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 = 65 nm in a pH 8.5 KH(2)PO(4)-NaOH buffer solution. The detected wavelengths of quantitative analysis were set at 410 nm for xanthopterin and 325 nm for isoxanthopterin, respectively. Pretreatment of urine samples only was filtrated through a 0.45 μm membrane filter, which was free from the tedious separation procedures. Under optimized conditions, the limits of detection (LOD) were 0.94 ng/mL for xanthopterin and 0.48 ng/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.

Development of a high-performance liquid chromatography – Tandem mass spectrometry urinary pterinomics workflow

“…More recent efforts have seen an attempt to develop robust analytical methodologies using advanced instrumental platforms and oxidative pretreatments to enable sensitive detection of biologically significant pteridine derivatives. To this end, a multitude of competent analytical techniques have emerged over the past several years using an array of sensitive instrumental platforms including capillary electrophoresislaser-induced fluorescence (CE-LIF) [35], high-performance liquid chromatographyfluorescence detection (HPLC-FD) [28,[36][37][38][39][40], high-performance liquid chromatographymass spectrometry (HPLC-MS) [41], high-performance liquid chromatographytandem mass spectrometry (HPLC-MS/MS) [29,[42][43][44], hydrophilic interaction chromatographytandem mass spectrometry (HILIC-MS/MS) [45], and synchronous fluorescence spectroscopy [46,47], which have been summarized in Table 1.…”

Steady state and time-resolved fluorescence spectroscopic characterization of normal and cancerous urine

“…For example, the native fluorescence of collagen, elastin and more generally proteins is related to the structural arrangement of cells and tissues. On the other hand, the native fluorescence from pyridoxine derivatives, reduced form of nicotinamide adenine dinucleotide (NADH), flavin, porphyrin and pteridine and its derivatives are related to metabolic changes [2,3]. Based on the report of Alfano and his group on the detection of dental carries and subsequently cancer using native fluorescence, the field of optical biopsy has been given importance in diagnostic oncology [4,5].…”

“…As these fluorophores distribution may be altered in urine under different pathological and metabolic alterations, determination of these metabolites were carried out using various tedious techniques, such as highperformance liquid chromatography, capillary electrophoresis and radioimmunoassay. For rapid characterisation, native fluorescence spectroscopic techniques have been considered in the characterisation of urine under different diseased conditions [3,10]. However, no detailed report on the characterisation and discrimination of urine of cancer patients from normal subjects using simple native fluorescence emission spectroscopy.…”

Characterization and Diagnosis of Cancer by Native Fluorescence Spectroscopy of Human Urine

“…Anwer et al had reported on the use of autofluorescence in discriminating the individuals with bacteriuria with the normal subjects and subsequently Sandeep Perinchery et al reported the autofluorescence of urine in the diagnosis of human urinary tract infection (20,22). Recently, Yi-Qun Wan made use of synchronous fluorescence spectroscopy and reported that the variation in isoxantopterin levels is significant in the urine of stomach cancer patients when compared with the normal subjects (24).…”

“…reported the autofluorescence of urine in the diagnosis of human urinary tract infection. Recently, Yi‐Qun Wan made use of synchronous fluorescence spectroscopy and reported that the variation in isoxantopterin levels is significant in the urine of stomach cancer patients when compared with the normal subjects .…”