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Abstract

A technique has been developed for the determination of the main products of tryptophan metabolism occurring usually in small amounts in the urine of healthy persons on a free diet. With a single column of cation exchange resin (Amberlite IR-120) and volatile buffers, metabolites present in 1–2% of 24-h urine were fractionated by elution with 100 ml of 0.1 M pyridine-formic acid buffer pH 2.60, and 140 ml of same buffer 0.3 M, pH. 5.60.The effluent was pooled in 6 large fractions which were lyophilized. Each residue was subjected to two-dimensional paper chromatography for a further purification.The spots corresponding to kynurenine, 3-hydroxykynurenine, Nα-acetylkynurenine, Nα-acetyl-3-hydroxykynurenine, kynurenic acid, xanthurenic acid, xanthurenic acid 8-methyl ether, anthranilic acid, 3-hydroxyanthranilic acid and 3-methoxyanthranilic acid were eluted and determined fluorometrically or colorimetrically.Since o-aminohippuric acid is partially destroyed, the method of Price and Brown was preferred for the first step of elution followed by paper chromatography of the residue to achieve complete specificity.Recoveries were quantitative over a range of 10 μg to 100 μg for all tested metabolites in each sample.Normal occurrence of the derivatives was determined in urines from 20 healthy young men and 17 healthy young women. No significant difference was noted between the two groups, where excretion showed similar values. The spontaneous excretion of tryptophan metabolites by healthy human beings ranges from trace amounts to a few milligrams daily.