STRUCTURE-BIODEGRADABILITY RELATIONSHIPS IN P Y R E T H R O I D INSECTICIDES 1 JOHN E. CASIDA, KENZO UEDA2, LORETTAC. GAUGHAN, LIEN T. JAOa, and DAVID M. SODERLUND Division of Entomology and Parasitology, University of California, Berkeley, California 94720
The metabolism of 20 pyrethroids has been examined to evaluate the contribution of detoxification in their selective action between insects and mammals. The studies utilized living houseflies, mice, or rats, or esterase and oxidase systems derived from these organisms. Pyrethroid-hydrolyzing esterases cleave the primary alcohol transsubstituted-cyclopropanecarboxylates much faster than the corresponding cis-isomers but are ineffective in hydrolyzing secondary alcohol esters. Microsomal enzymes oxidize the (+)-trans-chrysanthemate moiety at the trans-methyl group of the isobutenyl substituent and at one of the gem-dimethyl groups whereas the (+)-cisisomer is attacked at either of the isobutenyl methyl groups. Products isomerized at C3 of the cyclopropane are also detected but only after ester cleavage and oxidation of an isobutenyl methyl group. Each alcohol moiety has its own unique sites for oxidation involving pentadienyl, allyl, benzylic methylene, and aromatic substituents. An enhancement of insecticidal activity is expected on replacement of the biodegradable groupings with substituents relatively resistant to metabolism but this may also increase the mammalian toxicity. Knowledge of the pathways a n d rates of pyrethroid insecticide metabolism is useful in designing potent compounds and in understanding the basis for selective toxicity and synergist action. This discussion o f the metabolism o f pyrethroids includes the natural constituents, recently reviewed (Casida 1973), and synthetic analogs, particularly chrysanthemates of primary alcohols (Figure 1). The available information indicates that the initial metabolic attack detoxifies the compound, a finding consistent with the very specific structures needed for toxicity. Pyrethroids are metabolized by both hydrolytic and oxidative processes. The relative rates o f these reactions are important in selective toxicity phenomena and in determining the metabolites persisting in or excreted by mammals.
Metabolic studies with living mammals and insects Pyrethroid biodegradability is often evaluated by comparing the susceptibility of normal organisms with those treated with esterase inhibitors (e.g. S , S , S - t r i b u t y l tPresented at Third International Congress of Pesticide Chemistry (IUPAC), Helsinki, Finland, July 3-9, 1974. ZCurrent address: Pesticide Division, Institute for Biological Science, Sumitomo Chemical Co., Ltd., 4-2-I Takatsukasa, Takarazuka-shi, Hyogo-ken, Japan. ZCurrent address: Pharmacology and Toxicology Branch, National Institute of Environmental Health Sciences, Research Triangle Park, North Carolina 27709 Archives of Environmental Contamination and Toxicology Vol. 3, 491-500 (1975/76) 9 1976 by Springer-Verlag New York Inc.
491
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phosphorotrithioate) or oxidase inhibitors (e.g. piperonyl butoxide or sesamex). The results show the relative importance for selective toxicity of esteratic and oxidative metabolism of pyrethroids in insects (Elliot 1971, Jao and Casida 1974a, Miyamoto and Suzuki 1973, Yamamoto 1973) and mammals (Abernathy and Casida 1973, Abernathy et. al. 1973, Jao and Casida 1974a) and that small differences in structure often produce large changes in biodegradability. Radiotracer studies in living mammals and insects with standard techniques for metabolite isolation and characterization are necessary to support the deductions from bioassays with and without synergists. The urinary and fecal metabolites isolated from rats dosed orally are identified and residues in tissues measured. In houseflies treated topically or by injection, metabolic rates and pathways are correlated with synergist treatment or strain dependent differences in resistance to poisoning. Although the complete metabolic pathway has not been defined for any pyrethroid, the critical initial reactions and more than half of .the terminal in vivo metabolites have been identified in several cases. Each pyrethroid studied is attacked at several sites to give some relatively polar metabolites including conjugates that are often included among the "unidentified" fractions.
Metabolic studies with esterase and oxidase systems Esterases and oxidases acting on pyrethroid substrates are generally more active with mammalian than with insect enzyme preparations. However, studies with both mammals and insects are important in defining the basis for selective toxicity. Assay of the esterases does not require addition of a cofactor. The number and individual properties of the esterases are unknown since they have not been purified. Pyrethroids which are detoxified primarily by esterase action are often increased in their persistence by insecticides and related compounds that phosphorylate and carbamoylate these esterases (Abernathy and Casida 1973, Abernathy et al. 1973, Jao and Casida 1974a and b, Miyamoto and Suzuki 1973, Suzuki and Miyamoto 1974). Pyrethroid oxidations are assayed with microsomal preparations from mouse or rat liver or houseflies incubated with reduced nicotinamide-adenine dinucleotide phosphate (NADPH). Some of these oxidations are inhibited by piperonyl butoxide and other pyrethroid synergists (Jao and Casida 1974a, Miyamoto and Suzuki 1973, Yamamoto et al. 1969). O-Propyl O-(2,propynyl) phenylphosphonate and related compounds probably inhibit both the esterases and oxidases (Jao and Casida 1974a, Miyamoto and Suzuki 1973, Suzuki and Miyamoto 1974, Yamamoto 1973). Esteratic and oxidative metabolism are easily differentiated by in vitro procedures using suitable enzyme preparations or by additions to the incubation mixtures (Abernathy and Casida 1973, Abernathy et al. 1973, Jao and Casida 1974b,
Structure-Biodegradability Relationships in Pyrethroids
493
Soderlund and Casida 1974, Suzuki and Miyamoto 1974, Yamamoto et al. 1969). Two methods have been used to assay esterases only: either with acetone powders because oxidases are destroyed as they are prepared; or with microsomes or other fresh tissue preparations without adding the oxidase cofactor (NADPH). T o confirm that only esterases are being assayed in these systems, an appropriate inhibitor (paraoxon or tepp) is added to phosphorylate the esteratic sites and block the metabolism. The nature of the metabolic products also shows whether oxidation has been suppressed. Investigation of oxidase reactions requires suitable fresh microsomal preparations fortified with NADPH. However, the fresh microsomal preparations also possess esteratic activity. Accordingly, esterase inhibitors such as tepp are added to suppress any possible esterase action so only the oxidase reactions are measured. Finally, fresh microsomes and NADPH give a measure of overall biodegradability, i.e. the summation of esterase and oxidase attack.
Structure-biodegradability relationships The sites of metabolic attack on a variety of pyrethroids in living organisms and enzyme systems determined by the methods described above are given in Figure 1. These reactions are discussed on a generalized basis to show the emerging structure-biodegradability relationships. Several metabolic modifications are established for the cyclopropanecarboxylate moiety of pyrethroids. (+)-trans- and ( + ) - c i s - C h r y s a n t h e m a t e s are most readily oxidized at the trans (E)-methyl group of the isobutenyl substituent, giving in succession the corresponding alcohol, aldehyde, and carboxylic acid (Ueda et al. 1974a, Yamamoto and Casida 1966, Yamamoto et al. 1969); the c i s ( Z ) - m e t h y l group is oxidized in the ( + ) - c i s - c h r y s a n t h e m a t e but this site of oxidation is not detected in the ( + ) - t r a n s - c h r y s a n t h e m a t e (Ueda et al. 1974a). Although the isobutenyl group of chrysanthemates is epoxidized photochemically (Ueda et al. 1974b), there is little or no metabolic epoxidation at this site (Ueda et al. 1974a). With both (+)-trans- and ( + ) - c i s - r e s m e t h r i n , isomerization at Ca occurs during formation of metabolites by ester cleavage and oxidation (Ueda et al. 1974a). The pyrethrate methoxycarbonyl group is readily hydrolyzed (Casida et al. 1971, Elliott et al. 1972). Although the cyclopropane ring is intact in all identified metabolites, the small amount of ~4carbon dioxide liberated from carboxylate-labeled cyclopropane acids indicates that the ring may be cleaved to a small extent (Elliott et al. 1972, Ueda et al. 1974a). One methyl of the gem-dimethyl group of S-bioallethrin is oxidized to the corresponding alcohol, after oxidation of the E-methyl in the isobutenyl group (Elliott et al. 1972). Oxidation of the acid moiety at the C3 substituent is greatly suppressed in compounds with the dichlorovinyl in place of the isobutenyl group (Elliott et al. 1974)." The diverse alcoholic components of pyrethroids undergo a variety of oxidation reactions before or after ester hydrolysis. Pentadienyl and allyl groups in the pyrethrins and allethrin, respectively, are readily oxidized in mammals (Casida et al. 1971, Elliott et al. 1972) and probably in insects (Yamamoto 1973). Phenyl
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Rethronyl Esters rho
re O
H~'~,~0 '0~1~/r
rho
H ~ ' , gO O ~ / r
pyrethrin I
H"'~" r ~C)('u ' 0~ ' ~
pyrethrin II
S-bioallethrin
5-Benzyl-3-fu rylmethyl Esters
ro
re ro-.~ 9
~
ro-=f
(+)-trans-resmethrin
H,
o
HIL
/-~
~
-O
~-~ ~.-ro
e
\ro
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NRDC 108
bioresmethrin e
9
s~_-%~.,~x--m. 9 (o ~;.,A,~,~
J
(+)-trans-ethanomethrin
(+)-cis-ethanomet h rin
RU-15525
Fig. I. Pyrethroids subjected to metabolism studies and confirmed sites of metabolic attack. The metabolic systems are rats or rabbits (r), houseflies (h), microsome-NADPH oxidase systems of mouse or rat liver or houseflies (o), and esterases of mouse or rat liver microsomes or insects (e). The designated isomer was used with the following exceptions: (+--)-trans,cis-chrysanthemate with barthrin, dimethrin, and proparthrin for studies in living mammals; (+.)-trans-chrysanthemate with tetramethrin for studies in living mammals and a portion of the studies in houseflies; (__.)-trans-chrysanthemate with resmethrin in one study in living mammals. Two of the pyrethroids, permethrin (Elliott et al. 1973) and RU-15525 (Lhoste and Rauch, 1974), are not referred to in other literature cited. References to the metabolism studies on individual compounds are as follows: barthrin (Masri et al. 1964) S-bioallethrin (Abernathy and Casida 1973, Abernathy et al. 1973, Casida 1973, Casida et a/. 1971, Elliott et al. 1972, Jao and Casida 1974b, Soderlund and Casida 1974, Yamamoto 1973, Yamamoto and Casida 1966, Yamamoto et al. 1969) (+)-trans-dimethrin (Elliott et al. 1972, Masri et al. 1964, Yamamoto and Casida 1966, Yamamoto et al. 1969) (+)-trans- and (+)-cis-ethanomethrin (Abernathy and Casida 1973, Abernathy et al. 1973,
Soderlund and Casida 1974) (+)-trans- and (+)-cis-furamethrin (Abernathy and Casida 1973, Abernathy et al.
NRDC 108 (Abernathy and Casida 1973, Abernathy et al. 1973) (+)-trans- and (+)-cis-permethrin (Elliott et al. 1974, Soderlund and Casida 1974)
1973)
495
Structure-Biodegradability Relationships in Pyrethroids BenzyI Esters re
e
r
ho.~
/r
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el--
(+)-trans-di meth ri n
(+)-cis-phenothrin
(+)-trans-phenothrin r
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~"
)=_,.
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(+)~is-permeth rin
(+)-trans-permeth ri n
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./
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u
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(+)-trans-phenothrin (Abemathy and Casida 1973, Abernathy et al.
1973, Miyamoto et al. 1974, Soderlund and Casida 1974) (+)-cis-phenothrin (Abemathy and Casida 1973, Abemathy et al. 1973, Soderlund and Casida 1974) proparthrin (Nakanishi et al. 1971) pyrethrin I (Casida 1973, Casida et al. 1971, Elliott et al. 1972, Yamamoto 1973, Yamamoto and Casida 1966, Yamamoto et al. 1969) pyrethrin II (Casida 1973, Casida et al. 1971, Elliott et al. 1972, Yamamoto 1973) (+)-trans-resmethrin or bioresrnethrin (Abemathy and Casida 1973, Abernathy et al. 1973, Jao and Casida 1974a and b, Miyamoto et al. 1971, Soderlund and Casida 1974, Ueda et al. 1974a)
(+)-cis-resmethrin (Abernathy and Casida 1973, Abemathy et al. 1973, Jao and Casida 1974a and b, Soderlund and Casida 1974, Ueda et al. 1974a) RU-15525 (Soderlund and Casida 1974) (+)-trans-tetramethrin (Abemathy and Casida 1973, Abernathy et al. 1973, Elliott et al. 1972, Jao and Casida 1974a and b, Miyamoto and Suzuki 1973, Miyamoto et al. 1968, Suzuki and Miyamoto 1974, Yamamoto 1973, Yamamoto and Casida 1966, Yamamoto et al. 1969) ( + ) - c i s - t e t r a m e t h r i n (Abernathy and Casida 1973, Abernathy et al. 1973, Jao and Casida
1974a and b).
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J.E. Casida et al.
substituents of resmethrin, phenothrin, and permethrin undergo ring hydroxylation (Elliott et al. 1974, Miyamoto et al. 1971, Miyamoto et al. 1974, Ueda et al. 1974a) and the tetrahydrophthalimido group of tetramethrin appears to yield a 3-hydroxycyclohexane- 1,2-dicarboximide derivative (Miyamoto et al. 1968). The benzylic methylene group in resmethrin isomers is oxidized to give alcoholic and ketonic derivatives (Miyamoto et al. 1971, Ueda et al. 1974a). There is some evidence, but no conclusive proof, that ester cleavage occurs not only by esterase action but also by oxidative processes (Soderlund and Casida t974, Ueda et al. 1974a). The furylmethanol and benzyl alcohol fragments released are either conjugated as glucuronides or oxidized to the corresponding carboxylic acids, which in turn undergo conjugation with glucuronic acid or glycine (Elliott et al. 1974, Masri et al. 1964, Miyamoto et al. 1971, Miyamoto et al. 1974, Nakanishi et al. 1971, Ueda et aI. 1974a). The glucuronides of 3-hydroxycyclohexane-1,2- dicarboximide and 5-(4'-hydroxybenzyl) -3-furoic acid and the sulfate of the latter compound are reported as urinary metabolites of tetramethrin and resmethrin, respectively (Miyamoto et al. 1968, Miyamoto et al. 1971). Enzyme studies on the rate of pyrethroid metabolism in esterase, oxidase, and esterase plus oxidase systems have contributed greatly to the understanding of the influence of molecular structure on biodegradability. It appears that, in general, esterases are most important in metabolizing the trans-chrysanthemates of primary alcohols whereas oxidases are more important with the cis-chrysanthemates of primary alcohols and with rethronyl chrysanthemates (Abernathy and Casida 1973, Abernathy et al. 1973, Casida 1973, Casida et al. 1971, Elliott et al. 1972, Elliott et al. 1974, Jao and Casida 1974b, Miyamoto et al. 1974, Soderlund and Casida 1974, Suzuki and Miyamoto 1974, Ueda et al. 1974a). These conclusions are based on studies with fresh mouse liver microsomes (Soderlund and Casida 1974, Table I), supported by investigations with acetone powders of mouse liver microsomes (Abernathy and Casida 1973, Abernathy et al. 1973) and rat liver enzyme preparations (Miyamoto et al. 1974). The esterases of liver microsomes display a remarkable degree of substrate specificity, readily hydrolyzing (+)-trans-chrysanthemates of primary alcohols but not the corresponding cis-chrysanthemates or the (+)-trans-chrysanthemates of secondary alcohols. This esterase specificity has important toxicological implications since the more readily hydrolyzed (+)-trans-chrysanthemates are less toxic to mammals than the (+)-cis-chrysanthemates of primary alcohols (Abernathy and Casida 1973, Abernathy et al. 1973, Elliott 1971, Verschoyle and Barnes 1972). The rate difference between the trans- and cis-chrysanthemates is 2.6- > 50-times with acetone powders of mouse microsomes (Abernathy and Casida 1973, Abernathy et al. 1973) and with fresh microsbme preparations (Soderlund and Casida 1974, Table I). The optical configuation at cyclopropane C~ or the cyclopentylidenemethyl or dichlorovinyl replacements for the isobutenyl group are also influential on the hydrolysis rate (Abernathy and Casida 1973, Abernathy et al. 1973, Miyamoto et al. 1974, Soderlund and Casida 1974). The rate of cleavage depends also on the nature of the primary alcohol moiety, (+)-trans-furamethrin
Structure-Biodegradability Relationships in Pyrethroids
497
undergoing the most rapid hydrolysis (Abernathy and Casida 1973, Abernathy et al. 1973). Insect pyrethroid-hydrolyzing esterases differ from the mouse liver esterases in several respects: the insect esterases appear to be less active; the specificity for hydrolyzing trans- versus cis-chrysanthemates varies among five insect species; there are differences in the sensitivity to esterase inhibitors (Jao and Casida 1974b). These species differences in the esterase properties are important in designing synergists (Jao and Casida 1974a and b). Oxidation by the microsome-NADPH system is important in metabolism of the rethronyl chrysanthemates (S-bioallethrin) and of the cis-substituted cyclopropaneTable I. Metabolism rates for various pyrethroids incubated with mouse liver microsomes (Soderlund and Casida, 1974). Metabolism rate relative to (+)-trans-resmethrina Compound
Esterase only
Oxidase
Esterase
only
+ oxidase
44
43
(+)-Allethronyl ester S-Bioallethrin (+)-trans
< 2
5-Benzyl-3-furylmethyl esters Resmethrin (+)-trans (-)-trans (+)-cis (-)-cis
79 47 < 3 < 4
20 20 29 26
100 69 29 26
Ethanomethrin (+)-trans (+)-cis
19 < 3
12 21
33 19
RU-15525 (+)-cis
< 3
51
52
3-Phenoxybenzyl esters Phenothrin (+)-trans (+)-cis
59 < 4
27 37
78 37
Permethrin (+)-trans (+)-cis
78 < 2
30 26
112 29
aAmount of substrate, 100 nmoles; protein content of microsomes, 1.3 + 0.2 mg; incubation medium, 2.5 ml 0.05 M tn's-HCl, pH 7.5; esterase only assays involve no other components; oxidase only assays involve addition of 2.2/~moles NADPH and 300 nmoles tepp; esterase plus oxidase assays involve addition of 2.2 /~moles O NADPH; incubations of 0 to 30 min at 37 C; the esterase plus oxxdase rate for (+)-trans-resmethrin is 9.8 + 1.2 nmoles/min/mg protein (pseudo first order rate, calculated on the basis of 50% metabolism time).
498
J.E. Casida et al.
carboxylates of primary alcohols (resmethrin, ethanomethrin, RU-15525, phenothrin, and permethrin) (Table I). Although the cis-pyrethroids are generally oxidized as rapidly as or more rapidly than the trans-pyrethroids, the overall rate of metabolism is not as great as for the trans-isomers which are more readily hydrolyzed. The (+)- and (-)-isomers of c/s-resmethrin are metabolized at similar rates whereas the (+)-trans-isomer is metabolized more rapidly than the ( - ) - t r a n s isomer (Soderlund and Casida 1974). The oxidative and overall metabolism rates are reduced on replacing the isobutenyl group with the cyclopentylidene-methyl group (Abernathy et al. 1973, Soderlund and Casida 1974) but not with the dichlorovinyl group (Soderlund and Casida 1974) whereas the incorporation of a thiolactone moiety as in RU-15525 increases the metabolism rate (Soderlund and Casida 1974). Bioresmethrin is hydrolyzed by esterases of mouse brain (Jao and Casida 1974b) and S-bioallethrin is oxidized by lung microsomes (Casida 1973) but much more information is needed on both the hydrolytic and oxidative capacities of various tissues in pyrethroid metabolism.
Other considerations on pyrethroid metabolism Synergists to retard pyrethroid metabolism in insects must be selected with care to maintain the favorable safety factor for mammals (Casida 1970). Thus, high injected doses of the synergist piperonyl butoxide or of certain other esterase and oxidase inhibitors markedly increase the toxicity to mice of (+)-cis-resmethrin, ( + )-trans- and ( + )-cis-ethanomethrin, and ( + )-cis-furamethrin (Abernathy and Casida 1973, Abernathy et al. 1973, Jao and Casida 1974a). Although this phenomenon does not appear to pose any human hazard with the commercial pyrethroid-synergist combinations, it may become a problem if more effective synergists or pyrethroids more toxic to mammals enter use. Ideal pyrethroids should not need synergists. Several aspects of pyrethroid biodegradation warrant further study: metabolic fate in man; identity and quantity of metabolites in milk and meat from exposed animals; the metabolism after inhalation and dermal exposure; toxicology and metabolic fate of photoproducts, particularly because several of these differ from pyrethroid metabolites, e.g., epoxides, cyclic peroxides, and rearranged derivatives; fate in different types of plants and soils to lay the background for agricultural pest control. Thus, the information available on pyrethroid metabolism is a small portion of that needed for a group of insecticides likely to be used in increasing amounts in the future.
Acknowledgment Study supported in part by grants from: The National Institutes of Health (2 PO1 ES00049); The Rockefeller Foundation; S. B. Penick and Co., Orange, NJ; Agricultural Chemical Div., FMC Corp., Middleport, NY; S. C. Johnson and Son, Inc.,
Structure-Biodegradability Relationships in Pyrethroids
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Racine, Wis.; Cooper, McDouga11, and Robertson, Ltd., Berkhamsted, Herts, England; National Research Development Corp., London, England; Roussel-UclafProcida, Paris, France.
References Abernathy, C. O., and J. E. Casida: Pyrethroid insecticides: esterase cleavage in relation to selective toxicity. Science 179, 1235 (1973). Abernathy, C. O., K. Ueda, J. L. Engel, L. C. Gaughan, and J. E. Casida: Substrate-specificity and toxicological significance of pyrethroid-hydrolyzing esterases of mouse liver microsomes. Pesticide Biochem. Physiol. 3, 300 (1973). Casida, J. E.: Mixed-function oxidase involvement in the biochemistry of insecticide synergists. J. Agr. Food Chem. 18, 753 (1970). Casida, J. E.: Biochemistry of pyrethrins. In J. E. Casida (ed.): Pyrethrum the Natural Insecticide, pp. 101-120. New York: Academic Press (1973). Casida, J. E., E. C. Kimmel, M. Elliott, and N. F. Janes: Oxidative metabolism of pyrethrins in mammals. Nature 230, 326 (1971). Elliott, M.: The relationship between the structure and the activity of pyrethroids. Bull . W.H.O. 44, 315 (1971). Elliott, M., A. W. Farnham, N. F. Janes, P. H. Needham, D. A. Pulman, and J. H. Stevenson: A photostable pyrethroid. Nature 246, 169 (1973). Elliott, M., N. F. Janes, E. C. Kimmel, and J. E. Casida: Metabolic fate of pyrethrin I, pyrethrin II, and allethrin administered orally to rats. J. Agr. Food Chem. 20, 300 (1972). Elliott, M., K. Ueda, L. C. Gaughan, and J. E. Casida: Unpublished results (1974). Jao, L. T., and J. E. Casida: Esterase inhibitors as synergists for (+)-trans-chrysanthemate insecticide chemicals. Pesticide Biochem. Physiol. 4,456 (1974a). Jao, L. T., and J. E. Casida: Insect pyrethroid-hydrolyzing esterases. Pesticide Biochem. Physiol. 4, 465 (1974b). Lhoste, J. and F. Rauch: A new pyrethroid with a very strong knock-down effect. Paper presented at Third International Congress of Pesticide Chemistry (IUPAC), Helsinki, Finland, July 3-9, 1974. Masri, M. S., F. T. Jones, R. E. Lundin, G. F. Bailey, and F. DeEds: Metabolic fate of two chrysanthemumic acid esters: barthrin and dimethrin. Toxicol. Appl. Pharmacol. 6, 711 (1964). Miyamoto, J., and T. Suzuki: Metabolism of tetramethrin in houseflies in vivo. Pesticide Biochem. Physiol. 3, 30 (1973). Miyamoto, J., T. Nishida, and K. Ueda: Metabolic fate of resmethrin, 5-benzyl-3furylmethyl dl-trans-chrysanthemate in the rat. Pesticide Biochem. Physiol. 1, 293 (1971).
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Miyamoto, J., Y. Sato, K. Yamamoto, M. Endo, and S. Suzuki: Biochemical studies on the mode of action of pyrethroidal insecticides. Part I. Metabolic fate of phthalthrin in mammals. Agr. Biol. Chem. 32, 628 (1968). Miyamoto, J., T. Suzuki, and C. Nakae: Metabolism of phenothrin or 3-phenoxybenzyl d-trans-chrysanthemumate in mammals. Pesticide Biochem. Physiol. 4, 438 (1974). Nakanishi, M., Y. Kato, T. Furuta, and S. Miura: Metabolic fate of proparthrin. Studies on insecticide. VIII. Botyu-Kagaku 36, 116 (1971). Soderlund, D. M., and J. E. Casida: Unpublished results (1974). Suzuki, T., and J. Miyamoto: Metabolism of tetramethrin in houseflies and rats in vitro. Pesticide Biochem. Physiol. 4, 86 (1974). Ueda, K., L. C. Gaughan, and J. E. Casida: Unpublished results (1974a). Ueda, K., L. C. Gaughan, and J. E. Casida: Photodecomposition of resmethrin and related pyrethroids. J. Agr. Food Chem. 22, 212 (1974b). Verschoyle, R. D., and J. M. Barnes: Toxicity of natural and synthetic pyrethrins to rats. Pesticide Biochem. Physiol. 2, 308 (1972). Yamamoto, I.: Mode of action of synergists in enhancing the insecticidal activity of pyrethrum and pyrethroids. In J. E. Casida (ed.): Pyrethrum the Natural Insecticide, pp. 195-210. New York: Academic Press (1973). Yamamoto, I., and J. E. Casida: O-Demethyl pyrethrin II analogs from oxidation of pyrethrin I, allethrin, dimethrin, and phthalthrin by a house fly enzyme system. J. Econ. Entomol. 59, 1542 (1966). Yamamoto, I., E. C. Kimmel, and J. E. Casida: Oxidative metabolism of pyrethroids in houseflies. J. Agr. Food Chem. 17, 1227 (1969).
Manuscript received April 7, 1975; accepted April 15, 1975
ERRATUM Archives of Environmental Contamination and Toxicology, Vol. 3, No. 3 (1975): Page 358, Table I, column 5 (14 days after spraying, low airflow-rate sprayer) at tree site no. 4, reads 0.1264--should be 0.264.
The metabolism of 20 pyrethroids has been examined to evaluate the contribution of detoxification in their selective action between insects and mammals. The studies utilized living houseflies, mice, or rats, or esterase and oxidase systems derived from these organisms. Pyrethroid-hydrolyzing esterases cleave the primary alcohol transsubstituted-cyclopropanecarboxylates much faster than the corresponding cis-isomers but are ineffective in hydrolyzing secondary alcohol esters. Microsomal enzymes oxidize the (+)-trans-chrysanthemate moiety at the trans-methyl group of the isobutenyl substituent and at one of the gem-dimethyl groups whereas the (+)-cisisomer is attacked at either of the isobutenyl methyl groups. Products isomerized at C3 of the cyclopropane are also detected but only after ester cleavage and oxidation of an isobutenyl methyl group. Each alcohol moiety has its own unique sites for oxidation involving pentadienyl, allyl, benzylic methylene, and aromatic substituents. An enhancement of insecticidal activity is expected on replacement of the biodegradable groupings with substituents relatively resistant to metabolism but this may also increase the mammalian toxicity. Knowledge of the pathways a n d rates of pyrethroid insecticide metabolism is useful in designing potent compounds and in understanding the basis for selective toxicity and synergist action. This discussion o f the metabolism o f pyrethroids includes the natural constituents, recently reviewed (Casida 1973), and synthetic analogs, particularly chrysanthemates of primary alcohols (Figure 1). The available information indicates that the initial metabolic attack detoxifies the compound, a finding consistent with the very specific structures needed for toxicity. Pyrethroids are metabolized by both hydrolytic and oxidative processes. The relative rates o f these reactions are important in selective toxicity phenomena and in determining the metabolites persisting in or excreted by mammals.
Metabolic studies with living mammals and insects Pyrethroid biodegradability is often evaluated by comparing the susceptibility of normal organisms with those treated with esterase inhibitors (e.g. S , S , S - t r i b u t y l tPresented at Third International Congress of Pesticide Chemistry (IUPAC), Helsinki, Finland, July 3-9, 1974. ZCurrent address: Pesticide Division, Institute for Biological Science, Sumitomo Chemical Co., Ltd., 4-2-I Takatsukasa, Takarazuka-shi, Hyogo-ken, Japan. ZCurrent address: Pharmacology and Toxicology Branch, National Institute of Environmental Health Sciences, Research Triangle Park, North Carolina 27709 Archives of Environmental Contamination and Toxicology Vol. 3, 491-500 (1975/76) 9 1976 by Springer-Verlag New York Inc.
491
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J.E. Casida et al.
phosphorotrithioate) or oxidase inhibitors (e.g. piperonyl butoxide or sesamex). The results show the relative importance for selective toxicity of esteratic and oxidative metabolism of pyrethroids in insects (Elliot 1971, Jao and Casida 1974a, Miyamoto and Suzuki 1973, Yamamoto 1973) and mammals (Abernathy and Casida 1973, Abernathy et. al. 1973, Jao and Casida 1974a) and that small differences in structure often produce large changes in biodegradability. Radiotracer studies in living mammals and insects with standard techniques for metabolite isolation and characterization are necessary to support the deductions from bioassays with and without synergists. The urinary and fecal metabolites isolated from rats dosed orally are identified and residues in tissues measured. In houseflies treated topically or by injection, metabolic rates and pathways are correlated with synergist treatment or strain dependent differences in resistance to poisoning. Although the complete metabolic pathway has not been defined for any pyrethroid, the critical initial reactions and more than half of .the terminal in vivo metabolites have been identified in several cases. Each pyrethroid studied is attacked at several sites to give some relatively polar metabolites including conjugates that are often included among the "unidentified" fractions.
Metabolic studies with esterase and oxidase systems Esterases and oxidases acting on pyrethroid substrates are generally more active with mammalian than with insect enzyme preparations. However, studies with both mammals and insects are important in defining the basis for selective toxicity. Assay of the esterases does not require addition of a cofactor. The number and individual properties of the esterases are unknown since they have not been purified. Pyrethroids which are detoxified primarily by esterase action are often increased in their persistence by insecticides and related compounds that phosphorylate and carbamoylate these esterases (Abernathy and Casida 1973, Abernathy et al. 1973, Jao and Casida 1974a and b, Miyamoto and Suzuki 1973, Suzuki and Miyamoto 1974). Pyrethroid oxidations are assayed with microsomal preparations from mouse or rat liver or houseflies incubated with reduced nicotinamide-adenine dinucleotide phosphate (NADPH). Some of these oxidations are inhibited by piperonyl butoxide and other pyrethroid synergists (Jao and Casida 1974a, Miyamoto and Suzuki 1973, Yamamoto et al. 1969). O-Propyl O-(2,propynyl) phenylphosphonate and related compounds probably inhibit both the esterases and oxidases (Jao and Casida 1974a, Miyamoto and Suzuki 1973, Suzuki and Miyamoto 1974, Yamamoto 1973). Esteratic and oxidative metabolism are easily differentiated by in vitro procedures using suitable enzyme preparations or by additions to the incubation mixtures (Abernathy and Casida 1973, Abernathy et al. 1973, Jao and Casida 1974b,
Structure-Biodegradability Relationships in Pyrethroids
493
Soderlund and Casida 1974, Suzuki and Miyamoto 1974, Yamamoto et al. 1969). Two methods have been used to assay esterases only: either with acetone powders because oxidases are destroyed as they are prepared; or with microsomes or other fresh tissue preparations without adding the oxidase cofactor (NADPH). T o confirm that only esterases are being assayed in these systems, an appropriate inhibitor (paraoxon or tepp) is added to phosphorylate the esteratic sites and block the metabolism. The nature of the metabolic products also shows whether oxidation has been suppressed. Investigation of oxidase reactions requires suitable fresh microsomal preparations fortified with NADPH. However, the fresh microsomal preparations also possess esteratic activity. Accordingly, esterase inhibitors such as tepp are added to suppress any possible esterase action so only the oxidase reactions are measured. Finally, fresh microsomes and NADPH give a measure of overall biodegradability, i.e. the summation of esterase and oxidase attack.
Structure-biodegradability relationships The sites of metabolic attack on a variety of pyrethroids in living organisms and enzyme systems determined by the methods described above are given in Figure 1. These reactions are discussed on a generalized basis to show the emerging structure-biodegradability relationships. Several metabolic modifications are established for the cyclopropanecarboxylate moiety of pyrethroids. (+)-trans- and ( + ) - c i s - C h r y s a n t h e m a t e s are most readily oxidized at the trans (E)-methyl group of the isobutenyl substituent, giving in succession the corresponding alcohol, aldehyde, and carboxylic acid (Ueda et al. 1974a, Yamamoto and Casida 1966, Yamamoto et al. 1969); the c i s ( Z ) - m e t h y l group is oxidized in the ( + ) - c i s - c h r y s a n t h e m a t e but this site of oxidation is not detected in the ( + ) - t r a n s - c h r y s a n t h e m a t e (Ueda et al. 1974a). Although the isobutenyl group of chrysanthemates is epoxidized photochemically (Ueda et al. 1974b), there is little or no metabolic epoxidation at this site (Ueda et al. 1974a). With both (+)-trans- and ( + ) - c i s - r e s m e t h r i n , isomerization at Ca occurs during formation of metabolites by ester cleavage and oxidation (Ueda et al. 1974a). The pyrethrate methoxycarbonyl group is readily hydrolyzed (Casida et al. 1971, Elliott et al. 1972). Although the cyclopropane ring is intact in all identified metabolites, the small amount of ~4carbon dioxide liberated from carboxylate-labeled cyclopropane acids indicates that the ring may be cleaved to a small extent (Elliott et al. 1972, Ueda et al. 1974a). One methyl of the gem-dimethyl group of S-bioallethrin is oxidized to the corresponding alcohol, after oxidation of the E-methyl in the isobutenyl group (Elliott et al. 1972). Oxidation of the acid moiety at the C3 substituent is greatly suppressed in compounds with the dichlorovinyl in place of the isobutenyl group (Elliott et al. 1974)." The diverse alcoholic components of pyrethroids undergo a variety of oxidation reactions before or after ester hydrolysis. Pentadienyl and allyl groups in the pyrethrins and allethrin, respectively, are readily oxidized in mammals (Casida et al. 1971, Elliott et al. 1972) and probably in insects (Yamamoto 1973). Phenyl
494
J. E. Casida et al.
Rethronyl Esters rho
re O
H~'~,~0 '0~1~/r
rho
H ~ ' , gO O ~ / r
pyrethrin I
H"'~" r ~C)('u ' 0~ ' ~
pyrethrin II
S-bioallethrin
5-Benzyl-3-fu rylmethyl Esters
ro
re ro-.~ 9
~
ro-=f
(+)-trans-resmethrin
H,
o
HIL
/-~
~
-O
~-~ ~.-ro
e
\ro
(+)-ci$-resmeth rin
NRDC 108
bioresmethrin e
9
s~_-%~.,~x--m. 9 (o ~;.,A,~,~
J
(+)-trans-ethanomethrin
(+)-cis-ethanomet h rin
RU-15525
Fig. I. Pyrethroids subjected to metabolism studies and confirmed sites of metabolic attack. The metabolic systems are rats or rabbits (r), houseflies (h), microsome-NADPH oxidase systems of mouse or rat liver or houseflies (o), and esterases of mouse or rat liver microsomes or insects (e). The designated isomer was used with the following exceptions: (+--)-trans,cis-chrysanthemate with barthrin, dimethrin, and proparthrin for studies in living mammals; (+.)-trans-chrysanthemate with tetramethrin for studies in living mammals and a portion of the studies in houseflies; (__.)-trans-chrysanthemate with resmethrin in one study in living mammals. Two of the pyrethroids, permethrin (Elliott et al. 1973) and RU-15525 (Lhoste and Rauch, 1974), are not referred to in other literature cited. References to the metabolism studies on individual compounds are as follows: barthrin (Masri et al. 1964) S-bioallethrin (Abernathy and Casida 1973, Abernathy et al. 1973, Casida 1973, Casida et a/. 1971, Elliott et al. 1972, Jao and Casida 1974b, Soderlund and Casida 1974, Yamamoto 1973, Yamamoto and Casida 1966, Yamamoto et al. 1969) (+)-trans-dimethrin (Elliott et al. 1972, Masri et al. 1964, Yamamoto and Casida 1966, Yamamoto et al. 1969) (+)-trans- and (+)-cis-ethanomethrin (Abernathy and Casida 1973, Abernathy et al. 1973,
Soderlund and Casida 1974) (+)-trans- and (+)-cis-furamethrin (Abernathy and Casida 1973, Abernathy et al.
NRDC 108 (Abernathy and Casida 1973, Abernathy et al. 1973) (+)-trans- and (+)-cis-permethrin (Elliott et al. 1974, Soderlund and Casida 1974)
1973)
495
Structure-Biodegradability Relationships in Pyrethroids BenzyI Esters re
e
r
ho.~
/r
"H", ~ o 0 ~ 0 "~ re
el--
(+)-trans-di meth ri n
(+)-cis-phenothrin
(+)-trans-phenothrin r
v/
~"
)=_,.
r
ctf-/.. barthrin
(+)~is-permeth rin
(+)-trans-permeth ri n
Other Esters ho
rhe /~",
./
-HJo~ N
(+)-trans-tet ra meth ri n
(+) ~is-tet rameth ri n e
H
u
(+ )-tran$-fu rameth d n
H
r
proparthrin
H
"-"
(+)~is-furamethrin
(+)-trans-phenothrin (Abemathy and Casida 1973, Abernathy et al.
1973, Miyamoto et al. 1974, Soderlund and Casida 1974) (+)-cis-phenothrin (Abemathy and Casida 1973, Abemathy et al. 1973, Soderlund and Casida 1974) proparthrin (Nakanishi et al. 1971) pyrethrin I (Casida 1973, Casida et al. 1971, Elliott et al. 1972, Yamamoto 1973, Yamamoto and Casida 1966, Yamamoto et al. 1969) pyrethrin II (Casida 1973, Casida et al. 1971, Elliott et al. 1972, Yamamoto 1973) (+)-trans-resmethrin or bioresrnethrin (Abemathy and Casida 1973, Abernathy et al. 1973, Jao and Casida 1974a and b, Miyamoto et al. 1971, Soderlund and Casida 1974, Ueda et al. 1974a)
(+)-cis-resmethrin (Abernathy and Casida 1973, Abemathy et al. 1973, Jao and Casida 1974a and b, Soderlund and Casida 1974, Ueda et al. 1974a) RU-15525 (Soderlund and Casida 1974) (+)-trans-tetramethrin (Abemathy and Casida 1973, Abernathy et al. 1973, Elliott et al. 1972, Jao and Casida 1974a and b, Miyamoto and Suzuki 1973, Miyamoto et al. 1968, Suzuki and Miyamoto 1974, Yamamoto 1973, Yamamoto and Casida 1966, Yamamoto et al. 1969) ( + ) - c i s - t e t r a m e t h r i n (Abernathy and Casida 1973, Abernathy et al. 1973, Jao and Casida
1974a and b).
496
J.E. Casida et al.
substituents of resmethrin, phenothrin, and permethrin undergo ring hydroxylation (Elliott et al. 1974, Miyamoto et al. 1971, Miyamoto et al. 1974, Ueda et al. 1974a) and the tetrahydrophthalimido group of tetramethrin appears to yield a 3-hydroxycyclohexane- 1,2-dicarboximide derivative (Miyamoto et al. 1968). The benzylic methylene group in resmethrin isomers is oxidized to give alcoholic and ketonic derivatives (Miyamoto et al. 1971, Ueda et al. 1974a). There is some evidence, but no conclusive proof, that ester cleavage occurs not only by esterase action but also by oxidative processes (Soderlund and Casida t974, Ueda et al. 1974a). The furylmethanol and benzyl alcohol fragments released are either conjugated as glucuronides or oxidized to the corresponding carboxylic acids, which in turn undergo conjugation with glucuronic acid or glycine (Elliott et al. 1974, Masri et al. 1964, Miyamoto et al. 1971, Miyamoto et al. 1974, Nakanishi et al. 1971, Ueda et aI. 1974a). The glucuronides of 3-hydroxycyclohexane-1,2- dicarboximide and 5-(4'-hydroxybenzyl) -3-furoic acid and the sulfate of the latter compound are reported as urinary metabolites of tetramethrin and resmethrin, respectively (Miyamoto et al. 1968, Miyamoto et al. 1971). Enzyme studies on the rate of pyrethroid metabolism in esterase, oxidase, and esterase plus oxidase systems have contributed greatly to the understanding of the influence of molecular structure on biodegradability. It appears that, in general, esterases are most important in metabolizing the trans-chrysanthemates of primary alcohols whereas oxidases are more important with the cis-chrysanthemates of primary alcohols and with rethronyl chrysanthemates (Abernathy and Casida 1973, Abernathy et al. 1973, Casida 1973, Casida et al. 1971, Elliott et al. 1972, Elliott et al. 1974, Jao and Casida 1974b, Miyamoto et al. 1974, Soderlund and Casida 1974, Suzuki and Miyamoto 1974, Ueda et al. 1974a). These conclusions are based on studies with fresh mouse liver microsomes (Soderlund and Casida 1974, Table I), supported by investigations with acetone powders of mouse liver microsomes (Abernathy and Casida 1973, Abernathy et al. 1973) and rat liver enzyme preparations (Miyamoto et al. 1974). The esterases of liver microsomes display a remarkable degree of substrate specificity, readily hydrolyzing (+)-trans-chrysanthemates of primary alcohols but not the corresponding cis-chrysanthemates or the (+)-trans-chrysanthemates of secondary alcohols. This esterase specificity has important toxicological implications since the more readily hydrolyzed (+)-trans-chrysanthemates are less toxic to mammals than the (+)-cis-chrysanthemates of primary alcohols (Abernathy and Casida 1973, Abernathy et al. 1973, Elliott 1971, Verschoyle and Barnes 1972). The rate difference between the trans- and cis-chrysanthemates is 2.6- > 50-times with acetone powders of mouse microsomes (Abernathy and Casida 1973, Abernathy et al. 1973) and with fresh microsbme preparations (Soderlund and Casida 1974, Table I). The optical configuation at cyclopropane C~ or the cyclopentylidenemethyl or dichlorovinyl replacements for the isobutenyl group are also influential on the hydrolysis rate (Abernathy and Casida 1973, Abernathy et al. 1973, Miyamoto et al. 1974, Soderlund and Casida 1974). The rate of cleavage depends also on the nature of the primary alcohol moiety, (+)-trans-furamethrin
Structure-Biodegradability Relationships in Pyrethroids
497
undergoing the most rapid hydrolysis (Abernathy and Casida 1973, Abernathy et al. 1973). Insect pyrethroid-hydrolyzing esterases differ from the mouse liver esterases in several respects: the insect esterases appear to be less active; the specificity for hydrolyzing trans- versus cis-chrysanthemates varies among five insect species; there are differences in the sensitivity to esterase inhibitors (Jao and Casida 1974b). These species differences in the esterase properties are important in designing synergists (Jao and Casida 1974a and b). Oxidation by the microsome-NADPH system is important in metabolism of the rethronyl chrysanthemates (S-bioallethrin) and of the cis-substituted cyclopropaneTable I. Metabolism rates for various pyrethroids incubated with mouse liver microsomes (Soderlund and Casida, 1974). Metabolism rate relative to (+)-trans-resmethrina Compound
Esterase only
Oxidase
Esterase
only
+ oxidase
44
43
(+)-Allethronyl ester S-Bioallethrin (+)-trans
< 2
5-Benzyl-3-furylmethyl esters Resmethrin (+)-trans (-)-trans (+)-cis (-)-cis
79 47 < 3 < 4
20 20 29 26
100 69 29 26
Ethanomethrin (+)-trans (+)-cis
19 < 3
12 21
33 19
RU-15525 (+)-cis
< 3
51
52
3-Phenoxybenzyl esters Phenothrin (+)-trans (+)-cis
59 < 4
27 37
78 37
Permethrin (+)-trans (+)-cis
78 < 2
30 26
112 29
aAmount of substrate, 100 nmoles; protein content of microsomes, 1.3 + 0.2 mg; incubation medium, 2.5 ml 0.05 M tn's-HCl, pH 7.5; esterase only assays involve no other components; oxidase only assays involve addition of 2.2/~moles NADPH and 300 nmoles tepp; esterase plus oxidase assays involve addition of 2.2 /~moles O NADPH; incubations of 0 to 30 min at 37 C; the esterase plus oxxdase rate for (+)-trans-resmethrin is 9.8 + 1.2 nmoles/min/mg protein (pseudo first order rate, calculated on the basis of 50% metabolism time).
498
J.E. Casida et al.
carboxylates of primary alcohols (resmethrin, ethanomethrin, RU-15525, phenothrin, and permethrin) (Table I). Although the cis-pyrethroids are generally oxidized as rapidly as or more rapidly than the trans-pyrethroids, the overall rate of metabolism is not as great as for the trans-isomers which are more readily hydrolyzed. The (+)- and (-)-isomers of c/s-resmethrin are metabolized at similar rates whereas the (+)-trans-isomer is metabolized more rapidly than the ( - ) - t r a n s isomer (Soderlund and Casida 1974). The oxidative and overall metabolism rates are reduced on replacing the isobutenyl group with the cyclopentylidene-methyl group (Abernathy et al. 1973, Soderlund and Casida 1974) but not with the dichlorovinyl group (Soderlund and Casida 1974) whereas the incorporation of a thiolactone moiety as in RU-15525 increases the metabolism rate (Soderlund and Casida 1974). Bioresmethrin is hydrolyzed by esterases of mouse brain (Jao and Casida 1974b) and S-bioallethrin is oxidized by lung microsomes (Casida 1973) but much more information is needed on both the hydrolytic and oxidative capacities of various tissues in pyrethroid metabolism.
Other considerations on pyrethroid metabolism Synergists to retard pyrethroid metabolism in insects must be selected with care to maintain the favorable safety factor for mammals (Casida 1970). Thus, high injected doses of the synergist piperonyl butoxide or of certain other esterase and oxidase inhibitors markedly increase the toxicity to mice of (+)-cis-resmethrin, ( + )-trans- and ( + )-cis-ethanomethrin, and ( + )-cis-furamethrin (Abernathy and Casida 1973, Abernathy et al. 1973, Jao and Casida 1974a). Although this phenomenon does not appear to pose any human hazard with the commercial pyrethroid-synergist combinations, it may become a problem if more effective synergists or pyrethroids more toxic to mammals enter use. Ideal pyrethroids should not need synergists. Several aspects of pyrethroid biodegradation warrant further study: metabolic fate in man; identity and quantity of metabolites in milk and meat from exposed animals; the metabolism after inhalation and dermal exposure; toxicology and metabolic fate of photoproducts, particularly because several of these differ from pyrethroid metabolites, e.g., epoxides, cyclic peroxides, and rearranged derivatives; fate in different types of plants and soils to lay the background for agricultural pest control. Thus, the information available on pyrethroid metabolism is a small portion of that needed for a group of insecticides likely to be used in increasing amounts in the future.
Acknowledgment Study supported in part by grants from: The National Institutes of Health (2 PO1 ES00049); The Rockefeller Foundation; S. B. Penick and Co., Orange, NJ; Agricultural Chemical Div., FMC Corp., Middleport, NY; S. C. Johnson and Son, Inc.,
Structure-Biodegradability Relationships in Pyrethroids
499
Racine, Wis.; Cooper, McDouga11, and Robertson, Ltd., Berkhamsted, Herts, England; National Research Development Corp., London, England; Roussel-UclafProcida, Paris, France.
References Abernathy, C. O., and J. E. Casida: Pyrethroid insecticides: esterase cleavage in relation to selective toxicity. Science 179, 1235 (1973). Abernathy, C. O., K. Ueda, J. L. Engel, L. C. Gaughan, and J. E. Casida: Substrate-specificity and toxicological significance of pyrethroid-hydrolyzing esterases of mouse liver microsomes. Pesticide Biochem. Physiol. 3, 300 (1973). Casida, J. E.: Mixed-function oxidase involvement in the biochemistry of insecticide synergists. J. Agr. Food Chem. 18, 753 (1970). Casida, J. E.: Biochemistry of pyrethrins. In J. E. Casida (ed.): Pyrethrum the Natural Insecticide, pp. 101-120. New York: Academic Press (1973). Casida, J. E., E. C. Kimmel, M. Elliott, and N. F. Janes: Oxidative metabolism of pyrethrins in mammals. Nature 230, 326 (1971). Elliott, M.: The relationship between the structure and the activity of pyrethroids. Bull . W.H.O. 44, 315 (1971). Elliott, M., A. W. Farnham, N. F. Janes, P. H. Needham, D. A. Pulman, and J. H. Stevenson: A photostable pyrethroid. Nature 246, 169 (1973). Elliott, M., N. F. Janes, E. C. Kimmel, and J. E. Casida: Metabolic fate of pyrethrin I, pyrethrin II, and allethrin administered orally to rats. J. Agr. Food Chem. 20, 300 (1972). Elliott, M., K. Ueda, L. C. Gaughan, and J. E. Casida: Unpublished results (1974). Jao, L. T., and J. E. Casida: Esterase inhibitors as synergists for (+)-trans-chrysanthemate insecticide chemicals. Pesticide Biochem. Physiol. 4,456 (1974a). Jao, L. T., and J. E. Casida: Insect pyrethroid-hydrolyzing esterases. Pesticide Biochem. Physiol. 4, 465 (1974b). Lhoste, J. and F. Rauch: A new pyrethroid with a very strong knock-down effect. Paper presented at Third International Congress of Pesticide Chemistry (IUPAC), Helsinki, Finland, July 3-9, 1974. Masri, M. S., F. T. Jones, R. E. Lundin, G. F. Bailey, and F. DeEds: Metabolic fate of two chrysanthemumic acid esters: barthrin and dimethrin. Toxicol. Appl. Pharmacol. 6, 711 (1964). Miyamoto, J., and T. Suzuki: Metabolism of tetramethrin in houseflies in vivo. Pesticide Biochem. Physiol. 3, 30 (1973). Miyamoto, J., T. Nishida, and K. Ueda: Metabolic fate of resmethrin, 5-benzyl-3furylmethyl dl-trans-chrysanthemate in the rat. Pesticide Biochem. Physiol. 1, 293 (1971).
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Miyamoto, J., Y. Sato, K. Yamamoto, M. Endo, and S. Suzuki: Biochemical studies on the mode of action of pyrethroidal insecticides. Part I. Metabolic fate of phthalthrin in mammals. Agr. Biol. Chem. 32, 628 (1968). Miyamoto, J., T. Suzuki, and C. Nakae: Metabolism of phenothrin or 3-phenoxybenzyl d-trans-chrysanthemumate in mammals. Pesticide Biochem. Physiol. 4, 438 (1974). Nakanishi, M., Y. Kato, T. Furuta, and S. Miura: Metabolic fate of proparthrin. Studies on insecticide. VIII. Botyu-Kagaku 36, 116 (1971). Soderlund, D. M., and J. E. Casida: Unpublished results (1974). Suzuki, T., and J. Miyamoto: Metabolism of tetramethrin in houseflies and rats in vitro. Pesticide Biochem. Physiol. 4, 86 (1974). Ueda, K., L. C. Gaughan, and J. E. Casida: Unpublished results (1974a). Ueda, K., L. C. Gaughan, and J. E. Casida: Photodecomposition of resmethrin and related pyrethroids. J. Agr. Food Chem. 22, 212 (1974b). Verschoyle, R. D., and J. M. Barnes: Toxicity of natural and synthetic pyrethrins to rats. Pesticide Biochem. Physiol. 2, 308 (1972). Yamamoto, I.: Mode of action of synergists in enhancing the insecticidal activity of pyrethrum and pyrethroids. In J. E. Casida (ed.): Pyrethrum the Natural Insecticide, pp. 195-210. New York: Academic Press (1973). Yamamoto, I., and J. E. Casida: O-Demethyl pyrethrin II analogs from oxidation of pyrethrin I, allethrin, dimethrin, and phthalthrin by a house fly enzyme system. J. Econ. Entomol. 59, 1542 (1966). Yamamoto, I., E. C. Kimmel, and J. E. Casida: Oxidative metabolism of pyrethroids in houseflies. J. Agr. Food Chem. 17, 1227 (1969).
Manuscript received April 7, 1975; accepted April 15, 1975
ERRATUM Archives of Environmental Contamination and Toxicology, Vol. 3, No. 3 (1975): Page 358, Table I, column 5 (14 days after spraying, low airflow-rate sprayer) at tree site no. 4, reads 0.1264--should be 0.264.
