Open AccessNew enzymatic assay for serum urea nitrogen using urea amidolyase
Author(s) -
Kimura Shigeki,
Iyama Shigeru,
Yamaguchi Yoshihisa,
Kanakura Yuzuru
Publication year - 2003
Publication title -
journal of clinical laboratory analysis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.536
H-Index - 50
eISSN - 1098-2825
pISSN - 0887-8013
DOI - 10.1002/jcla.10067
Subject(s) - chemistry , urea , glutamate dehydrogenase , ammonium , chromatography , absorbance , enzyme , nitrogen , nuclear chemistry , biochemistry , organic chemistry , glutamate receptor , receptor
Abstract We established an enzymatic assay for measurement of serum urea nitrogen using urea amidolyase (EC 3.5.1.45) from yeast species. The method is based on hydrolysis of urea by the enzyme. In this assay, we eliminated endogenous ammonium ion by use of glutamate dehydrogenase (EC 1.4.1.4). Then in the presence of urea amido‐lyase, ATP, bicarbonate, magnesium, and potassium ions, ammonium ion was produced proportionally to urea concentration in serum. The concentra‐tion of ammonium ion formed was determined by adding GLDH to produce NADP + in the presence of 2‐oxoglutarate and NADPH. We then monitored the change of absorbance at 340 nm. The inhibitory effect of calcium ion on this assay was eliminated by adding glyco‐letherdiamine‐N, N, N′, N′‐tetraacetic acid to the reaction system. The with‐in‐assay coefficient of variations (CVs) of the present method were 1.80–3.76% (n = 10) at 2.8–19.0 mmol/L, respectively. The day‐to‐day CVs were 2.23–4.59%. Analytical recovery was 92–115%. The presence of ascorbic acid, bilirubin, hemoglobin, lipemic material, ammo‐nium ion, or calcium ion did not affect this assay system. The correlation be‐tween values obtained with the present method (y) and those by another enzy‐matic method (x) was 0.997 (y = 1.02x − 0.10 mmol/L, Sy/x = 0.841, n = 100), with a mean difference of –0.18 ± 0.86 mmol/L [(values by reference method – that of present method) ± SD] using the Bland‐Altman technique. J. Clin. Lab. Anal. 17:52–56, 2003. © 2003 Wiley‐Liss, Inc.