Additional three metabolites of methylhexabital (MHB, 5-cyclohexenyl-3, 5-dimethylbarbituric acid) were isolated from the urine of rabbits receiving MHB and chemical structures of two of these compounds were established as 5-(3-oxo-1-cyclo-hexenyl)-5-methylbarbituric acid and β-5-(3-hydroxy-1-cyclohexenyl)-3, 5-dimethyl-barbituric acid. The main metabolites obtained to date were diastereoisomeric 3-OH-MHB[α-and β-5-(3-hydroxy-1-cyclohexenyl)-3, 5-dimethylbarbituric acid, in which α-form was considerably predominant]and 3-keto-MHB[5-(3-oxo-1-cyclohexenyl)-3, 5-dimethyl-barbituric acid].