Identification of the new UV filter compound cysteine-L-3-hydroxykynurenine O-beta-d-glucoside in human lenses.

UV filters protect the human lens and retina from UV light-induced damage. Here, we report the identification of a new UV filter, cysteine-l-3-hydroxykynurenine O-beta-d-glucoside, which is present in older normal human lenses. Its structure was confirmed by independent synthesis. It is likely this novel UV filter is formed in the lens by nucleophilic attack of cysteine on the unsaturated ketone derived from deamination of 3-hydroxykynurenine O-beta-d-glucoside. Quantitation studies revealed considerable variation in normal lens levels that may be traced to the marked instability of the cysteine adduct. The novel UV filter was not detected in advanced nuclear cataract lenses.

Lenticular levels of amino acids and free UV filters differ significantly between normals and cataract patients.

PURPOSE: To determine the levels of free UV filters and selected amino acids in cataract lenses compared with normal lenses. METHODS: Indian cataract lenses (n=39) and normal lenses (n=6) were examined by HPLC to quantify levels of UV filter compounds, the UV filter precursor amino acid tryptophan (Trp), as well as tyrosine (Tyr) and uric acid. RESULTS: The levels of the two major primate UV filters, 3-hydroxykynurenine glucoside (3OHKG) and 4-(2-amino-3-hydroxyphenyl)-4-oxobutanoic acid glucoside (AHBG), in cataract lenses were markedly decreased compared with levels in normal lenses. By contrast, the levels of Trp were greatly increased. Mean Trp concentrations were an order of magnitude higher than in normal lenses, with 86% of dark-colored cataract lens nuclei having Trp concentrations greater than the mean level in the normal lenses. The concentrations of Tyr were also higher in cataract lenses. The levels of Kyn, however, were unchanged, and the uric acid levels were substantially lower. CONCLUSIONS: The levels of the free UV filter compounds 3OHKG and AHBG, and also of Trp, Tyr, and uric acid were different in cataract lenses compared to normal lenses. These data suggest that the metabolism of a large proportion of patients with cataract may be substantially different than in persons with normal lenses. Although the mechanism of such metabolic defects are unknown, the authors speculate that an amino acid transporter system may be upregulated in patients with cataract. Because kynurenine levels in cataract were not significantly different from those of normal lenses, there may be a defect in the lenticular UV filter pathway at one, or both, of the steps that convert kynurenine to 3OHKG.

A fluorescence-based assay for indoleamine 2,3-dioxygenase.

A rapid and sensitive fluorescence-based bioassay for determination of indoleamine 2,3-dioxygenase (IDO) activity has been developed. This assay relies on the quantification of the amount of kynurenine produced in the assay medium by fluorescence and complements the standard absorbance and high-performance liquid chromatography (HPLC) assay methods. The fluorescence method has limits of detection similar to those of the standard assay methods. Measured activities of IDO, including in the presence of tryptophan-based inhibitors, were in statistical agreement with the absorbance and HPLC assay methods. The fluorescence-based assay was also suitable for assessment of IDO inhibition by compounds that are incompatible with the absorbance method.