Akyilmaz Erol, Erdoğan Ali, Oztürk Ramazan, Yaşa Ihsan
Department of Biochemistry, Faculty of Science, Ege University, 35100 Bornova-Izmir, Turkey.
Biosens Bioelectron. 2007 Jan 15;22(6):1055-60. doi: 10.1016/j.bios.2006.04.023. Epub 2006 Jun 8.
A new amperometric microbial biosensor based on Saccharomyces cerevisiae NRRL-12632 cells, which had been induced for lysine oxidase enzyme and immobilized in gelatin by a cross-linking agent was developed for the sensitive determination of L-lysine amino acid. To construct the microbial biosensor S. cerevisiae cells were activated and cultured in a suitable culture medium. By using gelatine (8.43 mg cm(-2)) and glutaraldehyde (0.25%), cells obtained in the logarithmic phase of the growth curve at the end of a 14 h period were immobilized and fixed on a pretreated oxygen sensitive Teflon membrane of a dissolved oxygen probe. The assay procedure of the microbial biosensor is based on the determination of the differences of the respiration activity of the cells on the oxygenmeter in the absence and the presence of L-lysine. According to the end point measurement technique used in the experiments it was determined that the microbial biosensor response depended linearly on L-lysine concentrations between 1.0 and 10.0 microM with a 1 min response time. In optimization studies of the microbial biosensor, the most suitable microorganism quantities were found to be 0.97x10(5)CFU cm(-2). In addition phosphate buffer (pH 7.5; 50 mM) and 30 degrees C were obtained as the optimum working conditions. In characterization studies of the microbial biosensor some parameters such as substrate specificity, interference effects of some substances on the microbial biosensor responses, reproducibility of the biosensor and operational and storage stability were investigated.
基于酿酒酵母NRRL - 12632细胞构建了一种新型安培型微生物生物传感器，该细胞经赖氨酸氧化酶诱导，并通过交联剂固定在明胶中，用于灵敏测定L - 赖氨酸氨基酸。为构建微生物生物传感器，酿酒酵母细胞在合适的培养基中活化并培养。使用明胶（8.43 mg cm(-2)）和戊二醛（0.25%），将在14小时培养期结束时处于生长曲线对数期获得的细胞固定并附着在经预处理的溶解氧探头的氧敏感聚四氟乙烯膜上。微生物生物传感器的测定程序基于在氧测定仪上测定有无L - 赖氨酸时细胞呼吸活性的差异。根据实验中使用的终点测量技术，确定微生物生物传感器的响应在1.0至10.0 microM的L - 赖氨酸浓度范围内呈线性，响应时间为1分钟。在微生物生物传感器的优化研究中，发现最合适的微生物数量为0.97x10(5)CFU cm(-2)。此外，获得磷酸盐缓冲液（pH 7.5；50 mM）和30℃作为最佳工作条件。在微生物生物传感器的表征研究中，研究了一些参数，如底物特异性、某些物质对微生物生物传感器响应的干扰效应、生物传感器的重现性以及操作和储存稳定性。
