Metabolic Detoxification of Linear and Angular Furanocoumarins by

Chapter 41

Metabolic Detoxification of Linear and Angular Furanocoumarins by Caterpillars of the Black Swallowtail Butterfly Implications in Host Plant Selection Phenomena

Downloaded by UNIV OF MASSACHUSETTS AMHERST on May 24, 2018 | https://pubs.acs.org Publication Date: January 8, 1987 | doi: 10.1021/bk-1987-0330.ch041

G. Wayne Ivie, Don L. Bull, Ross C. Beier, and Nan W. Pryor Veterinary Toxicology and Entomology Research Laboratory, Agricultural Research Service, U.S. Department of Agriculture, College Station, TX 77841

Midgut and body tissues of caterpillars of the black swallowtail butterfly (Papilio polyxenes Fabr.) possess high enzymatic activity that catalyzes the detoxification of linear furanocoumarins, thus explaining the tolerance of P. polyxenes to these phototoxins. Observations from nature indicate that P. polyxenes caterpillars are less tolerant toward the presence of angular furanocoumarins in potential host plants, and our studies with a commonly occurring angular furanocoumarin suggest that metabolic detoxification of such compounds by P. polyxenes occurs at a relatively slower rate than with the linear analogs. The capacity to detoxify dietary furanocoumarins is a major determinant of host plant acceptability by P. polyxenes; furthermore, this phenomenon represents a clear example of herbivore circumvention of a normally effective host-plant-resistance mechanism. Furanocoumarins are present as secondary constituents in hundreds of plant species from at least eight families, but appear to be most predominant in species of Umbelliferae and Rutaceae (_1»_2)· Furanocoumarins occur as linear or angular analogs, which differ in the angle of furan ring fusion to the coumarin moiety (Figure 1). Well over 100 such compounds have thus far been isolated from plant sources; most of these arise by alkyl or alkoxy substitution at the available aromatic positions, at either of the two olefinic carbons of the furan ring, or, less frequently, at either of the two olefinic carbons of the coumarin lactone ring. Many if not most furanocoumarin derivatives possess potent photoactive properties. It is generally accepted that the biological actions of furanocoumarins are attributable to the fact that these compounds easily intercalate into the double helix of DNA, where they produce cyclobutane adducts with This chapter not subject to U.S. copyright. Published 1987 American Chemical Society

Waller; Allelochemicals: Role in Agriculture and Forestry ACS Symposium Series; American Chemical Society: Washington, DC, 1987.


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ALLELOCHEMICALS: ROLE IN AGRICULTURE AND FORESTRY

p y r i m l d i n e bases upon a c t i v a t i o n by l o n g - w a v e l e n g t h u l t r a v i o l e t light (3-5)· A c t i v e s i t e s a r e t h e f u r a n and coumarin l a c t o n e r i n g double bonds; l i n e a r furanocoumarins produce b o t h monoadducted and c r o s s l i n k e d DNA, but t h e c o n f i g u r a t i o n of t h e a n g u l a r furanocoumarins i s such t h a t o n l y monoadducts can be generated. Superoxide r a d i c a l s and/or s i n g l e t oxygen may a l s o be i n v o l v e d i n some of t h e b i o l o g i c a l a c t i o n s of t h e s e c h e m i c a l s (6-8). Furanocoumarins have a number o f s c i e n t i f i c a l l y i n t e r e s t i n g and even e c o n o m i c a l l y and m e d i c i n a l l y important a c t i o n s . They a r e e f f e c t i v e l y used i n human m e d i c i n e i n the treatment of v i t i l i g o ( s k i n d e p i g m e n t a t i o n , leukoderma) ( 9 ) and p s o r i a s i s ( 1 0 , 1 1 ) , and have shown promise a g a i n s t c e r t a i n o t h e r human maladies (12-15). P l a n t s t h a t c o n t a i n furanocoumarins a r e known t o cause a c u t e p h o t o s e n s i t i z a t i o n ( p h y t o p h o t o d e r m a t i t i s ) i n man (2,16). I t seems l i k e l y t h a t t h e b i o s y n t h e s i s of furanocoumarins by p l a n t s s e r v e s as a defense mechanism a g a i n s t p l a n t pathogens and h e r b i v o r o u s a n i m a l s , and t o improve c o m p e t i t i v e n e s s a g a i n s t o t h e r plant species. Furanocoumarins a r e i n f a c t known t o be a c t i v e as p h o t o t o x i n s a g a i n s t l i v e s t o c k (17) and as p h y t o a l e x i n s ( 1 6 , 1 8 ) . Furanocoumarins a r e a l s o seed g e r m i n a t i o n (19) and p l a n t growth (20) i n h i b i t o r s , a l t h o u g h l i g h t a c t i v a t i o n i s presumably not r e q u i r e d f o r t h e s e i n t e r a c t i o n s because such a c t i v i t i e s a r e expressed i n the s o i l environment. Most i n s e c t h e r b i v o r e s appear t o be r a t h e r e f f e c t i v e l y r e p e l l e d by f u r a n o c o u m a r i n - c o n t a i n i n g p l a n t s ( 2 1 - 2 4 ) . A notable e x c e p t i o n t o t h i s g e n e r a l i z a t i o n o c c u r s among some b u t t e r f l i e s of t h e f a m i l y P a p i l i o n i d a e , whose c a t e r p i l l a r s a r e adapted t o feed s u c c e s s f u l l y and i n f a c t p r e f e r e n t i a l l y on p l a n t s t h a t c o n t a i n l i n e a r , but not a n g u l a r , furanocoumarins ( 2 2 ) . These c i r c u m s t a n c e s prompted us t o undertake s t u d i e s w i t h t h e b l a c k s w a l l o w t a i l b u t t e r f l y ( P a p i l i o p o l y x e n e s ) and r a d i o l a b e l e d furanocoumarins i n attempts t o e l u c i d a t e t h e n a t u r e of t h e insect/furanocoumarin i n t e r a c t i o n s involved. Metabolic Basis f o r P a p i l i o polyxenes Furanocoumarins

Resistance to Linear

Our i n i t i a l s t u d i e s ( 2 5 , 2 6 ) d e t e r m i n e d the comparative f a t e of a r a d i o c a r b o n - l a b e l e d p r e p a r a t i o n of t h e commonly o c c u r r i n g l i n e a r furanocoumarin, xanthotoxin (8-methoxypsoralen) i n black s w a l l o w t a i l c a t e r p i l l a r s and i n f a l l armyworm ( S p o d o p t e r a f r u g i p e r d a J . E . Smith) l a r v a e . Black swallowtail c a t e r p i l l a r s are known not t o be a d v e r s e l y a f f e c t e d by l i n e a r furanocoumarins ( 2 2 ) , w h i l e Spodoptera s p p . a v o i d such p l a n t s as food s o u r c e s (21) . E q u i v a l e n t doses of [ C ] x a n t h o t o x i n (5 pg/g) a d m i n i s t e r e d o r a l l y t o l a s t - s t a g e l a r v a e of e i t h e r s p e c i e s was f o l l o w e d by d r a m a t i c a l l y d i f f e r e n t d i s p o s i t i o n p a t t e r n s . E l i m i n a t i o n of r a d i o c a r b o n i n t h e e x c r e t a was much more r a p i d i n P. p o l y x e n e s ( T a b l e I ) , and [ C] r e s i d u e s i n body t i s s u e s of S. f r u g i p e r d a accumulated t o much h i g h e r l e v e l s and p e r s i s t e d much l o n g e r t h a n i n P. polyxenes ( T a b l e I I ) . S t u d i e s of t h e n a t u r e o f the r a d i o c a r b o n r e s i d u e s i n body t i s s u e s showed t h a t l e v e l s of u n m e t a b o l i z e d x a n t h o t o x i n i n body t i s s u e s of


41.

IVIEETAL. Table

Metabolic

Detoxification

of

457

Furanocoumarins

I· R a d i o c a r b o n E x c r e t i o n A f t e r O r a l Treatment of Stage Larvae o f JP. p o l y x e n e s and S. f r u g i p e r d a w i t h [ C ] X a n t h o t o x i n at 5 ug/g

Last-

% of Administered Radiocarbon i n E x c r e t a (X + S . D . )


Hours A f t e r Treatment

P.

1.5 3 6 12 24 L

50.3 62.4 77.6 77.6 100.9

Data adapted from I v i e et (1984) ( 2 6 ) .

S.

polyxenes

al.

+ + + + +

—

0.9 18.3 41.6 59.7 87.8

6.8 4.2 0.8 6.0 5.6

(1983)

frugiperda

(25)

+ + + + +

—

and B u l l et

1.1 2.7 16.5 8.0 6.7

al.

Table I I . R a d i o c a r b o n R e s i d u e s i n Body T i s s u e s ( E x c l u s i v e and C o n t e n t s ) A f t e r O r a l Treatment o f L a s t - S t a g e Larvae P. p o l y x e n e s and S. f r u g i p e r d a w i t h l C]Xanthotoxin at 5 y g / g lU

of Gut of

a

% of A d m i n i s t e r e d R a d i o c a r b o n i n Body T i s s u e s (X ± S . D . ) Hours

After

Treatment 1.5 3 6 12 24 a

D a t a adapted from I v i e (1984) ( 2 6 ) .

P. p o l y x e n e s 3.6 3.3 0.2 0.1 0.2 et a l .

+ 1.0 + 1.8 + 0.1 + 0.1 + 0.2 (1983) (25)

S.

frugiperda

54.6 36.7 15.0 4.3 2.4 and B u l l et

+ 4.1 + 4.7 + 4.6 + 2.2 + 0.9 al.

ί>. f r u g i p e r d a were at e v e r y s a m p l i n g i n t e r v a l >50 times as h i g h as i n body t i s s u e s o f P. p o l y x e n e s . These data are h i g h l y s u p p o r t i v e of the h y p o t h e s i s t h a t r a p i d m e t a b o l i c d e t o x i f i c a t i o n of l i n e a r f u r a n o c o u m a r i n s , c o u p l e d w i t h r a p i d e x c r e t i o n of t h e m e t a b o l i t e s , a c c o u n t f o r the i n s e n s i t i v i t y of such i n s e c t s to the a d v e r s e e f f e c t s of t h e s e potent p h o t o t o x i n s ( 2 5 , 2 6 ) . Subsequent s t u d i e s by us (27) have shown t h a t h i g h t i t e r s of mixed f u n c t i o n o x i d a s e enzymes i n midgut and body t i s s u e s o f _P. p o l y x e n e s account f o r the r a p i d d e t o x i f i c a t i o n phenomena o b s e r v e d , which r e s u l t p r i m a r i l y i n c l e a v a g e of the f u r a n r i n g ( F i g u r e 2 ) .


458

ALLELOCHEMICALS: ROLE IN AGRICULTURE AND

FORESTRY

Comparative Fate of Linear and Angular Furanocoumarins i n P. polyxenes


Because the resistance of P. polyxenes to the toxic effects of linear furanocoumarins apparently does not extend to the angular furanocoumarins (22), we have undertaken comparative metabolic fate studies with a representative of each of these furanocoumarin classes. T r i t i a t e d psoralen or isopsoralen (Figure 1) was administered as before to last stage .P. polyxenes c a t e r p i l l a r s , and the d i s t r i b u t i o n , elimination, and biochemical fate of the compounds determined (28).

Table I I I . Excretion of Radioactivity After Oral Treatment of Last-Stage C a t e r p i l l a r s of P. polyxenes with Either [ H]Psoralen or [ H]Isopsoralen at 5 yg/g a

% of AdministeredJRadioactivity (X j S.D.) Hours After Treatment 0.75 1.5 3 6 12 a

Psoralen-treated 28.5 44.8 71.1 83.2 93.0

+ + + + +

5.7 8.8 4.3 6.8 0.9

i n Excreta

Isopsoralen-treated 19.4 40.9 67.6 77.7 90.8

+ 2.9 + 10.7 + 8.9 + 5.9 + 1.5

Data adapted from Ivie et a l . (28).

The disposition patterns of psoralen and isopsoralen i n IP. polyxenes under the parameters studied were not dramatically d i f f e r e n t . As indicated i n Table I I I , there were no appreciable differences i n the rate of excretion of r a d i o a c t i v i t y by c a t e r p i l l a r s treated with the two compounds. In body tissues, however, levels of t o t a l r a d i o a c t i v i t y in isopsoralen-treated c a t e r p i l l a r s were consistently about twice those observed i n psoralen-treated insects (Table IV). Further, levels of unmetabolized parent compounds retained i n body tissues (where toxic effects would be expressed) were on the order of 3 times as high i n c a t e r p i l l a r s treated with the angular furanocoumarin, isopsoralen (Table V). Analysis of excreta samples indicated that both psoralen and isopsoralen are metabolized extensively by P. polyxenes c a t e r p i l l a r s , primarily by the same furan ring cleavage reactions observed i n our e a r l i e r studies with xanthotoxin (Figure 3). Discussion Our studies with _P. polyxenes c a t e r p i l l a r s and radiolabeled furanocoumarins have provided what appears to be a d e f i n i t i v e explanation of the apparent t o t a l i n s e n s i t i v i t y of these insects to the toxic effects of linear furanocoumarins present i n t h e i r



41.

IVIE ET AL.

Metabolic

Detoxification

PSORALEN

of

459

Furanocoumarins

ISOPSORALEN

Figure 1. Structures of a linear furanocoumarin and an angular furanocoumarin (isopsoralen).

(psoralen)

Figure 2. Major metabolites of xanthotoxin (8-methoxypsoralen) i n last-stage larvae of the black swallowtail butterfly (Papilio polyxenes) and the f a l l armyworm (Spodoptera frugiperda).

Figure 3. Major metabolites of psoralen and isopsoralen i n last-stage c a t e r p i l l a r s of the black swallowtail b u t t e r f l y (Papilio polyxenes).


460


Table IV. Tritium Residues i n Body Tissues (Exclusive of Gut and Contents) After Oral Treatment of Last-Stage C a t e r p i l l a r s of P. polyxenes with Either [ H]Psoralen or [ H]Isopsoralen at 5 yg/g a

% of Administered Radioactivity i n Body Tissues (X ± S.D.)


Hours After Treatment 0.75 1.5 3 6 12 a


+ + + + +_


1.5 1.2 1.2 0.4 0.2

+ + + + +

3.0 5.1 1.7 1.4 0.6


Table V. Unmetabolized Psoralen or Isopsoralen i n Body Tissues (Exclusive of Gut and Contents) of Last Stage ^. polyxenes C a t e r p i l l a r s T r e a t e d Orally with Either^[ H]Psoralen or [ H]Isopsoralen at 5 yg/g a

% of Administered Radioactivity as Unmetabolized Parent Compound i n Body Tissues Hours After Treatment 0.75 1.5 3 6 12 a


+ + + + +

1.0 0.4 0.6 0.1 0.1


+ + + + +

2.0 4.7 2.0 0.3 0.1


normal host plants. P. polyxenes has evolved highly active d e t o x i f i c a t i o n mechanisms against linear furanocoumarins such that levels of the intact photosensitizers do not accumulate i n body tissues (where detrimental light-induced reactions would be expected to occur). On the other hand, at least one furanocoumarin-susceptible insect (Î5. frugiperda) metabolizes these compounds at a much lower rate, with the result that under constant dietary exposure, toxic levels of the intact phototoxins would presumably accumulate and be retained i n the general body circulation. Differences i n metabolic d e t o x i f i c a t i o n rates between l i n e a r and angular furanocoumarins observed i n our studies were not great. However, the fact that the angular furanocoumarin under study accumulated to appreciably higher levels i n body tissues of P. polyxenes c a t e r p i l l a r s than did the linear analog i s



41.

IVIE ET AL.

Metabolic

Detoxification

of

Furanocoumanns

461

supportive of the hypothesis that a reduced d e t o x i f i c a t i o n rate accounts, at least i n part, for the s u s c e p t i b i l i t y of P. polyxenes c a t e r p i l l a r s to the detrimental effects of angular furanocoumarins. This conclusion d i f f e r s from our e a r l i e r interpretation of data obtained from limited studies on the interaction of unlabeled psoralen and isopsoralen with P^. polyxenes (26). Plant furanocoumarins occur widely i n nature and provide formidable obstacles to grazing by herbivorous animals. Some insect species have nevertheless adapted to circumvent this powerful host-plant-resistance mechanism. It has been proposed that the l e a f - r o l l i n g habit of some insect species may be an evolutionary adaptation to avoid light and thus avoid the toxic effects of furanocoumarins (21). Also, evidence has recently been obtained that the capacity of at least one leaf-mining insect species to detoxify furanocoumarins allows the u t i l i z a t i o n of furanocoumarin-containing plants as hosts (29). Our studies indicate that rapid metabolic d e t o x i f i c a t i o n of linear furanocoumarins i s an effective resistance mechanism for P. polyxenes against the toxic e f f e c t s of these compounds. It has been postulated that the adaptation of some plants to produce angular furanocoumarins was i n response to the reduced effectiveness of the linear furanocoumarins as deterrents f o r herbivores such as P^. polyxenes (22). Such may indeed be true, but our studies on the comparative d e t o x i f i c a t i o n of linear and angular furanocoumarins suggest that, at best, the presence of angular furanocoumarins i n plants confers only a tenuous margin of relative "safety" against P. polyxenes.

Literature Cited 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13.

Berenbaum, M. Evolution 1983, 37, 163. Pathak, Μ. Α.; Daniels, F., Jr.; Fitzpatrick, T. B. J. Invest. Dermatol. 1962, 39, 225. Scott, B. R.; Pathak, Μ. Α.; Mohn, G. R. Mutat. Res. 1976 39, 29. Hearst, J. E. Ann. Rev. Biophys. Bioeng. 1981, 10, 69. Hearst, J. E. Stud. Biophys. 1983, 94, 25. Joshi, P. C.; Pathak, M. A. Biochem. Biophys. Res. Commun. 1983, 112, 638. Tamaro, M.; Babudri, N.; Pani, B.; Baccichetti, F.; Rodighiero, P. Med. Biol. Environ. 1983, 11, 493. Vedaldi, D.; Dall'Acqua, F.; Rodighiero, G. Med. Biol. Environ. 1983, 11, 507. Nordlund, J. J.; Lerner, A. B. Arch. Dermatol. 1982, 118, 5. Reshad, H.; Challoner, F.; Pollock, D. J.; Baker, H. Brit. J. Dermatol. 1984, 110, 299. Stern, R. S.; Laird, N.; Melski, J.; Parrish, J. Α.; Fitzpatrick, T. B.; Bleich, H. L. N. Engl. J. Med. 1984, 310, 1156. James, M. P. Clin. Exp. Dermatol. 1982, 7, 311. Adams, R.; Boyle, J.; Lever, R.; McQuillan, I.; Mackie, R. Scott. Med. J. 1982, 27, 264.



462


14. Wantzin, G. L.; Thomsen, K. Brit. J. Dermatol. 1982, 107, 687. 15. Edelson, R.; Berger, C.; Gasparro, F.; Lee, K.; Taylor J. Clin. Res. 1983, 31, 467A. 16. Austad, J.; Kalvi, G. Contact Dermatitis 1983, 9, 448. 17. Ivie, G. W. In "Effects of Poisonous Plants on Livestock"; Keeler, R.; Van Kampen, K.; James, L., Eds.; Academic: New York, 1978; p. 475. 18. Johnson, C.; Brannon, D. R.; Kuc, J. Phytochemistry 1973, 12, 2961. 19. Friedman, J.; Rushkin, E.; Waller, G. R. J. Chem. Ecol. 1982, 8, 55. 20. Shimomura, H.; Sashida, Y.; Nakata, H.; Kawasaki, J.; Ito, Y. Phytochemistry 1973, 21, 2213. 21. Berenbaum, M. Science 1978, 201, 532. 22. Berenbaum, M.; Feeny, P. Science 1981, 212, 927. 23. Muckensturm, B.; Duplay, D.; Robert, P. C.; Simonis, M. T.; Kienlen, J-C. Biochem. Syst. Ecol. 1981, 9, 289. 24. Gebreyesus, T.; Chapya, A. Curr. Themes Trop. Sci. 1983, 2, 237. 25. Ivie, G. W.; Bull, D. L.; Beier, R. C., Pryor, N. W.; Oertli, Ε. H. Science 1983, 221, 374. 26. Bull, D. L.; Ivie, G. W.; Beier, R. C.; Pryor, N. W.; Oertli, Ε. H. J. Chem. Ecol. 1984, 10, 893. 27. Bull, D. L.; Ivie, G. W.; Beier, R. C.; Pryor, N. W. J. Chem. Ecol. (in press). 28. Ivie, G. W.; Bull, D. L.; Beier, R. C.; Pryor, N. W. J. Chem. Ecol. (in press). 29. Ashwood-Smith, M. J.; Ring, R. Α.; Liu, M.; Phillips, S.; Wilson, M. Can. J. Zool. 1984, 62, 1971. RECEIVED December 23, 1985


Metabolic Detoxification of Linear and Angular Furanocoumarins by

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