The nature of the seasonal colour dimorphism in the scorpion fly,Panorpa japonica thunberg

The scorpion fly, Panorpa japonica, displays a seasonal colour dimorphism by changing from black to yellow in the adult state. The yellow pigment in the integument and haemolymph of the adult fly was identified as sepiapterin, while the black integument pigment was found to be melanin. After analysis of sepiapterin content by high performance liquid chromatography and determination of total haemolymph volume by [carboxyl-¹⁴C]inulin, it was shown that sepiapterin levels in the haemolymph and integument varied greatly both in the two colour types of insects and in the two sexes. Photometric analysis of melanin content showed that melanin levels correlated negatively with sepiapterin levels. These quantitative differences in sepiapterin and melanin fully explain the colour dimorphism in the insect.     

Incorporation of various putative precursors into cuticular 3,4-dihydroxyphenylacetic acid of adult Tenebrio molitor

Post-emergence levels of 3,4-dihydroxyphenylacetic acid (DOPAC) and ketocatechol were determined in cuticle from adult Tenebrio molitor. Possible pathways for biosynthesis of DOPAC were studied by comparing the incorporation of injected [U-¹⁴C]tyrosine, [7-¹⁴C]dopamine, [7-¹⁴C]DOPA, [7-¹⁴C]tyramine, [U-¹⁴C]p-hydroxyphenylpyruvic acid (p-HPPA) and [ring-³H]p-hydroxyphenylacetic acid (p-HPAA) into cuticular DOPAC during its period of maximal increase 1–3 days after adult emergence. Increased incorporation of [U-¹⁴C]tyrosine between days 0 and 3 suggests rapid de novo biosynthesis of DOPAC from this primary precursor. Of the putative intermediates tested, only p-HPPA had a pattern of incorporation similar to that seen with tyrosine. Since p-HPAA was poorly incorporated into both cuticle and DOPAC, a tentative pathway tyrosine → p-HPPA → 3,4-dihydroxyphenylpyruvic acid → DOPAC is proposed.     

Biosynthesis of internally branched monomethylalkanes in the cockroach Periplaneta fuliginosa.

Sodium [1-¹⁴C]acetate, sodium [1-¹⁴C]propionate, sodium [2-¹⁴C]propionate, sodium [3-¹⁴C]propionate and sodium [methyl-¹⁴C]methylmalonate were readily incorporated into the cuticular hydrocarbons of nymphal stages of the cockroach Periplaneta fuliginosa both in vivo and in vitro, whereas no incorporation of [methyl-¹⁴C]methionine was observed. The alkanes of the nymphal stages of this insect are 25+% n-alkanes, 14% 3-methylalkanes, and 59+% internally branched monomethylalkanes, principally 13-methylpentacosane. Sodium [1-¹⁴C]acetate was incorporated into each class of alkane at about its percentage composition. In contrast, labeled sodium propionate and sodium methylmalonate were preferentially incorporated into the branched fractions. Radio-gas-liquid chromatography showed that sodium [1-¹⁴C]propionate was incorporated almost exclusively into 3-methyltricosane and 13-methylpentacosane, whereas sodium [1-¹⁴C]acetate was incorporated into each glc peak at about its percentage composition. These data suggest that propionate, incorporated during chain elongation, serves as the branching methyl group donor for both the 3-methyl and the internally branched monomethylalkanes in insects. The location of hydrocarbon synthesis in P. fuliginosa was studied using an in vitro tissue slice system. Excised cuticle slices, with adhering fat body tissue removed, gave good incorporation of labeled substrates into the hydrocarbon fraction. No hydrocarbon synthesis was observed in fat body preparations.     

Mineralization of Parathion in the Rice Rhizosphere

We studied ¹⁴CO₂ evolution from ring-labeled [2,6-¹⁴C]parathion (O,O-diethyl-O-p-nitrophenyl phosphorothioate) in the rhizosphere of rice seedlings. The soil samples (nonflooded [60% water-holding capacity] and flooded) were treated first with technical parathion (20 μg/g) and then after 10 days with ring-labeled [¹⁴C]parathion. In unplanted soil, less than 5.5% of the ¹⁴C in the parathion was evolved as ¹⁴CO₂ in 15 days under both flooded and nonflooded conditions. In soil planted with rice, 9.2% of the radiocarbon was evolved as ¹⁴CO₂ under nonflooded conditions, and 22.6% was evolved under flooded conditions. These results suggest that soil planted with rice permits significant ring cleavage, especially under flooded conditions.     

NIACIN AND NIACINAMIDE TRANSPORT IN THE CENTRAL NERVOUS SYSTEM. IN VIVO STUDIES

Abstract– The concentration ol niacinamide in plasma and CSF was 0.5 and 0.7 μm respectively. The, mechanisms by which niacin and niacinamide, which are not synthesized in brain, enter brain, CSF and choroid plexus were investigated by injecting [¹⁴C]niacin or [¹⁴C]niacinamide intravenously and intraventricularly. [¹⁴C]Niacin or [¹⁴C]niacinamide, with or without unlabeled niacin or niacinamide, were infused intravenously at a constant rate into conscious rabbits. At 3 h, [¹⁴C]niacinamide, but not [¹⁴C]niacin, readily entered CSF, choroid plexus and brain. The addition of 4.1 mmol/kg niacinamide to the infusate markedly depressed the relative entry of [¹⁴C]niacinamide into choroid plexus and brain but not into CSF. After intraventricular injection, [¹⁴C]niacin was rapidly cleared from CSF and readily entered brain and choroid plexus. The addition of unlabeled niacin to the intraventricular injectate decreased the clearance of [¹⁴C]niacin from CSF and the entry of [¹⁴C]niacin into choroid plexus and brain. Unlike niacin, carrier niacinamide (82 μmol) in the injectate did not depress the extremely rapid clearance of intraventricularly injected [¹⁴C]niacinamide from CSF but did decrease the entry of [¹⁴C]niacinamide into brain. These results show that the control of entry and exit of niacinamide and niacin is the mechanism, at least in part, by which total niacin and NAD levels in brain cells are regulated. In the case of niacinamide which readily passes between CSF and plasma, the regulation of entry of niacinamide into brain cells by a high affinity accumulation system is an integral part of the homeostatic system. In the case of niacin, penetration into CSF and the extracellular space of brain from plasma as well as regulation of entry into brain cells by a saturable accumulation system are two distinct parts of the homeostatic system. In vivo, niacin that enters the central nervous system is converted to the principal plasma vitamer, niacinamide, in its free or bound forms such as NAD.     

Catechol conjugation with hemolymph proteins and their incorporation into the cuticle of the American cockroach,Periplaneta americana

Newly ecdysed American cockroaches, Periplaneta americana (sixth to last instar) were injected with radioactive dopamine (DA) and hemolymph was collected at 10–60 min post-ecdysis. Size-exclusion chromatography established the presence of at least three proteins that serve as catecholamine carriers. Reinjection of the smaller radiolabeled phenol-bound proteins into newly ecdysed animals results in in vivo aggregation, with the radiolabel bound to large MW proteins (30->200 kDa). In addition, the reinjection of radiolabeled protein of any size resulted in the incorporation of the label into the newly sclerotized cuticle. Hemolymph proteins were synthesized in vivo using [¹⁴C]leucine and subsequently double labeled in vivo with [³H]dopamine. After sclerotization (7 h post-ecdysis) the cuticle was extirpated, hydrolyzed and counted. An identical ratio of ¹⁴C to ³H was found in cuticle extracts as in the double-labeled hemolymph proteins, suggesting that the phenol-bound protein was incorporated in the cuticle unchanged. It appears that the catechol bound to the proteins exists as a β-glucoside.     

The incorporation of precursors into Drosophila larval cuticle

Incorporation of tritiated leucine, tyrosine and glucosamine into the integument of larval Drosophila melanogaster was followed by electron-microscope autoradiography. Tritiated leucine, tyrosine, and glucosamine were incorporated into the endocuticle by apposition, giving rise to a distinct band of label in the endocuticle at a level which depended on the time between labelling and fixation. The labelled amino acids, but not glucosamine, were also detected in the epicuticle and both above and below the distinct labelled band in the endocuticle. The results indicate that the epicuticle grows within the third instar by intussusception of new materials which are transported from the epidermal cells through the endocuticle to the epicuticle. Breakdown of cuticle which was radioactively labelled by feeding larvae tritiated precursors was also followed by autoradiography. The results indicate that the breakdown products from the old cuticle may be reutilized in the synthesis of new cuticle.     

An ultrastructural study of the invasion of Culex quinquefasciatus larvae by Leptolegnia chapmanii (Oomycetes: Saprolegniales)

Transmission electron microscopy of the invasion of Culex quinquefasciatus by Leptolegnia chapmanii confirmed that it is a primary pathogen and revealed several differences between penetration via the gut and penetration via the integument. The latter often involved aggregations of zoospores, appressoria-like swellings of the invasive hyphae, and lateral growth of hyphae between the epicutical and endocuticle. These features were not detected in the case of gut invasion, but hyphal septa at the point of entry were apparently peculiar to this route. There was no evident tissue specificity, and death presumably resulted from generalized destruction of tissues.     

Glycine,Serine, and Leucine Metabolism in Different Regions of Rat Central Nervous System

We have investigated the glycine, serine and leucine metabolism in slices of various rat brain regions of 14-day-old or adult rats, using [1-¹⁴C]glycine, [2-¹⁴C]glycine, L-[3-¹⁴C]serine and L-[U-¹⁴C]leucine. We showed that the [1-¹⁴C]glycine oxidation to CO₂ in all regions studied occurs almost exclusively through its cleavage system (GCS) in brains of both 14-day-old and adults rats. In 14-day-old rats, the highest oxidation of [1-¹⁴C]glycine was in cerebellum and the lowest in medulla oblongata. In these animals, the L-[U-¹⁴C]leucine oxidation was lower than the [1-¹⁴C]glycine oxidation, except in medulla oblongata where both oxidations were the same. Serine was the amino acid that showed lowest oxidation to CO₂ in all structure studied. In adult rats brains, the highest oxidation of [1-¹⁴C]glycine was in cerebral cortex and the lowest in medulla oblongata. We have not seen difference in the lipid synthesis from both glycine labeled, neither in 14-day-old rats nor in adult ones, indicating that the lipids formed from glycine were not neutral. Lipid synthesis from serine was significantly high than lipid synthesis and from all other amino acids studied in all studied structures. Protein synthesis from L-[U-¹⁴C]leucine was significantly higher than that from glycine in all regions and ages studied.     

Distinct patterns of entry of two non-metabolizable amino acids into brain and other organs of infant guinea pigs

Abstract— Entry of [3-¹⁴C] α-aminoisobutyric acid (AIB) and [1-¹⁴C] 1-aminocyclopentanecarboxylic acid (cycloleucine) into the brain and other organs of the infant guinea pig has been investigated in vivo. The entry of [¹⁴C]AIB into brain was markedly restricted in comparison to its entry into other organs. The mean distribution ratio (¹⁴C in tissue water/¹⁴C in plasma water) achieved in brain at 45 min after administration of a pulse of [¹⁴C]AIB was 0.3. All other organs studied concentrated [¹⁴C]AIB from the blood stream, with the greatest uptake occurring in liver and kidney, in which distribution ratios reached values of 5–10. In contrast to AIB, [¹⁴C]cycloleucine entered the brain at a rate approximately the same as that into other organs. Distribution ratios for [¹⁴C]cycloleucine ranged between 0.5 and 2.0 for all organs. During the first few days of postnatal life, there was a sharp increase of concentrative uptake of [¹⁴C]AIB into liver and kidney. The entry of [¹⁴C]AIB into brain remained unchanged during this period. There was a small (35 percent) decrease in the rate of entry of [¹⁴C]cycloleucine into brain during the first 3 days of postnatal life. Since [¹⁴C]AIB is known to be concentrated from the surrounding medium by brain slices in vitro, we concluded that the locus of restriction of the entry of [¹⁴C]AIB into the brain in vivo is at the blood-brain barrier. We hypothesize that this property of the barrier is important in preventing concentrative uptake of pharmacologically active and potentially harmful amino acids by brain tissue.     

No effect of maternal niacin defficiency on niacin metabolism in newborn brain

The effects of maternal niacin and tryptophan deficiency on: (1) total niacin levels and (2) niacinnamide entry into brain, blood, and liver of newborn rabbits were studied. The deficient maternal diet produced a decreased concentration of the oxidized niacinamide-containing vitamers in the liver (73% of controls;P<0.05) but not in the brain of newborn rabbits. In both deficient and control newborn rabbits, the entry of [¹⁴C]niacinamide into brain and liver was saturable with an increasing [¹⁴C]niacinamide concentration in plasma. Also, the formation of [¹⁴C] NAD in brain and liver and [¹⁴C]niacinamide mononucleotide in blood was saturable. In vitro, the affinity of the saturable accumulation system for [¹⁴C]niacinamide in both newborn and adult rabbit brain slices was comparable (0.9 M). The avid saturable, uptake system for niacinamide in rabbit brain contributes to total niacin homeostasis in brain.     

Riboflavin Homeostasis in the Central Nervous System

Abstract: The mechanisms by which riboflavin, which is not synthesized in mammals, enters and leaves brain, CSF, and choroid plexus were investigated by injecting [¹⁴C]riboflavin intravenously or intraventricularly. Tracer amounts of [¹⁴C]riboflavin with or without FMN were infused intravenously at a constant rate into normal, starved, or probenecid-pretreated rabbits. At 3 h, [¹⁴C]riboflavin readily entered choroid plexus and brain, and, to a much lesser extent, CSF. Over 85% of the [¹⁴C]riboflavin in brain and choroid plexus was present as [¹⁴C]FMN and [¹⁴C]FAD. The addition of 0.2 mmol/kg FMN to the infusate markedly depressed the relative entry of [¹⁴C]riboflavin into brain, choroid plexus, and, less so, CSF, whereas starvation increased the relative entry of [¹⁴C]riboflavin into brain and choroid plexus. After intraventricular injection (2 h), most of the [¹⁴C]riboflavin was extremely rapidly cleared from CSF into blood. Some of the [¹⁴C]riboflavin entered brain, where over 85% of the ¹⁴C was present as [¹⁴C]FMN plus [¹⁴C]FAD. The addition of 1.2³μmol FAD (which was rapidly hydrolyzed to riboflavin) to the injectate decreased the clearance of [¹⁴C]riboflavin from CSF and the phosphorylation of [¹⁴C]riboflavin in brain. Probenecid in the injectate also decreased the clearance of [¹⁴C]riboflavin from CSF. These results show that the control of entry and exit of riboflavin is the mechanism, at least in part, by which total riboflavin levels in brain cells and CSF are regulated. Penetration of riboflavin through the blood-brain barrier, saturable efflux of riboflavin from CSF, and saturable entry of riboflavin into brain cells are three distinct parts of the homeostatic system for total riboflavin in the central nervous system.     

Hydrocarbon accumulation and lipid biosynthesis during larval development in the cabbage looper,Trichoplusia ni

Larvae of the cabbage looper, Trichoplusia ni, were analyzed for the accumulation and biosynthesis of cuticular and internal hydrocarbon at closely spaced and accurately timed intervals during the fourth and fifth stadia. Large differences in the incorporation of [1-¹⁴C]acetate into hydrocarbon were observed at different times during larval development. Much higher incorporation was observed during feeding stages as compared to wandering stages, while lowest rates of biosynthesis occurred just prior to ecdysis. Fourth stadia wanderers accumulated increased amounts of internal hydrocarbon, which is apparently used to cover the newly forming cuticle. During the fourth to fifth stadium moult insects lost all cuticular hydrocarbon that was present on the old cuticle (about 8 μg/insect) and had about 8 μg/insect on the surface of the newly exposed cuticle. During the fourth stadium incorporation of [1-¹⁴C]acetate into total lipid declined between feeding and wandering stages from 24% of injected radiolabel to 7%. Similar decreases in lipid biosynthesis were observed between feeders and wanderers in fifth stadium larvae with the greatest decrease found in the triacylglycerol fraction. These results document dramatic changes in the accumulation and biosynthesis of hydrocarbon and other lipids during larval development.     

Observations on the integument of the water mite Arrenurus major (Acari: Parasitengona)

A study of the integument of the aquatic mite Arrenurus major Marshall is presented. When the cuticle is examined with the unaided eye and the light microscope, it appears to possess numerous tiny pits. However, scanning electron micrographs of the cuticle reveal that it is a solid surface with topographical sculpturing of the epicuticle, indicating that the “pits” are an internal phenomenon. In cuticle which has been sectioned, areas devoid of cuticular material beneath the thin exocuticle are revealed. These areas are the pits which are goblet-shaped. The integument consists of five major strata. These are from the outside to the inside: (1) a superficial layer with a maximum observed thickness of 725 Å, (2) an epicuticle with a thickness of about 900 Å and composed of at least four sublayers, (3) an exocuticle with a thickness of about 1.5 Å. Fibers of the exocuticle are arranged in a Bouligand pattern and exhibit a regularly occurring discontinuity with a spacing of 200 Å. (4) An endocuticle ranging from 15 to 20 μ in thickness. The endocuticle is characterized by bandings which superficially resemble the lamellae of insects but are not homologous, microfibers which exhibit a preferred orientation, and the presence of the pits; and (5) an epidermis lying beneath the endocuticle and extending into the pits. Pore canals are present only in the exocuticle and have their origin at the apices of the pits. The pore canals contain a central filament, and a plug is present just beneath the epicuticle.     

On the pronymphal stage in the migratory locust (Locusta migratoria, Orthoptera, Acrididae)

The structure of the integument, somatic and visceral muscles, midgut, and Malpighian tubules were investigated at the late stages of the embryonic and early postembryonic development of the migratory locust, Locusta migratoria, to assess the organization of its pronymphal stage. In its morphogenetic features, the vermiform locust larva sometimes called the pronymph corresponds to the first nymphal instar covered with the second embryonic cuticle which has not been shed. Since the first-instar locust nymphs before and after the shedding of this embryonic cuticle differ significantly in many morphological characters, two consecutive phases of this nymphal instar can be distinguished: the first phase existing from the moment of development of the third embryonic cuticle to the shedding of the second one; the second phase existing from the shedding of the second embryonic cuticle to the formation of the cuticle of the second nymphal instar. Since the pronymphal stage should precede the nymph stage, it may be concluded that the pronymph of the locust is fully embryonized and covered with the second embryonic cuticle, which is also typical of other insects with hemimetabolous development (Konopová and Zrzavý, 2005). Therefore, it would be erroneous to refer to the vermiform first-instar nymph as the pronymph, because the two stages are separated by molting and formation of a new cuticle.     

Biosynthesis of the hydrocarbon components of the sex pheromone of the housefly,Musca domestica L.

Biosynthesis of hydrocarbons, including components of the sex pheromone of the housefly Musca domestica L., was investigated. In vitro studies with isolated tissues from adult flies showed that the hydrocarbon components of the pheromone were synthesized primarily by the epidermal cells in abdominal segments two to seven. The incorporation of [³H or ¹⁴C]-labelled acetate, palmitate, stearate and oleate into the saturated and unsaturated hydrocarbon components showed that (Z)-9-tricosene (muscalure) was synthesized de novo by female insects and the distribution of label was consistent with a pathway in which oleic acid was elongated and then decarboxylated. A comparison of the incorporation and distribution of labelled acetate, propionate and succinate into hydrocarbons indicated that the mono- and dimethylalkanes were formed by the substitution of a methylmalonyl-CoA for malonyl-CoA during chain elongation. The incorporation of radioactivity from [1-¹⁴C]-propionate increased dramatically in female insects two days after adult emergence, which corresponds in time to the production of methyl branched alkanes. In contrast, this substrate was not efficiently incorporated at any time into male insects.     

Turnover of protein and diacylglycerol components of lipophorin in insect haemolymph

Lipophorin, radiolabelled in the protein or diacylglycerol moiety, was purified from adult locusts injected previously with [¹⁴C]protein hydrolysate or sodium[1-¹⁴C]palmitate. The radiolabelled lipophorin was injected into adult male locusts and haemolymph samples taken periodically to determine the rate of disappearance of radioactivity from the haemolymph. Lipophorin was also purified from locusts that had been injected four days previously with ([¹⁴C]protein)-lipophorin to demonstrate that the radioactivity observed in the haemolymph at this time is due to radiolabelled lipophorin. The results indicate that the half-life of the protein component of lipophorin in resting insects is about 5–6 days whereas that of the diacylglycerol component is only about 2–3 hr.The results are consistent with the hypothesis that lipophorin functions as a “reusable shuttle” to transport a variety of lipid classes between sites of absorption, storage and utilisation.     

Electron microscopical localization of chitin in the cuticle of Halicryptus spinulosus and Priapulus caudatus (Priapulida) using gold-labelled wheat germ agglutinin: phylogenetic implications for the evolution of the cuticle within the Nemathelminthes

 The ultrastructure of the cuticle of adult and larval Priapulus caudatus and Halicryptus spinulosus is investigated and new features of cuticle formation during moulting are described. For the localization of chitin by TEM wheat germ agglutinin coupled to colloidal gold was used as a marker. Proteinaceous layers of the cuticle are revealed by digestion with pronase. The cuticle of larval and adult specimens of both species consists of three main layers: the outer, very thin, electron-dense epicuticle, the electron-dense exocuticle and the fibrillar, electron-lucent endocuticle. Depending on the body region, the exocuticle comprises two or three sublayers. The endocuticle can be subdivided into two sublayers as well. In strengthened parts such as the teeth, the endocuticle becomes sclerotized and appears electron-dense. Only all endocuticular layers show an intense labelling with wheat germ agglutinin-gold conjugates in all investigated specimens. Additional weak labelling is observed in the exocuticle III layer of the larval lorica of P. caudatus. All other cuticular layers remain unlabelled. Chitinase dissolves the unsclerotized endocuticular layers almost completely, but also exocuticle II and partly the loricate exocuticle III. The epicuticle, the homogeneous exocuticle I and the sclerotized endocuticle are not affected by chitinase. The labelling is completely prevented in all layers after incubation with chitinase. Pronase dissolves all exocuticular layers, but not evenly. The presumably sclerotized regions of exocuticle I are not affected as well as the complete epicuticle and the endocuticle. All cuticular features of the Priapulida are compared with the cuticle of each high-ranked taxon within the Nemathelminthes with special regard to the occurrence of chitin. Based on this out-group comparison it can be concluded that: (1) a two-layered cuticle with a trilaminate epicuticle and a proteinaceous basal layer represents an autapomorphic feature of the Nemathelminthes, (2) the stem species of the Cycloneuralia have already evolved an additional basal chitinous layer, (3) such a three-layered cuticle is maintained as a plesiomophy in the ground pattern of the Scalidophora and (4) in the Nematoida, the chitinous basal layer is replaced by a collagenous one at least in the adults; the synthesis of chitin is restricted to early developmental phases or the pharyngeal cuticle. Accepted: 12 March 1998     

The uptake of organic molecules by the ventral tube of Tomocerus flavescens (Tullberg, 1871) (Insecta: Collembola)

The net influx of water-soluble organic molecules by the ventral tube of Tomocerus flavescens, a soil litter-inhabiting Collembolan, was investigated. The following substances were tested: [¹⁴C]urea, [¹⁴C]glycerol, [¹⁴C]erythritol, [¹⁴C]l-leucine, [¹⁴C]d-glucose, [³H]inulin. The animals were exposed to moist filter paper containing a specific test solution. When they evert their ventral-tube vesicles, they absorb water and solutes through the cuticle and the transport epithelium into the body haemolymph. Contamination by radioactive substances and oral solute uptake was avoided by an experimental device. It is evident that the uptake rates decrease with increasing molecular mass especially in a range of 100–200. Further, the rates correlate with the radius of hydrated molecules and their lipid solubility. Significant differences in urea uptake have been shown for animals less than 10 days in culture (“field” animals) and more than 10 days in culture (“laboratory” animals). Whether changes in cuticle permeability could be affected by abrasion is discussed. There is a high deviation amongst uptake values in all experimental series. It seems probably that, besides individual differences caused by abrasion, the animals differ physiologically, e.g. during the moulting cycle and seasonally. A nutritive function of the ventral tube seems to be unlikely. Calculation reveals that the absorbed glucose provides only 0.013% of the amount the animals need for respiration.     

Integumental esteratic activity in Triatoma infestans and its contribution to the degradation of organophosphorus insecticides

1. Esteratic activity was found in the integument of adult Triatoma infestans, principally located in the epidermis. Specific activity was on average 29.3 nM phenylthioacetate/hr per anatomical unit.2. Esteratic activity depends on the insect's age and its starvation state. A dramatic increase was observed 1 week after moulting followed by a slight decay as a function of the age. A significant decrease of the activity was observed with a longer fasting time.3. Integumental esterases were characterized as carboxylesterases, butyrylcholinesterases and aryl plus acetylesterases by using eserine and paraoxon as inhibitors and acetylthiocholine, butyrylthiocholine and phenylthioacetate as substrates.4. Epidermis homogenates were able to hydrolyse OP insecticides when incubated in vitro. Hydrolysis of the carboxyester linkage by malathion was established and cleavage of the P—S—C bond by parathion.