An optical fiber biosensor consisting of acetylcholinesterase (AChE) and bromothymol blue (BTB) doped sol–gel film was employed to detect organophosphate pesticide chlorpyrifos. The main advantage of this optical biosensor is the use of a single sol–gel film with immobilized AChE and BTB. The compatibility of this mixture (AChE and BTB) with the sol–gel matrix has prevented leaching of the film. The immobilization of the enzyme and indicator was simple without chemical modification. The biosensing element on single sol–gel film has been placed inside the flow-cell for flow system. In the presence of a constant AChE, a color change of the BTB and the measured reflected signal at wavelength 622 nm could be related to the pesticide concentration in the sample solutions. The performance of optical biosensor in the flow system has been optimized, including chemical and physical parameters. The response time of the biosensor is 8 min. A linear calibration curve of chlorpyrifos against the percentage inhibition of AChE was obtained from 0.05 to 2.0 mg/L of chlorpyrifos (18–80% inhibition, R 2 = 0.9869, n = 6). The detection limit for chlorpyrifos was 0.04 mg/L. The results of the analysis of 0.5–1.5 mg/L of chlorpyrifos using this optical biosensor agreed well with chromatographic method.
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