Development and validation of a liquid-chromatographic procedure for serum creatinine.

We evaluated and compared three analytical procedures for measuring creatinine by testing pooled sera and sera from individual human subjects. In two of the methods, an enzymic and a conventional colorimetric procedure, creatinine was separated from serum proteins with tungstic acid and detected at 520 nm after reaction with alkaline picrate (Jaff#{233} reagent). The amount of enzymaticaIly active creatinine was estimated by computing the difference between the creatinine concentrations in specimens incubated with and without creatinine deiminase (EC 3.5.4.21; creatinine iminohydrolase). For the other procedure we measured creatinine at 236 nm in serum ultrafiltrates injected on a reversed-phase high-performance liquid-chromatographic column, with allopurinol as an internal standard. Studies with serum pools showed that the chromatographic procedure was more precise than the enzymic and conventional tests. The routine Jaff#{233} method was not specific and produced erroneously high creatinine results for serum pools supplemented with ascorbate, pyruvate, acetone, and glucose. There was little or no interference with the chromatographic or enzymic techniques. Recovery of creatinine added to serum specimens was about 95% for the chromatographic procedures, but only 82% for both Jaff#{233} methods.