Determination of Xanthine and Uric Acid in Xanthinuric Urine and Extracellular Fluid of Porcine Endothelial Cells of the Pulmonary Artery by High Performance … (original) (raw)
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Simultaneous determination of purine derivatives in urine by high-performance liquid chromatography
Journal of Animal and Feed Sciences
A high-performance liquid chromatography method for the analysis of allantoin, uric acid, hypoxanthine and xanthine in urine of sheep is described. Urine samples were analyzed directly after dilution. The combination of HPLC reversed phase C 18 columns with monitoring the effluent at 205 nm provides a relatively rapid and simple analytical tool for studying purine derivatives in the urine of ruminants. Separation and quantification of purine derivatives was achieved using three C 18 columns connected in a series together with a NH 4 H,PC" 4-NH 4 H 2 P0 4 and methanol (95:5) gradient. The average recoveries of standard compounds added to urine samples were satisfaction (96-103%). The low value of the intra-assay coefficient of variation and high recovery point to the satisfactory precision and accuracy of the reported method.
Determination of purine derivatives in bovine urine using rapid chromatographic techniques
Archiva Zootechnica, 2009
In this study, two techniques (HPLC-UV and TLC) were assessed as alternatives to the analysis of purine derivatives (allantoin, uric acid, xanthine and hypoxanthine) using classical spectrometry method, still largely used in SouthEastern Europe. With HPLC technique, very good calibration curves were obtained for all four purine derivatives (r>0.999). Also, detection limits were satisfactory: ≥ 7.5 µg/mL for allantoin, ≥ 2.7 µg/mL for uric acid, ≥ 5.5 µg/mL for xanthine and ≥ 10.02 µg/mL for hypoxanthine. The analysis of urine samples taken from dairy cows gave results which were in the range of literature data for allantoin and uric acid, but not for hypoxanthine (whose concentration was much higher than values reported in literature). TLC technique allowed good separation of standards and elaboration of good calibration curves (r>0.984). Detection limits were too high, if the whole range of values reported in literature for the concentrations purine derivatives in ruminants' urine is taken into account. Moreover, on urine samples the spots of creatinie overlapped the spots of uric acids, impairing the quantitative estimation of purine derivatives. It is concluded that HPLC technique can be used to assess the concentration of allantoin and uric acid in the urine of cows. The TLC technique can be used qualitatively, further studies on samples preparation and procedure being needed in order to use it for quantitative determinations.
Journal of chromatography. B, Biomedical sciences and applications, 2000
A HPLC method for the determination of allantoin, uric acid, hypoxanthine and xanthine (purine metabolites) in ovine urine without the disadvantages inherent in derivatization is described. After dilution 1:6 with water, urine samples were injected onto the column. Separation and quantification of purine metabolites was achieved using two Nova-Pak C18 columns (4 microm, 250x4.6 mm, Waters). A binary gradient program and UV detection were used for purine metabolites analysis. Clear resolution of purine metabolites was obtained in less than 15 min. Allantoin, uric acid, hypoxanthine and xanthine in the effluent were monitored at 225, 284, 250 and 267 nm, respectively. The average recoveries of purine metabolite standards added to urine samples were satisfactory (100.2-102.9%). The low coefficients of variation (0.29-0.73%) as well as the low detection (0.16-0.70 nmol) and quantification (0.52-2.32 nmol) limits indicate satisfactory precision, reproducibility and sensitivity of the pro...
Determination of urinary methylated purine pattern by high-performance liquid chromatography
Journal of chromatography. B, Biomedical sciences and applications, 2001
We describe the group selective separation and quantification of unmodified and modified purines in human urine by high-performance reverse phase liquid chromatography. The pattern of oxypurines and methylated purines: hypoxanthine (Hx), xanthine (X), 1-methyl hypoxanthine (1-MHx), 1-methyl guanine (1-MG), 3-methyl guanine (3-MG), 7-methyl guanine (7-MG), 1-methyl xanthine (1-MX), 3-methyl xanthine (3-MX), 7-methyl xanthine (7-MX), 1,7-dimethyl guanine (1,7-dMG), 1,3-dimethyl xanthine (1,3-dMX), 1,7-dimethyl xanthine (3,7-dMX) and 1,3,7-trimethyl xanthine (1,3,7-tMX) were determined in a single run in urine of a healthy subject and a gout patient before and after treatment with allopurinol. This method may be useful to investigate the urinary pattern of methylated bases in diseases involving purine metabolism.
Physiologic Levels of Uric Acid Inhibit Xanthine Oxidase in Human Plasma
Pediatric Research, 1993
A B S T R A n. Xanthine osidase, a key source of reactive oxygen species, and purine substrates are detected in the circulation after ischemia-reperfusion. Iiigh levels of uric acid, produced by a santhine osidase-catalyzed reaction, are found in human plasma. \\'e studied whether uric acid could alter xanthine osidase activity in plasma obtained from eight adults and eight neonates. Known amounts of uric acid were added to santhine and santhine osidasesupplemented buffer and plasma, and the production of uric acid and superoside was determined. Uric acid, 150 and 300 phi, decreased the oxidation of santhine to uric acid in adult plasma by 37.5 f 5.6 and 48.9 f 6.1% and formation of superoside by 23.2 f 1.9 and 32.0 f 2.3%,
HPLC method for measurement of purine nucleotide degradation products in cerebrospinal fluid
Clinical chemistry, 1996
We describe a convenient method for the separation and quantification of xanthine, hypoxanthine, and uric acid in 20 microL of cerebrospinal fluid (CSF) with use of HPLC and ultraviolet detection. The analysis is performed on a Sepharon SGX C18 column and the elution system consists of potassium phosphate buffer, pH 5.1, with 20 mL/L methanol. The lower limit of detection was 4 pmol for hypoxanthine and xanthine and 6 pmol for uric acid. Analytical recoveries of purine metabolites ranged from 98.6% to 102.9%. The intra- and interassay CVs were <3%. The applicability of the method is illustrated with the determination of micromolar concentrations of xanthine, hypoxanthine, and uric acid in CSF samples obtained from 113 patients with various neurological disorders.
Journal of Trace and Microprobe Techniques, 2001
We describe a convenient method for the separation and quantification of xanthine, hypoxanthine, and uric acid in 20 µL samples of amniotic fluid using HPLC and UV detection. The analysis is performed on a Sepharon SGX C 18 column and the elution system consists of a 60-mmol/L potassium phosphate buffer, pH 4.6. The limit of detection is 7 pmol for hypoxanthine and xanthine and 10 pmol for uric acid. The intra-and interassay coefficients of variation are less than 5%. The applicability of the method is illustrated with the determination of micromolar concentrations of oxopurines in amniotic fluid samples obtained from 204 trimester pregnancies with an apparent increased risk of chromosome disorders.