A new enzymatic assay of urinary guanidinoacetic acid.
We describe a new enzymatic determination of urinary guanidinoacetic acid (GAA) with guanidinoacetate kinase (ATP: guanidinoacetate N-phosphotransferase, EC 2.7.3.1), which does not require a blank to correct for endogenous constituents (ADP and pyruvate). In the first step, pyruvate kinase (ATP: pyruvate 2-O-phosphotransferase, EC 2.7.1.40) and lactate dehydrogenase (L-lactate: NAD+ oxidoreductase, EC 1.1.1.27) were used to eliminate endogenous constituents (ADP and pyruvate) in the presence of phosphoenolpyruvate and NADH. In the second step, urinary GAA was phosphorylated in the presence of ATP by guanidinoacetate kinase to form phosphoguanidinoacetate and ADP. The resultant ADP was sequentially measured at 340 nm in a coupled reaction catalyzed by pyruvate kinase and lactate dehydrogenase. The standard curve was linear up to 20 mg/dl for standard solutions of GAA. Analytical recovery of GAA added to normal urines ranged from 97.0 to 103.2% (mean 100.7%). The within-run and between-run studies gave CV values of less than or equal to 3.6% and less than or equal to 4.8%, respectively. No significant interference by endogenous urinary compounds were observed with the proposed method under this study. The results obtained by the present method correlated well with those obtained by a high-performance liquid chromatographic method. This method is accurate and simple, and less time-consuming than those previously reported. We determined the concentrations of GAA in 24-h urine samples by the proposed method, and observed that the urinary excretion of GAA decreased markedly in patients with renal failure.
Shirokane
Y
Y
Research & Development Division of Kikkoman Corporation, Chiba-ken, Japan.
Nakajima
M
M
Mizusawa
K
K
D000328:Adult; D000368:Aged; D015415:Biomarkers / Q000652:urine; D005260:Female; D005998:Glycine / Q000031:analogs & derivatives* / Q000652:urine; D006801:Humans; D007677:Kidney Function Tests; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008297:Male; D008875:Middle Aged; D010770:Phosphotransferases / Q000378:metabolism; D017852:Phosphotransferases (Nitrogenous Group Acceptor); D011770:Pyruvate Kinase / Q000378:metabolism; D012016:Reference Values; D013056:Spectrophotometry, Ultraviolet / Q000379:methods
D015415:Biomarkers; D007770:L-Lactate Dehydrogenase; D010770:Phosphotransferases; D011770:Pyruvate Kinase; D017852:Phosphotransferases (Nitrogenous Group Acceptor); C074685:guanidinoacetate kinase; C004946:glycocyamine; D005998:Glycine