Selective inhibition of separate esterases in rat and mouse liver microsomes hydrolyzing malathion,trans-permethrin,and cis-permethrin

Separate esterase activities of rat and mouse liver microsomes hydrolyzing malathion, trans-permethrin, and cis-permethrin were differentiated on the basis of their sensitivities to inhibition by paraoxon and α-naphthyl N-propylcarbamate (NPC). In rat liver microsomes, the malathionhydrolyzing activity was more sensitive to both inhibitors and showed a different time course of NPC inhibition than the activities hydrolyzing the permethrin isomers. Paraoxon completely inhibited trans-permethrin hydrolysis, but only partially inhibited that of cis-permethrin. The paraoxonsensitive trans- and cis-permethrin-hydrolyzing activities were not differentially inhibited, but separate inhibition curves were obtained for the inhibition of trans- and cis-permethrin hydrolysis by NPC. The mouse liver esterase activity hydrolyzing trans-permethrin showed a similar paraoxon sensitivity to that of rat liver, but that the paraoxon-sensitive portion of the cis-permethrinhydrolyzing activity was 5.5-fold less sensitive to paraoxon than the corresponding rat liver activity and was clearly differentiated from the mouse liver trans-permethrin-hydrolyzing activity. The mouse liver malathion-hydrolyzing activity was 100-fold less sensitive to paraoxon and 14-fold less sensitive to NPC than the corresponding rat liver activity. Rat and mouse liver esterase activities hydrolyzed trans- and cis-permethrin at similar rates under standard assay conditions, but mouse liver esterases were 10-fold less active in hydrolyzing malathion. The higher specific activity of rat liver malathion-hydrolyzing esterases resulted from the greater apparent affinity and maximum velocity for malathion hydrolysis. These results demonstrate that the hydrolysis of malathion, trans-permethrin, and cis-permethrin by rat and mouse liver microsomal preparations involves several esterases with differing substrate specificities and inhibitor sensitivities.     

The toxicological significance of paraoxon deethylation

The in vivo formation of deethylation and hydrolytic products of paraoxon degradation after parathion or paraoxon administration was nearly equal in control male rats, and the relative abundance of metabolites was not appreciably altered by pretreatment of rats with enzymeinducing agents. However, pretreatment with inducers dramatically increased the oxidative paraoxon O-deethylase of male rat liver while having little effect on hydrolytic enzymes. Prior to induction, the hepatic O-deethylase activity was greatly inferior to the various hydrolytic enzymes, but nearly equal levels of both enzyme systems were found after induction. These results indicate that a large portion of the hepatic hydrolases detected in vitro is not active in vivo. It also appears that the majority of the induced hepatic deethylase was not involved in vivo at the dosage levels employed. The in vivo metabolism of monoethyl paraoxon was also demonstrated. The predominant metabolite of ethyl-[1-¹⁴C]monoethyl paraoxon is ¹⁴CO₂, while phenyl-[1-¹⁴C]monoethyl paraoxon yielded 4-nitro[1-¹⁴C]phenol. Paraoxon deethylation was also shown to be an important detoxication mechanism in female rats and male mice and must be considered in interpreting the toxicological properties of parathion and paraoxon.     

Effect of liver enzyme induction on paraoxon metabolism in the rat

Orally administered [1-¹⁴C]ethyl paraoxon, O,O-diethyl-O-p-nitrophenyl phosphate, is readily absorbed from the gastrointestinal tract of male albino rats. Radioactivity is essentially eliminated in 72 hr by excretion into urine and feces and by expiration as ¹⁴CO₂. Compounds with radioactivity in the urine are tentatively identified as diethyl phosphoric acid, desethyl paraoxon, ethanol, metabolites conjugated with amino acids, and paraoxon; the first compound is the predominant radioactive metabolite. Intraperitoneally injected phenobarbital, DDT, dieldrin, and endrin are inducers of microsomal enzymes that degrade paraoxon. The aryl phosphate-cleaving activity in vitro is not dependent on the addition of NADPH. O-Dealkylation of paraoxon is catalyzed by microsomal enzymes that require NADPH and oxygen and are inhibited by carbon monoxide. Microsomal enzymes from rats pretreated with enzyme inducers give an increased rate of O-dealkylation of paraoxon. Reduced glutathione has little or no effect on paraoxon degradation by either microsomal or soluble enzymes. Actinomycin D inhibits O-dealkylation of paraoxon in vivo, as indicated by reduction of ¹⁴CO₂ formation, and in vitro, as indicated by decreased activity of microsomal O-dealkylase. The role of microsomal mixed-function oxidases and NADPH-dependent O-dealkylase in the metabolism of organophosphorus insecticides is discussed.     

Toxicity in the brain of organophosphate insecticides: Comparison of the toxicities of metabolites with parent compounds using an intracerebral injection method

The toxicity in the brain of several parathion, fenthion, and fensulfothion insecticides and their toxic metabolites was determined by a technique of directly injecting the compounds into the region of the third ventricle of conscious mice, an area rich in cholinesterase activity. The results were compared on a body weight basis to the toxicity of these compounds when given by ip and oral routes. The results show that there is a direct relationship between the relative inhibition of cholinesterase activity in the brain by the organophosphates (e.g., methyl paraoxon, Sumioxon, and some members of the fenthion series) and the toxicity of these compounds in the brain. Methyl paraoxon and Sumioxon were found to be very toxic in the brain, Sumioxon being three to four times less toxic than methyl paraoxon. This is of the same order of effect of these compounds in inhibiting cholinesterases. It is concluded that any selective effects of Sumithion compared with methyl parathion must be due to the greater rate of metabolism of Sumithion to less toxic metabolites as well as to the lower toxicity of the oxon metabolite and not due to the relative rates of penetration of the toxic oxygen metabolites as previously suggested [J. Miyamoto, Agr. Biol. Chem.28, 422 (1964)]. A gas-liquid chromatographic method was employed to assess the distribution in the brain following intracerebral injection of the parathion-type compounds. The results suggest that there may be intracerebral metabolism of thionophosphates in vivo.     

Species differences in the in vitro inhibition of brain acetylcholinesterase and carboxylesterase by mipafox,paraoxon, and soman

Acute intraperitoneal toxicity of mipafox, paraoxon, and soman was highest in chicken, followed by rat, and lowest in frog. Species differences in organophosphorus toxicity were not related to differences in the specific activities of either acetylcholinesterase or carboxylesterase in brain. The sensitivity to inhibition of brain acetylcholinesterase in vitro by the organophosphorus compounds was closely related to the susceptibility of the species to acute organophosphorus poisoning. Both the acute toxicity and the sensitivity of brain acetylcholinesterase to inhibition in vitro by organophosphorus compounds in all the species were in the following order of increasing activity: mipafox < paraoxon < soman. The sensitivity of brain carboxylesterase to inhibition by the organophosphorus compounds were less than that of acetylcholinesterase and it could not be related to species susceptibility to acute organophosphorus toxicity. Paraoxon-insensitive phenyl valerate hydrolase in chicken brain was more sensitive to inhibition by mipafox and soman compared to rat; in chicken the sensitivity of paraoxon-insensitive phenyl valerate hydrolase to inhibition by soman was 9000 times more than that by mipafox, while in rat it was 100 times more. Frog brain had no paraoxon-insensitive phenyl valerate hydrolase activity. No evidence of age dependence was noticed in the specific activities of brain acetylcholinesterase, carboxylesterase, neurotoxic esterase, and paraoxon-insensitive phenyl valerate hydrolase or in the sensitivity of these enzymes to in vitro inhibition by organophosphorus compounds in both rats and chickens.     

Role of a microsomal carboxylesterase in reducing the action of malathion in eggs of Triatoma infestans

A microsomal malathion carboxylesterase present in Triatoma infestans eggs was active from the first day of embryonic development. This microsomal egg malathionase (MEM) showed a unique band in polyacrilamide gel electrophoresis (PAGE) when malathion was used as substrate. In vivo metabolism of [¹⁴C]malathion during all embryonic development showed a 10% degradation due to carboxylesterases. The in vitro evaluation of the same metabolic pathway catalyzed by the microsomal fraction of T. infestans eggs showed partial inhibition by paraoxon. α- and β-malathion monoacids were identified as the main metabolites of the in vivo and in vitro metabolic pathways. The carboxylesterase band that appeared in PAGE (MEM) from the first day of embryonic development could be the main cause of malathion tolerance in T. infestans eggs.     

Paraoxon-induced release of β-glucuronidase from rat hepatocytes in vitro

A suspension culture of isolated rat hepatocytes was used to reproduce in vitro the paraoxon-induced release of hepatic β-glucuronidase observed in vivo. After a short latent period, exposure of hepatocytes to paraoxon at 10^?7 to 10^?4M resulted in a typical dose-dependent response, with highest release occurring at 10^?4M paraoxon. With 10^?3M paraoxon, however, response was anomalous with a much-decreased enzyme release. As expected from earlier results in vivo, SV₁-oxon exhibited less effect than paraoxon.     

Multiple forms of esterases in mouse,rat, and rabbit liver,and their role in hydrolysis of organophosphorus and pyrethroid insecticides

Six to seven esterases from mouse, rat, and rabbit liver microsomes were resolved by chromatofocusing in the pH range 7–4. Each esterase peak showed a different substrate specificity pattern with the substrates evaluated. Malathion and paraoxon hydrolysis always corresponded with p-nitrophenyl acetate and methylthiobutyrate hydrolysis, whereas the pattern of fenvalerate hydrolysis was more complicated. Phosphorotriester hydrolase activity was isolated, and was found to be more specific toward paraoxon than toward the other insecticides. Time-course studies of paraoxon hydrolysis indicated that the hydrolysis of paraoxon by carboxylesterase was an inhibitory reaction. This reaction and phosphorotriester hydrolase activity can serve as a detoxication reaction toward organophosphate insecticides.     

Hepatic disposition of parathion: Uptake by isolated hepatocytes and chromatographic translobular migration

Isolated rat hepatocytes suspended in Waymouth's medium absorbed parathion rapidly and reversibly until the intracellular parathion concentration reached more than 300 times the ambient concentration. The distribution quotient q, defined as the ratio of intra- and extracellular concentrations at equilibrium, decreased when horse serum was added to the medium. The high hepatocyte affinity and rapid uptake of parathion suggested that this compound might be almost completely absorbed by periportal hepatocytes in the perfused liver and migrate downstream chromatographically through the lobule. Parathion infusion experiments verified this prediction and showed that the migration rate is dependent on the q value. The chromatographic feature may be useful as a basis for nonhistological investigation of intralobular hepatocyte heterogeneity. The lobule may function as a reverse-phase chromatograph also for many other unionized xenobiotics. Implications of the findings in the hepatic disposition of xenobiotics in vivo are discussed.     

The biochemical basis of resistance to organophosphorus insecticides in the sheep blowfly, Lucilia cuprina

The metabolism in vivo and in vitro of [¹⁴C]parathion and [¹⁴C]paraoxon was studied in a susceptible (LS) and an organophosphorus-resistant (Q) strain of the sheep blowfly, Lucilia cuprina. Both strains detoxified the insecticides in vivo via a number of pathways, but the resistant strain produced more of the metabolites diethyl phosphate and diethyl phosphorothionate. No difference was found between strains in the rate of penetration of the compounds used. Also, in vitro studies showed no difference between strains in the sensitivity of head acetylcholinesterase to inhibition by paraoxon. Both the microsomal and the 100,000g supernatant fractions degraded paraoxon, but resistance in Q could be explained by the eightfold greater rate of diethyl phosphate production with or without added NADPH. Parathion was also degraded to diethyl phosphorothionate by an NADPH-requiring enzyme in microsomal preparations from both strains. However, Q produced significantly more diethyl phosphorothionate in vivo than LS. It was concluded that organophosphorus resistance in Q was due mainly to a microsomal phosphatase hydrolyzing phosphate but not phosphorothionate esters, probably enhanced by a microsomal oxidase detoxifying the latter.     

Quantitative structure-activity studies of benzoylphenylurea larvicides: I. Effect of substituents at aniline moiety against Chilo suppressalis walker

The larvicidal activity of a series of N-2,6-difluoro- and N-2,6-dichlorobenzoyl-N′-(4-substituted phenyl)ureas against nondiapause larvae of the rice stem borer, Chilo suppressalis Walker, was measured by topical application and oral administration methods under conditions with and without piperonyl butoxide as an inhibitor of oxidative metabolism. The effects of substituents at the anilide moiety on the larvicidal activity were analyzed quantitatively using physicochemical substituent parameters and regression analysis. The results indicate that the oxidative metabolism in the larval body which is favored by electron-donating substituents is significant in determining the activity. The activity, when the metabolic factor is eliminated, is enhanced by electron-with-drawing and hydrophobic substituents and lowered by bulky groups. The inhibitory activity against new cuticle formation of the same series of compounds was also measured using cultured integument of the rice stem borer diapause larva. The comparison of the quantitative analyses between larvicidal and integument-level activities shows that inhibition of cuticular development is the most important factor governing larvicidal activity.     

Enantioselective metabolism and cytotoxicity of the chiral herbicide ethofumesate in rat and chicken hepatocytes

We investigated the metabolic kinetics and toxicity of ethofumesate (ETO) in rat and chicken hepatocytes using a chiral high-performance liquid chromatographic (HPLC) method. The metabolic of ETO in rat hepatocytes was enantioselective, whereas it was not in chicken hepatocytes. The T_1/2 of (−)-ETO was about two times longer than that of (+)-ETO after the rat hepatocytes had been incubated with 20 μM rac-ETO. There was no chiral conversion or transformation during their incubation with the hepatocytes. Toxicity differences were observed between the two enantiomers of ETO, reflected in their EC₅₀ values in rat and chicken hepatocytes. The stereoselective cytotoxicity of the two enantiomers was opposite in rat and chicken hepatocytes. We have developed a method of studying the toxicokinetics and cytotoxicity of chiral agrochemicals in hepatocytes isolated from mammals (rats) and chicken. The data presented here allow a more thorough understanding of this pesticide and should be useful in its full environmental assessment.     

Comparative effects of organophosphorus insecticides on the activities of acetylcholinesterase,diacylglycerol kinase,and phosphatidylinositol phosphodiesterase in rat brain microsomes

The activities of acetylcholinesterase, diacylglycerol kinase, and phosphatidylinositol phosphodiesterase in rat brain microsomes were measured in the presence and absence of the organophosphorus insecticides, parathion and diazinon, and their respective oxon analogs, paraoxon and diazoxon. Marked inhibition of acetylcholinesterase (by 45–99%) was observed in the presence of paraoxon (10^?2–10^?6M) and diazoxon (10^?2–10^?4M). Reduction of acetylcholinesterase activity (by 22–33%) was achieved with the parent insecticides at high concentrations only (10^?2M). In most cases, diacylglycerol kinase was insensitive to the pesticides. Marked stimulation of phosphatidylinositol phosphodiesterase (by 10–57%) was observed in the presence of all pesticides (10^?2–10^?3M). The phosphodiesterase exhibited slightly greater sensitivity to the parent compounds compared to the oxon derivatives. Stimulation of the phosphodiesterase by the insecticides was not correlated with acetylcholinesterase inhibition. Accordingly, the increase in phosphodiesterase activity was judged not to be acetylcholine mediated, but rather represented a direct effect of the pesticides on the enzyme or its microenvironment. Based on the present in vitro observations, it is proposed that certain organophosphorus pesticides may interfere with the normal process of synaptic transmission through both the inhibition of acetylcholinesterase and the stimulation of phosphatidylinositol phosphodiesterase. In view of the high concentrations of pesticides required to elicit the latter effect, interpretation of its physiological significance must await results from further studies performed in vivo.     

Biochemical studies on sheep body louse Bovicola ovis (Schrank) (Phthiraptera: Trichodectidae): comparison of pyrethroid and organophosphate resistant and susceptible strains

Strains of sheep louse Bovicola ovis (Schrank) with various levels of resistance to pyrethroid and one strain with high degree of resistance to organophosphate (OP) insecticides were used to investigate the biochemical mechanisms of insecticide resistance, i.e., enhanced levels of general esterases, specific acetylcholinesterases (AChE), glutathione S-transferase (GST), and mixed function oxidases. Native gel electrophoresis combined with quantitative enzyme assays showed analogous expression profiles of several esterase isozymes in all the strains tested. The determination of the sensitivity of each esterase isozyme to five inhibitors (acetylthiocholine iodide, butyrylthiocholine iodide, paraoxon eserine sulfate, and pCMB) led to the identification of nine esterases in the B. ovis strain. Gel electrophoresis results are supported by enzyme assay studies where, except for the OP resistant strain, no differences in esterase activities were detected in all the pyrethroid resistant and susceptible strains assayed. Statistical analyses demonstrated that some strains have elevated GST activities compared to the susceptible reference strain.     

Metabolism of tetramethrin in houseflies and rats in vitro

Degradation of radiolabeled tetramethrin or 3,4,5,6-tetrahydrophthalimidomethyl dl-trans chrysanthemumate was tested in vitro by using abdomens of SK, lab-em-7-em, R_HOKOTA and P_y strains of houseflies and rat liver. The effect of NADH₂ and NADPH₂ on the metabolism of tetramethrin by housefly abdomen homogenate was slight, but phosphorothioates, their oxygen analogs, carbamate insecticides, NIA 16388, p-chloromercuribenzoic acid, and mercuric chloride showed marked inhibition. The enzyme activity was localized mainly in the microsomal fraction, where the major metabolites were 3,4,5,6-tetrahydrophthalimide (TPI) (a nonenzymatic reaction from N-(hydroxymethyl) 3,4,5,6-tetrahydrophthalimide, MTI) and chrysanthemumic acid. Smaller amounts of oxidized tetramethrins and chrysanthemumic acid were also produced. The cleavage of tetramethrin into MTI and chrysanthemumic acid was inhibited by such compounds as paraoxon, carbaryl, PCMB, NIA16388, and mercuric chloride. NADPH₂ or NADPH₂ plus carbon monoxide produced little effect. Similar results were obtained with rat liver microsomal fraction. It is presumed from the above findings that the cleavage is catalyzed either by a carboxyesterase or a hydrolase, and that some pyrethroids are metabolized in insects primarily through hydrolytic pathways. Metabolites from oxidative pathways (as in mammals) are formed in minor quantities.     

Insecticide cyclic nucleotide interactions: I. Quinoxalinedithiol derivatives: A new group of potent phosphodiesterase inhibitors

The direct effects of tepp, methyl paraoxon, DDT, dieldrin, aldicarb, dimetilan, rotenone, allethrin, and oxythioquinox were surveyed on cockroach brain adenyl cyclase and phosphodiesterase in vitro. The most striking result of this survey was the observation that oxythioquinox is a potent inhibitor of phosphodiesterase. The inhibitory activities of seven different quinoxalinedithiol derivatives were compared with those of methyl-xanthines and SQ 65,442 on phosphodiesterases from cockroach brain, rat brain, and beef heart. Although I₅₀ values of the quinoxaline inhibitors were found to be in the μM range, solubility deficiencies apparently limit their effectiveness with inhibition reaching limiting values of about 70–90% as concentrations are increased. Evaluation of the quinoxaline inhibitors to enhance the accumulation of cyclic AMP in the assay of adenyl cyclase did not demonstrate any significant advantage over the use of aminophylline, a standard inhibitor for this purpose. A new assay for phosphodiesterase, involving separation of substrate from product on aluminum oxide columns, was developed by modification of similar techniques utilized in the assay of adenyl cyclase.     

Insensitive acetylcholinesterase,high glutathione-S-transferase,and hydrolytic activity as resistance factors in a tetrachlorvinphos-resistant strain of house fly

The gene producing insensitive acetylcholinesterase in a tetrachlorvinphos-resistant strain of house fly was introduced into the genome of a susceptible strain. The resistance to a number of organophosphorus insecticides, which was high in the original strain (234- and more than 800-fold for paraoxon and tetrachlorvinphos, respectively), was much lower in the resulting strain, ranging from 53-fold for paraoxon to 3.9-fold for tetrachlorvinphos. Synergists further decreased these factors. The reduction in sensitivity of the acetylcholinesterase in vitro was about 20-fold for both inhibitors. The Michaelis parameters for hydrolysis of acetylcholine and acetylthiocholine of the mutant and normal acetylcholinesterase showed only minor differences. An increased rate of metabolism of the insecticides was found in the original strain. In vitro, glutathione-dependent detoxication was from 9- to 120-fold that of a susceptible strain, depending on the insecticide and the conditions used. The enzyme de-alkylated parathion and methylparaoxon, but there was no de-arylation of parathion, although it is important in other strains. In vitro, paraoxon and methylparaoxon were also hydrolysed, and this is not so in the susceptible strain. The high resistance present in the original strain seems to be due to joint action of at least three mechanisms.     

Paraoxon formation and hydrolysis by mammalian liver

In vitro studies of the desulfuration of parathion at 37°C by hepatic tissue from males and females of nine mammalian species revealed sex and species variation in initial rates of parathion desulfuration and arylesterase-catalyzed hydrolysis of the oxygen analogue, paraoxon. Double reciprocal plots of initial rates of parathion activation for representative males and females of each species gave K_m values ranging from 0.2 × 10^?4?1.0 × 10^?4M parathion. Guinea pigs and rats were the only animals showing sex differences in activation, males possessing higher desulfurating abilities than the corresponding females. Based upon the sex possessing the higher desulfurating ability, the species pattern of decreasing activity was hamster > guinea pig > mouse > rat > rabbit > bovine > dog > porcine > cat. Studies of paraoxon hydrolysis indicated that only rats showed sex differences in hydrolysis, males possessing higher arylesterase activity than females. The species pattern of decreasing hydrolytic activity was in the order mouse > bovine > rat > guinea pig > rabbit > hamster > cat > dog > porcine.     

Significance of phosphorylation in organophosphate-mediated β-glucuronidase release from the rat liver

The elevation of rat blood β-glucuronidase caused in vivo by O,O-dialkyl O-phenyl phosphates and phosphorothioates correlated well with the electron-withdrawing tendency (σ^?) of leaving group substituents indicating the importance of a phosphorylation mechanism in the enzyme release. Hydrophobic bonding of these compounds may facilitate the phosphorylation since hydrophobicity (π) of substituents also correlated with the enzyme release. SKF 525-A decreased the elevation of β-glucuronidase by parathion through the suppression of paraoxon production. Pretreatment of rats with phenobarbital or DDE resulted in lower and delayed enzyme release caused by parathion.     

Biochemical characteristics of insecticide resistance in the fall armyworm, Spodoptera frugiperda (J.E. Smith)

A strain of the fall armyworm, Spodoptera frugiperda (J.E. Smith), collected from corn in Citra, Florida, showed high resistance to carbaryl (562-fold) and methyl parathion (354-fold). Biochemical studies revealed that various detoxification enzyme activities were higher in the field strain than in the susceptible strain. In larval midguts, activities of microsomal oxidases (epoxidases, hydroxylase, sulfoxidase, N-demethylase, and O-demethylase) and hydrolases (general esterase, carboxylesterase, β-glucosidase) were 1.2- to 1.9-fold higher in the field strain than in the susceptible strain. In larval fat bodies, various activities of microsomal oxidases (epoxidases, hydroxylase, N-demethylase, O-demethylases, and S-demethylase), glutathione S-transferases (CDNB, DCNB, and p-nitrophenyl acetate conjugation), hydrolases (general esterase, carboxylesterase, β-glucosidase, and carboxylamidase) and reductases (juglone reductase and cytochrome c reductase) were 1.3- to 7.7-fold higher in the field strain than in the susceptible strain. Cytochrome P450 level was 2.5-fold higher in the field strain than in the susceptible strain. In adult abdomens, their detoxification enzyme activities were generally lower than those in larval midguts or fat bodies; this is especially true when microsomal oxidases are considered. However, activities of microsomal oxidases (S-demethylase), hydrolases (general esterase and permethrin esterase) and reductases (juglone reductase and cytochrome c reductase) were 1.5- to 3.0-fold higher in the field strain than in the susceptible strain. Levels of cytochrome P450 and cytochrome b₅ were 2.1 and 1.9-fold higher, respectively, in the field strain than in the susceptible strain. In addition, acetylcholinesterase from the field strain was 2- to 85-fold less sensitive than that from the susceptible strain to inhibition by carbamates (carbaryl, propoxur, carbofuran, bendiocarb, thiodicarb) and organophosphates (methyl paraoxon, paraoxon, dichlorvos), insensitivity being highest toward carbaryl. Kinetics studies showed that the apparent K_m value for acetylcholinesterase from the field strain was 56% of that from the susceptible strain. The results indicated that the insecticide resistance observed in the field strain was due to multiple resistance mechanisms, including increased detoxification of these insecticides by microsomal oxidases, glutathione S-transferases, hydrolases and reductases, and target site insensitivity such as insensitive acetylcholinesterase. Resistance appeared to be correlated better with detoxification enzyme activities in larval fat bodies than in larval midguts, suggesting that the larval fat body is an ideal tissue source for comparing detoxification capability between insecticide-susceptible and -resistant insects.