Measuring phosphate accurately in the field — without a lab, without interference from oxygen or complex sample matrices — has long been a challenge. A collaborative research team has published a second-generation electrochemical phosphate biosensor that changes that, with NECi's purine nucleoside phosphorylase (PNP) enzyme at its core. The result: reliable, point-of-use phosphate quantification even in the hands of non-trained users.
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Highlights
- Second generation electrochemical phosphate biosensor
- Bioelectrocatalytic–electrochemical cascade bypasses O₂ interferences
- Phosphate quantification possible in complex sample matrices
- Practical point-of-use phosphate sensing, even for non-trained users
Plain-Language Summary
The biosensor uses NECi's PNP enzyme alongside xanthine oxidase (XOx) in a two-enzyme cascade. When phosphate is present, the enzymes convert inosine into hypoxanthine and then uric acid, generating up to 6 electrons per phosphate molecule — a strong, measurable signal. A secondary electrochemical step using uric acid oxidation further amplifies the signal and specifically counteracts oxygen interference, making the sensor reliable even at low phosphate concentrations in real-world samples.
Abstract
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 O₂. 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 O₂, the uric acid production and therefore the delivery of electrons through the subsequent electrochemical process are independent of the presence of O₂. Consequently, the electrochemical process counterbalances the O₂ 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.
Authors
Gabriel Kopiec, Karolina Starzec, Jolanta Kochana, Troy P. Kinnunen-Skidmore, Wolfgang Schuhmann, Wilbur H. Campbell, Adrian Ruff, Nicolas Plumeré
Contributing Organizations
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
The Nitrate Elimination Co., Inc. (NECi), Lake Linden, MI 49945, United States
