An electrochemical biosensor for the detection of dichlorophenoxyacetic acid (2,4-D) based on a screen-printed carbon paste electrode (SPE) was developed. The biosensor film deposited on the SPE was a sol–gel/chitosan film containing immobilized alkaline phophatase enzyme (ALP). The 2,4-D herbicide was determined by the quantitative inhibition on the enzyme alkaline phosphatase. Ascorbic acid 2-phosphate (AA2P) was used as a substrate to indicate the inhibition by 2,4-D. The enzymic reaction with the substrate was monitored amperometrically at a potential of +600 mV and at optimum pH of 8.5. The linear response range towards the substrate AA2P was 10–80 M. For 2,4-D determination, the maximum inhibition of the alkaline phosphatase was achieved after 80 ppb (60% inhibition) 2,4-D with a linear response range of 2–80 ppb.
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