Novel spectrophotometric method for the quantitation of urinary xanthurenic acid and its application in identifying individuals with hyperhomocysteinemia associated with Vitamin B6 deficiency.

A novel spectrophotometric method for the quantification of urinary xanthurenic acid (XA) is described. The direct acid ferric reduction (DAFR) procedure was used to quantify XA after it was purified by a solid-phase extraction column. The linearity of proposed method extends from 2.5 to 100.0 mg/L. The method is precise, yielding day-to-day CVs for two pooled controls of 3.5% and 4.6%, respectively. Correlation studies with an established HPLC method and a fluorometric procedure showed correlation coefficients of 0.98 and 0.98, respectively. Interference from various urinary metabolites was insignificant. In a small-scale screening of elderly conducted at Penghu county in Taiwan (n = 80), we were able to identify a group of twenty individuals having hyperhomocysteinemia (>15 µ mole/L). Three of them were found to be positive for XA as analyzed by the proposed method, which correlated excellently with the results of the activation coefficient method for RBC's AST/B6 functional test. These data confirm the usefulness of the proposed method for identifying urinary XA as an indicator of vitamin B6 deficiency-associated hyperhomocysteinemic condition.

Transamination of 3-hydroxykynurenine to produce xanthurenic acid: a major branch pathway of tryptophan metabolism in the mosquito, Aedes aegypti, during larval development.

An electrochemically active compound was detected in the larvae of Aedes aegypti mosquitoes and progressive accumulation of this compound was observed during larval development. The compound was purified from mosquito larvae using various chromatographic techniques and spectral analysis of the purified compound resulted in its identification as xanthurenic acid. Production of xanthurenic acid results from the transamination of 3-hydroxykynuorenine, and analysis of the biochemical pathway in xanthurenic acid production revealed the presence of a particular transaminase that has a much higher specific activity to 3-hydroxykynurenine than to kynurenine in the mosquito larvae. Concentration of xanthurenic acid is closely related to the level of this transaminase activity. Results suggest that this particular transaminase plays an important role in regulating the level of 3-hydroxykynurenine in the mosquito, A. aegypti during larval development.

Deamination of 3-hydroxykynurenine in bovine lenses: a possible mechanism of cataract formation in general.

BACKGROUND: 3-Hydroxykynurenine, a metabolite of tryptophan, acts as UV filter in the human lens. In this study, we looked for this substance and its metabolites in young and old bovine lenses, because of their possible role in the formation of cataract. METHODS: The substances were detected by HPLC analysis. The fluorescent substance formed from 3-hydroxykynurenine was characterized by thin-layer chromatography followed by reaction with ninhydrin, UV and fluorescence spectrum analysis, and atom bombardment for molecular mass determination. The kynurenine aminotransferase activity was determined by the method of Tobes. RESULTS: 3-Hydroxykynurenine was detected at concentrations of 0.07, 0.19, and 1.14 micrograms/g of tissue in the bovine iris/ciliary body, retina, and transparent bovine lenses respectively. 3-Hydroxykynurenine was deaminated in old bovine eyes but not in calf eyes. In old eyes, kynurenine aminotransferase activity was 2.7, 3.5, and 9.6 mumol/g of tissue per h in retina, iris/ciliary body, and lens respectively. CONCLUSION: The deamination of 3-hydroxykynurenine resulted in the formation of a fluorescent substance which was identified as oxidized xanthurenic acid. This substance, accumulating in the bovine lens and interacting with lens proteins, could induce cataract formation.

Endogenous inhibitor of ecdysone synthesis in crabs.

Attempts to isolate the molt-inhibiting hormone (MIH) of crustaceans from crab eyestalks (ES) resulted in the characterization of xanthurenic acid as an inhibitor of ecdysone biosynthesis in the cultured Y-organ-complex (YOC) homogenate. It was also found that 3-hydroxy-L-kynurenine present in the ES is transformed into xanthurenic acid in the YOC and body fluid. Its mode of inhibitory action in ecdysone biosynthesis is probably inactivation of cytochrome P-450.

Xanthurenic acid 8-O-beta-D-glucoside, a novel tryptophan metabolite in eye-color mutants of Drosophila melanogaster.

An unknown fluorescent metabolite has been isolated from heads of eye-color mutants of Drosophila melanogaster. Only a few mutations cause it to accumulate, viz. cardinal (cd), dark red brown (drb), Henna-recessive (Hnr), purple (pr), Punch2 (Pu2), Punch-Grape (PuGr), and scarlet (st). After purification by ion-exchange chromatography, the spectroscopic, chemical, and enzymatic analyses revealed that it is a novel quinoline derivative: xanthurenic acid 8-O-beta-D-glucoside. Feeding experiments suggest that this glucoside is synthesized from 3-hydroxykynurenine and that free xanthurenic acid is not a precursor. The results from the analysis for its occurrence in double mutants, together with the fact that xanthurenic acid 8-glucoside share the same precursor as xanthurenic acid and xanthommatin, suggest that xanthurenic acid 8-glucoside formation is closely related to the regulation of the last step in the biosynthesis of xanthommatin.

Purification of xanthurenic acid-insuline complex.

Tritium labeled xanthurenic acid (XA)-insulin complexes A and B were obtained. These were shown to be one component by the disc electrophoresis, and bound XA was not liberated by the electrophoresis from the paper electrophoresis experiment. The result of the gel filtration showed that the molecular weight of the complex A and B was 24,000. But there is still a possibility that complex B might be contaminated with free insulin.