Student Research: Yutaka Aoki

MS, , 1993
Faculty Advisor: Matthew C. Keifer

The Influence of Anticoagulant, Temperature, and Time on In Vitro Recovery of Organophosphate Inhibited Cholinesterase


Depression of plasma (PChE) and erythrocyte (AChE) cholinesterase is used to monitor exposure to organophosphate (OP) insecticides. Lab-based cholinisterase (ChE) testing necessitates sample transportation and storage and thus assumes no considerable time-dependent change in ChE activities. Recovery of ChE activity during such intervals may generate flase negative (artificially higher) measurements. Such recovery is conceivable considering possible OP removal by plasma arylesterase, the reactivation rate of OP-inhibited ChE and the length of the aforementioned intervals.

To examine this possibility, normal blood samples were spiked with OP and incubated to simulate blood from OP-exposed subjects. ChE activity was tesed after 0, 4, 24, and 48 hour storage at 0 and 30 degrees Celsius. Oxon metabolites of commonly-used thioate OPs, parathion, chlorpyrifos, and their methyl analogs, were used since oxons are primarily responsible for in vivo ChE depression following exposure to thioates. The effects of two anticoagulants, heparin and EDTA, were examined since inhibition of arylesterase by EDTA may hinder recovery.

The results suggested that the recovery is possible under certain conditions. AChE inhibited by dimethoxy OPs recovered significantly during storage presumably because reactivation half lives of such AChE are less than a few hours. Contrary to the original prediction, this recovery was enhanced by EDTA. Under other conditions EDTA hindered recovery to some extent. Storage at 0 degrees Celsius resulted in smaller change than at 30 degress Celsius but 0 degrees storage did not completely preclude recovery. In some cases false positive (artificially lower) AChE and PChE measurements were observed.

These results should be confirmed by epidemiological studies. Assuming that significant change in ChE activities in either direction is possible under real-world situations, field-based ChE testing would be preferred to lab-based testing since field-based testing eliminates the time interval. The secondary goal of the present study was to compare a field-based ChE kit to a lab-based spectrophotometric method. Correlation coefficients between ChE values obtained with the two methods were more than 0.8 The kit provides an important alternative to conventional lab-based ChE testing methods.