J E Leklem

Direct and steam distillation autoanalyzer methods for assay of diazotizable aromatic amine metabolites of tryptophan in urine and in serum.

Abstract A procedure is described for the assay of aromatic amine metabolites of tryptophan by an automated diazotization and coupling procedure using Technicon Autoanalyzer equipment. The assay is reliable and sensitive and if many samples are to be analyzed, will afford a several-fold saving in time and labor. This automated procedure is also satisfactory for assay of the enzymes tryptophan oxygenase and kynureninase. In addition, a procedure is described for automated steam distillation of aromatic amines in urine and serum and for their simultaneous assay by means of diazotization and coupling. This steam distillation method is specific for kynurenine and acetylkynurenine and is suitable for the routine screening of large numbers of urine or blood serum samples for elevated levels of these metabolites as a measure of abnormal tryptophan metabolism.

Effect of oral contraceptives on urinary metabolite excretions after administration of L-tryptophan or L-kynurenine sulfate☆☆☆

Abstract The metabolism of orally administered doses of L-tryptophan and L-kynurenine sulfate has been compared in 18 women using an estrogen-containing oral contraceptive, and 12 controls who were not taking any steroid preparations. Both compounds produced higher urinary excretions of several intermediates of the tryptophan-nicotinic acid ribonucleotide pathway in the oral contraceptive users than in the controls, indicating that these steroids have an effect beyond the tryptophan oxygenase-mediated step. Increases in the ratio of 3-hydroxykynurenine to 3-hydroxyanthranilic acid excreted were consistent with an inhibition of kynureninase, due to impaired pyridoxal phosphate function, but only tryptophan loading caused an elevated xanthurenic acid excretion. This difference may have been due to an influence of the kidney on kynurenine metabolism.

Abnormal tryptophan metabolism in patients with scurvy-type skin.

Abstract South African Bantu subjects with a scurvy-type skin, which appears after a prolonged hemosiderosis caused by a high dietary iron intake, were studied with the oral 2-g load of tryptophan test. A majority of the subjects excreted statistically significantly elevated levels of anthranilic acid glucuronide, acetylkynurenine, kynurenine, and hydroxykynurenine, after the 2-g load of l -tryptophan. The data suggest that tryptophan metabolism in these subjects may be inhibited at some step beyond hydroxykynurenine as a result of lowered tissue pyridine nucleotide activity or tissue enzyme poisoning by the high iron levels that are present.

Tryptophan: abnormal metabolism in pellagra patients.

Abstract South African Bantu subjects with pellagra excreted abnormally elevated levels of kynurenine, acetylkynurenine, 3-hydroxykynurenine and several other metabolites of the kynurenine-niacin pathway after an oral load of 2 Gm. of L-tryptophan. The data suggest that tryptophan pyrrolase activity may be elevated in these subjects either as the result of a stress-mediated induction or as the result of lack of negative feedback by tissue pyridine nucleotides.

A mechanism for the formation in vivo of the 8-methyl ether of xanthurenic acid in humans

Abstract The mechanism of formation of the tryptophan metabolite, xanthurenic acid 8-methyl ether, has been open to question since it was shown to occur in the urine of humans. To investigate possible pathways, human subjects were given l -methionine-methyl- 14 C, l -kynurenine-keto- 14 C, 3-hydroxy- l -kynurenine-keto- 14 C, or xanthurenic acid-4- 14 C. Analysis of urine by ion exchange chromatography, paper chromatography and carier isolations showed that the methyl group of xanthurenic acid 8-methyl ether came from l -methionine-methyl- 14 C. The xanthurenic acid 8-methyl ether accounted for 0.005–0.038% of the dose of l -methionine-methyl- 14 C. Keto - 14 C labeled l -kynurenine and 3-hydroxy- l -kynurenine were both converted to xanthurenic acid 8-methyl ether, but accounted for only 0.015 and 0.028% of the dose, respectively. Xanthurenic acid-4- 14 C also gave rise to labeled xanthurenic acid 8-methyl ether in four human subjects, with this metabolite accounting for 0.045–0.15% of the dose. No urinary 3-methoxykynurenine was detected, but labeled 3-methoxyanthranilic acid (from l -methionine-methyl- 14 C) was detected. These data indicate that xanthurenic acid may be methylated directly to form xanthurenic acid 8-methyl ether in humans. Since 3-methoxykynurenine could not be detected, it is suggested that 3-methoxyanthranilic acid is formed by methylation of 3-hydroxyanthranilic acid rather than from 3-methoxykynurenine, although the transient presence of the latter cannot be excluded.