A gas chromatographic method was applied to the determination of the urinary glycine conjugates, hippuric, o-, m- and p-methylhippuric acids. These were extracted with ethyl acetate from urine after acidification with hydrochloric acid. The internal standard solution (heptadecanoic acid methanol solution) was added before extraction and a diazomethane-ether-ethanol solution was subsequently added to the dried extracts. The methylated residues were dissolved in methanol and injected into a gas chromatograph as described by Buchet and Lauwerys (1973). By the combined use of gas chromatography and mass spectrometry the methyl esters of hippuric acid and m-methylhippuric acid were identified in the urine of a volunteer who had been exposed to toluene and m-xylene vapours. When the urine specimen contained salicyluric acid (a urinary metabolite of salicylic acid) two sharp peaks were observed. The faster peak coincided with m- or p-methylhippuric acid. The upper limit of urinary hippuric acid concentration in healthy subjects with no occupational exposure was calculated by this method to be 1.026 microgram/ml (fiducial limit 5%) after correction to 1.024 for variation in urinary density.
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