A MICROCOLORIMETRIC DETERMINATION OF CREATINE IN URINE BY THE JAFFE REACTION

Abstract

In 1945, a method was described by Bonsnes and the writer (1) for the determination of urinary creatine which involved only the addition of picric acid to dilute urine solutions and heating in a boiling water bath. An 80 per cent conversion of creatine to creatinine was obtained. The method has now been modified to achieve 100 per cent conversion, on the basis of Clark and Thompson’s recent studies (2) on the effect of pH on the quantitative conversion of creatine to creatinine. They have shown that 80 per cent of creatine is converted at pH 1, 85 per cent at pH 1.5, 98 to 100 per cent at pH 2 to 2.5, and that with increasing pH the per cent conversion is considerably decreased, falling below 50 per cent at pH 4.5. In the light of Clark and Thompson’s observations, the pH at the start and finish of the heating procedure employed in the original method of Bonsnes and Taussky was reinvestigated. It was found to be slightly more acid than requisite to achieve complete conversion of creatine to creatinine. By increasing the initial total volume it was possible to obtain a mixture of the appropriate pH range for complete conversion. Optimal conditions were established for the quantitative conversion of 10 to 80 y of creatine, a convenient range for a wide variety of urine concentrations. Under these experimental conditions glucose, which normally interferes with the determination of creatine by the Benedict, method (3) does not, do so in amounts up to 60 gm. per liter of urine. Preliminary reaction of the urine with iodine was introduced to eliminate the interference of acetone, acetoacetic ester, and ascorbic acid.