Enzymatic measurement of creatine in erythrocytes

Abstract

Abstract We describe an automated enzymatic procedure for quantifying creatine in erythrocytes. In this assay, after sample clean-up, creatine kinase (CK; EC 2.7.3.2) and pyruvate kinase (EC 2.1.7.40) are used as auxiliary enzymes; lactate dehydrogenase (EC 1.1.1.27) is used in the indicator reaction. CK is also used as the starting reagent. Data obtained with the present method (y) correlated as follows with those from the comparison method (x) for creatine measurement in erythrocytes (colorimetric, diacetyl-alpha-naphthol): y = 1.017x - 27.5 (S(y/x) = 11.9, r = 0.997). Precision data were obtained by analyzing three different samples of erythrocytes nine times per day for 25 days with both methods. The total CVs determined by our method and the comparison method were respectively 3.0-4.8% (measured creatine range, 171-463 micromol/L) and 5.1-7.1% (creatine, 185-485 micromol/L). We established the reference interval (mean +/- 2SD) for creatine in erythrocytes from healthy individuals as 194-538 micromol/L for men (n = 71) and 241-677 micromol/L for women (n = 100). Using erythrocytes from patients with various hematological abnormalities and chronic renal failure, who were being treated with continuous ambulatory peritoneal dialysis or hemodialysis, we performed a clinical evaluation and obtained results in agreement with existing data on biochemical markers for the mean age of erythrocytes.