ANTICHOLINESTERASE ACTIVITY MEASUREMENT BY A CHOLINE BIOSENSOR - APPLICATION IN WATER ANALYSIS

A choline amperometric biosensor was assembled and used to measure the anticholinesterase activity due to compounds (which have the property to inhibit cholinesterase enzymes) present in water samples. This par ameter can be used as a 'toxicological index', defined as the amount o f compound which causes a certain percentage of cholinesterase inhibit ion equivalent to a known amount of a reference compound causing the s ame percentage inhibition. The organophosphorus insecticide Paraoxon, which has proved to be a strong inhibitor of cholinesterase enzymes, w as chosen as the reference compound. The analysis was carried out by m onitoring the decrease of cholinesterase activity in the presence of a pesticide and a substrate specific for the enzyme whose reaction prod uces choline. The decrease in choline production was measured by the c holine sensor and correlated to the concentration of anticholinesteras e compound present in the solution. Parameters such as buffer, pH, tem perature and incubation time were optimized. The rate constant K(i) wa s calculated experimentally for Paraoxon and used in the anticholinest erase activity measurements at different fixed incubation times. The p robe was calibrated with different standard solutions of Paraoxon. The effect of Paraoxon and heavy metals on the choline probe was evaluate d. This probe was then used for the determination of anticholinesteras e activity of some organophosphorus pesticides, and heavy metals in sp iked waters. Samples were also analysed by liquid/liquid extraction an d GC determination. Results seem to correlate with acute toxicity expr essed as LD50 (oral, rat). Analysis of water samples from different so urces in central Italy were analysed for total anticholinesterase acti vity (TAA) and compared with a reference procedure.