Design and preparation of novel antibody system and application for the determination of heroin metabolites in urine by capillary electrophoresis

Abstract

A novel multi-target antibody to morphine and derivatives was induced by designed morphine-3-site substituted and the polyclonal antibody was prepared with immunizing rabbits. A simple, specific and accurate method for the determination of morphine and related compounds, codeine, acetylcodeine, 6-monoacetylmorphine and morphine-3-glucuronide in urine of heroin abusers, has been developed using the multi-target immunoaffinity column (IAC) prior to capillary electrophoresis separation. The analytes were extracted from the urine of drug addicts with the column, which was made by coupling CNBr-activated Sepharose 4B and multi-target polyclonal antibodies. The analytes of interest were extracted with a methanol/water mixture in one step. Baseline separation of these analogs was achieved by CE using β-cyclodextrin as additive and they were monitored at 214 nm. The assay presented very good reproducibility and precision with the recovery and detection limit between 91–105% and 10–20 ng/mL based on S/N = 2, respectively. The inter-day and intra-day variation, capacity and elution conditions of the immunoaffinity column were also discussed. The metabolites in five heroin addicts’ urine were measured by the present method. The experimental results indicated that the combination of IAC and CE was a useful technique for determination of heroin metabolites from urine samples.