Electrochemical Trimethylamine N-Oxide Biosensor with Enzyme-Based Oxygen-Scavenging Membrane for Long-Term Operation under Ambient Air

Abstract

An amperometric trimethylamine N-oxide (TMAO) biosensor is reported, where TMAO reductase (TorA) and glucose oxidase (GOD) and catalase (Cat) were immobilized on the electrode surface, enabling measurements of mediated enzymatic TMAO reduction at low potential under ambient air conditions. The oxygen anti-interference membrane composed of GOD, Cat and polyvinyl alcohol (PVA) hydrogel, together with glucose concentration, was optimized until the O2 reduction current of a Clark-type electrode was completely suppressed for at least 3 h. For the preparation of the TMAO biosensor, Escherichia coli TorA was purified under anaerobic conditions and immobilized on the surface of a carbon electrode and covered by the optimized O2 scavenging membrane. The TMAO sensor operates at a potential of −0.8 V vs. Ag/AgCl (1 M KCl), where the reduction of methylviologen (MV) is recorded. The sensor signal depends linearly on TMAO concentrations between 2 µM and 15 mM, with a sensitivity of 2.75 ± 1.7 µA/mM. The developed biosensor is characterized by a response time of about 33 s and an operational stability over 3 weeks. Furthermore, measurements of TMAO concentration were performed in 10% human serum, where the lowest detectable concentration is of 10 µM TMAO.

Nanomaterial-Based Electrochemical Nanodiagnostics for Human and Gut Metabolites Diagnostics: Recent Advances and Challenges

“…Through quantitative investigation, the limit of quantitation (LOQ) for TMAO of <6.7 μM was achieved with wide linearity ranging from 15.6–500 μM as proof of concept [ 171 ]. Waffo et al [ 172 ] also fabricated an amperometric TMAO biosensor in which glucose oxidase (GOD), TMAO reductase (TorA), and catalase (Cat) were attached to the surface of the electrode, allowing enzymatic TMAO reduction measurements, as shown in Figure 8 iii. The sensor depicted linearity for concentrations of TMAO that range between 2–15 mM, having sensitivity 2.75 1.7 μA/mM, 33 s of response time, and 3 weeks stability.…”

“… Metabolite Nanomaterial Based Biosensing Platform Electrochemical Technique Limit of Detection (LOD) Linear Detection Range (LDR) Real Sample Ref. TMAO MIP/ITO DPV 1 ppm 1–15 ppm Urine [ 15 ] PAH@MnO 2 - <6.7 μM 15.6 to 500 μM Urine [ 171 ] TorA/GOD/Cat Amperometry 10 µM 2 µM–15 mM Human serum [ 172 ] S.loihica PV-4 Chronoamperometry 5.96 lM 0 to 250 µM Real serum [ 173 ] Indoxyl sulfate (IS) GR-SPE SWV 0.064 μM 0.5–80 μM Human serum and urine [ 181 ] Carbon composite film electrode Voltammetric ...…”

“…[ 171 ], copyright 2021 MDPI]; ( iii ) TMAO biosensor with various components [Adapted with permission from ref. [ 172 ], copyright 2021 MDPI]; and ( iv ) schematic sketch of microbial electrochemical technology established for the TMAO detection [Adapted with permission from ref. [ 173 ], copyright 2022 Elsevier].…”

Application of an Imprint‐and‐Report Sensor Array for Detection of the Dietary Metabolite Trimethylamine N‐Oxide and Its Precursors in Complex Mixtures

“…Several electrochemical and fluorescence detection methods for TMAO have been reported, [18–23] but these sensors are limited in that they do not address differentiation of TMAO from other related metabolites or test for interference from these metabolites. Additionally, many of them have high limits of detection that are not relevant to analysis of TMAO in plasma samples [18–20, 23] . Thus, we sought to develop a high‐throughput sensor for this class of metabolites using fluorescence detection that functions at physiological plasma concentrations.…”

“…Existing methods for measuring TMAO and its associated metabolites are typically timeconsuming and expensive, such as tandem MS-MS with isotope-labeled internal standards and stable isotope dilution LCMS. [13][14][15][16][17] Several electrochemical and fluorescence detection methods for TMAO have been reported, [18][19][20][21][22][23] but these sensors are limited in that they do not address differentiation of TMAO from other related metabolites or test for interference from these metabolites. Additionally, many of them have high limits of detection that are not relevant to analysis of TMAO in plasma samples.…”