Simultaneous Determination of Xanthopterin and Isoxanthopterin in Human Urine by Synchronous Fluorescence Spectroscopy

Abstract

A simple, rapid, sensitive and selective method for simultaneously determining xanthopterin and isoxanthopterin content in human urine has been developed using synchronous fluorescence spectroscopy based on their intrinsic fluorescence. The synchronous fluorescence spectra were obtained with Δλ = 65nm in a pH 8.5 KH(2)PO(4)-NaOH buffer solution. The detected wavelengths of quantitative analysis were set at 410nm for xanthopterin and 325nm for isoxanthopterin, respectively. Pretreatment of urine samples only was filtrated through a 0.45μm membrane filter, which was free from the tedious separation procedures. Under optimized conditions, the limits of detection (LOD) were 0.94ng/mL for xanthopterin and 0.48ng/mL for isoxanthopterin. The recoveries ranged from 88.0% to 103.8 % for healthy and cancer urine samples, with coefficient of variation between 2.09% and 7.06%. The proposed method has been successfully applied to the simultaneous analysis for xanthopterin and isoxanthopterin in human urine. The results showed that the average level of isoxanthopterin was significantly elevated in urine excreted by stomach cancer patients (P < 0.01), while no significant change of xanthopterin level was found between stomach cancer patients and healthy individuals. This potentially indicates that an increase in amounts of isoxanthopterin can be associated with the presence of stomach cancer.