Biocatalysis and Biosensor Laboratory, Department of Food Science and Technology, BOKU-University of Natural Resources and Life Sciences, Muthgasse 18, 1190 Vienna, Austria.
Analytical Chemistry-Center for Electrochemical Sciences (CES), Faculty of Chemistry and Biochemistry, Ruhr University Bochum, Universitätsstraße 150, 44780 Bochum, Germany.
Anal Chem. 2021 Jun 1;93(21):7732-7738. doi: 10.1021/acs.analchem.1c01182. Epub 2021 May 20.
Enzymatic hydrolysis of lignocellulosic biomass for biofuel production relies on complex multi-enzyme ensembles. Continuous and accurate measurement of the released key products is crucial in optimizing the industrial degradation process and also investigating the activity and interaction between the involved enzymes and the insoluble substrate. Amperometric biosensors have been applied to perform continuous cellobiose measurements during the enzymatic hydrolysis of pure cellulose powders. The oxygen-sensitive mediators used in these biosensors restricted their function under physiological or industrial conditions. Also, the combined measurements of the hydrolysis products cellobiose and glucose require a high selectivity of the biorecognition elements. We employed an [Os(2,2'-bipyridine)Cl]Cl-modified polymer and cellobiose dehydrogenase to fabricate a cellobiose biosensor, which can accurately and specifically detect cellobiose even in the presence of oxygen and the other main product glucose. Additionally, a glucose biosensor was fabricated to simultaneously measure glucose produced from cellobiose by β-glucosidases. The cellobiose and glucose biosensors work at applied potentials of +0.25 and +0.45 V versus Ag|AgCl (3 M KCl), respectively, and can selectively detect their substrate. Both biosensors were used in combination to monitor the hydrolysis of pure cellulose of low crystallinity or industrial corncob samples. The obtained results correlate with the high-performance liquid chromatography pulsed amperometric detection analysis and demonstrate that neither oxygen nor the presence of redox-active compounds from the lignin fraction of the corncob interferes with the measurements.
用于生物燃料生产的木质纤维素生物质的酶解依赖于复杂的多酶复合物。在优化工业降解过程以及研究所涉及的酶与不溶性底物之间的活性和相互作用时，释放的关键产物的连续和准确测量至关重要。电流型生物传感器已被应用于在纯纤维素粉末的酶解过程中进行连续的纤维二糖测量。这些生物传感器中使用的氧敏介体限制了它们在生理或工业条件下的功能。此外，水解产物纤维二糖和葡萄糖的联合测量需要生物识别元件具有高选择性。我们采用[Os（2,2'-联吡啶）Cl]Cl 修饰的聚合物和纤维二糖脱氢酶来制备纤维二糖生物传感器，即使在存在氧气和另一种主要产物葡萄糖的情况下，该生物传感器也可以准确且特异性地检测纤维二糖。此外，还制备了葡萄糖生物传感器来同时测量β-葡萄糖苷酶从纤维二糖产生的葡萄糖。纤维二糖和葡萄糖生物传感器的工作电位分别为+0.25 和+0.45 V 相对于 Ag|AgCl（3 M KCl），并且可以选择性地检测其底物。这两个生物传感器结合使用，用于监测低结晶度的纯纤维素或工业玉米芯样品的水解。所得结果与高效液相色谱脉冲安培检测分析相关联，并且表明，无论是氧气还是玉米芯木质素部分中的氧化还原活性化合物的存在都不会干扰测量。
