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Evaluation of a fluorescent method for measuring cholinesterase activity in mammalian blood and tissue.

Authors
Stamler-CJ; Arrieta-DE; Basu-N; Henderson-JD; Wilson-BW; Chan-HM
Source
Bull Environ Contam Toxicol 2006 Dec; 77(6):785-792
NIOSHTIC No.
20031822
Abstract
Cholinesterase (ChE) activity has been widely used as a biochemical marker of cholinergic system function in laboratory, wildlife, and human studies (Fulton and Key 2001; Padila et.al. 1996). Acetylcholinesterase (AChE, EC 3,1,1.7) prefers to hydrolyze acetylcholine (ACh), butyrlcholinesterase or non-specific cholinesterase prefers other choline esters (BChE, EC 3,1,1,8). Acute exposures to organophosphate and carbamate pesticides inhibit ChE activities, causing accumulation of ACh at synapses, and their disruption in organisms. Although critical to cholinergic transmission in the brain, ChEs are also present in several other tissues, including blood. Both red blood cell (RC) and plasma ChE activities have been widely used as bio-indicators of exposure and intoxication to these pesticides (Wilson et al 1997). Reduced ChE activities in blood have been reported among spray-workers as a result of improper handling of pesticides (Baker et al 1978; Wilson et al 1997). Exposures to other environmental pollutants, including mercury, have been shown to cause alterations in ChE activity in humans (Zabinski et al 2000), wildlife (Lionetto et al 2003) and laboratory animals (Gill et al1990; Hastings et al 1975; Lakshmana et al. 1993). There are several methods to measure ChE activity (St. Omer and Rottinghaus 1992, Wilson 2001). The most widely used is a colorimetric assay developed by Ellman et al (1961). While the Ellman assay is simple and inexpensive, the absorbance peak of hemoglobin (415nm) can interfere with detection of its reaction end-product, 2-nitro-5-benzoic acid (405nm). Recently, a fluorescent based ChE assay kit was marketed for measurement of the purified enzyme (A12217, Molecular Probes, Eugene, OR, USA ( Zhou et al 2000). It detects choline, the ChE reaction product, though a multi-step process in which the oxidation of choline by choline oxidase results in the formation of H202, which is detected though a horseradish peroxidase-coupled reaction with 10-acetyl-3,7- dihydroxyphenoxazine (Amplex Red). The final product, resorufin, is detected by its fluorescence (ex. 563, em. 587) (Zhou et al 1997). It is said to provide high sensitivity (0.002 units/ml of purified AChE from electric eel, as reported by Molecular Probes) without hemoglobin interference. The objective of this study was to evaluate this fluorescent assay as a tool to measure ChE activities in biological specimens collected from humans and wildlife.
Keywords
Exposure-levels; Sampling; Sampling-methods; Mammalian-cells; Blood-analysis; Blood-sampling; Tissue-disorders; Hemodynamics; Biochemical-tests; Biomarkers; Agricultural-chemicals; Pesticides-and-agricultural-chemicals; Agricultural-workers; Pollutants; Work-environment; Work-practices; Humans; Animals
Contact
School of Dietetics and Human Nutrition, Macdonald Campus, McGill University, 21,111 Lakeshore Road, Ste. Anne de Bellevue, Quebec, H9X 3V9, Canada
CODEN
BECTA6
Publication Date
20061201
Document Type
Journal Article
Funding Type
Agriculture; Cooperative Agreement
Fiscal Year
2007
NTIS Accession No.
NTIS Price
Identifying No.
Cooperative-Agreement-Number-U50-OH-007550
Issue of Publication
6
ISSN
0007-4861
Source Name
Bulletin of Environmental Contamination and Toxicology
State
CA
Performing Organization
University of California - Davis
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