Quaternary salts of 2-[(hydroxyimino)methyl]imidazole. 2. Preparation and in vitro and in vivo evaluation of 1-(alkoxymethyl)-2-[(hydroxyimino)methyl]-3-methylimida zolium halides for reactivation of organophosphorus-inhibited acetylcholinesterases

A series of structurally related mono- and bis-1,3-disubstituted 2-[(hydroxyimino)methyl]imidazolium halides were evaluated in vitro for their ability to reactivate electric eel, bovine, and human erythrocyte (RBC) acetylcholinesterases (AChE) inhibited by ethyl p-nitrophenyl methylphosphonate (EPMP) and 3,3-dimethyl-2-butyl methyl-phosphonofluoridate (soman, GD). All new compounds were characterized for (hydroxyimino)methyl acid dissociation constant, nucleophilicity, octanol-buffer partition coefficient, reversible AChE inhibition, and kinetics of reactivation of EPMP-inhibited AChEs. For GD-inhibited AChEs, maximal reactivation was used to compare compounds since rapid phosphonyl enzyme dealkylation "aging" complicated interpretation of kinetic constants. For comparison, we also evaluated three known pyridinium therapeutics, 2-PAM, HI-6, and toxogonin. In vivo evaluation in mice revealed that when selected imidazolium compounds were coadministered with atropine sulfate, they were effective in providing lifesaving protection against both GD and EPMP challenges. This was a major accomplishment in the search for effective anticholinesterase therapeutics--the synthesis and preliminary evaluation of the first new monoquaternary soman antidotes with potencies superior to 2-PAM. Significantly, there was an apparent inverse relationship between in vitro and in vivo results; the most potent in vivo compounds proved to be the poorest in vitro reactivators. These results suggested that an alternative and possibly novel antidotal mechanism of protective action may be applicable for the imidazolium aldoximes. Selected compounds were also evaluated for their inhibition of AChE phosphorylation by GD and antimuscarinic and antinicotinic receptor blocking effects.     

Synthesis of novel 6-substituted-3(2H)-pyridazinone-2-acetyl-2-(substituted/-nonsubstituted benzal)hydrazone derivatives and acetylcholinesterase and butyrylcholinesterase inhibitory activities in vitro

In this study thirteen 6-substituted-3(2H)-pyridazinone-2-acetyl-2-(substituted/nonsubstituted benzal)hydrazone V derivatives were synthesized as acetylcholinesterase and butyrylcholinesterase inhibitors. Ten of the synthesized compounds were synthesized for the first time in this study and the rest of them had been synthesized in a previous study. The structures of compounds V were elucidated by IR, 1H-NMR and MASS spectra. The acetylcholinesterase (AChE) and butyrylcholinesterase (BChe) inhibitory activities of V derivatives were measured using Ellman's method. While some of the 6-substituted-3(2H)-pyridazinone-2-propyl-3-(substituted/-nonsubstituted benzal)hydrazone V derivatives exhibited significant AChE inhibitory activity, none of the compoundsshowed BChE inhibitory activity. Theseresults indicate that V derivatives were AChE inhibitors with AChE/ BChE selectivity.     

Quaternary salts of 2-[(hydroxyimino)methyl]imidazole. 3. Synthesis and evaluation of (alkenyloxy)-, (alkynyloxy)-, and (aralkyloxy)methyl quaternarized 2-[(hydroxyimino)methyl]-1-alkylimidazolium halides as reactivators and therapy for soman intoxication

A series of structurally related monosubstituted 1-[(alkenyloxy)methyl]-, 1-[(alkynyloxy)methyl]-, and 1-[(aralkyloxy)methyl]-2-[(hydroxyimino)methyl]-3-methyli midazolium halides were prepared and evaluated. All new compounds were characterized with respect to (hydroxyimino)methyl acid dissociation constant, nucleophilicity, and octanol-buffer partition coefficient. The alkynyloxy-substituted compounds were also evaluated in vitro with respect to reversible inhibition of human erythrocyte (RBC) acetylcholinesterase (AChE) and kinetics of reactivation of human AChE inhibited by ethyl p-nitrophenyl methylphosphonate (EPMP). In vivo evaluation in mice revealed that coadministration of alkynyloxy-substituted imidazolium compounds with atropine sulfate provided significant protection against a 2 x LD50 challenge of GD. For the alkynyloxy-substituted imidazolium drugs there is a direct relationship between in vitro and in vivo activity: the most potent in vivo compounds against GD proved to be potent in vitro reactivators against EPMP-inhibited human AChE. These results differ from the observations made on the sterically hindered imidazolium compounds (see previous article) and suggest that several antidotal mechanisms of protective action may be applicable for the imidazolium aldoxime family of therapeutics. The ability of the alkynyloxy substituents to provide life-saving protection against GD intoxication was not transferable to the pyridinium or triazolium heteroaromatic ring systems.     

Apoptosis induced by β-amyloid25-35 in acetylcholinesteraseoverexpressing neuroblastoma cells

AIM: To examine the relationship between apoptosis induced by β-amyloid fragment 25-35 (Aβ25-35) and the activ-ity of acetylcholinesterase (ACHE) in AChE over-expresser-SC42 cells. METHODS: Cell survival was measuredby microscopy and MTT reduction; DNA laddering was observed by electrophoresis; AChE activity was determined by spectrophotometry. RESULTS: Aβ25-35 1 μmol/L exposure for 24-48 h caused a significant decrease in cell viability, along with changes in morphology and DNA fragmentation. AChE activity was affected in an inversemanner, increasing gradually to a level that was 1.7-fold higher than control at the 48-h time point. No change in thecytotoxicity of Aβ25-35 was observed when the increased AChE activities were effectively inhibited by huperzine Athroughout the 48-h exposure period. CONCLUSION: Although Aβ25-35 can induce apoptosis in SC42 cells andsimultaneously increase AChE activity, the capacity of AChE to hydrolyze acetylcholine is not involved in thisapoptosis model.     

Bis-(-)-nor-meptazinols as novel nanomolar cholinesterase inhibitors with high inhibitory potency on amyloid-beta aggregation

Bis-(-)-nor-meptazinols (bis-(-)-nor-MEPs) 5 were designed and synthesized by connecting two (-)-nor-MEP monomers with alkylene linkers of different lengths via the secondary amino groups. Their acetylcholinesterase (AChE) inhibitory activities were more greatly influenced by the length of the alkylene chain than butyrylcholinesterase (BChE) inhibition. The most potent nonamethylene-tethered dimer 5h exhibited low-nanomolar IC 50 values for both ChEs, having a 10 000-fold and 1500-fold increase in inhibition of AChE and BChE compared with (-)-MEP. Molecular docking elucidated that 5h simultaneously bound to the catalytic and peripheral sites in AChE via hydrophobic interactions with Trp86 and Trp286. In comparison, it folded in the large aliphatic cavity of BChE because of the absence of peripheral site and the enlargement of the active site. Furthermore, 5h and 5i markedly prevented the AChE-induced Abeta aggregation with IC 50 values of 16.6 and 5.8 microM, similar to that of propidium (IC 50 = 12.8 microM), which suggests promising disease-modifying agents for the treatment of AD patients.     

Insulin-like effect of (-)epicatechin on erythrocyte membrane acetylcholinesterase activity in type 2 diabetes mellitus

1. Changes in the activity of acetylcholinesterase (AChE) have been reported in diabetes mellitus that have been linked to certain brain defects. The erythrocyte membrane AChE is reported to be similar to AChE present in the brain. 2. Epicatechin, a member of a group of polyphenolic compounds collectively known as "catechins" that are present in tea and belong to the flavonoid family, has been reported to possess insulin-like activity. 3. In the present study, the in vitro effect of (-)epicatechin and/or insulin was tested on erythrocyte membrane AChE in normal and type 2 diabetic patients. The aim of the study was to test the efficacy of (-)epicatechin to mimic insulin in its effect on erythrocyte membrane AChE. 4. Acetylcholinesterase activity was significantly lower in type 2 diabetic patients than in normal controls and in vitro insulin treatment restored this activity to normal levels. Epicatechin (1 mmol/L) also caused an elevation in AChE activity in diabetic erythrocytes, an effect that was similar to the effect of insulin. 5. Epicatechin has a pronounced insulin-like effect on erythrocyte membrane-bound AChE in type 2 diabetic patients; however, the mechanism of action of epicatechin remains speculative.     

3-(4-[[Benzyl(methyl)amino]methyl]phenyl)-6,7-dimethoxy-2H-2-chromenone (AP2238) inhibits both acetylcholinesterase and acetylcholinesterase-induced beta-amyloid aggregation: a dual function lead for Alzheimer's disease therapy

In recent years, the investigation of acetylcholinesterase (AChE) inhibitors has gained further interest, because the involvement of the peripheral site of the enzyme in the beta-amyloid (Abeta) aggregation process has been disclosed. We present here, for the first time, a direct evidence of the Abeta antiaggregating action of an AChE inhibitor (AP2238) purposely designed to bind at both the catalytic and the peripheral sites of the human enzyme.     

Recent advances in acetylcholinesterase Inhibitors and Reactivators: an update on the patent literature (2012-2015)

Introduction: Acetylcholinesterase (AChE) is the major enzyme that hydrolyzes acetylcholine, a key neurotransmitter for synaptic transmission, into acetic acid and choline. Mild inhibition of AChE has been shown to have therapeutic relevance in Alzheimer’s disease (AD), myasthenia gravis, and glaucoma among others. In contrast, strong inhibition of AChE can lead to cholinergic poisoning. To combat this, AChE reactivators have to be developed to remove the offending AChE inhibitor, restoring acetylcholine levels to normal.

Areas covered: This article covers recent advances in the development of acetylcholinesterase modulators, including both inhibitors of acetylcholinesterase for the efforts in development of new chemical entities for treatment of AD, as well as re-activators for resurrection of organophosphate bound acetylcholinesterase.

Expert opinion: Over the past three years, research efforts have continued to identify novel small molecules as AChE inhibitors for both CNS and peripheral diseases. The more recent patent activity has focused on three AChE ligand design areas: derivatives of known AChE ligands, natural product based scaffolds and multifunctional ligands, all of which have produced some unique chemical matter with AChE inhibition activities in the mid picomolar to low micromolar ranges. New AChE inhibitors with polypharmacology or dual inhibitory activity have also emerged as highlighted by new AChE inhibitors with dual activity at L-type calcium channels, GSK-3, BACE1 and H3, although most only show low micromolar activity, thus further research is warranted. New small molecule reactivators of organophosphate-inhibited AChE have also been disclosed, which focused on the design of neutral ligands with improved pharmaceutical properties and blood-brain barrier (BBB) penetration. Gratifyingly, some research in this area is moving away from the traditional quaternary pyridinium oximes AChE reactivators, while still employing the necessary reactivation group (oximes). However, selectivity over inhibition of native AChE enzyme, effectiveness of reactivation, broad-spectrum reactivation against multiple organophosphates and reactivation of aged-enzyme continue to be hurdles for this area of research.     

Design, synthesis and evaluation of novel 2-(aminoalkyl)-isoindoline-1,3-dione derivatives as dual-binding site acetylcholinesterase inhibitors

A new series of 2-(diethylaminoalkyl)-isoindoline-1,3-dione derivatives intended as dual binding site cholinesterase inhibitors were designed using molecular modeling and evaluated as inhibitors of acetylcholinesterase (AChE), butyrylcholinesterase (BuChE), and the formation of the β-amyloid (Aβ) plaques. For AChE inhibitory activity, the spectrophotometric method of Ellman and the electrophoretically mediated microanalysis assay were used, giving good results. Most of the synthesized compounds had AChE inhibitory activity with IC(50) values ranging from IC(50) = 0.9 to 19.5 μM and weak Aβ anti-aggregation inhibitory activity. These results support the outcome of docking studies which tested compounds targeting both the catalytic active site (CAS) and the peripheral anionic site (PAS) of AChE. The most promising selective AChE inhibitors are compounds 10 (IC(50) = 1.2 μM) and 11 (IC(50) = 1.1 μM), with 6-7 methylene chains, which also inhibit Aβ fibril formation.     

Acetylcholinesterase inhibitors: a patent review (2008 - present)

INTRODUCTION: Both acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) are present in the body in large amounts. AChE is an important part of the cholinergic nervous system taking place in the central and peripheral nervous system. AChE is a target of several toxins such as insecticide carbofuran, nerve agents, sarin, soman, tabun and VX. Beside toxins, drugs for treatment of Alzheimer's disease and myasthenia gravis, such as galantamine, donepezil, rivastigmine, tacrine, huperzine, pyridostigmine and neostigmine, are known. AREAS COVERED: The review gives an overview of the importance of the cholinergic nervous system, the biochemistry of AChE and the role of AChE inhibitors. Current efforts to introduce potent drugs for Alzheimer's disease therapy and reduce toxicity, while keeping the maximal pharmacological effect, are also discussed. EXPERT OPINION: The current research effort into AChE inhibitors can be divided into two categories. First, new toxins useful for agricultural purposes and second, novel drugs that need to be prepared, although there is less interest in the new toxins. The research for drugs for Alzheimer's disease needs to focus on inhibitors that reduce the deposition of amyloid plaques, but do not initiate AChE expression.     

Two new 2-(2-Hydroxy-2-phenylethyl)chromens from agarwood originating from Aquilaria crassna

Two new 2-(2-hydroxy-2-phenylethyl)chromones (1?2), along with three known 2-(2-phenylethyl)chromones (3?5), were isolated from the agarwood originating from Aquilaria crassna Pierre ex Lecomte. Their structures were determined by the spectroscopic methods including 1D and 2D NMR analysis and comparison with reported data in the literature. All the compounds were isolated from agarwood of A. crassna for the first time. Compounds 1 and 2 exhibited inhibitory activity against acetylcholinesterase (AChE) with 17.4 ± 0.6 and 15.8 ± 0.7%, respectively, at a concentration of 50 μg/ml. Besides, Compound 3 expressed antibacterial activities against Ralstonia solanacearum with diameter of the inhibition zone of 6.80 ± 0.08 mm at a concentration of 10 mg/ml.     

3-(氨基烷氧基芳基)-7H-噻唑并[3,2-b]-1,2,4-三嗪-7-酮类化合物的设计、合成与乙酰胆碱酯酶抑制活性

目的初步探讨在7H-噻唑并[3,2-b]-1,2,4-三嗪-7-酮类化合物母核C-3位苯环侧链中引入二乙胺基团的3-(氨基烷氧基芳基)-7H-噻唑并[3,2-b]-1,2,4-三嗪-7-酮类化合物的合成及其乙酰胆碱酯酶抑制活性。方法以取代的苯甲醛和乙酰甘氨酸为初始原料,经Erlenmeyer-Plchl反应、缩合反应、水解反应、缩合反应,生成6-芳甲基-3-硫代-1,2,4-三嗪-5(2H)-酮类化合物,再与取代的α-氯代苯乙酮反应,得到6-芳甲基-3-(羟基芳基)-7H-噻唑并[3,2-b]-1,2,4-三嗪-7-酮类化合物;以芳基乙烯为原料,经温和的氧化反应、缩合反应得到3,4-二氢-6-芳基-3-硫代-1,2,4-三嗪-5(2H)-酮类化合物,再与取代的α-氯代苯乙酮在乙酸中反应得到6-芳基-3-(羟基芳基)-7H-噻唑并[3,2-b]-1,2,4-三嗪-7-酮类化合物。两条合成路线得到的3-(羟基芳基)-7H-噻唑并[3,2-b]-1,2,4-三嗪-7-酮类化合物进一步经Williamson反应制备得到10个3-(烷氧基芳基)-7H-噻唑并[3,2-b]-1,2,4-三嗪-7-酮类化合物。所有目标化合物结构均经质谱、红外光谱和核磁共振氢谱确证。采用Ellman法对目标化合物进行体外乙酰胆碱酯酶抑制活性筛选。结果根据前期已筛选化合物的活性数据和总结出的初步构效关系,设计并合成了10个C-3位苯环侧链中含有二乙胺基团的3-(氨基烷氧基芳基)-7H-噻唑并[3,2-b]-1,2,4-三嗪-7-酮类化合物。体外乙酰胆碱酯酶抑制活性筛选表明,所有目标化合物均具有乙酰胆碱酯酶抑制活性,其中7个化合物在10μmol.L-1浓度水平抑制活性超过了50%。结论根据体外重组人源AChE(rhAChE)抑制活性的测试结果,发现C-3位苯环侧链中含有二乙胺基团的7H-噻唑并[3,2-b]-1,2,4-三嗪-7-酮类化合物均具有较好的rh-AChE抑制活性。在这一位置的侧链中引入二乙胺基团,可以增强化合物对rhAChE的抑制活性。     

Design, synthesis, and structure-activity relationships of a series of 3-[2-(1-benzylpiperidin-4-yl)ethylamino]pyridazine derivatives as acetylcholinesterase inhibitors

Starting from the 3-[2-(1-benzylpiperidin-4-yl)ethylamino]-6-phenylpyridazine 1, we performed the design, the synthesis, and the structure-activity relationships of a series of pyridazine analogues acting as AChE inhibitors. Structural modifications were achieved on four different parts of compound 1 and led to the following observations: (i) introduction of a lipophilic environment in the C-5 position of the pyridazine ring is favorable for the AChE-inhibitory activity and the AChE/BuChE selectivity; (ii) substitution and various replacements of the C-6 phenyl group are possible and led to equivalent or slightly more active derivatives; (iii) isosteric replacements or modifications of the benzylpiperidine moiety are detrimental to the activity. Among all derivatives prepared, the indenopyridazine derivative 4g was found to be the more potent inhibitor with an IC(50) of 10 nM on electric eel AChE. Compared to compound 1, this represents a 12-fold increase in potency. Moreover, 3-[2-(1-benzylpiperidin-4-yl)ethylamino]-5-methyl-6-phenylpyridazine 4c, which showed an IC(50) of 21 nM, is 100-times more selective for human AChE (human BuChE/AChE ratio of 24) than the reference compound tacrine.     

Effects of atrazine and chlorpyrifos on acetylcholinesterase and Carboxylesterase in brain and muscle of common carp

The aim of this study was to investigate the effects of chlorpyrifos (CPF), atrazine (ATR) and the mixture of them on acetylcholinesterase (AChE) and carboxylesterase (CbE) in brain and muscle of common carp, respectively. 220 carps were averagely divided into 11 groups according to the different treatments and concentration, including the exposure and recovery experiments. The activities of AChE and CbE of the brain and muscle were determined at the end of the exposure and the recovery. The results showed that in the control group, the specific enzymatic activities in the brain were higher than that in the muscle. The activities of AChE and CbE in the exposure groups were significantly lower than that in the control group except for the CbE activity in the ATR low-dose group. There was a negative dose-response relationship between the activities of AChE and CbE and the pesticides concentration. The activities of AChE and CbE in the recovery groups were significantly higher than that in the exposure group except for the CbE activity in the ATR low-dose group, AChE activity in the high-dose group of the mixture of ATR and CPF, and AChE activity of the brain in the CPF high-dose group. The results suggested that: (1) brain AChE may be considered as a very sensitive and early biomarker of exposure to CPF, ATR, or the mixture of ATR and CPF, (2) brain CbE may be used as a secondary biomarker for evaluating the exposure to CPF, ATR, or the mixture of ATR and CPF and (3) the change of the AChE and CbE activities caused by the mixture of ATR and CPF was more sensitive than that caused by the ATR or CPF alone.     

合成左旋石杉碱甲对胆碱酯酶活性和小鼠水迷宫操作的作用

目的：比较合成和天然礤旋右杉碱甲（Ｈｕｐ Ａ）对胆碱酯酶的抑制作用以及对东莨菪碱所致记忆障碍的改善作用。方法：比色法用于测定胆碱酯酶活性，小鼠水迷宫用于评价促智作用。结果：合成ＨｕｐＡ抑制大鼠皮层和红细胞膜乙酰胆碱脂酶的ＩＣ５０值分别为６４．７（５２．６－７９．５），５３．９（４３．６－６６．６）ｎｍｏｌ·Ｌ＾－１，抑制大鼠血清丁酰胆碱酯酶的ＩＣ５０为５３．６（４４．９－０．４８）μｍｏｌ·Ｌ＾－     

含氧叔丁基三唑醇类化合物的合成及其体外抗真菌活性

目的设计合成含氧取代的叔丁基三唑醇类化合物并研究其体外抗真菌活性。方法以一氯频那酮为起始原料经多步反应合成目标化合物,化合物结构经1H-NMR谱、IR谱确证;选择8种真菌为实验菌株,按国际标准抗真菌敏感性实验方法测定体外抑菌活性。结果与结论合成了12个新化合物。所有目标化合物对8种真菌均具有一定的抑制作用。可以将现有的三唑醇类抗真菌药物结构中的2,4-二氟苯基替换成其他疏水性基团来设计抗真菌化合物。     

Role of B-esterases in assessing toxicity of organophosphorus (chlorpyrifos, malathion) and carbamate (carbofuran) pesticides to Daphnia magna

In this study, the cladoceran Daphnia magna was exposed to two model organophosphorous and one carbamate pesticides including malathion, chlorpyrifos and carbofuran to assess acetylcholinesterase (AChE) and carboxylesterase (CbE) inhibition and recovery patterns and relate those responses with individual level effects. Our results revealed differences in enzyme inhibition and recovery patterns among the studied esterase enzymes and pesticides. CbE was more sensitive to organophosphorous than AChE, whereas both CbE and AChE showed equivalent sensitivities to the carbamate carbofuran. Recovery patterns of AChE and CbE activities following exposure to the studied pesticides were similar with 80-100% recoveries taking place 12 and 96 h after exposure to organophosphorous and carbamates pesticides, respectively. The physiological role of AChE and CbE inhibition patterns in Daphnia was examined by using organophosphorous and carbamate compounds alone and with specific inhibitors of CbE. Under exposure to organophosphorous pesticides, survival of Daphnia juveniles was impaired at AChE inhibition levels higher than 50% whereas under exposure to the carbamate carbofuran low levels of AChE inhibition affected mortality. Inhibition of CbE by 80-90% increased toxicity to organophosphorous and carbamate pesticides by up to two- and four-fold, respectively. Our results suggest that both AChE and CbE enzymes are involved in determining toxicity of Daphnia to the studied chemicals and that AChE inhibition levels higher than 50% can be considered of environmental concern to Daphnia species.     

Antioxidant and antiacetylcholinesterase activities of Sorbus torminalis (L.) Crantz (wild service tree) fruits

In this study, the antioxidant and antiacetylcholinesterase activities of Sorbus torminalis (L.) Crantz fruits were evaluated. Total phenolic and flavonoid compounds, 2,2′-azino-bis (3-ethylbenzothioazoline-6-sulfonic acid), 2,2′-diphenyl-1-picrylhydrazyl, and superoxide anion radicals scavenging activities and ferric-reducing antioxidant power of water, ethyl acetate, acetone, and methanol extracts were determined for the measurement of the antioxidant activity. Quercetin and α-tocopherol were used as standard antioxidants. The inhibitory effect of the water extract on acetylcholinesterase (AChE) was evaluated using the Ellman method and galantamine was used as a standard. Water extract had the highest total phenolic concentration and the strongest antioxidant activity followed by ethyl acetate and acetone extracts whereas methanol extract has the lowest phenolics and weakest antioxidant activity. Moreover, water extract showed moderate ability to inhibit AChE. It was concluded that fruits of S. torminalis have antioxidant and anti-AChE activities and that the plant might be a natural source of antioxidants and AChE inhibitors.     

Roles of uptake, biotransformation, and target site sensitivity in determining the differential toxicity of chlorpyrifos to second to fourth instar Chironomous riparius (Meigen)

Early life stages of aquatic organisms tend to be more sensitive to various chemical contaminants than later life stages. This research attempted to identify the key biological factors that determined sensitivity differences among life stages of the aquatic insect Chironomous riparius. Specifically, second to fourth instar larvae were exposed in vivo to both low and high waterborne concentrations of chlorpyrifos to examine differences in accumulation rates, chlorpyrifos biotransformation, and overall sensitivity among instars. In vitro acetylcholinesterase (AChE) assays were performed with chlorpyrifos and the metabolite, chlorpyrifos-oxon, to investigate potential target site sensitivity differences among instars. Earlier instars accumulated chlorpyrifos more rapidly than later instars. There were no major differences among instars in the biotransformation rates of chlorpyrifos to the more polar metabolites, chlorpyrifos-oxon, and chlorpyridinol (TCP). Homogenate AChE activities from second to fourth instar larvae were refractory to chlorpyrifos, even at high concentrations. In contrast, homogenate AChE activities were responsive in a dose-dependent manner to chlorpyrifos-oxon. In general, it appeared that chlorpyrifos sensitivity differences among second to fourth instar C. riparius were largely determined by differences in uptake rates. In terms of AChE depression, fourth instar homogenates were more sensitive to chlorpyrifos and chlorpyrifos-oxon than earlier instars. However, basal AChE activity in fourth instar larvae was significantly higher than basal AChE activity in second to third instar larvae, which could potentially offset the apparent increased sensitivity to the oxon.     

Differential sensitivity of blood, peripheral, and central cholinesterases in beagle dogs following dietary exposure to chlorpyrifos

Two studies were performed to find out whether exposure limits that protect brain acetylcholinesterase (AChE) will protect peripheral tissue AChE after exposure to chlorpyrifos (CPF), an organophosphate insecticide. In a methods-development study, male dogs (3/dose) were exposed to 0.0, 0.3, 0.6, or 1.2mg/kg/day CPF in their diets for 4 weeks. Mixed cholinesterase (mChE), AChE, and butyrylcholinesterase (BuChE) activities were measured in plasma, RBC, brain, left atrium and ventricle, diaphragm, quadriceps, and nodose ganglia. Plasma, brain and peripheral tissue BuChE was inhibited at all dose levels. While RBC AChE was inhibited at all doses, brain and peripheral AChE activities were unaffected. In the main study, dogs (4/sex/dose) were exposed to 0.0, 0.5, 1.0, or 2.0mg/kg/day CPF in their diets for six weeks and RBC AChE was significantly inhibited at all doses in both sexes. Diaphragm, quadriceps, and nodose ganglia AChE was unaffected by treatment. Brain AChE was decreased by approximately 6% compared to controls in high-dose groups, and this was considered a threshold effect. Left atrium AChE in high-dose dogs was 25.5% less (males) and 32.1% greater (females) than controls; these differences were attributed to chance. While peripheral tissue and brain AChE were not affected following exposure to 1.0mg/kg/day, RBC AChE was inhibited at all doses. These results show that RBC AChE is more sensitive than brain or peripheral tissue AChE to inhibition by CPF, and that protection of brain AChE would protect peripheral tissue AChE.