Abstract
A method utilizing reverse-phase liquid chromatography/electrochemistry ( LC EC ) was developed for the simultaneous determination of aniline and its hydroxylated derivatives, p-aminophenol, o-aminophenol, m-aminophenol, and N-phenylhydroxylamine. To achieve separation of these compounds, a mobile phase of 3.0% dimethylformamide and 97.0% 0.05 m piperazine acetate, pH 5.4, containing 0.05 m KNO 3 was developed. A procedure is also presented for the determination of p-nitrophenol, nitrobenzene, and nitrosobenzene, possible aniline metabolites in higher N-oxidation states, using reductive amperometric detection. The hydroxylated compounds, including the hydroxylamine, and nitrosobenzene are easily detected as metabolites of aniline in mouse liver slice or microsomal preparations. No prior extraction, preconcentration, or derivatization steps are needed for the determinations, which can be accomplished by a direct injection of the incubation mixture. The K m value for the hepatic aniline 4-hydroxylase activity in male Cox-Swiss mice microsomal preparations has been determined to be 0.52 m m; the V max value is 2.90 ± 0.64 nmol min −1 mg microsomal protein −1. Detection limits for all compounds of interest are in the picomole range.
Published Version
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