Determination of urinary 4-pyridoxic acid using high performance liquid chromatography

A high performance liquid chromatographic method is presented for the determination of the vitamin B6 metabolite, 4-pyridoxic acid, in urine. Urine samples are treated with trichloroacetic acid to precipitate protein. An aliquot of the supernatant is chromatographed using 0.033 M phosphate buffer containing 5% (v/v) methanol (pH 2.2), a fluorometric detector and a commercial reverse phase octadecylsilica column. The high precision of the method and the absence of interfering compounds have been demonstrated. This method provides a rapid, sensitive, and quantitative technique for the measurement of urinary 4-pyridoxic acid for use in metabolic and nutritional studies.