kikkoman.co.jp, as data

Img. Ref.:NAR 46: W296–W303 (2018)### アプリケーション### 分析手順#### PrincipleThe appearance of urea is measured spectrophotometrically at 435 nm.#### Definition of unitOne unit (U) is defined as the amount of enzyme which produces 1 μmol of urea per min at 37℃ and pH 7.7 under the conditions described below.#### Reagents* Potassium phosphate buffer, 0.3 M; pH 7.7: mix 0.3 M KH2PO4 solution and 0.3 M K2HPO4 solution to make a pH 7.7 solution.* Creatine solution, 0.1 M: 1.49 g of creatine monohydrate/100 mL of distilled water.* p-Dimethylaminobenzaldehyde (DMAB) solution, 0.87%: dissolve 2.0 g of DMAB in 100 mL of ethanol (99.5%) and add 15 mL of conc. HCl and 115 mL of distilled water.* Enzyme dilution buffer: mix 10 mM KH2PO4 solution and 10 mM K2HPO4 solution to make a pH 8.0 solution and add 2-mercaptoethanol (0.16 mL/liter of the buffer).Sample: dissolve the lyophilized enzyme to a volume activity of 1.4–2.8 U/mL with ice-cold enzyme dilution buffer (Reagent D) immediately before measurement.#### Procedure1. Pipette the following reagents into a test tube.Potassium phosphate buffer2. Equilibrate at 37℃ for about 5 min.3. Add 0.1 mL of sample and incubate for 10 min at 37℃.4. Add 2.0 mL of DMAB solution (Reagent C) and allow to stand for about 30 min at 25℃.5. Read the absorbance at 435 nm in a cuvette (light path: 1 cm) (AS).The blank solution is prepared by reversing the sequence of addition of sample and DMAB solution (Reagent C) (A0).#### CalculationActivity can be calculated by using the following formula:### 実験データ### Line-up### FAQSuzuki, M, *Medical Technology*,** 7**, 945–950 (1979).A new enzymatic determination of serum creatine*Clinica Chimica Acta*, **140,** 289–294.A new enzymatic serum creatinine measurement based on an endogenous creatine-eliminating system*Clinica Chimica Acta*, **143**, 147–155.