High-performance liquid chromatographic determination of 3-hydroxykynurenine with fluorimetric detection; comparison of preovulatory phase and postovulatory phase urinary excretion.
A high-performance liquid chromatographic assay for 3-hydroxykynurenine in human urine is described. A fluorescent derivative of 3-hydroxykynurenine was prepared, based on the reaction of the compound with p-toluenesulphonyl chloride in a basic medium. The analytical method for the measurement of the fluorescent compound employed a Tosoh ODS 80 column eluted with 10 mM potassium dihydrogenphosphate (pH 4.5) and acetonitrile (3:2, v/v) and detection at an excitation wavelength of 375 nm (10 nm bandpass) and an emission wavelength of 455 nm (10 nm bandpass). The column temperature was maintained at 25 degrees C. The detection limit was 3 pmol (673 pg) at a signal-to-noise ratio of 5:1. The fluorescent derivative of 3-hydroxykynurenine was eluted at ca. 12.5 min. The technique was applied to the analysis of human urine. The total analysis time was ca. 15 min.
Shibata
K
K
Department of Food Science and Nutrition, Teikoku Women's University, Osaka, Japan.
Onodera
M
M
D000328:Adult; D002851:Chromatography, High Pressure Liquid; D005260:Female; D005498:Follicular Phase; D006801:Humans; D007737:Kynurenine / Q000031:analogs & derivatives* / Q000652:urine; D010060:Ovulation; D013050:Spectrometry, Fluorescence
C005045:3-hydroxykynurenine; D007737:Kynurenine