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[Studies on development of radioimmunoassay for diazepam and it's medicolegal application].

A sensitive radioimmunoassay was developed for the determination of diazepam. Detection of diazepam in tissues of rats and in blood from unnatural death victims and traffic accident victims was examined by this procedure. Antisera capable of binding [3H] diazepam were obtained by repeated immunization of rabbits with temazepam (oxydiazepam)-3-hemisuccinate conjugated to bovine serum albumin. The antibody of a 1: 10000 dilution could bind approximately 50% of the added radioactivity. The level at least as low as 1 pg of diazepam could be detected by this procedure. This antibody strictly recognized the existence of the methyl group at the position-1 and of the keto group at the position-2 of the benzodiazepine ring, and was highly specific for temazepam and diazepam among benzodiazepine derivatives. But other benzodiazepine derivatives had no cross-reactivity with this antibody. The distribution of diazepam in biological fluids and tissues of rats 1, 2, 4 and 8 hr after intraperitoneal injection was examined. The level of diazepam in serum, saliva, brain and bone marrow declined within 2 hr, while the level in liver and kidney was maximum 2 hr after administration. The concentration of diazepam in each tissue leveled off 4 hr after administration. Among these results the remarkable finding was that the concentration in the bone marrow is much higher than that in serum, saliva and brain, appearing to be due to a high accumulation of diazepam in this tissue. This result indicates that the bone marrow is a very useful material for the detection of diazepam in skeletonized remains. There ware good correlations (gamma = 0.8595-0.9973) in the concentrations of diazepam between serum, saliva and brain, and bone marrow. Detection of diazepam in blood from unnatural death victims and traffic accident victims were examined. Diazepam was identified in 3 cases of 50 unnatural death victims and in 4 cases of 48 traffic accident victims.