Simultaneous Determination of Creatinine and Uric Acid in Human Urine by High-Performance Liquid Chromatography

Abstract

An environmentally friendly reversed-phase HPLC method for simultaneous determination of creatinine and uric acid in human urine samples has been developed. Human urine samples were pretreated by dilution, protein precipitation, centrifugation and filtration, followed by HPLC separations using a reversed-phase C18 column with an aqueous mobile phase of phosphate buffer. The retention loss of a C18 column when using the highly aqueous mobile phases was avoided by employing a gradient elution using a small volume (<0.23 mL) of acetonitrile and phosphate buffer at pH 4.75. This developed method provides a simple, rapid separation and sensitive detection for the species of interest in 10 min with UV detection at 205 nm. Quantitation was carried out by relating the peak areas of the identified compounds to that of hypoxanthine as an internal standard. The detection limits for creatinine and uric acid were 0.045 and 0.062 mg mL -1 , respectively. The recoveries of the standards added to urine samples were 87.3 -102.2% for creatinine and 97.3 -108.6% for uric acid, and the relative standard deviation for both analytes was less than 1.0%. This method has been successfully applied to estimating of creatinine and uric acid in human urine.

Validation of a multi-analyte HPLC-DAD method for determination of uric acid, creatinine, homovanillic acid, niacinamide, hippuric acid, indole-3-acetic acid and 2-methylhippuric acid in human urine

“…So far only a few methods covering some of the above mentioned analytes in urine were described. For example Zuo et al described a HPLC-UV method for CREA and UA, while Shirao et al analyzed CREA, HVA and UA using miscellar electrokinetic chromatography [17,18]. Antunes et al validated a HPLC-UV method for CREA and UA in dried urine spots [11].…”

Changes in urinary nanocrystallites in calcium oxalate stone formers before and after potassium citrate intake

“…9,10 Using a spectrophotometer, the UA content was determined by its ability to reduce Fe (III) to Fe (II), which can then coordinate with phenanthroline to produce orangered complexes. 11 …”

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“…Zuo et al 12 developed an environmentally friendly reversed-phase HPLC method for simultaneous determination of creatinine and UA in human urine samples, which were pretreated by dilution, protein precipitation, centrifugation and filtration, followed by HPLC separations using a reversed-phase C18 column with an aqueous mobile phase of phosphate buffer. This developed method provided a simple, rapid separation and sensitive detection for the species of interest in 10 min with UV detection at 205 nm.…”