A SENSITIVE METHOD FOR THE DETERMINATION AND ISOLATION OF PENTOBARBITAL-C¹⁴ METABOLITES AND ITS APPLICATION TO IN VITRO STUDIES OF DRUG METABOLISM

¹ Wellcome Research Laboratories, Burroughs Wellcome & Co. (U.S.A.), Inc., Tuckahoe, New York, and The Department of Pharmacology, Albert Einstein College of Medicine, New York, New York

A method is described for measuring the in vitro metabolism of pentobarbital-C¹⁴ to l- and d-pentobarbital alcohol (5-ethyl-5-(1-methyl-3-hydroxybutyl)-barbituric acid). Since the method utilizes paper chromatography and measures product formation, it is extremely sensitive and can be used for kinetic studies of microsomal drug metabolism. A more rapid method for measuring the formation of pentobarbital metabolites is also presented which can be used when enzyme activity is high. In addition, it is shown that the metabolism of pentobarbital is stimulated by chronic phenobarbital treatment and that the l-alcohol metabolite of pentobarbital, which is formed by the microsomal fraction of liver, can be further metabolized by an enzyme system present in the 100,000 x g supernatant fraction of liver which requires nicotinamide adenine dinucleotide for activity.