Validation of a conductometric bienzyme biosensor for the detection of proteins as marker of organic matter in river samples

作     者：KHADRO Basma NAMOUR Philippe BESSUEILLE Franois LEONARD Didier JAFFREZIC-RENAULT Nicole 

作者机构：University of Lyon Laboratory of AnalyticalCNRS UMR 5180 69622 Villeurbanne France Cemagref of Lyon 

出 版 物：《Journal of Environmental Sciences》 (环境科学学报（英文版）)

年 卷 期：2009年第21卷第4期

页      面：545-551页

核心收录：

学科分类：082803[工学-农业生物环境与能源工程] 08[工学] 0828[工学-农业工程] 0804[工学-仪器科学与技术] 

基　　金：supported by CEMAGREF Laboratory of Lyon 

主　　题：conductometric biosensor proteins enzyme natural water analyses total organic carbon organic nitrogen 

摘      要：This article describes a conductometric bi-layer based bienzyme biosensor for the detection of proteins as a marker of organic matter in rivers. Proteins were chosen to be used as indicators of urban pollution. The working mechanism of the bienzyme biosensor is based on the enzymatic hydrolysis of proteins into several fractions （peptides and amino acids）, which results in a local conductivity change depending of the concentration of proteins. In this work, we began with the optimization of biosensor response using bovine serum albumin （BSA） as standard protein. For this objective seven enzymatic biosensors were prepared： four enzymatic sensors with only one layer of enzyme （proteinase K, trypsin, pronase or protease X） and three other enzymatic sensors with two layers （first layer： membrane containing proteinase K; second layer： one of the three other enzymes： trypsin, pronase or protease X）. The biosensors were obtained through the deposition of enzymatic layers and the cross-linking process between enzymes and BSA in saturated glutaraldehyde vapour. The response of the various biosensors, described previously, were compared with the values of total organic carbon （TOC）, and those of organic nitrogen （No~）, as determined by the laboratory accredits （CEMAGREF of Lyon） using the traditional method of analysis （NF EN 1484, infrared spectroscopy） and （NF EN 25663, mineralization/colorimetry assay） respectively for each water sample obtained from different sites in Lyon （France）. The linear correlations obtained with the response of the seven biosensors showed the most important indices of correlations for the biosensor with two enzymatic layers： proteinase K ＋ pronase （pkp）. The optimum conditions for the preparation of the pkp biosensor increased the sensitivity and gave a limit of quantification of 0.583 μg/L for TOC and 0.218 μg/L for Norg in water samples. This sensor shows good reproducibility （2.28%）, a capacity to be used at temperatures range 10-- 30℃ （dependi