Development of Enzyme Immunoassay for Captan and Its Degradation Product Tetrahydrophthalimide in Foods

Abstract

A simple, sensitive, and precise enzyme-linked immunosorbent assay (ELISA) is described for the quantitation of captan as its degradation product tetrahydrophthalimide (THPI) in foods using polyclonal antibodies. Three hapten analogues of THPI with different alkyl spacer arm lengths were synthesized. Immunogens and coating proteins were prepared by coupling these haptens to human serum albumin and ovalbumin, respectively. A 5-carbon spacer arm appeared to be optimum for the production of antibodies. Heterologous coating proteins did not improve the sensitivity, but reduction of homologous coating protein concentration did improve the sensitivity, resulting in a concentration of test compound required to inhibit binding by 50% of 15.5 ng/mL. The antiserum is specific for captan, captafol, and THPI, but not other structurally related compounds. The minimum detection limit was 1 ng/mL; the linearity was 1-200 ng/mL. The overall recoveries of captan and THPI from 11 commodities spiked at 4 levels were 92 and 100%, respectively. The intra-assay and interassay coefficients of variation were 9.1 and 16.8% for apple blanks and 5.9 and 4.2% for apple spiked with 3 ppm THPI, respectively. The ELISA described is suitable for measuring captan and THPI at levels comparable to those typically found in fruit.