Simple high-performance liquid chromatographic method to analyze serum creatinine has several advantages over the Jaffé picric acid reaction as demonstrated with a cimetidine dose response in rhesus monkeys.
A simple method for creatinine determination was developed using high-performance liquid chromatography (HPLC) to more accurately monitor serum creatinine levels in experimental animal models when compared to the Jaffé method. The new HPLC procedure will replace the traditional Jaffé method for rhesus monkey kidney function studies. We developed an isocratic method using a polymeric, hydrophilic, silica-based strong cation-exchange bed with a 5.0 mmol/l lithium acetate matrix, pH 4.9, which isolates creatinine with no detectable impurities as determined by three-dimensional ultraviolet-visible spectral analysis. Sample preparation includes deproteination with acetonitrile, evaporation, and resolubilization in mobile phase followed by quantitation with UV detection at 234 nm. Extraction efficiency across the measured range was 96 +/- 2%. From numerous extracted rhesus monkey creatinine curves (n=38) a slope of 251,100 +/- 756 (95% CI) and an intercept of 675.6 +/- 712.7 (95% CI) was calculated. Extraction efficiency and peak purity tests with human plasma were cross-compared with rhesus monkey serum producing equivalent results. An average of 120 samples can be run daily.
Johns
B A
BA
Department of Pharmacology, Merck Research Laboratories, West Point, PA 19486, USA.
Broten
T
T
Stranieri
M T
MT
Holahan
M A
MA
Cook
J J
JJ
D000818:Animals; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D002927:Cimetidine / Q000008:administration & dosage*; D003404:Creatinine / Q000097:blood*; D005260:Female; D006801:Humans; D008253:Macaca mulatta; D008297:Male; D010851:Picrates / Q000737:chemistry*; D012680:Sensitivity and Specificity; D013056:Spectrophotometry, Ultraviolet
D010851:Picrates; D002927:Cimetidine; C005858:picric acid; D003404:Creatinine