Radioimmunoassay of chlordiazepoxide in plasma

A simple, rapid, and sensitive radioimmunoassay was developed for the determination of chlordiazepoxide (7-chloro-2-methylamino-5-phenyl-3H-1,4-benzodiazepine 4-oxide) in plasma. Antiserum capable of binding chlordiazepoxide-2-14C was obtained from rabbits following immunization with an antigen prepared by coupling the reactive acyl azide of 7-chloro-5-(4-hydrazinocarbonylmethoxyphenyl)-2-methylamino-3H-1,4-benzodiazepine 4-oxide to bovine serum albumin. The radioimmunoassay allows for the specific determination of chlordiazepoxide directly in plasma without extraction and was compared with a differential spectrofluorometric assay for chlordiazepoxide. Both methods gave satisfactory agreement for the plasma levels of chlordiazepoxide in human subjects resulting from single and chronic oral doses of chlordiazepoxide hydrochloride.