Crude extract of ChE from the liver of Puntius javanicus was purified using procainamide-sepharyl 6B. S-Butyrylthiocholine iodide (BTC) was selected as the specific synthetic substrate for this assay with the highest maximal velocity and lowest biomolecular constant at 53.49 µmole/min/mg and 0.23 mM, respectively, with catalytic efficiency ratio of 0.23. The optimum parameter was obtained at pH 7.5 and optimal temperature in the range of 25 to 30°C. The effect of different storage condition was assessed where ChE activity was significantly decreased after 9 days of storage at room temperature. However, ChE activity showed no significant difference when stored at 4.0, 0, and -25°C for 15 days. Screening of heavy metals shows that chromium, copper, and mercury strongly inhibited P. javanicus ChE by lowering the activity below 50%, while several pairwise combination of metal ions exhibited synergistic inhibiting effects on the enzyme which is greater than single exposure especially chromium, copper, and mercury. The results showed that P. javanicus ChE has the potential to be used as a biosensor for the detection of metal ions.
A bnormal variants of plasma cholinesterase (ChE, EC. 3.1.1.8) are a rarity in this region and to date there is no reported case of suxamethonium sensitivity in the Malaysian population. We now report a case of a Malaysian Indian patient who received suxamethonium, developed prolonged apnoea and on investigation was found to be a homozygote for the silent gene. His family was screened for abnormal variants of plasma cholinesterase. The results are discussed.
The spectrophotometric method of Ellman was used to determine cholinesterase (ChE) levels in plasma and whole blood in elderly patients and umbilical cord blood of newborn infants. The mean +/- SD for plasma and whole blood ChE levels were 2.24 +/- 0.58 micromol/min/ml and 4.38 +/- 0.65 micromol/min/ml respectively in a group of healthy elderly patients (n = 25). The mean +/- SD for plasma and whole blood ChE levels were 2.23 +/- 0.77 and 3.31 +/- 0.56 respectively in cord blood of healthy full term newborn infants (n = 25). In a group of healthy adult blood donors the general mean +/- SD for plasma and whole blood ChE have been shown to be 2.71 +/- 0.75 and 4.87 +/- 0.73 micromol/min/ml respectively. Statistical analysis revealed that the mean levels of plasma and whole blood ChE in elderly, newborn infants and adults were different (p < 0.001). However, there was no statistical difference between the mean level of plasma ChE in elderly and cord blood but each shared statistically significant difference when compared to the mean level in adults. The mean activity of whole blood ChE in the 3 groups was different from one another. Theoretically, decreased levels of ChE would suggest a careful assessment of drugs that are hydrolysed by ChE.
Data on blood cholinesterase levels in the Malaysian population is lacking. The spectrophotometric method of Ellman was used to determine the red cell, plasma and whole blood cholinesterase (ChE) levels in 407 Malaysian blood donors. The mean+1SD for plasma ChE in females (n = 48) was 2.37 + 0.70 umol/min/ml and 2.76 + 0.75 umol/min/ml in males (n = 359). The mean plasma ChE in males was higher than in females (p < 0.001). The mean+1SD for red cell ChE in females was 9.01 + 1.20 umol/min/ml whereas in males it was 7.69 +1.30 umol/min/ml (the mean red cell ChE in females was higher than in males, p < 0.0001). The mean+1SD for whole blood ChE for females was 4.31+ 0.58 umol/min/ml and for males it was 4.95 + 0.71 umol/min/ml. The mean whole blood ChE in males was higher than in females (p < 0.0001). Sex influenced the plasma, red cell and whole blood ChE. In males the plasma ChE was affected by the race factor. The mean+1SD plasma ChE for the Malay, Chinese and Indian were 2.92 + 0.80, 2.73 + 0.71 and 2.61+ 0.73 respectively (p < 0.002). The age factor in males affected the red cell ChE with 7.88 + 1.32 in the (30-69) age group and 7.47 + 1.23 in the (15-29) age group (p < 0.005). The whole blood ChE in females was affected by blood groups. The mean+1SD whole blood ChE for blood groups A,B and O were 4.19 + 0.42, 3.93 + 0.46 and 4.49 + 0.62 respectively (p < 0.03). The significant difference is between the ChE of group B and O, but the ChE of group A could not be determined to be different from group B or O. These results serve as guidelines for our local population in the evaluation of cholinesterase levels with regard to pesticide poisoning, liver biosynthetic capacity and unusual sensitivity to succinylcholine.
The plasma cholinesterase baseline levels in Malaysian workers are within the guidelines used. There is a need to carry out close monitoring of workers handling organophosphate s even in a well-run, modern factory where there are no hazards in the plant operation. Abnormal results are mainly because many of our workers are unaware of the health hazards and do not follow orders properly, ie personal hygiene, such as washing of hands before eating, etc. The use of temporary workers should be discouraged because of poor compliance with rules and difficulty in follow up. Product stewardship and education of farmers, sprayers and end users are of paramount importance. Finally there is a great need for educaiton of everyone from factory workers, farmers, sprayers to the general public.
The aim of this study was to find out the indicator as a marker of person who are exposed to pesticides (organophospate). Up to now the serum cholinesterase is used as a marker. This new method will not taking blood vein since this be the obstacle in conducting research in a rural area.
In this study whole blood cholinesterase activities were determined (tintometric method) of agricultural pesticide users exposed to organophosphorus compounds in Indonesia, Malaysia, Sri Lanka and Thailand. Analysis of the data demonstrated a universal relationship between cholinesterase levels and the time between the last exposure to organophosphorus compounds and the day of blood collection for cholinesterase determination. It is suggested that it takes approximately 5 days before whole blood cholinesterase levels revert to their normal values. No relationship between whole blood cholinesterase levels and the sex of pesticide users was demonstrated except in Sri Lanka where female sprayers had lower values. It is suggested that these lower values are associated with the anaemic status of female agricultural workers. No relationship between whole blood cholinesterase level and age was demonstrated. The measurement of pre-exposure cholinesterase values is essential for comparison of values after pesticide application.
Abnormal variants of plasma cholinesterase are a rarity in this region and to date there is only one reported case of suxamethonium sensitivity in a Malaysian population. We now report a case of a Malaysian Chinese patient who received suxamethonium, developed prolonged apnoea and on investigation found to be a homozygote for the silent gene. His family was screened for abnormal variants of plasma cholinesterase. The results are discussed.
Surveys in a number of European and American populations have found the frequency of occurrence of the heterozygotes for the gene for the dibucaine-resistant variant of cholinesterase (E.C.3.1.1.8) to be relatively constant. Similar surveys in Oriental population have shown low incidence of the same gene. This study done on the multi-racial population consisting of 3 major groups shows an absence of the gene for the dibucaine resistant variant of cholinesterase. This is supported by the clinical experience in the use of suxamethonium as a single dose in more than 25,000 individuals.
Heavy metals with high chemical activity from sludge and waste release, agriculture, and
mining activity are a major concern. They should be carefully managed before reaching the
main water bodies. Excessive exposure to heavy metal may cause toxic effect to any types of
organism from the biomolecular to the physiological level, and ultimately cause death. Monitoring is the best technique to ensure the safety of our environment before a rehabilitation is
needed. Nowadays, enzyme-based biosensors are utilised in biomonitoring programmes as
this technique allows for a real-time detection and rapid result. It is also inexpensive and easy
to handle. Enzyme-based biosensors are an alternative for the preliminary screening of
contamination before a secondary screening is performed using high-performance technology.
This review highlights the current knowledge on enzyme-based biosensors, focusing on
cholinesterase for toxic metal detection in the environment.
Paddy (unmilled rice), milled rice and maize-bound 14C residues were prepared using 14C-succinate-labelled malathion at 10 and 152 ppm. After 3 months, the bound residues accounted for 12%, 6.5% and 17.7% of the applied dose in paddy, milled rice and maize respectively in the grains treated at 10 ppm. The corresponding values for the 152 ppm were 16.6%, 8.5% and 18.8%. Rats fed milled rice - bound 14C-residues eliminated 61% of the 14C in the faeces and 28% in the urine. The corresponding percentages for paddy and maize were 72%, 9% and 53%, 41% respectively; indicating that bound residues from milled rice and maize were moderately bioavailable. When rice-bound malathion residues (0.65 ppm in feed) were administered to rats in a 5 week feeding study, no signs of toxicity were observed. Plasma and RBC cholinesterase activities were slightly inhibited: blood urea nitrogen was significantly elevated in the test animals. Other parameters examined showed no or marginal changes.
Fogging activity has become one of the important methods in tackling dengue outbreaks nowadays. Despite this, it is an occupation that has known hazards namely noise and pesticides which can affect the body organs. Based on this knowledge, a cross sectional study was conducted to determine the association between hearing impairment and serum cholinesterase level among foggers from five randomly selected Health District Offices in Selangor. A total of 200 workers comprising of 100 workers exposed to fogging activity and another 100 workers not exposed were randomly selected for this study. Data was obtained through self administered questionnaire, audiometric testing and serum cholinesterase level measurement. The overall prevalence of hearing impairment was 16% with foggers having significant higher prevalence (24%) compared to the non foggers (8%) (χ2=9.52, p=0.002). Measurement of serum cholinesterase showed that exposed workers had lower level compared to the non exposed group. Similarly, those who have hearing impairment had also lower serum cholinesterase level although it was not statistically different. Among all studied factors, age and duration of exposure to fogging activities were statistically different with hearing impairment (p
Acetylcholinesterase (AChE) is usually used as an inhibitive assay for insecticides. A lesser
known property of AChE is its inhibition by heavy metals. In this work we evaluate an AChE
from brains of striped snakehead (Channa striatus) wastes from aquaculture industry as an
inhibitive assay for heavy metals. We discovered that the AChE was inhibited almost completely
by Hg2+, Ag2+ and Cu2+ during an initial screening. When tested at various concentrations, the
heavy metals exhibited exponential decay type inhibition curves. The calculated IC50 for the
heavy metals Hg2+, Ag2+, Pb2+, Cu2+ and Cr6+ were 0.08432, 0.1008, 0.1255, 0.0871, and 0.1771,
respectively. The IC50 for these heavy metals are comparable and some are lower than the IC50
values from the cholinesterases from previously studied fish. The assay can be carried out in less
than 30 minutes at ambient temperature.
Acetylcholinesterase (AChE) is usually used as an inhibitive assay for insecticides. A lesserknown
property of AChE is its inhibition by heavy metals. In this work, we evaluate an AChE
from brains of Clarias batrachus (catfish) exposed to wastes from aquaculture industry as an
inhibitive assay for heavy metals. We discovered that the AChE was inhibited completely by
Hg2+, Ag2+, Pb2+, Cu2+, Cd2+, Cr6+ and Zn2+ during initial screening. When tested at various
concentrations, the heavy metals exhibited exponential decay type inhibition curves. The
calculated IC50 (mg/L) for the heavy metals Ag2+, Cu2+, Hg2+, Cr6+ and Cd2+ were 0.088, 0.078,
0.071, 0.87 and 0.913, respectively. The IC50 for these heavy metals are comparable, and some
are lower than the IC50 values from the cholinesterases from previously studied fish. The assay
can be carried out in less than 30 minutes at ambient temperature.
A phytochemical investigation on the ethyl acetate soluble fraction of Lonicera quinquelocularis (whole plant) led to the first time isolation of one new phthalate; bis(7-acetoxy-2-ethyl-5-methylheptyl) phthalate (3) and two new benzoates; neopentyl-4-ethoxy-3, 5-bis (3-methyl-2-butenyl benzoate (4) and neopentyl-4-hydroxy-3, 5-bis (3-methyl-2-butenyl benzoate (5) along with two known compounds bis (2-ethylhexyl phthalate (1) and dioctyl phthalate (2). Their structures were established on the basis of spectroscopic analysis and by comparison with available data in the literature. All the compounds (1-5) were tested for their acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) inhibitory activities in dose dependent manner. The IC50 (50% inhibitory effect) values of compounds 3 and 5 against AChE were 1.65 and 3.43 µM while the values obtained against BChE were 5.98 and 9.84 µM respectively. Compounds 2 and 4 showed weak inhibition profile.
Children are the vulnerable group in the agricultural community due to their early exposure to pesticides through the dynamic interplay between genetic predisposition, environment, and host-related factors. This study aims to identify the possible association between the depression in blood cholinesterase level and genotoxic effect among farm children. The results of micronuclei assay and comet assay showed that the reduced blood cholinesterase level from organophosphate pesticide exposure is significantly associated with an increase in chromosome breakage and DNA strand breaks. These genotoxicity end points suggest that farm children's cells experience early DNA damage that may lead to uncontrolled cell proliferation during their adulthood. Thus, farm children who grow up near pesticide-treated farmland have a higher probability of developing cancer than children with minimal or zero exposure to pesticides.
Two series of novel acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) inhibitors containing benzimidazole core structure were synthesized by a four-step reaction pathway starting from 4-fluoro-3-nitrobenzoic acid as the basic compound. The structure of the novel benzimidazoles was characterized and confirmed by the elemental and mass spectral analyses as well as (1)H NMR spectroscopic data. Of the 34 novel synthesized compounds, three benzimidazoles revealed AChE inhibition with IC50<10 μM. The highest inhibitory activity (IC50=5.12 μM for AChE and IC50=8.63 μM for BChE) corresponds to the compound 5IIc (ethyl 1-(3-(1H-imidazol-1-yl)propyl)-2-(4-nitrophenyl)-1H-benzo[d]imidazole-5-carboxylate). The relationship between lipophilicity and the chemical structures as well as their limited structure-activity relationship was discussed.
Series of hitherto unreported piperidone grafted pyridopyrimidines synthesized through ionic liquid mediated multi-component reaction. These compounds were evaluated for their inhibitory activities against AChE and BChE enzymes. All the compounds displayed considerable potency against AChE with IC50 values ranging from 0.92 to 9.11 μM, therein compounds 6a, 6h and 6i displayed superior enzyme inhibitory activities compared to standard drug with IC50 values of 0.92, 1.29 and 2.07 μM. Remarkably, all the compounds displayed higher BChE inhibitory activity compared to galantamine with IC50 values of 1.89-8.13 μM. Molecular modeling, performed for the most active compounds using three dimensional crystal structures of TcAChE and hBChE, disclosed binding template of these inhibitors into the active site of their respective enzymes.
Poisoning is one of the most important causes of morbidity and mortality in developing countries like India. Anticholinesterase compounds like organophosphates (OP) and carbamates account for the majority of these poisoning cases because of their easy availability and agricultural use. Carbamates are as popular as OPs as insecticides that often go undiagnosed. A fatal case of carbofuran poisoning is presented where serial cholinesterase estimation played a major role in the diagnosis of the same. The pertinent medical literature on carbofuran poisoning is reviewed. The establishment of poison information center in each state is needed for proper diagnosis and management of poisoning cases.
The release of pollutants, especially heavy metals, into the aquatic environment is known to have detrimental effects on such an environment and on living organisms including humans when those pollutants are allowed to enter the food chain. The aim of this study is to analyse the damage to Clarias gariepinus' liver caused by exposure to different concentrations of copper. In the present study, samples of C. gariepinus were exposed to sub-lethal copper sulphate (CuSO4) concentrations (from 0.2 to 20.0 mg/L) for 96 h. Physiological and behavioural alterations were observed with respect to their swimming pattern, mucus secretion and skin colour. Mortality was also observed at high concentrations of copper. Histopathological alterations of the liver were analysed under light, transmission and scanning electron microscopies. The liver of the untreated group showed normal tissue structures, while histopathological abnormalities were observed in the treated fish under light and electron microscopes with increased copper concentrations. Histopathological abnormalities include necrosis, melanomacrophage, hepatic fibrosis and congested blood vessels. In addition, the enzyme activity of liver cholinesterase (ChE) was also found to be affected by copper sulphate, as 100% of cholinesterase activity was inhibited at 20.0 mg/L. Thus, liver enzyme activity and histopathological changes are proven to be alternative sources for biomarkers of metal toxicity.