  pest  m i g r a t o r i a , was s u b s t a n t i a l l y  more s e n s i t i v e t o a wider range of compounds than f o u r l e p i d o p t e r a n s p e c i e s t e s t e d (Simmonds e t a l . 1985).  The  anthranoid,  harunganine, was  phagodeterrent t o the polyphagous c o t t o n leafworm Spodoptera when presented  on cabbage, whereas the same compound was  the h o s t p l a n t was et  pest a  littoralis  i n e f f e c t i v e when  wheat, d e s p i t e cabbage b e i n g a p r e f e r r e d p l a n t (Simmonds  a l . 1985). The  d i f f e r e n c e i n response t o a treatment can depend more on the  method than on the product  tested.  For example, l a r v a e f e d  d i e t s t r e a t e d w i t h moderate amounts of a l l e l o c h e m i c a l s may l a r v a l growth response o f t r e a t e d v e r s u s c o n t r o l l a r v a e . f a c t o r t o c o n s i d e r when u s i n g b i o a s s a y s  i s the number of c h o i c e s g i v e n an i n s e c t .  bioassay  r e s u l t e d i n 100%  19% d e t e r r e n c e  deterrency  i n a s i n g l e choice bioassay  Benz 1984).  exaggerate the Another  important  inhibitory  A dual choice  feeding  of P i e r i s b r a s s i c a e compared w i t h  c o n c e n t r a t i o n of an A r t e m i s i a a b s i n t h i u m ( A b i v a r d i and  supra-optimal  to s c r e e n p e s t i c i d a l or  products  test  when f e d an  equivalent  extract applied to l e a f discs  R e s u l t s from l a b o r a t o r y s c r e e n i n g s  should  be  only  78 s u b s t a n t i a t e d i n the t a r g e t a r e a t e s t i n g candidates of the b i o a s s a y The  ( e . g . , greenhouse or f i e l d ) .  s e l e c t e d i n the l a b o r a t o r y may  Field  determine the  relevance  f o r s c r e e n i n g s u i t a b l e c o n t r o l agents.  relevance  of u s i n g r e s u l t s from l a b o r a t o r y r e a r e d i n s e c t s to  calculate concentration l e v e l s for f i e l d screening i n s e c t c o n t r o l products. d i f f e r e n t l y than f i e l d  s t a b l e under f i e l d  Laboratory  populations  1986), j u s t as products  trials  should  be addressed when  r e a r e d i n s e c t s may  of the same s p e c i e s ( B r a t t s t e n e t a l .  t h a t f u n c t i o n w e l l i n the l a b o r a t o r y may  conditions.  respond  not  I have shown ( T a b l e V I I I ) t h a t JP. s a u c i a  l a r v a e from an F^ g e n e r a t i o n o f a f i e l d  c o l l e c t e d p o p u l a t i o n were 2.5  h e a v i e r than l a r v a e from a two-year-old  laboratory reared colony  same a r t i f i c i a l d i e t w i t h 50% extract.  Therefore,  be  fed  times the  (dwt/dwt) of an e t h a n o l i c _A. t r i d e n t a t a  a realistic  e s t i m a t i o n of the e x t r a c t c o n c e n t r a t i o n  needed should be performed on f i e l d  c o l l e c t e d i n s e c t s or t h e i r o f f s p r i n g .  Another p a r t of t h i s experiment showed the a c c u r a c y  of the  bioassay  There was  f o r r e p o r t i n g r e l a t i v e growth i n h i b i t i o n .  laboratory no  s i g n i f i c a n t growth d i f f e r e n c e s (p=0.05) between the l a b o r a t o r y r e a r e d field  or  c o l l e c t e d p o p u l a t i o n s of JP. s a u c i a l a r v a e ( T a b l e V I I I ) f e d d i e t  c o n t a i n i n g _A. t r i d e n t a t a e x t r a c t when compared to t h e i r r e s p e c t i v e controls. N a t u r a l p l a n t defenses may of p e s t p o p u l a t i o n s .  be of use i n a g r i c u l t u r e f o r the management  Several researchers  crude p l a n t e x t r a c t s s h o u l d not p r e s e n t 1983,  have noted t h a t the s p r a y i n g of  insurmountable problems (Jacobson  Jermy e t a l . 1981), p a r t i c u l a r l y f o r underdeveloped c o u n t r i e s where  variable e f f i c a c y i s acceptable. feasibility ethanol.  of f i e l d  The  r e s u l t s r e p o r t e d h e r e i n show the  s p r a y i n g _A. t r i d e n t a t a e x t r a c t s f o r m u l a t e d  in  30%  79 My  field  t r i a l with an _A. t r i d e n t a t a e x t r a c t a g a i n s t cabbage i n s e c t  pests using a s i n g l e a p p l i c a t i o n r e s u l t e d and  lower  l a r v a l pest counts  i n a higher q u a l i t y cabbage y i e l d  than the s o l v e n t t r e a t e d c o n t r o l s ( T a b l e V I I ) . TM  The  standard i n s e c t i c i d e spray of d e l t a m e t h r i n and  h i g h e s t q u a l i t y cabbage.  The  Superspred  gave the  b e n e f i t s of the _A. t r i d e n t a t a e x t r a c t were  e v i d e n t f o r the f i r s t week a f t e r s p r a y i n g , but i n t e g r a t i n g other o p t i o n s such as s p r a y i n g more f r e q u e n t l y or combining c o n t r o l may  control  strategies  make the _A. t r i d e n t a t a e x t r a c t more e f f e c t i v e . Cabbage l o o p e r e q u i v a l e n t s (CLEs) have been used f o r measuring pest  damage on cabbage from the t h r e e major l e p i d o p t e r a n cabbage p e s t s ( S h e l t o n et  a l . 1982).  In the p r e s e n t t h e s i s , _A. t r i d e n t a t a sprayed  cabbage  s i g n i f i c a n t l y l e s s damaged by major i n s e c t p e s t s , e s t i m a t e d as CLEs, the s o l v e n t t r e a t e d c o n t r o l s ( F i g . A ) .  D e l t a m e t h r i n was  r e p o r t e d t o decrease  Basedow e t a l . 1985,  t h a t one  lower  s p r a y s , however, are  the p o p u l a t i o n s of s e v e r a l p r e d a t o r y arthropods  as c a r a b i d s , s t a p h i l i n i d s , s p i d e r s and 1985,  Deltamethrin  p h y t o s e i i d mites  Samsoe-Petersen 1985).  than  shown t o be a good  c h o i c e as an i n s e c t i c i d e s t a n d a r d as i t r e s u l t e d i n s i g n i f i c a n t l y CLEs than any of the o t h e r t r e a t m e n t s .  was  such  (Matcham and Hawkes  Bernays (1983) suggests  of the p o t e n t i a l advantages of s p r a y i n g p l a n t s w i t h growth  i n h i b i t o r y compounds i s t h a t they may An important  r e s u l t of the f i e l d  s i g n i f i c a n t l y lower cabbage ( F i g . 6 ) .  l a r v a l ICW The  lower  a v o i d damage t o n o n - t a r g e t experiment was  observed  i n the  counts on the _A. t r i d e n t a t a e x t r a c t sprayed  counts of l a r v a l P_. rapae may  i n d i r e c t mode of a c t i o n of the _A. t r i d e n t a t a e x t r a c t . counts c o u l d e x p l a i n most, i f not a l l , of the reduced i n f e s t i n g the cabbage f o l i a g e .  organisms.  be due  to an  Reduced P_. rapae larval  populations  egg  In p l a n t s , p h y t o c h e m i c a l s t h a t d e t e r c h e m i c a l defense a g a i n s t  herbivorous i n s e c t s .  p l a n t s , they determine l a r v a l food generation. and  o v i p o s i t i o n are the f i r s t  l i n e of  When females o v i p o s i t on  c h o i c e and s u r v i v a l o f t h e succeeding  To o p t i m i z e l a r v a l s u r v i v o r s h i p , a d u l t o v i p o s i t i o n p r e f e r e n c e  l a r v a l food  s u i t a b i l i t y s h o u l d be s y n c h r o n i z e d .  Non-host  plant  e x t r a c t s a p p l i e d t o c r o p p l a n t s may d e t e r o v i p o s i t i o n , and thus p r o t e c t t h e c r o p from h e r b i v o r y , rapae .  i f l a r v a e have l i m i t e d m o b i l i t y as i n t h e case o f P_.  Lundgren (1975) t e s t e d s e v e r a l p l a n t e x t r a c t s , i n c l u d i n g  a b s i n t h i u m and A^. abrotanum, f o r t h e i r a b i l i t y three P i e r i s species  t o deter  Artemisia  o v i p o s i t i o n of  i n two-choice t e s t s , and found s i g n i f i c a n t l y  eggs l a i d on e x t r a c t t r e a t e d cabbage l e a v e s . tridentata extract applied  to f i e l d  fewer  In t h i s t h e s i s an _A.  grown cabbage r e s u l t e d i n s i g n i f i c a n t l y  fewer (p<0.05) P_. rapae eggs compared t o cabbage t r e a t e d w i t h t h e c a r r i e r s o l v e n t a l o n e and t h e d e l t a m e t h r i n - s u r f a c t a n t deltamethrin-surfactant  treatment ( F i g . 7 ) . The  t r e a t e d cabbage r e c e i v e d  t h e most P_. rapae eggs.  TM (The  surfactant, Triton-X-100  , has been shown t o i n c r e a s e  x y l o s t e l l a o v i p o s i t i o n on B r u s s e l the  surfactant  Plutella  s p r o u t s [ P e r r i n and P h i l l i p s 1978], but  e f f e c t was n o t i s o l a t e d i n t h i s experiment.)  Laboratory  experiments confirmed t h e o v i p o s i t i o n d e t e r r e n c y o f t h e _A. t r i d e n t a t a extract.  O b s e r v a t i o n s i n d i c a t e d t h e mode o f o v i p o s i t i o n d e t e r r e n c y o f t h e  A- t r i d e n t a t a t r e a t e d cabbage was due t o a c o n t a c t than r e p e l l e n c y ,  chemoreception  rather  because cabbage b u t t e r f l i e s were not i n h i b i t e d from  a l i g h t i n g on t h e t r e a t e d  plants.  However, whether t h e P_. rapae females were d e t e r r e d  from o v i p o s i t i n g ,  or i f t h e p l a n t s were u n r e c o g n i z a b l e as h o s t p l a n t s , i s n o t c l e a r . F o r example, c u t i c u l a r components i n t o b a c c o have been shown t o s t i m u l a t e o v i p o s i t i o n o f R. v i r e s c e n s  ( C u t l e r e t a l . 1986).  T a r s a l contact  cabbage f o l i a g e was found t o have i m p o r t a n t i n f l u e n c e s  with  on t h e o v i p o s i t i o n  behavior of P_. rapae, whereas h o s t - p l a n t  odor, o v i p o s i t o r t i p c o n t a c t  p r e v i o u s l y l a i d eggs showed no i n f l u e n c e ( T r a y n i e r l a b o r a t o r y experiments r e p o r t e d whether s p r a y i n g  1979).  and  In the f i e l d  i n t h i s t h e s i s i t remains to be  and  determined  the cabbage masked o v i p o s i t i o n s t i m u l a t i n g c u t i c u l a r  c h e m i c a l s , blocked  P_. rapae chemoreceptors, or a c t i v a t e d  deterrent  receptors. Non-host p l a n t e x t r a c t s and as o v i p o s i t i o n d e t e r r e n t s and  non-cruciferous  for  P. rapae (Renwick and  s p e c i f i c p h y t o c h e m i c a l s have been examined  t o i n s e c t p e s t s of cabbage.  e x t r a c t s have been r e p o r t e d Radke 1985).  coumarin sprayed cabbage d e t e r r e d field  experiment r e p o r t e d  herein  Non-host c r u c i f e r o u s  as o v i p o s i t i o n  deterrents  Tabashnik (1985) showed t h a t  o v i p o s i t i o n by P_. x y l o s t e l l a . ( F i g . A), the l a r g e p o p u l a t i o n  In of  the £.  x y l o s t e l l a l a r v a e on A., t r i d e n t a t a sprayed p l a n t s i n d i c a t e d t h a t i f coumarins were p r e s e n t at  the c o n c e n t r a t i o n Field  are r a r e .  trials  The  i n s e c t s may  of cages and  laboratory reared  have l i t t l e and  bearing  and  the other  three lepidopterans,  f o r the s e s q u i t e r p e n e  Williams  laboratory  Y e l l o w w o o l l y b e a r , Spilosoma the cabbage l o o p e r , T_. n i ,  plants containing  armyworms, Spodoptera f r u g i p e r d a , a preference  i n s e c t s (e.g.,  l a c t o n e s , showed t h a t o v i p o s i t i o n  v i r g i n i c a , showed no o v i p o s i t i o n p r e f e r e n c e , f o r the two  have  cage e x p e r i m e n t s w i t h Vernonia  depended on the s p e c i e s o f moth.  showed a p r e f e r e n c e  cages  on the a c t u a l f i e l d s i t u a t i o n .  Jones (1978) u s i n g  p l a n t s , w i t h and without s e s q u i t e r p e n e preference  field  e f f e c t s of the c o n t r o l l e d environments and  Nonetheless, Burnett  field  w i t h o u t the a i d of f i e l d  Most i n v e s t i g a t o r s t h a t have ventured i n t o the  a l . 1986).  reared  f a c t o r s i n the  sprayed.  of o v i p o s i t i o n d e t e r r e n t s  r e s o r t e d t o the use et  i n the e x t r a c t , they were not  (fall,  s o u t h e r n and  S^. e r i d a n i a , and lactone  sesquiterpene  lactones  yellowstriped  S^. o r n i t h o g a l l i ) showed  l a c k i n g Vernonia  species.  82 B u r n e t t and  Jones (1978) a l s o showed t h a t the  significantly  i n h i b i t e d from o v i p o s i t i n g on  sesquiterpene lactones The  when 1% g l a u c o l i d e - A  sesquiterpene lactones  responsible  f a l l armyworm  was  the V e r n o n i a s p e c i e s was  applied  to the f o l i a g e .  i n the A^. t r i d e n t a t a e x t r a c t c o u l d  f o r the o v i p o s i t i o n d e t e r r e n c y r e p o r t e d  lacking  therefore  i n t h i s t h e s i s on  be  £.  rapae . Combining c o n t r o l s t r a t e g i e s ( i e . i n s e c t i c i d e and have advantages over the use i n s e c t i c i d e s may  of e i t h e r agent a l o n e .  p r o l o n g the use  of e x i s t i n g and  by s l o w i n g the r a t e of i n s e c t r e s i s t a n c e  plant extract)  could  Combinations of  novel c o n t r o l  (Georghiou 1983).  techniques  Reduced  s y n t h e t i c i n s e c t i c i d e use would lower the i n s e c t i c i d e l o a d on the c r o p i n the environment and  may  Growth i n h i b i t o r s and  permit the r e t u r n of b e n e f i c i a l organisms. o v i p o s i t i o n deterrents  n a t u r a l enemies i f they are not a d v e r s e l y use  of growth i n h i b i t o r s c o u l d ,  may  enhance the a c t i o n of  a f f e c t e d by t h e s e compounds.  f o r example, be used i n c o n j u n c t i o n  the r e l e a s e of i n s e c t p a r a s i t e s and  predators.  The  with  Weseloh e t a l . (1983) have  shown t h a t r e l e a s e of the p a r a s i t i c wasp, A p a n t e l e s m e l a n o s c e l u s , and s p r a y s of the l e p i d o p t e r a n  and  pathogen, B a c i l l u s t h u r i n g i e n s i s ,  field  acted  s y n e r g i s t i c a l l y t o c o n t r o l gypsy moth because the b a c t e r i a maintained  the  larvae longer  by  the  parasite.  looper,  i n the  second i n s t a r , which i s the host stage p r e f e r r e d  V e l v e t bean c a t e r p i l l a r , A n t i c a r s i a gemmatalis, and  Pseudoplusia includens,  pathogen, Nomuraea r i l e y i ,  are more s u s c e p t i b l e to the  entomophagous  i n the e a r l y i n s t a r s (Boucias e t a l . 198A).  growth i n h i b i t o r s m a i n t a i n i n s e c t s i n s t a g e s v u l n e r a b l e p a r a s i t e s , they may  soybean  to predators  p r o v i d e another t o o l f o r p r o t e c t i n g c r o p s i n  i n t e n s i v e l y managed a g r i c u l t u r a l systems.  and  If  83 Mammalian t o x i c i t y of a p e s t i c i d e .  While  i s an important f a c t o r when c o n s i d e r i n g the m e r i t s t h e r e a r e no s t u d i e s on mammalian t o x i c i t y of  e t h a n o l i c ti. t r i d e n t a t a e x t r a c t s , b i g sagebrush f o r a g e by pronghorn  i s used as a major winter  a n t e l o p e ( C r o n i n e t a l . 1978), mule deer  Reid 1975), and pygmy r a b b i t s (White e t a l . 1982).  (Hansen and  In a d d i t i o n ,  s e s q u i t e r p e n e l a c t o n e s , one of the p r i n c i p l e c l a s s e s of secondary i n _A. t r i d e n t a t a , have been i n v e s t i g a t e d as antitumor agents  compounds  (Lee e t a l .  1977). P l a n t - d e r i v e d c h e m i c a l s a r e most o f t e n b i o d e g r a d a b l e and  thus they  might prove t o be p r e f e r r e d a l t e r n a t i v e s t o s y n t h e t i c c h e m i c a l s p e r s i s t i n the environment.  that  _A. t r i d e n t a t a i s l i s t e d as one of f o u r weeds  i n the U n i t e d S t a t e s w i t h the most p o t e n t i a l f o r c r o p development and c o m m e r c i a l i z a t i o n f o r s o u r c e s o f i n s e c t a t t r a c t a n t s , r e p e l l e n t s or t o x i c a n t s (Jacobson 1983). Warren 1981) 1981). new  T h i s s p e c i e s i s drought  and can support a w i n t e r shoot removal  t o l e r a n t ( R i c k a r d and o f about  50% ( F e t c h e r  The r e s i l i e n c e o f _A. t r i d e n t a t a shrubs adds t o t h e i r p o t e n t i a l as a  c r o p f o r e x p l o i t i n g s e m i - a r i d m a r g i n a l lands (Jacobson  1983).  Insects that are r a p i d l y developing r e s i s t a n c e to s y n t h e t i c i n s e c t i c i d e s c r e a t e problems f o r p e s t c o n t r o l , w h i l e p l a n t s , w i t h t h e i r m u l t i c h e m i c a l d e f e n s e s , may  c o n t a i n s o l u t i o n s t o some of t h e s e  problems.  R e l i a n c e on mono-chemical pest c o n t r o l i s inadequate and thus i t i s an opportune  time t o study pest c o n t r o l methods t h a t have e v o l v e d i n p l a n t s .  84 VI. The  CONCLUSIONS  o b j e c t i v e s of t h i s t h e s i s were t o s e l e c t a potent growth  i n h i b i t o r y e x t r a c t from an a s t e r a c e o u s weed, to a s s e s s the e x t r a c t f o r d e l e t e r i o u s e f f e c t on i n s e c t s , to e v a l u a t e potent growth i n h i b i t o r and inhibitory The  to e x p l o r e  the  field  e f f i c a c y of the most  the c h e m i s t r y of i n s e c t growth  activity.  r e s u l t s show t h a t :  1. Of  s i x a s t e r a c e o u s weeds e x t r a c t e d  i n EtOH and  p e t r o l , s i x of  12 e x t r a c t s i n h i b i t e d P_. s a u c i a l a r v a l growth by more than 90%  the  compared  to  the growth of c o n t r o l l a r v a e ( a t f i v e times n a t u r a l l y o c c u r r i n g concentrations). 2. N a t u r a l l y o c c u r r i n g c o n c e n t r a t i o n s l e a v e s and  flowers  of _A. t r i d e n t a t a and  two  of an e t h a n o l i c e x t r a c t from of i t s chromatographic  f r a c t i o n s s i g n i f i c a n t l y i n h i b i t e d e a r l y l a r v a l growth of P_. 3. F e e d i n g b i o a s s a y s  saucia.  showed a s i g n i f i c a n t dose-response by P_.  and _A. c a l i f o r n i c a l a r v a e t o _A. t r i d e n t a t a and no  the  saucia  C_. s u a v e o l e n s e x t r a c t s ,  s i g n i f i c a n t d i f f e r e n c e s were found between these e x t r a c t s .  but,  Extracts  from both p l a n t s i n h i b i t e d growth more i n A_. c a l i f o r n i c a l a r v a e than i n P_. s a u c i a l a r v a e and in  the _A. t r i d e n t a t a e t h a n o l i c e x t r a c t i n h i b i t e d growth more  A. c a l i f o r n i c a l a r v a e than the C. 4.  suaveolens e x t r a c t s .  Second i n s t a r P_. s a u c i a l a r v a e were l e s s s e n s i t i v e t o the  i n h i b i t o r y e f f e c t s of the e x t r a c t s than n e o n a t a l P_. 5. R e s u l t s  obtained  from the  field  t r i d e n t a t a e t h a n o l i c e x t r a c t a t 0.2  g/ml  trial  growth  saucia.  suggest t h a t the 30%  protected  cabbage from i n s e c t  damage s i g n i f i c a n t l y b e t t e r than the water or EtOH c o n t r o l s . I n s e c t damage t o cabbage, however, was spray a t 17.9  s i g n i f i c a n t l y l e s s w i t h the  u g / l than i n a l l other  treatments.  aq  A_. pest  pest  deltamethrin  6.  R e s i d u a l o v i p o s i t i o n d e t e r r e n c y t o P_. rapae was suggested  r e s u l t s o b t a i n i n the f i e l d  trial.  Laboratory  from  experiments w i t h caged J?.  rapae appear t o c o n f i r m a c o n t a c t o v i p o s i t i o n d e t e r r e n c y due t o the A^. t r i d e n t a t a e t h a n o l i c e x t r a c t a t 0.2 7.  g/ml on cabbage.  An F-^ g e n e r a t i o n o f f i e l d - c o l l e c t e d P. s a u c i a grew s i g n i f i c a n t l y  b e t t e r than the l a r v a e from t h e l a b o r a t o r y c o l o n y .  However, the growth  i n h i b i t i o n of P_. s a u c i a l a r v a e by t h e _A. t r i d e n t a t a e x t r a c t was not s i g n i f i c a n t l y d i f f e r e n t between the two p o p u l a t i o n s r e l a t i v e t o t h e i r respective  controls.  My f i n d i n g s r e v e a l e d t h a t u s i n g i n s e c t growth i n h i b i t o r y e x t r a c t s , s e l e c t e d i n l a b o r a t o r y bioassays, should c o n s t i t u t e only the f i r s t the development o f n o v e l b o t a n i c a l pest c o n t r o l s .  stage i n  The next stage  should  i n v a r i a b l y be t e s t i n g t h e products on t a r g e t p l a n t s and i n s e c t s i n the field.  F u r t h e r i n v e s t i g a t i o n s u s i n g more i n t e n s i v e  t e c h n i q u e s may h e l p e l u c i d a t e the s p e c i f i c c h e m i c a l s  phytochemical or c h e m i c a l  r e s p o n s i b l e f o r both t h e growth and o v i p o s i t i o n i n h i b i t o r y The  mixtures  activity.  e x t r a c t i o n and s c r e e n i n g o f a s t e r a c e o u s weeds has advanced t h e  p o t e n t i a l use o f these n a t u r a l p r o d u c t s i n i n s e c t p e s t c o n t r o l  programmes.  E f f e c t i v e i n s e c t growth i n h i b i t o r s and o v i p o s i t i o n d e t e r r e n t s may be used in  combination  w i t h other pest c o n t r o l o p t i o n s p r o v i d e d they a r e n o n - t o x i c  to  humans, economical  t o produce, and harmless i n the environment.  86  VII.  BIBLIOGRAPHY  A b i v a r d i , C. and G. Benz. 1984. T e s t s with e x t r a c t s o f 21 m e d i c i n a l p l a n t s f o r a n t i f e e d a n t a c t i v i t y a g a i n s t l a r v a e of P i e r i s b r a s s i c a e L. B u l l . Soc. Entomol. S u i s s e 57:383-392. Adams, C. M. and E . A. Bernays. 1978. The e f f e c t of combinations of d e t e r r e n t on the f e e d i n g behavior o f L o c u s t a m i g r a t o r i a . Entomol. Exp. A p p l . 23:101-109. A g o s i n , M. and A. S. P e r r y . 1974. Microsomal m i x e d - f u n c t i o n o x i d a s e s . V.5, pp.538-596 i n M. R o c k s t e i n , ed. Biochemisty o f I n s e c t s . Academic P r e s s , N.Y. Ahmad, S. 1986. Enzymatic a d a p t a t i o n s of h e r b i v o r o u s i n s e c t s and mites to p h y t o c h e m i c a l s . J . Chem. E c o l . 12:533-559. Basedow, T. H. Rzehak, and K. Vob. 1985. S t u d i e s on the e f f e c t of d e l t a m e t h r i n sprays on the number of e p i g e a l p r e d a t o r y a r t h r o p o d s o c c u r r i n g i n a r a b l e f i e l d s . P e s t i c . S c i . 16:325-331. B a s o l , M. S. 1980. Comparative t o x i c i t y of some p e s t i c i d e s on human h e a l t h and some a q u a t i c s p e c i e s . J . E n v i r o n . S c i . H e a l t h [B] B15_:993-1004. Beck, S. D. 1960. The European c o r n b o r e r , P y r a u s t a n u b i l a l i s (Hubn.), and i t s p r i n c i p a l host p l a n t . V I I . L a r v a l f e e d i n g behavior and host p l a n t r e s i s t a n c e . Ann. Entomol. Soc. Am. 53:206-212. B e i r n e , B. P. 1971. P e s t i n s e c t s o f annual crop p l a n t s i n Canada I . L e p i d o p t e r a I I . D i p t e r a I I I . C o l e o p t e r a . Memoirs Entomol. Soc. Canada, No. 78. Ottawa, Canada. 124 pp. B e l l , W. J . and R. T. Carde, eds. 1984. Chemical Ecology of I n s e c t s . S i n a u e r Assoc., I n c . , Sunderland, Mass. 524 pp. B e n t l e y , M. D. and 8 o t h e r s . 1982. E f f e c t of some n a t u r a l l y o c c u r r i n g c h e m i c a l s and e x t r a c t s of non-host p l a n t s on f e e d i n g by spruce budworm l a r v a e ( C h o r i s t o n e u r a f u m i f e r a n a ) L i f e S c i e n c e s and A g r i c . Exp. S t a . , U n i v . o f Maine (Orono), Tech. B u l l . 107. Berenbaum, M. 1985. Brementown r e v i s i t e d i n t e r a c t i o n s among a l l e l o c h e m i c a l s i n p l a n t s . Rec. Adv. Phytochem. 19:139-169 Berenbaum, M. and J . J . N e a l . 1985. Synergism between m y r i s t i c i n and x a n t h o t o x i n , a n a t u r a l l y c o o c c u r r i n g p l a n t t o x i c a n t . J . Chem. E c o l . 11:1349-1358. Bernays, E . A. 1983. A n t i f e e d a n t s i n crop pest management, pp. 259-269 in_ D. L. Whitehead and W. S. Bower, e d s . N a t u r a l Products f o r I n n o v a t i v e P e s t Management, [Current Themes i n T r o p i c a l S c i e n c e ] . V o l . 2. Pergamon P r e s s , Oxford. 583 pp.  87 Bernays, E. A. and R. F. Chapman. 1977. D e t e r r e n t c h e m i c a l s as a b a s i s of o l i g o p h a g y i n L o c u s t a m i g r a t o r i a ( L . ) . E c o l . Entomol. 2_:1-18. Bernays, E. A. and R. F. Chapman. 1978. P l a n t c h e m i s t r y and a c r i d o i d f e e d i n g b e h a v i o r , pp.99-141 in_ J.B. Harborne, ed. B i o c h e m i c a l Aspects of P l a n t and Animal C o e v o l u t i o n . Academic P r e s s , London. Bernays, E. A. and S. J . Simpson. 1982. I n s e c t P h y s i o l . 16:59-118.  C o n t r o l of food i n t a k e .  B i g n e l l , D. E. 1978. E f f e c t s of c e l l u l o s e i n the d i e t s of Entomol. Exp. A p p l . 24:254-257.  Adv.  cockroaches.  Bjorkman, R. 1976. P r o p e r t i e s and f u n c t i o n of p l a n t m y r o s i n a s e s . pp.191205 i n J.G. Vaughan, A.J. MacLeod and B.M.G. Jones, eds. The B i o l o g y and Chemistry of the C r u c i f e r a e . Academic P r e s s , N.Y. 355 pp. B l a u , P. A., P. Feeny, P., L. Contardo, D. S. Robson. 1978. A l l y l g l u c o s i n o l a t e and h e r b i v o r o u s c a t e r p i l l a r s : A c o n t r a s t i n t o x i c i t y and t o l e r a n c e . S c i e n c e 200:1296-1298. B o i t e a u , G. King, R. R. and D. Levesque. 1985. L e t h a l and s u b l e t h a l e f f e c t s of a l d i c a r b on two potato aphids Myzus p e r s i c a e and Macrosiphum euphorbiae. J . Econ. Entomol. 78:41-44. B o u c i a s , D. G., D. L. B r a d f o r d , and C. S. B a r f i e l d . 1984. S u s c e p t i b i l i t y o f the v e l v e t b e a n c a t e r p i l l a r and soybean l o o p e r t o Nomuraea r i l e y i : E f f e c t s of pathotype, dosage, temperature, and h o s t age. J . Econ. Entomol 77:247-253. B r a t t s t e n , L. B. 1979. B i o c h e m i c a l defense mechanisms i n h e r b i v o r e s a g a i n s t p l a n t a l l e l o c h e m i c a l s . pp.199-270 i n G.E. Rosenthal and Janzen, eds. H e r b i v o r e s : T h e i r I n t e r a c t i o n with Secondary P l a n t M e t a b o l i t e s . Academic P r e s s , New York.  D.H.  B r a t t s t e n , L. B., C. W. Holyoke, J r . , J . R. Leeper, and K. F. R a f f a . I n s e c t i c i d e r e s i s t a n c e : C h a l l e n g e t o pest management and b a s i c r e s e a r c h . S c i e n c e 231:1255-1260.  1986.  Brown, D., F.0. Asplund and V. McMahon. 1975. P h e n o l i c c o n s t i t u e n t s of A r t e m i s i a t r i d e n t a t a spp. vaseyana. Phytochem. 14:1083-1084. B u r n e t t , W. C , J r . , and S. B. Jones. 1978. I n f l u e n c e of s e s q u i t e r p e n e l a c t o n e s o f V e r n o n i a (Compositae) on o v i p o s i t i o n p r e f e r e n c e s o f L e p i d o p t e r a . Amer. Mid. Nat. 100:242-245. B u r n e t t , W. C , J r . , S. B. Jones, J r . , T. J . Mabry and W. G. P a d o l i n a . 1974. S e s q u i t e r p e n e l a c t o n e s - i n s e c t f e e d i n g d e t e r r e n t s i n V e r n o n i a . Biochem. System. E c o l . J2:25-29. B u t t k u s , J . A., R. J . Bose, and D. A. Shearer. 1977. Terpenes i n the e s s e n t i a l o i l o f sagebrush. J . A g r i c . Food Chem. 25:288-291.  88 Campbell, B. C. and S. S. D u f f e y . 1981. A l l e v i a t i o n o f a l p h a - t o m a t i n e induced t o x i c i t y t o t h e p a r a s i t o i d , Hyposoter exiguae, by p h y t o s t e r o l s i n the d i e t o f the host H e l i o t h i s z e a . J . Chem. E c o l . 7^:927-944. C a r r o l l , C. F. and C. Hoffman. 1980. Chemical f e e d i n g d e t e r r e n t m o b i l i z e d to i n s e c t h e r b i v o r y and counter a d a p t a t i o n by Epiachna t r e d e c i m n o t a t a . S c i e n c e 209:414-416. Chapman, R. F. 1974. The c h e m i c a l i n h i b i t i o n o f f e e d i n g by phytophagous i n s e c t s : a r e v i e w . B u l l . Entomol. Res. 64:339-363. Chew, F. S. 1975. C o e v o l u t i o n o f p i e r i d b u t t e r f l i e s and t h e i r c r u c i f e r o u s f o o d p l a n t s . I I . The r e l a t i v e q u a l i t y o f a v a i l a b l e r e s o u r c e s . O e c o l o g i a 20:117-127. Chew, F. S. 1980. Foodplant 46:347-353.  preferences of P i e r i s c a t e r p i l l a r s .  Oecologia  Courtney, S. P. 1981. C o e v o l u t i o n o f p i e r i d b u t t e r f l i e s and t h e i r c r u c i f e r o u s f o o d p l a n t s I I I . A n t h o c h a r i s cardamines s u r v i v a l , development and o v i p o s i t i o n . O e c o l o g i a 51:91-96. Courtney, S. P. 1986. The ecology o f p i e r i d b u t t e r f l i e s : D y n a m i c s and i n t e r a c t i o n s . Adv. E c o l . Res. 15:51-131. C r o f t , B. A. 1982. Arthropod r e s i s t a n c e t o i n s e c t i c i d e s : a key t o p e s t c o n t r o l f a i l u r e s and s u c c e s s e s i n North American a p p l e o r c h a r d s . Entomol. Exp. A p p l . 31:88-110. C r o n i n , E . H., P. A. Ogden, J . A. Young and W. Laycock. 1978. The e c o l o g i c a l n i c h e o f poisonous p l a n t s i n range communities. J . Range Manage. 31:328-334. C r o t e a u , R. and R. C. Ronald. 1983. T e r p e n o i d s . Chapter 13, J o u r n a l o f Chromatography L i b r a r y , V o l . 22B:148-189 in_ E. Heftman, ed. Chromatography. Fundamentals and A p p l i c a t i o n s o f Chromatographic and E l e c t r o p h o r e t i c Methods. P a r t B : A p p l i c a t i o n s . E l s e v i e r S c i e n t i f i c Pub. Co., N.Y. 564 pp. C u t l e r , J . G., R. F. Severson, P. D. C o l e , D. M. Jackson, and A. W. Johnson. 1986. Secondary m e t a b o l i t e s from h i g h e r p l a n t s . T h e i r p o s s i b l e r o l e as b i o l o g i c a l c o n t r o l agents, pp. 178-196 i n M. B. Green, P. A. Hedin, e d s . N a t u r a l R e s i s t a n c e o f P l a n t s t o P e s t s . Roles of A l l e l o c h e m i c a l s . ACS Symp. S e r . 296, American Chemical S o c , Washington, D C. C z e r w i n s k i , C. and M. B. Isman. 1986. Urban p e s t management d e c i s i o n making and s o c i a l c o n f l i c t i n t h e c o n t r o l o f gypsy moth i n west-coast c i t i e s . B u l l . Entomol. Soc. Am. 32:36-41. Dadd, R. H. 1970. Arthropod n u t r i t i o n , pp. 35-95 i n M. F l o r k i n and B.T. Scheer, e d s . Chemical Zoology. Academic P r e s s , N.Y.  89 Dauterman, W. C. and E. Hodgson. 1978. D e t o x i f i c a t i o n mehanisms i n i n s e c t s , pp.541-577 In M. R o c k s t e i n , ed. B i o c h e m i s t r y of I n s e c t s . Academic P r e s s , N.Y. de Boer, G., V. G. D e t h i e r , and L. M. Schoonhoven. 1977. Chemoreceptors i n the p r e o r a l c a v i t y of the tobacco hornworm. Entomol. Exp. A p p l . 21:287-289. D e l l e Monache, F.,G. B. M a r i n i B e t t o l o , and E. A. Bernays. 1984. I s o l a t i o n of i n s e c t a n t i f e e d a n t a l k a l o i d s from Maytenus r i g i d a ( C e l a s t r a c e a e ) . Z. Angew. Entomol. 97:406-414. de P o n t i , 0. M. B. 1977. R e s i s t a n c e i n Cucumis s a t i v a L. t o T e t r a n y c h u s u r t i c a e Koch. 1. The r o l e o f p l a n t b r e e d i n g i n i n t e g r a t e d c o n t r o l . E u p h y t i c a 26:633-640. D e t h i e r , V. G. 1973. E l e c t r o p h y s i o l o g i c a l s t u d i e s of g u s t a t i o n i n l e p i d o p t e r o u s l a r v a e I I . T a s t e s p e c t r a i n r e l a t i o n to f o o d - p l a n t d i s c r i m i n a t i o n . J . Comp. P h y s i o l . 82:103-134. D e t h i e r , V. G. 1980. E v o l u t i o n of r e c e p t o r s e n s i t i v i t y t o secondary p l a n t s u b s t a n c e s w i t h s e p c i a l r e f e r e n c e to d e t e r r e n t s . Am. Nat. 115:45-66. D e v o n s h i r e , A. L. and G. D. Moore. 1982. A c a r b o x y l e s t e r a s e w i t h broad s u b s t r a t e d s p e c i f i c i t y causes organophosphorous, carbamate and p y r e t h r o i d r e s i s t a n c e i n peach-potato aphids (Myzus p e r s i c a e ) . P e s t i c . Biochem. P h y s i o l . 18:235-246. D o b r i n , G. C. and R. B. Hammond. 1985. The a n t i f e e d i n g a c t i v i t y of s e l e c t e d p y r e t h r o i d s towards the Mexican bean b e e t l e ( C o l e o p t e r a : C o c c i n e l l i d a e ) . J . Kansas Entomol. Soc. 58:422-427. Dover, J . W. 1985. The responses of some L e p i d o p t e r a to l a b i a t e herb white c l o v e r e x t r a c t s . Entomol. Exp. A p p l . 39:177-182. E i s n e r , T.  1964.  C a t n i p : i t s raison d'etre.  and  S c i e n c e 146:1318-1320.  E l l i o t , M. and N. F. J a n e s . 1973. Chemistry of the n a t u r a l p y r e t h r i n s . 55-100 i n J . E. C a s i d a , ed. Pyrethrum the N a t u r a l I n s e c t i c i d e . Academic P r e s s , Inc., N.Y. 329 pp. E l s e y , K. D. 1985. pickleworm and 78:1048-1051.  R e s i s t a n c e mechanisms i n C u r c u r b i t a moschata to melonworm ( L e p i d o p t e r a : P y r a l i d a e ) . J . Econ. Entomol.  E l - S h a z l y , A. M., A. H. Ahmed, A. H. El-Sebae, and E. A. Kadous. T o x i c e x t r a c t s of weeds I I I . F u n g i t o x i c i t y of weed e x t r a c t s . A l e x a n d r i a J . A g r i c . Res. 29:1559-1569.  1981.  F e t c h e r , N. E. D. 1981. E f f e c t s of g r a z i n g on c o l d d e s e r t shrubs:a s i m u l a t i o n model based on r e l a t i v e growth r a t e . E c o l . M o d e l l i n g 86. F i n n e y , D. J . 1971. London.  pp.  P r o b i t A n a l y s i s . T h i r d e d i t i o n . Cambridge Univ.,  13:49-  90 F r a e n k e l , G. S. 1959. The r a i s o n d ' e t r e of secondary S c i e n c e 129:1466-1470.  plant  substances.  Freedman, B., L. J . Nowak, W. F. Kwolek, E. C. B e r r y , and W. D. G u t h r i e . 1979. A bioassay f o r p l a n t d e r i v e d p e s t c o n t r o l agents u s i n g the european c o r n b o r e r . J . Econ. Entomol. 72:541-545. F u n a k i , E. and N. Motoyama. 1986. Cross r e s i s t a n c e t o v a r i o u s i n s e c t i c i d e s of the h o u s e f l i e s s e l e c t e d w i t h a p y r e t h r o i d s . J . P e s t i c . S c i . 11:219— 222. Georghiou, G. P. 1983. Management of r e s i s t a n c e i n a r t h r o p o d s , pp.769-792. i n G.P. Georghiou, T. S a i t o , eds. Pest R e s i s t a n c e t o P e s t i c i d e s . Plenum P r e s s , New York. Gould, F. 1986. S i m u l a t i o n models f o r p r e d i c t i n g d u r a b i l i t y o f i n s e c t r e s i s t a n t germ plasm:Hessian f l y ( D i p t e r a : C e c i o m y i i d a e ) - r e s i s t a n t winter wheat. E n v i r o n . Entomol. 15:11-23. Gould, F., C. R. C a r r o l l , and D. J . Futuyma. 1982. C r o s s - r e s i s t a n c e t o p e s t i c i d e s and p l a n t d e f e n s e s : A study o f the two-spotted s p i d e r - m i t e . Entomol. Exp. A p p l . 31:175-180. Greissman, T. A. and T. S. G r i f f i n . 1971. S e s q u i t e r p e n e l a c t o n e s : A c i d c a t a l y z e d c o l o r r e a c t i o n s as an a i d i n s t r u c t u r e d e t e r m i n a t i o n . Phytochem. 10:2475-2485. Groves, C. R. and J . E. Anderson. 1981. A l l e l o p a t h i c e f f e c t s o f _A. t r i d e n t a t a l e a v e s on g e r m i n a t i o n and growth o f two g r a s s s p e c i e s . Am. Nat. 106:73-79. Gupta,P. D. and A. J . T h o r s t e i n s o n . 1960. Food p l a n t r e l a t i o n s h i p s of the diamondback moth [ P l u t e l l a m a c u l i p e n n i s ( C u r t . ) ] . I I . Sensory r e g u l a t i o n o f o v i p o s i t i o n of t h e a d u l t female. Entomol. Exp. A p p l . 2:305-314. H a n i o t a k i s , G. E. and A. Voyadjoglou. 1978. O v i p o s t i o n r e g u l a t i o n i n Dacus o l e a e by v a r i o u s o l i v e f r u i t c h a r a c t e r s . Entomol. Exp. A p p l . 24:187192. Hansen, R. M. and L. D. R e i d . 1975. D i e t o v e r l a p o f deer, i n southern C o l o r a d o . J . Range Manage. 28:43-47. Harborne, J . B., ed. 1982. I n t r o d u c t i o n t o E c o l o g i c a l Academic P r e s s , London. 278 pp.  e l k , and c a t t l e  Biochemistry.  Hardman, J . A. and P. R. E l l i s . 1978. Host p l a n t f a c t o r s i n f l u e n c i n g the s u s c e p t i b i l i t y o f c r u c i f e r o u s c r o p s t o cabbage r o o t f l y a t t a c k . Entomol. Exp. A p p l . 24:393-397. Harmatha, J . and J . Nawrot. 1984. Comparison o f the f e e d i n g d e t e r r e n t a c t i v i t y of some s e s q u i t e r p e n e l a c t o n e s and a l i g n a n l a c t o n e towards s e l e c t e d i n s e c t s t o r a g e p e s t s . Biochem. System. E c o l . 12:95-98.  91 Haverty, M. I . , and J . L. Robertson. 1982. L a b o r a t o r y b i o a s s a y s f o r s e l e c t i n g c a n d i d a t e i n s e c t i c i d e s and a p p l i c a t i o n r a t e s f o r f i e l d t e s t s on the western spruce budworm. J . Econ. Entomol. _75_: 179-182. Hedin, P. A., ed. 1983. P l a n t R e s i s t a n c e t o I n s e c t s . ACS Amercia Chemical S o c i e t y , Washington, D.C. 375 pp.  Symp. S e r .  208.,  Herout, V. 1970. Chemotaxonomy of the f a m i l y Compositae ( A s t e r a c e a e ) . 93-110 in_ H. Wagner and L. Horhammer, eds. Pharmacognosy and P h y t o c h e m i s t r y . S p r i n g e r - V e r l a g , B e r l i n . 386 pp.  pp.  Heywood, V. H., J . B. Harborne, and B. L. T u r n e r , eds. 1977. The B i o l o g y and Chemistry of the Compositae. V o l . 1. Academic P r e s s , London, pp. 1118. H s i a o , T. H. 1969. Chemical b a s i s of h o s t s e l e c t i o n and p l a n t r e s i s t a n c e i n oligophagous i n s e c t s . Entomol. Exp. A p p l . 12:777-788. H s i a o , T. H. and G. F r a e n k e l . 1968. S e l e c t i o n and s p e c i f i c i t y of the Colorado potato b e e t l e f o r s o l a n a c e o u s and non-solanaceous p l a n t s . Ann. Entomol. Soc. Am. 61:493-503. H s i a o , T. H. and G. F r a e n k e l . 1968a. The r o l e of secondary p l a n t substances from the host p l a n t of the C o l o r a d o potato b e e t l e . Entomol. Soc. Am. 61_: 485-493. H u f f a k e r , C. B., ed. 1980. New Sons, Inc., N.Y. 500 pp.  Ann.  Technology o f Pest C o n t r o l . John Wiley  &  Hwang, Y., K. Wu, J . Kumamoto, H. A x e l r o d , and M. S. M u l l a . 1985. I s o l a t i o n and i d e n t i f i c a t i o n of mosquito r e p e l l e n t s i n A r t e m i s i a v u l g a r i s . J . Chem. E c o l . 11:1297-1306. I g n o f f o , C. M., B. P u t t i e r , N. L. Marston, D. L. H o s t e t t e r , and W. A. D i c k e r s o n . 1975. Seasonal i n c i d e n c e of the entomopathogenic fungus S p i c a r i a r i l e y i a s s o c i a t e d with n o c t u i d p e s t s of soybean. J . I n v e r t e b r . P a t h o l . 25:133-137. Ishikawa, S. 1966. E l e c t r i c a l response and f u n c t i o n o f a b i t t e r substance r e c e p t o r a s s o c i a t e d w i t h the m a x i l l a r y s e n s i l l a of the silkworm, Bombyx mori L. J . C e l l P h y s i o l . 67:1-11. Isman, M. B. and S. S. Duffey. 1982. T o x i c i t y of tomato p h e n o l i c compounds t o the tomato fruitworm, H e l i o t h i s z e a . Entomol. Exp. A p p l . 31:370376. Isman, M. B. and P. P r o k s c h . 1985. D e t e r r e n t and i n s e c t i c i d a l chromenes and benzofurans from E n c e l i a ( A s t e r a c e a e ) . Phytochem. 24:1949-1951. Isman, M. B. and E. R o d r i g u e z . 1983. L a r v a l growth i n h i b i t o r s from of Parthenium ( A s t e r a c e a e ) . Phytochem. 22:2709-2713.  species  Isman, M. B. and E. R o d r i g u e z . 1984. F e e d i n g and growth of n o c t u i d l a r v a e on f o l i a r m a t e r i a l and e x t r a c t s of guayule, r e l a t e d s p e c i e s of Parthenium, and F h y b r i d s . E n v i r o n . Entomol. 13:539-542. 1  92  Jacobson, M. 1958. I n s e c t i c i d e s from p l a n t s , a review of the l i t e r a t u r e , 1941-1953. Agr. Handbook No. 154, U.S. Dept. of A g r i c u l t u r e , Washington, D.C. 299 pp. Jacobson, M. 1983. I n s e c t i c i d e s , i n s e c t r e p e l l e n t s , and a t t r a c t a n t s from a r i d / s e m i a r i d l a n d p l a n t s , pp. 138-146 i n Plants:The P o t e n t i a l s f o r E x t r a c t i n g P r o t e i n , M e d i c i n e s , and Other U s e f u l Chemicals - Workshop P r o c e e d i n g s . U. S. Congress, O f f i c e of Technology Assessment, OTA-BPF-23, Sept. 1983, Washington, D.C. 252 pp. Jacobson, M. 1986. The neem t r e e : n a t u r a l r e s i s t a n c e par e x c e l l e n c e , pp. 220-232 i n M.B. Green and P.A. Hedin, eds. N a t u r a l R e s i s t a n c e of P l a n t s t o P e s t s . ACS Symp. Ser. 296, American Chemical S o c i e t y , Washington, D.C. 1986. Jacobson, M. and D. G. Crosby, ed. M a r c e l Dekker, I n c . , N.Y., 585  1971. pp.  Naturally Occurring  Insecticides.  Jacobson, M., D. K. Reed, M. M. C r y s t a l , D. S. Moreno and E. L. Soderstrom.. 1978. Chemistry and b i o l o g i c a l a c t i v i t y of i n s e c t f e e d i n g d e t e r r e n t s from c e r t a i n weed and crop p l a n t s . Entomol. Exp. A p p l . 24:448-457. J a v i d , A. M. and J . N. A l l . 1984. E f f e c t s of methomyl on weight and development of f a l l armyworm. Ann. Entomol. Soc. Am. 77:193-196. Jermy, T. 1966. F e e d i n g i n h i b i t o r s and food preference phytophagous i n s e c t s . Entomol. Exp. A p p l . 9^:1-12.  i n chewing  Jermy, T. 1983. M u l t i p l i c i t y of i n s e c t a n t i f e e d a n t s i n p l a n t s , pp. 223-234 i n D. L. Whitehead and W. S. Bower, eds. N a t u r a l P r o d u c t s f o r I n n o v a t i v e Pest Management, [ C u r r e n t Themes i n T r o p i c a l S c i e n c e ] . V o l . 2. Pergamon P r e s s , O x f o r d . 583 pp. Jermy, T., E. A. Bernays and A. S z e n t e s i . 1982. The e f f e c t of repeated exposure to d e t e r r e n t s on t h e i r a c c e p t a b i l i t y t o phytophagous i n s e c t s . i n J.H. V i s s e r and A.K. Minks, eds. P r o c . 5th I n t . Symp. I n s e c t - P l a n t R e l a t i o n s h i p , Wageningen, 1982. Jermy, T., B. A. B u t t , L. McDonough, D. L. Dreyer, and A. F. Rose. 1981. A n t i f e e d i n g f o r the C o l o r a d o p o t a t o b e e t l e . I . A n t i f e e d i n g c o n s t i t u e n t s o f some p l a n t s from the sagebrush community. I n s e c t S c i . A p p l . 1:237-242. Jones, C. G., J . R. A l d r i c h , and M. S. Blum. 1981. B a l d c y p r e s s a l l e l o c h e m i c s and the i n h i b i t i o n of silkworm e n t e r i c microorganisms. Some e c o l o g i c a l c o n s i d e r a t i o n s . J . Chem. E c o l . 7^:103-114. Jones, S. B., J r . , W. C. B u r n e t t , J r . , N. C. C o i l e , T. J . Mabry, and M. F. B e t k o u s k i . 1979. S e s q u i t e r p e n e l a c t o n e s of Vernonia - I n f l u e n c e of g l a u c o l i d e - A on the growth r a t e and s u r v i v a l of l e p i d o p t e r o u s l a r v a e . O e c o l o g i a 39:71-77. K e l s e y , R. G., and F. S h a f i z a d e h . 1979. S e s q u i t e r p e n e l a c t o n e s and s y s t e m a t i c s of the genus A r t e m i s i a . Phytochem. 18:1591-1611.  93 K e l s e y , R. G., J . R. Stephens, and F. S h a f i z a d e h . 1982. The chemical c o n s t i t u e n t s of sagebrush f o l i a g e and t h e i r i s o l a t i o n . J . Range Manage. 35:617-622. K e l s e y , R. G., W. E. Wright, F. Sneva, A. Winward, and C. B r i t t o n . 1983. The c o n c e n t r a t i o n and c o m p o s i t i o n of b i g sagebrush e s s e n t i a l o i l s from Oregon. Biochem. System. E c o l . 11:353-360. K i n o s h i t a , G. B. 1985. The economics of e n t o m o l o g i c a l e f f o r t : V i e w p o i n t the p e s t i c i d e i n d u s t r y . Can. Entomol. 117:909-921.  of  Kogan, M. and D. Cope. 1974. F e e d i n g and n u t r i t i o n of i n s e c t s a s s o c i a t e d w i t h soybeans. 3. Food i n t a k e , u t i l i z a t i o n , and growth i n the soybean l o o p e r , P s e u d o p l u s i a i n c l u d e n s . Ann. Entomol. Soc. Am. 67:66-72. K r i e g e r , R. I . , P. P. Feeny, and C. F. W i l k i n s o n . 1971. enzymes i n the guts of c a t e r p i l l a r s : a n e v o l u t i o n a r y defenses? S c i e n c e 172:579-581.  Detoxication answer to p l a n t  Kubo, I . , T. Matsumoto and J . A. K l o c k e . 1984. M u l t i c h e m i c a l r e s i s t a n c e s of the c o n i f e r Podocarpus g r a c i l i o r (Podocarpaceae) t o i n s e c t a t t a c k . J . Chem. E c o l . 10:547-559. Kubo, I . and K. N a k a n i s h i . 1977. I n s e c t a n t i f e e d a n t s and r e p e l l a n t s from A f r i c a n p l a n t s , pp.165-178 in_ P.A. Hedin, ed. Host P l a n t R e s i s t a n c e to P e s t s . ACS Symp.Ser. 62. American Chemical S o c i e t y , Washington, D.C. Kuc,  J . and L. S h a i n . 1977. A n t i f u n g a l compounds a s s o c i a t e d w i t h d i s e a s e r e s i s t a n c e i n p l a n t s , pp.497-535 i n M.R. S i e g e l and H.D. S i s l e r , eds. A n t i f u n g a l Compounds. V o l . 2. M a r c e l Dekker, N.Y. 674 pp.  Kumar, K. and R. B. Chapman. 1984. S u b l e t h a l e f f e c t s o f i n s e c t i c i d e s on the diamondback moth P l u t e l l a x y l o s t e l l a ( L . ) . P e s t i c . S c i . 15:344352. L a r s o n , R. 1985. Dec. 3. Lee,  S t a b l e a n t i - p e s t neem seed e x t r a c t . U.S.  Pat.  4,556,562.  K. H. and 8 o t h e r s . 1977. S e s q u i t e r p e n e antitumor agents: I n h i b i t o r s o f c e l l u l a r metabolism. S c i e n c e 196:533.  Luck, R. F., R. Vander Bosch, and R. G a r c i a . 1977. Chemical i n s e c t c o n t r o l - a t r o u b l e d pest management s t r a t e g y . B i o s c i e n c e 27:606-611. Lundgren, L. 1975. N a t u r a l p l a n t c h e m i c a l s a c t i n g as o v i p o s i t i o n d e t e r r e n t s on cabbage b u t t e r f l i e s ( P i e r i s b r a s i c a ( L . ) , P_. rapae and P. n a p i ( L . ) ) . Z o o l . S c r . 4:253-238. M a l l i s , A., ed. 1982. Handbook of Pest C o n t r o l , p.631. Franzak & Co., C l e v e l a n t , Ohio. 1101 pp.  (L.)  Foster  Mansour, M. H. 1982. The c h r o n i c e f f e c t s o f some a l l e l o c h e m i c s on the l a r v a l development and a d u l t r e p r o d u c t i v i t y of the c o t t o n leafworm, Spodoptera l i t t o r a l i s . Z. P f l a n z e n k r . P f f a n z e n s c h u t z 89:224-229.  94 Matcham, E. J . and C. Hawkes. 1985. F i e l d assessment of the e f f e c t s of d e l t a m e t h r i n on polyphagous p r e d a t o r s i n winter wheat. P e s t i c . S c i . 16:317-320. Mattson, W. J . , J r . 1980. H e r b i v o r y i n r e l a t i o n t o p l a n t n i t r o g e n Ann. Rev. E c o l . S y s t . 11:119-161. Maxwell, F. G. and P. Jennings, e d s . 1980. Breeding I n s e c t s . John Wiley & Sons,Inc.,N.Y. 683 pp.  content.  Plants Resistant to  Mclndoo, N. E . and S i e v e r s , A. F. 1924. P l a n t s t e s t e d f o r o r r e p o r t e d t o possess i n s e c t i c i d a l p r o p e r t i e s . U.S. Dept. o f A g r i c u l t u r e . B u l l . No. 1201, Washington, D.C. 61 pp. M c M i l l i a n , W. W., M. C. Bowman, R. L. Burton, K. J . S t a r k s and B. R. Wiseman. 1969. E x t r a c t of c h i n a b e r r y l e a f as a f e e d i n g d e t e r r e n t and growth r e t a r d a n t f o r l a r v a e of the c o r n earworm and f a l l armyworm. J . Econ. Entomol. 62:708-710. M e t c a l f , R. F. 1980. Changing r o l e of i n s e c t i c i d e s i n crop p r o t e c t i o n . Ann. Rev. Entomol. 25:219-256. M i t c h e l l , B. K. and J . F. S u t c l i f f e . 1984. Sensory i n h i b i t i o n as a mechanism o f f e e d i n g d e t e r r e n c e : e f f e c t s of t h r e e a l k a l o i d s on l e a f b e e t l e f e e d i n g . P h y s i o l . Entomol. j):57-64. M i t c h e l l , E. R. and R. R. Heath. 1985. I n f l u e n c e of Amaranthus h y b r i d u s a l l e l o c h e m i c s on o v i p o s i t i o n behavior of Spodoptera e x i g u a and S. e r i d a n i a . J . Chem. E c o l . 11:609-618.  L.  Munakata, K. 1970. I n s e c t a n t i f e e d a n t s i n p l a n t s , pp.179-187 i n D.L. Wood, R.M. S i l v e r s t e i n and M. Nakajima, eds. C o n t r o l of I n s e c t Behaviour by N a t u r a l P r o d u c t s . Academic P r e s s , N.Y. Murray, R. D. H., J . Medez, and S. A. Brown, eds. 1982. The N a t u r a l Coumarins. W i l e y - I n t e r s c i e n c e , N.Y. 702 pp. Nagy, J . G. and R. P. o i l s of Artemisia the v o l a t i l e o i l s aerobic bacteria.  Tengerdy. 1967. A n t i b a c t e r i a l a c t i o n of e s s e n t i a l as an e c o l o g i c a l f a c t o r I . A n t i b a c t e r i a l a c t i o n o f o f A r t e m i s i a t r i d e n t a t a and A r t e m i s i a nova on A p p l . M i c r o b i o l . 15:819-821.  Nawrot, J . , E. B l o s z y k , H. Grabarczyk, and B. Drozdz. 1982. Feeding d e t e r r e n t a c t i v i t y of the Compositae p l a n t e x t r a c t s f o r t h e s e l e c t e d s t o r a g e p e s t s . Prace Naukowe I n s t y t u t u Ochrony R o s l i n XXIV (1):37-43. Nawrot, J . , E. B l o s z y k , J . Harmatha and L. Novotny. 1984. The e f f e c t of b i s a b o l o a n g e l o n e , h e l e n a l i n and bakkenolide A on development and behavior o f some s t o r e d product b e e t l e s . Z. Angew. Entomol. 98:394398. N i e l s e n J . K. 1978. Host p l a n t d i s c r i m i n a t i o n w i t h i n c r u c i f e r a e : F e e d i n g responses o f four l e a f b e e t l e s ( C o e l p t e r a : C h r y s o m e l i d a e ) t o g l u c o s i n o l a t e s , c u c u r b i t a c i n s and c a r d e n o l i d e s . Entomol. Exp. A p p l . 24:41-54.  95 N i e l s e n , J . K. 1978a. Host p l a n t s e l e c t i o n of monophagous and oligophagous f l e a b e e t l e s f e e d i n g on c r u c i f e r s . Entomol. Exp. A p p l . 24:564-569. O b r y c k i , J . J . and M. J . Tauber. 1984. N a t u r a l enemy a c t i v i t y on g l a n d u l a r pubescent p o t a t o p l a n t s i n the greenhouse:An u n r e l i a b l e p r e d i c t o r of e f f e c t s i n f i e l d . E n v i r o n . Entomol. 13:679-693. P a i n t e r , R. H. 1951. Kansas, 520 pp.  Insect  r e s i s t a n c e i n crop p l a n t s . Univ. P r e s s of  P a t t e r s o n , C. G., D. E. K n o v e l , T. R. Kemp and J . G. Rodriquez. 1975. Chemical b a s i s f o r r e s i s t a n c e t o T e t r a n y c h u s u r t i c a e Koch i n tomatoes. E n v i r o n . Entomol. 4^:670-674. P e r r i n , R. M. and M. L. P h i l l i p s . 1978. Some e f f e c t s o f mixed c r o p p i n g on the p o p u l a t i o n dynamics of i n s e c t p e s t s . Entomol. Exp. A p p l . 24:385393. Pieman, A. K., R. L R a n i e r i , G. H. N. Towers and J . Lam. 1980. V i s u a l i z a t i o n reagents f o r s e s q u i t e r p e n e l a c t o n e s and polyacetylenes on t h i n - l a y e r chromatographs. J . Chromatog. 169:187-198. P r i e s t e r . T . M. and G. P. Georghiou. 1980. C r o s s - r e s i s t a n c e spectrum i n p y r e t h r o i d - r e s i s t a n t Culex q u i n q u e f a c i a t u s . P e s t i c . S c i . 11:617-624. Ramirez, C. 1969. A n t i b a c t e r i a l a c t i o n of n o n - v o l a t i l e substances from A r t e m i s i a t r i d e n t a t a N u t t . s s p . t r i d e n t a t a . Can. J . M i c r o b i o l . 15:1341. Reese, J . C. 1979. I n t e r a c t i o n of a l l e l o c h e m i c a l s w i t h n u t r i e n t c h e m i c a l s i n h e r b i v o r e food, i i i G.A R o s e n t h a l and D.H. Janzen, eds. H e r b i v o r e s : T h e i r I n t e r a c t i o n with Secondary P l a n t M e t a b o l i t e s . Academic P r e s s , New York. Reese, J . C. 1983. N u t r i e n t - a l l e l o c h e m i c a l i n t e r a c t i o n s i n h o s t p l a n t r e s i s t a n c e , pp. 231-243. in_ P.A. Hedin, ed. P l a n t R e s i s t a n c e t o I n s e c t s . American Chemical S o c i e t y , ASC Symp. Ser. 208, Washington, D.C. 375 pp. Reese, J . C. and Beck, S. D. 1976. E f f e c t s of a l l e l o c h e m i c s on the b l a c k cutworm A g r o t i s i p s i l o n : E f f e c t s of p-benzoquinone, hydroquinone and duroquinone on l a r v a l growth, development and u t i l i z a t i o n of f o o d . Ann. Entomol. Soc. Am. 69:59-67. Reese, J . C. and Beck, S. D. 1976a. E f f e c t of a l l e l o c h e m i c a l s on the black cutworm A g r o t i s i p s i l o n : E f f e c t of r e s o r c i n o l , p h l o r o g l u c i n o l and g a l l i c a c i d on l a r v a l growth, development and u t i l i z a t i o n of f o o d . Ann. Entomol. Soc. Am. 69:999-1003. Renwick, J . A. A. and C. D. Radke. 1985. C o n s t i t u e n t s of h o s t - and nonh o s t p l a n t s d e t e r r i n g o v i p o s i t i o n by the cabbage b u t t e r f l y , P i e r i s r a p a e . Entomol. Exp. A p p l . 39:21-26. Rickard, 1977  W. H. and J . L. Warren. 1981. Response of steppe shrubs to drought. Northwest S c i . 55:108-112.  the  96  R i s c h , S. J . 1985. E f f e c t s of induced c h e m i c a l changes on i n t e r p r e t a t i o n of f e e d i n g p r e f e r e n c e t e s t s . Entomol. Exp. A p p l . 39:81-84. Robert, P. C. and P. B l a i s i n g e r . 1978. Role of non-host p l a n t c h e m i c a l s i n the r e p r o d u c t i o n of an o l i g o p h a g o u s i n s e c t : the sugar beet moth S c r o b i p a l p a o c e l l a t e l l a . Entomol. Exp. A p p l . 24:432-436. Rodriguez, E., N. J . Carman, G. Vander V e l d e , J . H. McReynolds and T. J . Mabry. 1972. Methoxylated f l a v o n o i d s from A r t e m i s i a . Phytochem. 11:3509-3514. Roeske, C. N., J . N. S e i b e r , L. P. Brower and C. M. M o f f i t t . 1976. Milkweed c a r d e n o l i d e s and t h e i r comparative p r o c e s s i n g by monarch b u t t e r f l i e s (Danaus p l e x i p p u s L . ) . Rec. Adv. Phytochem. 10:93-167. R o s e n t h a l , G. A., C. G. Hughes and D. H. Janzen. 1982. L-Canavanine, a d i e t a r y n i t r o g e n s o u r c e f o r t h e seed p r e d a t o r Caryedes b r a s i l i e n s i s ( B r u c h i d a e ) . S c i e n c e 217:353-355. R o s e n t h a l , G. A. and D. H. Janzen. e d s . 1979. H e r b i v o r e s : T h e i r I n t e r a c t i o n s with Secondary P l a n t M e t a b o l i t e s . Academic P r e s s , 718 pp.  N.Y.  Samsoe-Petersen, L. 1985. L a b o r a t o r y t e s t s t o i n v e s t i g a t e the e f f e c t s of p e s t i c i d e s on two b e n e f i c i a l arthropods:A p r e d a t o r y mite P h y t o s e i u l u s p e r s i m i l i s and a rove b e e t l e A l e o c h a r a b i l i n e a t a . P e s t i c . S c i . 16:321331. Schmidt, D. J . and J . C. Reese. 1986. Sources of e r r o r i n n u t r i t i o n a l index s t u d i e s of i n s e c t s on a r t i f i c i a l d i e t . J . I n s e c t P h y s i o l . 32:193-198. Schoonhoven, L. M. 1982. A p p l . 31:57-69.  B i o l o g i c a l a s p e c t s of a n t i f e e d a n t s . Entomol.  Exp.  Schroeder, L. A. 1976. E f f e c t of food d e p r i v a t i o n on the e f f i c i e n c y of u t i l i z a t i o n of dry matter, energy and n i t r o g e n by the c h e r r y s c a l l o p moth C a l o c a l p e u n d u l a t a . Ann. Entomol. Soc. Am. 69:55-58. S c r i b e r , J . M. and F. S l a n s k y , J r . 1981. The n u t r i t i o n a l e c o l o g y immature i n s e c t s . Ann. Rev. Entomol. 26:183-211.  of  S c r i v e n , R. and C. E. Meloan. 1984. Determining the a c t i v e component i n 1 , 3 , 3 - t r i m e t h y l 2 - o x a b i c y c l o [2,2,2] octane ( c i n e o l e ) t h a t r e p e l s the American cockroach, P e r i p l a n e t a americana. Ohio J . S c i . 84:85-88. Seaman, D. L. 1982. S e s q u i t e r p e n e l a c t o n e s as taxonomic c h a r a c t e r s i n the A s t e r a c e a e . Bot. Rev. 48:121-595. S e l f , L. S., F. E. G u t h r i e , and E. Hodgson. 1964. Metabolism by t o b a c c o - f e e d i n g i n s e c t s . Nature 204:301-302.  of n i c o t i n e  S h a f i z a d e h , F., N. R. Bhadane, M. S. M o r r i s , F. G. K e l s e y , and S. N. Khanna. 1971. Sesquiterpene l a c t o n e s of b i g sagebrush. Phytochem. 10:2745-2754.  97 S h e l t o n , A. M., J . T. A n d a l o r o , and J . B a r n a r d . 1982. E f f e c t s o f cabbage l o o p e r , imported cabbageworm and diamondback moth on f r e s h market and p r o c e s s i n g cabbage. J . Econ. Entomol. 75:742-745. S h u r r , K. and F. G. Holdaway. 1970. O l f a c t o r y responses o f female O s t r i n i a n u b i l a l i s . Entomol. Exp. A p p l . 13:455-461. Simmonds, M. S. J . , W. M. B l a n e y , F. D e l l e Monache, M. Marquina MacQuahae, and G. B. M a r i n i B e t t o l o . 1985. I n s e c t a n t i f e e d a n t p r o p e r t i e s of a n t h r a n o i d s from t h e genus V i s m i a . J . Chem. E c o l . 11:1593-1599. Sinden, S. L., L. L. S a n f o r d , W. W. C a n t e l o , and K. L. Deahl. 1986. L e p t i n e g l y c o a l k a l o i d s and r e s i s t a n c e t o the Colorado p o t a t o b e e t l e i n Solanum chacoense. E n v i r o n . Entomol. 15:1057-1062. Smith, A. E. and D. M. Secoy. 1975. F o r e r u n n e r s o f p e s t i c i d e s i n c l a s s i c a l Greece and Rome. J . A g r i c . Food Chem. 23:1050-1055. Smith, C. M. 1978. F a c t o r s f o r c o n s i d e r a t i o n i n d e s i g n i n g s h o r t - t e r m i n s e c t host p l a n t b i o a s s a y s . B u l l . Entomol. Soc. Am. 24:393-395. Soo Hoo, C. F. and G. F r a e n k e l . 1966. The consumption, d i g e s t i o n , and u t i l i z a t i o n o f food p l a n t s by a polyphagous i n s e c t , P r o d e n i a e r i d a n i a Cramer. J . I n s e c t P h y s i o l . 12:711-730. Suomi, D., J . J . Brown, and R. D. Akre. 1986. Responses t o p l a n t e x t r a c t s of n e o n a t a l c o d l i n g moth l a r v a e C y d i a pomonella. J . Entomol. Soc. B r i t . Columbia. 83:12-18. Tabashnik, B. E. 1985. D e t e r r e n c e o f diamondback moth o v i p o s i t i o n by p l a n t compounds. E n v i r o n . Entomol. 14:575-578. T h i b o u t , E . and J . Auger. 1983. Substance chimiques s e c o n d a i r e s des A l l i u m determinant l e comportement de ponte de l a t t e i g e du p o i r e a u , A c r o l e p i o p s i s a s s i c t e l l a ( Z e l l . ) (Lepidoptera:Yponomeutidae). B u l l . Soc. Entomol. F r . 88:359-368. T h o r s t e i n s o n , A. J . 1960. Host s e l e c t i o n i n phytophagous i n s e c t s . Ann. Rev. Entomol. _5:193-218. T i n g l e , F. C. and E . R. M i t c h e l l . 1984. Aqueous e x t r a c t s from i n d i g e n o u s p l a n t s as o v i p o s i t i o n d e t e r r e n t s f o r H e l i o t h i s v i r e s c e n s ( F . ) . J . Chem. E c o l . 10:101-113. T r a y n i e r , R. M. M. 1979. Long-term changes i n the o v i p o s i t i o n b e h a v i o r o f the cabbage b u t t e r f l y , P i e r i s rapae , induced by c o n t a c t w i t h p l a n t s . P h y s i o l . Entomol. 4:87-96. T r i a l , H., J r . , and J . B. Dimond. 1979. Emodin i n buckthorn:A f e e d i n g d e t e r e n t t o phytophagous i n s e c t s . Can Entomol. 111:207-212. T u r n e r , N. J . 1979. P l a n t s i n B r i t i s h Columbia I n d i a n Technology. Handbook - B r i t i s h Columbia P r o v i n i c a l Museum No. 38., B. C. Prov. Museum, V i c t o r i a , B. C. 304 pp.  98 Verma, H. N. and M. M. Abid A l i Khan. 198A. A n t i v i r a l agent s t i m u l a t e d by l e a f e x t r a c t s . J . P l a n t D i s . P r o t . 91_: 266-272. V i l l a n i , M. and F. Gould. 1985. S c r e e n i n g o f crude p l a n t e x t r a c t s as f e e d i n g d e t e r r e n t s of the wireworm, Melanotus communis. Entomol. Exp. A p p l . 37:69-75. Waldbauer, G. P. 1968. The consumption and u t i l i z a t i o n of food by i n s e c t s . Adv. I n s e c t P h y s i o l . 2:229-288. Weseloh, R. M., T. G. Andreadis, R. E. B. Moore, J . F. Anderson, N. R. Dubois and F. B. Lewis. 1983. F i e l d c o n f i r m a t i o n o f a mechanism c a u s i n g synergism between B a c i l l u s t h u r i n g i e n s i s and the gypsy moth p a r a s i t o i d Apanteles melanoscelus. J . I n v e r t . P a t h o l . 41:99-103. White, S. M., J . T. F l i n d e r s and B. L. Welch. 1982. P r e f e r e n c e of pygmy r a b b i t s (Brachylagus i d a h o e n s i s ) f o r v a r i o u s p o p u l a t i o n s o f b i g sagebrush (_A. t r i d e n t a t a ) . J . Range Manage. 35:724-726. Whitehead, D. L. and W. S. Bower, eds. 1983. N a t u r a l Products f o r I n n o v a t i v e Pest Management, [ C u r r e n t Themes i n T r o p i c a l S c i e n c e ] . V o l . 2. Pergamon P r e s s , Oxford, pp. 583. W h i t t a k e r , R. H. 1970. The b i o c h e m i c a l ecology of h i g h e r p l a n t s , pp. 43-70 i n E. Sonoheimer and J.B. Simone, eds. Chemical E c o l o g y . Academic P r e s s , N.Y. W i l l i a m s . A. L., E. R. M i t c h e l l , R. R. Heath, and C. S. B a r f i e l d . 1986. O v i p o s i t i o n d e t e r r e n t s f o r f a l l armyworm from l a r v a l f r a s s , c o r n l e a v e s , and a r t i f i c i a l d i e t . E n v i r o n . Entomol. 15:327-330. Wisdom, C. S., J . T. Smiley, and E. Rodriguez. 1983. T o x i c i t y and d e t e r r e n c y of s e s q u i t e r p e n e l a c t o n e s and chromenes to the c o r n earworm. J . Econ Entomol. 76:993-998. Woodhead, S. and E. A. Bernays. 1978. The c h e m i c a l b a s i s o f r e s i s t a n c e of Sorghum b i c o l o r t o a t t a c k by L o c u s t a m i g r a t o r i a . Entomol. Exp, A p p l . 24:123-144. Yang, K. 1983. I n s e c t a n t i f e e d i n g p h e n y l a c t y l e n e s from growing buds of A r t e m i s i a c a p i l l a r i s . J . A g r i c . Food Chem. 31:667-668. Z i t t e r , T. A. 1984. E f f e c t s of c i t i z e n chemophobia on p l a n t P l a n t D i s . 68:655.  pathology.  Z u n i g a , G. E., M. S. Salgado and L. J . C o r c u e r a . 1985. Role of an i n d o l e a l k a l o i d i n t h e r e s i s t a n c e o f b a r l e y s e e d l i n g s t o a p h i d s . Phytochem. 24:945-947.  99  VIII.  APPENDICES  APPENDIX I A n a l y s i s o f V a r i a n c e T a b l e i n c l u d i n g the s e p a r a t i o n o f i n d i v i d u a l degrees of freedom u s i n g o r t h o g o n a l c o n t r a s t s f o r Cabbage Looper E q u i v a l e n t s , f o r the f o u r t r e a t m e n t s , f o u r b l o c k s and f i v e survey days from the f i e l d t r i a l on cabbage, J u l y 25, 1985  SOURCE  BLOCKS TREATMENTS D e l t a m e t h r i n v s . OTHERS* A. t r i d e n t a t a v s . CONS 30% aq EtOH v s . H20 DAYS TREATMENT * DAYS D e l t a m e t h r i n v s . OTHERS * LIN D e l t a m e t h r i n v s . OTHERS * QUA D e l t a m e t h r i n v s . OTHERS * DEV A. t r i d e n t a t a v s . CONS * LIN A. t r i d e n t a t a v s . CONS * "QUA A. t r i d e n t a t a v s . CONS * DEV 30% aq EtOH v s . H20 * LIN 30% aq EtOH v s . H20 * QUA 30% aq EtOH v s . H20 * DEV ERROR TOTAL  PROBABILITY  DF  SUM SQ  F-VALUE  3 3  0.928 27.646  0.897 26.715  0.4487 <0.0001  1 1 1  25.596 1.744 0.306  74.203 5.055 0.886  <0.0001 0.0284 0.3506  4 12  45.650 14.464  33.085 3.494  <0.0001 <0.0007  1 1 1 1 1 1 1 1  8.524 2.819 0.141 0.324 0.213 0.031 0.541 0.353 0.754  24.712 8.173 0.409 0.937 0.619 0.091 1.569 1.024 2.187  <0.0001 0.0059 0.5251 0.3367 0.4348 0.7646 0.2155 0.3159 0.1447  1  57 79  19.662 108.35  OTHERS = t h e t h r e e o t h e r t r e a t m e n t s , aq EtOH, 30% aq EtOH, and H 0  namely, _A. t r i d e n t a t a e x t r a c t i n 30%  2  2 CONS = t h e two c o n t r o l s , t h e c a r r i e r s o l v e n t 30% aq EtOH, and H 0 9  100  APPENDIX I I A n a l y s i s of Variance T a b l e i n c l u d i n g the s e p a r a t i o n o f i n d i v i d u a l degrees of freedom u s i n g o r t h o g o n a l c o n t r a s t s f o r imported cabbageworm, ]?. rapae, l a r v a l counts f o r f o u r t r e a t m e n t s , f o u r b l o c k s and f i v e survey days from the f i e l d t r i a l on cabbage, J u l y 25, 1985  SOURCE  BLOCKS TREATMENTS D e l t a m e t h r i n v s . OTHERS* A. t r i d e n t a t a v s . CONS 30% aq EtOH v s . H20 DAYS TREATMENT * DAYS D e l t a m e t h r i n v s . OTHERS * LIN D e l t a m e t h r i n v s . OTHERS * QUA D e l t a m e t h r i n v s . OTHERS * DEV A. t r i d e n t a t a v s . CONS * LIN A. t r i d e n t a t a v s . CONS * QUA A. t r i d e n t a t a v s . CONS * DEV 30% aq EtOH v s . H20 * LIN 30% aq EtOH v s . H20 * QUA 30% aq EtOH v s . H20 * DEV ERROR TOTAL  PROBABILITY  DF  SUM SQ  F-VALUE  3 3  2.475 19.790  2.986 23.880  0.0386 <0.0001  1 1 1  12.173 6.896 0.721  44.067 24.964 2.609  <0.0001 <0.0001 0.1118  4 12  34.009 10.984  30.779 3.314  <0.0001 <0.0007  1  4.283 2.001 0.031 1.319 0.784 0.012 0.129 0.546 0.593  15.505 7.245 0.111 4.775 2.840 0.045 0.468 1.964 2.148  0.0002 0.0093 0.7403 0.0330 0.0974 0.8327 0.4967 0.1653 0.1482  1 1 1 1 1 1 1 1  57 79  OTHERS = the t h r e e o t h e r t r e a t m e n t s , aq EtOH, 30% aq EtOH, and H„0 CONS = the two c o n t r o l s , t h e c a r r i e r  15.746 83.003  namely, A^. t r i d e n t a t a e x t r a c t i n 30%  s o l v e n t 30% aq EtOH, and H„0  101  APPENDIX I I I A n a l y s i s o f V a r i a n c e T a b l e i n c l u d i n g the s e p a r a t i o n of i n d i v i d u a l degrees of freedom u s i n g o r t h o g o n a l c o n t r a s t s f o r imported cabbageworm, P. rapae, egg counts, f o r the f o u r t r e a t m e n t s , f o u r b l o c k s and f i v e survey days from the f i e l d t r i a l on cabbage, J u l y 25, 1985  SOURCE  BLOCKS TREATMENTS D e l t a m e t h r i n v s . OTHERS* A. t r i d e n t a t a v s . CONS 30% aq EtOH v s . H20 DAYS TREATMENT * DAYS D e l t a m e t h r i n v s . OTHERS * LIN D e l t a m e t h r i n v s . OTHERS * QUA D e l t a m e t h r i n v s . OTHERS * DEV A. t r i d e n t a t a v s . CONS * LIN A. t r i d e n t a t a v s . CONS * QUA A. t r i d e n t a t a v s . CONS * DEV 30% aq EtOH v s . H20 * LIN 30% aq EtOH v s . H20 * QUA 30% aq EtOH v s . H20 * DEV ERROR TOTAL  DF  SUM SQ  F-VALUE  3 3  9.363 31.476  4.270 14.353  0.0094 <0.0001  1 1 1  17.901 12.287 1.288  24.488 16.809 1.763  <0.0001 0.0002 0.1906  4 12  77.694 23.006  26.572 2.6227  <0.0001 0.0089  1 1 1 1 1 1 1 1  0.008 8.195 0.599 0.563 5.515 3.808 0.675 0.149 0.037  1  57 79  0.010 11.211 0.819 0.771 7.544 5.209 0.924 0.204 0.051  PROBABILITY  0.9194 0.0016 0.3700 0.3843 0.0085 0.0269 0.3413 0.6536 0.8230  35.087 191.43  OTHERS = the t h r e e o t h e r t r e a t m e n t s , aq EtOH, 30% aq EtOH, and H 0  namely, _A. t r i d e n t a t a e x t r a c t i n 30%  o  CONS = the two c o n t r o l s , t h e c a r r i e r s o l v e n t 30% aq EtOH, and H 0 9  

Cite

Citation Scheme:

        

Citations by CSL (citeproc-js)

Usage Statistics

Share

Embed

Customize your widget with the following options, then copy and paste the code below into the HTML of your page to embed this item in your website.
                        
                            <div id="ubcOpenCollectionsWidgetDisplay">
                            <script id="ubcOpenCollectionsWidget"
                            src="{[{embed.src}]}"
                            data-item="{[{embed.item}]}"
                            data-collection="{[{embed.collection}]}"
                            data-metadata="{[{embed.showMetadata}]}"
                            data-width="{[{embed.width}]}"
                            async >
                            </script>
                            </div>
                        
                    
IIIF logo Our image viewer uses the IIIF 2.0 standard. To load this item in other compatible viewers, use this url:
http://iiif.library.ubc.ca/presentation/dsp.831.1-0097049/manifest

Comment

Related Items