Automated detection of chlorpyrifos and its metabolites by a continuous flow system-based enzyme sensor Original Research Article

This work is a continuation of the development of a previously described pesticide sensor and presents significant improvements including automation of the system and a new cell design [Biosens. Bioelectron. 13 (1998) 213]. These modifications to the continuous flow system permitted a reduction of analysis time and led to a better reproducibility of the delivery times of the different reagent solutions. Chlorpyrifos, methyl-chlopyrifos and their metabolites (oxon forms and trichloro pyridinol) were used for testing the continuous flow system. Kinetics using free acetylcholinesterase activity showed that pesticides were one 1000-fold less toxic than the oxon forms (bimolecular rate constants (ki) for chlorpyrifos-oxon and methyl-chlorpyrifos-oxon were 3.0×106 and 1.3×106 M−1 min−1, respectively). Trichloro-pyridinol did not inhibit free acetylcholinesterase activity. In continuous flow conditions, chlorpyrifos, methyl-chlopyrifos and trichloropyridinol did not significantly decrease the activity of the immobilised enzyme. The biosensor detection of chlorpyrifos-oxon gave a calibration curve with a lowest detectable concentration ranging from 0.72×10−6 M (240 ppb) to 73×10−9 M (24 ppb) when the incubation time was increased from 1 to 16 min. The rinsing step after biosensor inhibition by methyl-chlorpyrifos-oxon led to a partial reactivation of the immobilised enzyme. Under these conditions, the lowest detectable concentration ranged from 2.3×10−6 M (700 ppb) to 0.89×10−6 M (270 ppb) when the incubation time was increased from 1 to 8 min.