Development and Validation of a Simple High Performance Liquid Chromatography/UV Method for Simultaneous Determination of Urinary Uric Acid, Hypoxanthine, and Creatinine in Human Urine
Uric acid and hypoxanthine are produced in the catabolism of purine. Abnormal urinary levels of these products are associated with many diseases and therefore it is necessary to have a simple and rapid method to detect them. Hence, we report a simple reverse phase high performance liquid chromatogra...
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| Language: | English |
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Wiley
2018-01-01
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| Series: | International Journal of Analytical Chemistry |
| Online Access: | http://dx.doi.org/10.1155/2018/1647923 |
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| author | Nimanthi Wijemanne Preethi Soysa Sulochana Wijesundara Hemamali Perera |
| author_facet | Nimanthi Wijemanne Preethi Soysa Sulochana Wijesundara Hemamali Perera |
| author_sort | Nimanthi Wijemanne |
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| description | Uric acid and hypoxanthine are produced in the catabolism of purine. Abnormal urinary levels of these products are associated with many diseases and therefore it is necessary to have a simple and rapid method to detect them. Hence, we report a simple reverse phase high performance liquid chromatography (HPLC/UV) technique, developed and validated for simultaneous analysis of uric acid, hypoxanthine, and creatinine in human urine. Urine was diluted appropriately and eluted with C-18 column 100 mm × 4.6 mm with a C-18 precolumn 25 mm × 4.6 mm in series. Potassium phosphate buffer (20 mM, pH 7.25) at a flow rate of 0.40 mL/min was employed as the solvent and peaks were detected at 235 nm. Tyrosine was used as the internal standard. The experimental conditions offered a good separation of analytes without interference of endogenous substances. The calibration curves were linear for all test compounds with a regression coefficient, r2>0.99. Uric acid, creatinine, tyrosine, and hypoxanthine were eluted at 5.2, 6.1, 7.2, and 8.3 min, respectively. Intraday and interday variability were less than 4.6% for all the analytes investigated and the recovery ranged from 98 to 102%. The proposed HPLC procedure is a simple, rapid, and low cost method with high accuracy with minimum use of organic solvents. This method was successfully applied for the determination of creatinine, hypoxanthine, and uric acid in human urine. |
| format | Article |
| id | doaj-art-4508e7eedabd4393943d747f2fd757e6 |
| institution | Kabale University |
| issn | 1687-8760 1687-8779 |
| language | English |
| publishDate | 2018-01-01 |
| publisher | Wiley |
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| series | International Journal of Analytical Chemistry |
| spelling | doaj-art-4508e7eedabd4393943d747f2fd757e62025-02-03T05:57:58ZengWileyInternational Journal of Analytical Chemistry1687-87601687-87792018-01-01201810.1155/2018/16479231647923Development and Validation of a Simple High Performance Liquid Chromatography/UV Method for Simultaneous Determination of Urinary Uric Acid, Hypoxanthine, and Creatinine in Human UrineNimanthi Wijemanne0Preethi Soysa1Sulochana Wijesundara2Hemamali Perera3Department Biochemistry and Molecular Biology, Faculty of Medicine, University of Colombo, Colombo, Sri LankaDepartment Biochemistry and Molecular Biology, Faculty of Medicine, University of Colombo, Colombo, Sri LankaDepartment Biochemistry and Molecular Biology, Faculty of Medicine, University of Colombo, Colombo, Sri LankaDepartment of Psychological Medicine, Faculty of Medicine, University of Colombo, Colombo, Sri LankaUric acid and hypoxanthine are produced in the catabolism of purine. Abnormal urinary levels of these products are associated with many diseases and therefore it is necessary to have a simple and rapid method to detect them. Hence, we report a simple reverse phase high performance liquid chromatography (HPLC/UV) technique, developed and validated for simultaneous analysis of uric acid, hypoxanthine, and creatinine in human urine. Urine was diluted appropriately and eluted with C-18 column 100 mm × 4.6 mm with a C-18 precolumn 25 mm × 4.6 mm in series. Potassium phosphate buffer (20 mM, pH 7.25) at a flow rate of 0.40 mL/min was employed as the solvent and peaks were detected at 235 nm. Tyrosine was used as the internal standard. The experimental conditions offered a good separation of analytes without interference of endogenous substances. The calibration curves were linear for all test compounds with a regression coefficient, r2>0.99. Uric acid, creatinine, tyrosine, and hypoxanthine were eluted at 5.2, 6.1, 7.2, and 8.3 min, respectively. Intraday and interday variability were less than 4.6% for all the analytes investigated and the recovery ranged from 98 to 102%. The proposed HPLC procedure is a simple, rapid, and low cost method with high accuracy with minimum use of organic solvents. This method was successfully applied for the determination of creatinine, hypoxanthine, and uric acid in human urine.http://dx.doi.org/10.1155/2018/1647923 |
| spellingShingle | Nimanthi Wijemanne Preethi Soysa Sulochana Wijesundara Hemamali Perera Development and Validation of a Simple High Performance Liquid Chromatography/UV Method for Simultaneous Determination of Urinary Uric Acid, Hypoxanthine, and Creatinine in Human Urine International Journal of Analytical Chemistry |
| title | Development and Validation of a Simple High Performance Liquid Chromatography/UV Method for Simultaneous Determination of Urinary Uric Acid, Hypoxanthine, and Creatinine in Human Urine |
| title_full | Development and Validation of a Simple High Performance Liquid Chromatography/UV Method for Simultaneous Determination of Urinary Uric Acid, Hypoxanthine, and Creatinine in Human Urine |
| title_fullStr | Development and Validation of a Simple High Performance Liquid Chromatography/UV Method for Simultaneous Determination of Urinary Uric Acid, Hypoxanthine, and Creatinine in Human Urine |
| title_full_unstemmed | Development and Validation of a Simple High Performance Liquid Chromatography/UV Method for Simultaneous Determination of Urinary Uric Acid, Hypoxanthine, and Creatinine in Human Urine |
| title_short | Development and Validation of a Simple High Performance Liquid Chromatography/UV Method for Simultaneous Determination of Urinary Uric Acid, Hypoxanthine, and Creatinine in Human Urine |
| title_sort | development and validation of a simple high performance liquid chromatography uv method for simultaneous determination of urinary uric acid hypoxanthine and creatinine in human urine |
| url | http://dx.doi.org/10.1155/2018/1647923 |
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