Hapten synthesis and development of an indirect competitive enzyme-linked immunosorbent assay for chlorpromazine in pork, chicken and swine liver

Abstract

The use of chlorpromazine (CPZ) as a sedative for livestock has been prohibited in the European Union and many other countries. In this study, a new hapten 7-amino-chlorpromazine sulfoxide (hapten 1) against CPZ was synthesized and coupled to ovalbumin (OVA) as an immunogen to produce polyclonal antibody (PAb) specific for CPZ. An heterologous hapten 7-(4-carboxyl-phenylazo)-chlorpromazine (hapten 2) was synthesized and coupled to bovine serum albumin (BSA) as coating antigen to improve the assay sensitivity. The results showed that the hapten-heterologous systems exhibited 20 times higher sensitivity than the hapten-homologous one. Based on the hapten-heterologous system, an indirect competitive enzyme-linked immunosorbent assay (ELISA) for CPZ was developed, the IC₅₀ value was 0.58 ng mL⁻¹ and the limit of detection was 0.03 ng mL⁻¹. The assay showed no cross-reaction with the CPZ analogues. The average recoveries of CPZ from spiked samples were estimated to range from 77.1% to 98.6%, with a coefficient of variation (CV) of less than 10.9%. Linear regression analysis showed a good correlation between the CPZ concentrations obtained from ELISA and HPLC analysis, which suggested that the ELISA is a convenient supplementary analytical tool for monitoring CPZ.