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Hindi Research Article
Spectrophotometric Determination of Uric Acid in Urine Based-Enzymatic Method Uricase with 4-Aminodiphenylamine Diazonium Sulfate (Variamine Blue RT Salt)
Hairul Hisham Hamzah1,3*, Zainiharyati Mohd Zain2 , Nor Lailatul Wahidah Musa3 , Yun-Chun Lin1 and Emma Trimbee1
1School of Chemistry, University of Southampton, Highfield Campus, SO17 1BJ, Southampton, UK
2Faculty of Applied Sciences, Universiti Teknologi MARA (UiTM), 40450, Shah Alam Selangor, Malaysia
3Faculty of Applied Sciences, Universiti Teknologi MARA Pahang, 26400, Jengka Pahang, Malaysia
- *Corresponding Author:
- Hairul Hisham Hamzah, PhD
School of Chemistry, University of Southampton
Highfield Campus, Southampton SO17 1BJ, UK
E-mail: hhh1v12@soton.ac.uk
Received date: March 30, 2013; Accepted date: June 05, 2013; Published date: June 07, 2013
Citation: Hamzah HH, Zain ZM, Musa NLW, Lin YC, Trimbee E (2013) Spectrophotometric Determination of Uric Acid in Urine Based-Enzymatic Method Uricase with 4-Aminodiphenylamine Diazonium Sulfate (Variamine Blue RT Salt). J Anal Bioanal Tech S7:011. doi: 10.4172/2155-9872.S7-011
Copyright: © 2013 Hamzah HH, et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Abstract
A simple spectrophotometric method based-uricase enzyme for the detection of uric acid in normal urine and gout patient’s urine samples has been developed based on the reaction of hydrogen peroxide (H2O2) with yellow color of 4-Aminodiphenylamine Diazonium Sulfate (variamine blue RT salt) to yield a pale yellow-green coloured solution. The absorbance of products as the output of enzymatic reaction and variamine blue RT salt were detected using uv-visible spectrophotometer with maximum absorbance at 269 nm. The response time of this proposed method was 20 minutes. The optimum pH activity on enzymatic and variamine blue RT salt reactions was foud to be pH 9. The relative standard deviation (RSD) of the reproducibility of this method was very good at 0.7%. The calibration plot of different concentrations of uric acid was found to be between 0.5-13 mM with a detection limit of 0.58 mM. The kinetic parameters the Michaelis-Menten constant (KM) and the maximum abosrbance (Absmax) in the presence of different concentrations of uric acid were evaluated using the Linewaver-Burk and Hofstee plots respectively. The enzymatic-spectrophotometric method was compared with established clinicalmethod and satisfactory agreement was achieved.
