Center for Electrochemical Sciences (CES), Ruhr-Universität Bochum, D-44780 Bochum, Germany.
Department of Analytical Chemistry, Faculty of Chemistry, Jagiellonian University, 30-060 Krakow, Poland.
Biosens Bioelectron. 2018 Oct 15;117:501-507. doi: 10.1016/j.bios.2018.06.047. Epub 2018 Jun 25.
Despite the availability of numerous electroanalytical methods for phosphate quantification, practical implementation in point-of-use sensing remains virtually nonexistent because of interferences from sample matrices or from atmospheric O. In this work, phosphate determination is achieved by the purine nucleoside phosphorylase (PNP) catalyzed reaction of inosine and phosphate to produce hypoxanthine which is subsequently oxidized by xanthine oxidase (XOx), first to xanthine and then to uric acid. Both PNP and XOx are integrated in a redox active Os-complex modified polymer, which not only acts as supporting matrix for the bienzymatic system but also shuttles electrons from the hypoxanthine oxidation reaction to the electrode. The bienzymatic cascade in this second generation phosphate biosensor selectively delivers four electrons for each phosphate molecule present. We introduced an additional electrochemical process involving uric acid oxidation at the underlying electrode. This further enhances the anodic current (signal amplification) by two additional electrons per analyte molecule which mitigates the influence of electrochemical interferences from the sample matrix. Moreover, while the XOx catalyzed reaction is sensitive to O, the uric acid production and therefore the delivery of electrons through the subsequent electrochemical process are independent of the presence of O. Consequently, the electrochemical process counterbalances the O interferences, especially at low phosphate concentrations. Importantly, the electrochemical uric acid oxidation specifically reports on phosphate concentration since it originates from the product of the bienzymatic reactions. These advantageous properties make this bioelectrochemical-electrochemical cascade particularly promising for point-of-use phosphate measurements.
尽管有许多用于磷酸盐定量的电化学分析方法，但由于样品基质或大气 O 的干扰，实际上在现场即时感测中几乎无法实施。在这项工作中，通过嘌呤核苷磷酸化酶 (PNP) 催化肌苷和磷酸盐的反应来实现磷酸盐的测定，产生次黄嘌呤，然后次黄嘌呤被黄嘌呤氧化酶 (XOx) 氧化，首先氧化为黄嘌呤，然后氧化为尿酸。PNP 和 XOx 都整合在一个氧化还原活性 Os 配合物修饰的聚合物中，该聚合物不仅作为双酶系统的支撑基质，还将电子从次黄嘌呤氧化反应转移到电极。第二代磷酸盐生物传感器中的双酶级联反应选择性地为每个存在的磷酸盐分子提供四个电子。我们引入了一个额外的电化学过程，涉及底层电极上的尿酸氧化。这通过每个分析物分子额外的两个电子进一步增强了阳极电流（信号放大），从而减轻了样品基质电化学干扰的影响。此外，虽然 XOx 催化的反应对 O 敏感，但尿酸的产生，因此通过随后的电化学过程传递电子是独立于 O 的存在的。因此，电化学过程平衡了 O 的干扰，特别是在低磷酸盐浓度下。重要的是，电化学尿酸氧化专门报告磷酸盐浓度，因为它起源于双酶反应的产物。这些有利的特性使这种生物电化学-电化学级联反应特别有前途用于现场即时磷酸盐测量。
