Simultaneous high-performance liquid chromatographic determination of urinary metabolites of benzene, nitrobenzene, toluene, xylene and styrene.

A high-performance liquid chromatographic method is described for the simultaneous determination of six urinary metabolites of several aromatic chemicals: phenol (from benzene), hippuric acid (from toluene), 3-methylhippuric acid (from xylene), mandelic and phenylglyoxylic acid (from styrene) and 4-nitrophenol (from nitrobenzene). Reversed-phase liquid chromatography was performed in an isocratic mode at 1 ml/min on a 5-microns C18 column using two mobile phases: (A) acetonitrile-1% phosphoric acid (10:90); (B) acetonitrile-1% phosphoric acid (30:70). Phase A separates the six metabolites well, but phase B allows to a more rapid and reproducible simultaneous determination of phenolic compounds than phase A. For these compounds a prior enzymic hydrolysis step using Helix pomatia juice is performed to hydrolyse their sulphate and glucuronate conjugates. The reproducibility and the specificity are both excellent. Furthermore, the method is rapid, economical and easily automated. The proposed method appears very suitable for the routine monitoring of workers exposed to these chemicals on the basis of the biological threshold limit values.