Inhibition Of Pseudocholinesterase Research Articles

Urine Carbamate Determination Compared with Plasma Pseudocholinesterase for Diagnosis of Insecticides Poisoning

Abstract 1.1.Introduction:Diagnosis of carbamate poisoning is frequently missed especially when patients present after the recovery of pseudocholinesterase inhibition. Confusion is due to the similarity of the clinical picture with that of food poisoning and other infectious syndromes. 1.2.Patients and methods:Patients (n=40) admitted within 4 hours of carbamate intoxication were included in the study. They were graded according to Poison severity score PSS. Normal volunteers (n=10) were included and served as negative control group. Normal subjects and all patients were tested for plasma pseudocholinesterase (PsChE) within 4 hours and 18 hours from carbamates exposure. Urine carbamate concentration was determined on the 18th hour and 2 days after exposure. Different PSS groups and normal volunteers were compared for their plasma PsChE and urine carbamate levels using student t test and correlation coefficient Method used for PsChE determination is the kinetic assay method. Urine carbamate concentrations were determined by spectrophotometry using modification of the method of Tamrakar. Carbamate determination method was tested for linearity, precision, recovery and limit of detection of the carbamates 1.3.Results:PsChE was significantly lower in PSS II compared to PSS I. No significant difference in PsChE was noticed between both patient’s groups and normal volunteers on the 18th hour from carbamate exposure. Urine carbamate concentration was significantly higher in PSS II compared to PSS I on the 18th hour and 2 days after exposure. No correlation was detected between the maximum urine carbamate concentration and plasma pseudocholinesterase. The method specificity increased after modification from 76.9 to 91%. Recovery ranged between 95 and 98%. Limit of detection is 0.1µg/ml and limit of quantification is 0.2 µg/ml. The technique is highly reproducible with inter-day precision RSD less than 8%. 1.4. Conclusion: A better evaluation and diagnosis of carbamate poisoning patients with delayed presentation can be achieved using urine carbamate determination rather than plasma pseudocholinesterase which is often normalized at the time of presentation.

Evaluation of acetylcholine esterase activity in the blood of workers exposed

Introduction:Organophosphate and carbamate insecticides pose major environmental pollution problems and health hazards to people and animals. These insecticides inhibit cholinesterase (ChE) activity in the nervous tissues and neuromuscular junctions. The measurement of blood ChE is a useful tool for monitoring exposure to organophosphate and carbamate insecticides. The purpose of the present study was to use a modified electrometric technique for measuring blood ChE in workers exposed to the organophosphate and carbamate insecticides in Kirkuk, Iraq. Method: A modified electrometric method was used to measure ChE activity in the whole blood of male workers (n = 40) exposed to organophosphate and carbamate insecticides, for a duration of not less than six years. Healthy male volunteers (n =12) not exposed to insecticides served as controls. Following in vitro inhibition of pseudo cholinesterase by quinidine sulfate, true cholinesterase activity was estimated in the blood of the subjects. After in vitro addition of the organophosphate (chlorpyrifos and methidathion, 0.5 and 1 µM) and carbamate (carbaryl, 5 and 10 µM) insecticides to the reaction mixtures, inhibitions of blood ChE were also determined. Results: Mean values of ChE activities (pH/20 min) in the whole blood of healthy non-exposed subjects and insecticide-exposed workers were 1.41and 1.2, respectively. Whole blood ChE activities of the exposed workers was significantly lower than those of healthy individuals. Conclusions: These findings indicate the usefulness of the modified electrometric method for monitoring blood ChE activity in insecticide-exposed workers and there was a significant effect of these Organophosphate and carbamate insecticides on the activity of Ach esterase in workers blood. .

Successful use of rocuronium and sugammadex in a patient with myasthenia

Successful use of rocuronium and sugammadex in a patient with myasthenia

Electrometric measurement of plasma, erythrocyte, and whole blood cholinesterase activities in healthy human volunteers

The measurement of blood cholinesterase activity is a useful tool for monitoring exposure to organophosphate and carbamate insecticides. Blood cholinesterase activity is measured colorimetrically or electrometrically. Recently, a simple and practical electrometric method has been described and validated for measuring blood cholinesterase activity in people and animals. The purpose of the present report was to use the modified electrometric technique for measuring blood (plasma, erythrocyte and whole blood) cholinesterase activities in apparently healthy human volunteers in Mosul, Iraq. Cholinesterase activities in the plasma, erythrocytes, and whole blood of healthy male (n = 72) and female (n = 31) volunteers were measured by an electrometric method; the method involved the addition of 0.2 ml of blood sample to 3 ml of distilled water followed by 3 ml of barbital-phosphate buffer solution (pH 8.1). The pH (pH1) of the mixture was measured, and then 0.1 ml of 7.5% of acetylcholine iodide, as a substrate, was added. The reaction mixture was incubated at 37 degrees C for 20 minutes. The pH (pH2) of the reaction mixture was measured after the end of the incubation period. Enzyme activity was expressed as DeltapH/20 min = pH1- pH2 - (DeltapH of the blank). The blank was without the blood sample. Following in vitro inhibition of pseudo cholinesterase by quinidine sulfate, true cholinesterase activity was estimated in the plasma of the subjects. After in vitro addition of the organophosphate (chlorpyrifos and methidathion, 0.5 and 1 microM) and carbamate (carbaryl, 5 and 10 microM) insecticides to the reaction mixtures, inhibitions of blood cholinesterases were measured. Mean reference cholinesterase activities (DeltapH/20 min) in the plasma, erythrocytes, and whole blood of male subjects were 0.98, 1.39, and 1.41, respectively. Females were 0.85, 1.22, and 1.23, respectively. Ten minutes after in vitro addition of quinidine sulfate to inhibit pseudo cholinesterase activity in the plasma, the estimated true cholinesterase activities in males and females were 0.08 and 0.07 DeltapH/20 min, respectively. The percentage of true cholinesterase in the plasma of males and females was 8.2. Using the modified electrometric method, various percentages of cholinesterase inhibitions in the plasma, erythrocytes, and whole blood were detected after in vitro addition of the organophosphate insecticides (chlorpyrifos and methidathion) and the carbamate insecticide (carbaryl) to the reaction mixtures. These findings are the first collective report of human plasma, erythrocyte, and whole blood cholinesterase activities as determined by the modified electrometric method, and they could serve as reference points for future studies that involve human exposure to anticholinesterase pesticides.

Temperature, the benzoylcholine substrate, and fluoride inhibition of pseudocholinesterase

To the Editor: The magnitude of sevoflurane’s (and enflurane’s) effect on fluoride inhibition of pseudocholinesterase (PCHE) deserves attention in modern anesthetic practice. Nearly all studies dealing with this topic1–3 refer to the study of Kambam et al.4 who reported an inhibition of PCHE between 16 to 73% with 10 to 100 μmol·L–1 fluoride, the concentrations usually obtained during sevoflurane (and enflurane) anesthesia. Since we could not confirm this inhibition with mivacurium and butyrylthiocholine as substrates and at temperatures between 28 to 37°C3,5 we investigated fluoride inhibition of PCHE with benzoylcholine at 25°C, the substrate and temperature Kambam et al. used in their study, and at 37°C, the temperature currently recommended for determination of enzymatic activity. After approval by our Local Ethics Committee and with written informed consent, serum samples of eight healthy volunteers were obtained, spiked with fluoride (0, 10, 50 and 100 μmol·L–1) and adjusted to 25°C and 37°C, respectively. Thereafter, 50 μmol·L–1 benzoylcholine were added, and at timed interval aliquots were withdrawn to measure the concentrations of benzoylcholine by high-performance liquid chromatography. After calculating the half-lives with KINETICA 2000 (InnaPhase Corp. Philadelphia, PA, USA) a one-factor (fluoride concentration) analysis of variance with a post hoc Dunnett test was performed at each temperature, using the fluoride-free sample as the control category. The results are presented in the Figure. At 25°C there was an increase of the benzoylcholine half-time of 9.3% with 10 μmol·L–1 fluoride, and a 36% increase with 50 μmol·L–1 fluoride. The 68% increase in benzoylcholine half-time was significant (P < 0.05) at a concentration of 100 μmol·L–1. In contrast, no change was observed over the same range of fluoride concentrations at 37°C. We thus confirm one aspect of the study of Kambam et al.4 who measured PCHE activity at 25°C with a photometric assay. In contrast, when using a high-performance liquid chromatography assay at clinically relevant body temperature (37°C) and with fluoride concentrations up to 100 μmol·L–1, fluoride inhibition of PCHE is not apparent, whether using the substrates mivacurium and butyrylthiocholine3,5 or benzoylcholine, as reported here. In conclusion, the influence of substrate and temperature on fluoride inhibition of PCHE must be taken into consideration when interpreting the results of investigations with sevoflurane and enflurane.

Anticholinesterase activity in an alkaloid extract of Huperzia saururus

Huperzia saururus (Lam.) Trevis. (Lycopodiaceae) is used widely in Argentinian traditional medicine as an aphrodisiac and for memory improvement. An aqueous extract from the aerial parts was obtained by decoction, revealing the presence of alkaloids, among other constituents. By partition with organic solvent in alkaline media, alkaloids were extracted and then purified by gel permeation. We studied the anticholinesterase activity in vitro of the alkaloid extract using erythrocyte membranes and human serum as sources of acetylcholinesterase and pseudocholinesterase, respectively. The results show a marked inhibition of true acetylcholinesterase with an IC50 value of 0.58 microg/ml. Low inhibition of pseudocholinesterase was observed (IC50 value = 191 microg/ml). This shows a selectivity of the extract for the true acetylcholinesterase. Furthermore, chemical study of the bioactive extract was performed by GC-MS, revealing the presence of seven Lycopodium alkaloids, including some not identified previously: sauroxine, 6-hydroxylycopodine, N-acetyllycodine, lycopodine, lycodine, N-methyllycodine, and clavolonine. Further investigations will be undertaken in order to discover which compound/s are responsible for the aqueous extract's acetylcholinesterase activity.

Effects of Edrophonium and/or Pseudocholinesterase for the Reversal of Mivacurium-Induced Paralysis in vitro

Background: Mivacurium is a nondepolarizing neuromuscular blocking agent hydrolyzed by pseudocholinesterase. Anticholinesterase used in the reversal of mivacurium-induced muscle relaxation may also inhibit plasma pseudocholinesterase, and delay hydrolysis of mivacurium. In this study, the effects of edrophonium and/or bovine pseudocholinesterase (BpChE) in the reversal of mivacurium were investigated with the rat phrenic nerve-diaphragm preparation. Methods: Fifty Sprague-Dawley rats (150 - 200 g) were randomly allocated into 10 groups based on the dosage of edrophonium and BpChE. Each animal was anesthetized with thiopental sodium (40 mg/kg J.P.). The phrenic nerve-diaphragm was dissected and mounted in a bath containing an oxygenated Krebs' solution at 32. The phrenic nerve was stimulated at supramaximal intensity and the single twitch responses and train of four (TOF) ratio were measured. After stabilization of the twitch responses, mivacurium (1 /ml) was administered incrementally to obtain more than 95% twitch inhibition. Reversal of the mivacurium-induced block by edrophonium (0.01, 0.1, 1, or 10 ,/ml) and/or BpChE (0.1 u, or 1.0 u/ml) were tested. A single twitch height more than 75% of the baseline value was considered an adequate reversal. Results: Mivacurium-induced paralysis was recovered more effectively by BpChE 1.0 u/ml than the other groups. Edrophonium improved a single twitch in a dose dependent manner. Conclusions: Mivacurium-induced paralysis can be more effectively reversed by BpChE than edrophonium. Inhibition of pseudocholinesterase was not observed by increasing the dose of edrophonium.

Prolonged succinylcholine-induced paralysis in organophosphate insecticide poisoning

A case of prolonged succinylcholine-induced paralysis in a child with organophosphate insecticide poisoning is presented. Three hours and 15 minutes of apnea after the administration of succinylcholine was attributed to a decreased rate of succinylcholine metabolism from inhibition of pseudocholinesterase by the insecticide. Only one similar case has been reported previously in the English medical literature. If succinylcholine is to be used in patients with organophosphate poisoning, a prolonged paralysis should be anticipated.

Influence of blood proteins on biomedical analysis. VII. Electrophoretic analysis of the interaction of pseudocholinesterase with fatty acid and/or human serum albumin.

In order to characterize the fatty acid (FA)-induced inhibition of pseudocholinesterase (pchE), the interaction of pchE with FA (n-capric acid and n-lauric acid) and/or human serum albumin (HSA) was studied by electrophoreses on a cellulose acetate membrane and a polyacrylamide gel thin-layer. It was demonstrated that pchE binds with n-[1-14C] lauric acid, suggesting that the FA-induced inhibition of pchE activity is based on the interaction of the enzyme with FA. The binding of pchE with n-[1-14C] lauric acid was depressed by HSA. Consequently, the present results suggest that the reduction of FA-induced inhibition of pchE activity by HSA is associated closely with the depression of pchE-FA binding by HSA. Moreover, no selectivity in the n-capric acid-induced inhibition of pchE isozymes was found in the present study, i. e., all isozymes (C1-C4) of pchE were equally inhibited by n-capric acid.

Effect of para substituents of 2-(N,N-dimethylamino)ethyl phenyl phenylphosphonates on inhibition of human pseudocholinesterase.

Human serum pseudocholinesterase inhibition by nineteen of the title phosphonates was studied and the effect of para substituents of the phenoxy and phenyl groups on inhibition rates was compared. The effect of substituents of the phenoxy group on the reaction rate is in some cases even 60 times as great as the effect of substituents of the phenyl group. Logarithms of rate constants correlate linearly with sigma po substituent constants of the phenoxy group, but no linear dependence is observed for para substituents of the phenyl group. The role of the apicophilicity of substituents of phosphorus in the enzyme--inhibitor complex is discussed.

Comparative Study of the Autonomic Innervation of the Mammalian Ovary, with Particular Regard to the Follicular System

The autonomic innervation of the ovary was studied in 12 mammalian species utilizing the cholinesterase method in combination with pseudocholinesterase inhibition for the cholinergic component, and glyoxylic acid histochemistry together with fluorometric determination of noradrenaline for the adrenergic component. Ovaries from cow, sheep, cat, and guinea pig were very richly supplied with adrenergic nerves in the cortical stroma, particularly enclosing follicles in various stages of development. In the follicular wall the nerve terminals were located in the theca externa, where they ran parallel to the follicular surface. Numerous adrenergic terminals also surrounded ovarian blood vessels. The adrenergic innervation was of intermediary density in the human ovary and in the pig, dog, cat, and opossum. Ovaries from rabbit, mouse and hamster had a sparse adrenergic nerve supply. The amount of intraovarian adrenergic nerves agreed well with the tissue concentration of noradrenaline in the various species. The cholinergic innervation was generally less well developed, but had the same distribution as the adrenergic system around blood vessels and in the ovarian stroma, including follicular walls.

Histochemical study on regional differences in the cholinergic nerve supply of the choroid plexus from various laboratory animals

The cholinergic nerve supply of the mammalian choroid plexus was investigated by histochemical demonstration of acetylcholinesterase following inhibition of pseudocholinesterase. Material from mouse, rat, hamster, guinea pig, rabbit, cat, dog, pig, cow, and baboon was included in the study. Positive staining was found in all animals except the hamster. The plexuses of all ventricles received parasympathetic cholinergic nerves, which were associated not only with the blood vessels but also with the plexus epithelium. Usually, the plexus of the third ventricle contained somewhat more nerves than those of the lateral ventricles, which were better supplied than the plexus of the fourth ventricle. The most well-developed cholinergic innervation was found in the plexus tissue from pig and cat, whereas mouse and dog plexuses received comparatively few nerve fibers. The choroid plexus of the other animals contained a cholinergic nerve plexus of intermediary density.

Correlation of Pseudocholinesterase Inhibition and Plant Growth Retardation by Quaternary Ammonium Derivatives of (+)-Limonene

THE terpene hydrocarbon (+)-limonene is widely distributed in nature and comprises more than 95 per cent of distilled citrus peel oils. As part of a study of limonene derivatives having possible biological activity, I reported the synthesis of a number of quaternary ammonium compounds which are plant growth retardants1,2. Several of these derivatives also inhibited human blood serum (pseudo) cholinesterase in tests for drug activity (my unpublished results). Here I report the discovery of a direct correlation between serum cholinesterase inhibition by these compounds and their ability to retard plant growth.

Measurement of hydrolysis of acetylthiocholine with the silver thiol electrode. Its use in the study of serum pseudocholinesterase inhibition

A micro method is described for the measurement with a silver thiol electrode of the rate of hydrolysis by serum pseudocholinesterase of acetylthiocholine; 0.005-ml samples of serum are adequate and the method may be used to determine the degree of inhibition of the esterase by inhibitors such as dibucaine, fluoride, and solanine. Because of the simplicity of the measuring system the method could be easily automated.

The function of true and pseudocholinesterase in the mammalian ileum

The responses of the mammalian intestine to nervously released and externally applied ACh was tested when either true cholinesterase (acetylcholinesterase) or pseudocholinesterase activity was inhibited. Nicotine was used to cause ACh release from nerve endings while ACh added to the bath represented ACh of non-nervous origin. The specific cholinesterase inhibitors used were BW 284C51 for true cholinesterase inhibition and isoOMPA for pseudocholinesterase inhibition. When true cholinesterase was inhibited in the rat, rabbit or guinea-pig, the responses to both nervously released ACh and ACh of non-nervous origin were potentiated; the former (nicotine responses) being much more increased than the latter. Pseudo-cholinesterase inhibition in the rat or guinea-pig potentiated responses to externally applied ACh only. However, the rabbit differed in that the action of both nervously released and added ACh were potentiated after pseudocholinesterase inhibition. Control experiments were done to eliminate possible direct effects of the inhibitors on the sensitivity of the preparation. It is concluded that true cholinesterase normally functions in the removal of both nervous and non-nervous ACh, while pseudocholinesterase, in the rat and guinea-pig, destroys only ACh of non-nervous origin.

The effect of selective inhibition of pseudocholinesterase on the spontaneous and evoked activity of the cat's cerebral cortex.

The Journal of PhysiologyVolume 136, Issue 1 p. 20-40 ArticleFree Access The effect of selective inhibition of pseudocholinesterase on the spontaneous and evoked activity of the cat's cerebral cortex J. E. Desmedt, J. E. DesmedtSearch for more papers by this authorG. La Grutta, G. La GruttaSearch for more papers by this author J. E. Desmedt, J. E. DesmedtSearch for more papers by this authorG. La Grutta, G. La GruttaSearch for more papers by this author First published: 03 April 1957 https://doi.org/10.1113/jphysiol.1957.sp005741Citations: 67AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinkedInRedditWechat Citing Literature Volume136, Issue1April 3, 1957Pages 20-40 RelatedInformation

Influence of vitamin-E on liveresterase and cholinesterase

1. 1. a- dl-tocopheryl phosphate (TPh) added in vitro inhibits crude rat liveresterase, purified horse liveresterase, horse serum pseudo cholinesterase and ox nucleus caudatus true cholinesterase. 2. 2. The percentage inhibition at a given concentration depends largely on the amount of enzyme preparation used. 3. 3. The inhibition of pseudo cholinesterase by TPh is difficult to reverse but a certain amount of reversion can be obtained by dialysis against distilled water during 7 days and by the addition of Tween 8o. 4. 4. The inhibition of pseudo cholinesterase and true cholinesterase by TPh is not (exclusively) due to the removal of Ca ++ions. 5. 5. The action of TPh is identical with that of the anionic detergents Na-lauryl sulphate and Na-cetyl sulphate. 6. 6. TPh, Na-lauryl sulphate and Na-cetyl sulphate are probably bound to the pseudo cholinesterase preparation in equal amounts, whereas about 15,000 times less of physostigmine is bound. 7. 7. a- dl-tocopherol added as an emulsion in vitro to liver homogenates of normal and E-deficient rats has no effect on liveresterase activity. 8. 8. It is concluded that TPh acts as an anionic detergents and that the in vitro effects bears no relation to the physiological action of vitamin E.

Reversibility of the inhibition of true cholinesterase by physostigmine

1. 1. The kinetics of a true cholinesterase preparation from rat brain were studied. Acetyl-β-methylcholine (amechol) was used as a substrate. It was found that, at a constant concentration of enzyme, the reaction is of a unimolecular type when the concentration of substrate is lower than 0.01 M; when the substrate concentration rises above 0.01 M the reaction assumes a zero order character. 2. 2. When the reaction velocity is determined in presence of physostigmine, it is found that a progressive inhibition takes place during the test, which is slowed down, but never reversed by the substrate amechol. 3. 3. When the enzyme is incubated for varying periods, prior to the activity test, with the same low concentration of physostigmine, it is found that the degree of inhibition increases with the duration of the incubation period. It is found that also under these conditions, in presence of a low concentration of physostigmine, the inhibition is progressive and is not reversed by the substrate, when this is added after the incubation period. In these experiments amechol was used as a substrate. 4. 4. Dialysis reverses the inhibition by physostigmine of true cholinesterase towards acetylcholine in 4 hours. The reversal of the inhibition of pseudo cholinesterase appears to proceed more slowly. 5. 5. Washing out of rat brain suspensions previously inhibited with physostigmine does not cause reversal of the inhibition. 6. 6. It is not possible to reverse part of the inhibition acutely by means of dilution after ten minutes of incubation. 7. 7. It is concluded that no competitive reversal by substrate of physostigmine inhibition takes place, but that it is possible to reverse this inhibition by dialysis for several hours. Within shorter observation periods no reversal is demonstrable by dialysis or dilution using acetylcholine as a substrate except after very short periods of incubation. 8. 8. It is suggested that, as a result of incubation with physostigmine, two types of inhibition of cholinesterase may occur simultaneously; one towards amechol, the other towards acetylcholine. The physostigmine inhibition probably affects two different active groups, both present in our preparation of true cholinesterase.