Simultaneous liquid chromatographic determination of vanillylmandelic acid, homovanillic acid, and 5-hydroxy-3-indoleacetic acid in urine, using isocratic elution and electrochemical detection.

A method for the simultaneous measurement of vanillylmandelic acid, homovanillic acid and 5-hydroxyindoleacetic acid in urine is described. Based on reversed-phase liquid chromatography with electrochemical detection, the procedure employs isocratic elution, thus making it suitable for use in the less well-equipped clinical or research laboratory. A simple extraction of the acids from acidified urine into ethyl acetate, is followed by evaporating to dryness a portion of the organic layer, and redissolving the residue in chromatographic mobile phase. Up to 20 samples can be analysed in a single working day. The method is validated and the results obtained are compared with reference methods. The cause of contamination of the glassy carbon surface of the working electrode is investigated, and a simple electrochemical pretreatment is described that overcomes this problem. Finally, the extra clinical information that can be derived from multi-metabolite assays is considered.