Determination of the herbicide chlorsulfuron by amperometric sensor based on separation-free bienzyme immunoassay

Abstract A separation-free bienzyme immunoassay system has been developed for the electrochemical determination of the herbicide chlorsulfuron. Screen printed electrode with horseradish peroxidase as an integral component of the carbon ink was used as the detector. A membrane with immobilised anti-chlorsulfuron antibodies was attached to the electrode. Free chlorsulfuron in the sample under test and a chlorsulfuron–glucose oxidase conjugate competed for the available binding sites of the membrane-immobilised antibodies. Addition of glucose induced the generation of hydrogen peroxide by the glucose oxidase conjugate, which was in turn reduced by the peroxidase. The latter process caused an electrical current change, due to direct re-reduction of peroxidase by a direct electron transfer mechanism, which was measured to determine the chlorsulfuron content in the sample. Any H 2 O 2 formed in the bulk solution by unbound chlorsulfuron–glucose oxidase conjugate was scavenged by catalase thus removing the need for a washing step. The optimal protocol for chlorsulfuron detection by the method described was found to be: Biodyne A membrane as a carrier, concentration of IgG during adsorption on the membrane—1 ng ml −1 , concentration of the chlorsulfuron–glucose oxidase conjugate—2.5 μg ml −1 (by enzyme), working potential +50 mV versus Ag/AgCl. The total assay time was 15 min. The measuring range for chlorsulfuron detection was 0.01–1 ng ml −1 . The method was suitable application for on-site ecological monitoring.