In Vitro Evaluation of Miniaturized Amperometric Enzyme Sensor Based on the Direct Electron Transfer Principle for Continuous Glucose Monitoring

2022 ◽  
pp. 193229682110706
Author(s):  
Yutaro Inoue ◽  
Yasuhide Kusaka ◽  
Kotaro Shinozaki ◽  
Inyoung Lee ◽  
Koji Sode
Keyword(s):  
Electron Transfer ◽  
Continuous Glucose Monitoring ◽  
Glucose Sensor ◽  
Direct Electron Transfer ◽  
Glucose Dehydrogenase ◽  
Glucose Monitoring ◽  
Reference Electrode ◽  
Enzyme Sensor

Background: The bacterial derived flavin adenine dinucleotide (FAD)-dependent glucose dehydrogenase (FADGDH) is the most promising enzyme for the third-generation principle-based enzyme sensor for continuous glucose monitoring (CGM). Due to the ability of the enzyme to transfer electrons directly to the electrode, recognized as direct electron transfer (DET)-type FADGDH, although no investigation has been reported about DET-type FADGDH employed on a miniaturized integrated electrode. Methods: The miniaturized integrated electrode was formed by sputtering gold (Au) onto a flexible film with 0.1 mm in thickness and divided into 3 parts. After an insulation layer was laminated, 3 openings for a working electrode, a counter electrode and a reference electrode were formed by dry etching. A reagent mix containing 1.2 × 10−4 Unit of DET-type FADGDH and carbon particles was deposited. The long-term stability of sensor was evaluated by continuous operation, and its performance was also evaluated in the presence of acetaminophen and the change in oxygen partial pressure (pO2) level. Results: The amperometric response of the sensor showed a linear response to glucose concentration up to 500 mg/dL without significant change of the response over an 11-day continuous measurement. Moreover, the effect of acetaminophen and pO2 on the response were negligible. Conclusions: These results indicate the superb potential of the DET-type FADGDH-based sensor with the combination of a miniaturized integrated electrode. Thus, the described miniaturized DET-type glucose sensor for CGM will be a promising tool for effective glycemic control. This will be further investigated using an in vivo study.

Bioconversion of Carbon Monoxide to Formate Using Artificially Designed Carbon Monoxide:Formate Oxidoreductase in Hyperthermophilic Archaea

2021 ◽  
Author(s):  
Jae Kyu Lim ◽  
Ji-In Yang ◽  
Yun Jae Kim ◽  
Yeong-Jun Park ◽  
Yong Hwan Kim
Keyword(s):  
Carbon Monoxide ◽  
Electron Transfer ◽  
Fusion Protein ◽  
Direct Electron Transfer ◽  
Co2 Reduction ◽  
Hyperthermophilic Archaea ◽  
Production Activity ◽  
Formate Production

Abstract Ferredoxin-dependent metabolic engineering of electron transfer circuits has been developed to enhance redox efficiency in the field of synthetic biology, e.g., for hydrogen production and for reduction of flavoproteins or NAD(P)+. Here, we present the bioconversion of carbon monoxide (CO) gas to formate via a synthetic CO:formate oxidoreductase (CFOR), designed as an enzyme complex for direct electron transfer between noninteracting CO dehydrogenase and formate dehydrogenase using an electron-transferring Fe-S fusion protein. The CFOR-introduced Thermococcus onnurineus mutant strains showed CO-dependent formate production in vivo and in vitro. The formate production rate from purified CFOR complex and specific formate productivity from the bioreactor were 348 ± 34 μmol/mg/min and 90.2 ± 20.4 mmol/g-cells/h, respectively. The CO-dependent CO2 reduction/formate production activity of synthetic CFOR was confirmed, indicating that direct electron transfer between two unrelated dehydrogenases was feasible via mediation of the FeS-FeS fusion protein.

Engineered PQQ Glucose Dehydrogenase‐Based Enzyme Sensor for Continuous Glucose Monitoring

2004 ◽  
Vol 37 (9) ◽  
pp. 1847-1857 ◽  
Author(s):  
Junko Okuda ◽  
Junko Wakai ◽  
Satoshi Igarashi ◽  
Koji Sode
Keyword(s):  
Continuous Glucose Monitoring ◽  
Glucose Dehydrogenase ◽  
Glucose Monitoring ◽  
Enzyme Sensor ◽  
Pqq Glucose Dehydrogenase

scholarly journals Engineered Glucose Oxidase Capable of Quasi-Direct Electron Transfer after a Quick-and-Easy Modification with a Mediator

2020 ◽  
Vol 21 (3) ◽  
pp. 1137 ◽  
Author(s):  
Nanami Suzuki ◽  
Jinhee Lee ◽  
Noya Loew ◽  
Yuka Takahashi-Inose ◽  
Junko Okuda-Shimazaki ◽  
...  
Keyword(s):  
Electron Transfer ◽  
Glucose Oxidase ◽  
Glucose Sensor ◽  
Direct Electron Transfer ◽  
Glucose Monitoring ◽  
Single Step ◽  
Redox Mediator ◽  
Electrochemically Active ◽  
Phenazine Ethosulfate ◽  
The Impact

Glucose oxidase (GOx) has been widely utilized for monitoring glycemic levels due to its availability, high activity, and specificity toward glucose. Among the three generations of electrochemical glucose sensor principles, direct electron transfer (DET)-based third-generation sensors are considered the ideal principle since the measurements can be carried out in the absence of a free redox mediator in the solution without the impact of oxygen and at a low enough potential for amperometric measurement to avoid the effect of electrochemically active interferences. However, natural GOx is not capable of DET. Therefore, a simple and rapid strategy to create DET-capable GOx is desired. In this study, we designed engineered GOx, which was made readily available for single-step modification with a redox mediator (phenazine ethosulfate, PES) on its surface via a lysine residue rationally introduced into the enzyme. Thus, PES-modified engineered GOx showed a quasi-DET response upon the addition of glucose. This strategy and the obtained results will contribute to the further development of quasi-DET GOx-based glucose monitoring dedicated to precise and accurate glycemic control for diabetic patient care.

scholarly journals Basement Membrane-Based Glucose Sensor Coatings Enhance Continuous Glucose Monitoring in Vivo

2015 ◽  
Vol 9 (5) ◽  
pp. 957-965 ◽  
Author(s):  
Ulrike Klueh ◽  
Yi Qiao ◽  
Caroline Czajkowski ◽  
Izabela Ludzinska ◽  
Omar Antar ◽  
...  
Keyword(s):  
Basement Membrane ◽  
Continuous Glucose Monitoring ◽  
Glucose Sensor ◽  
Glucose Monitoring

A Miniaturized Nafion-Based Glucose Sensor: in vitro and in vivo evaluation in dogs

1994 ◽  
Vol 17 (2) ◽  
pp. 88-94 ◽  
Author(s):  
F. Moussy ◽  
D.J. Harrison ◽  
R.V. Rajotte
Keyword(s):  
Glucose Sensor ◽  
Reference Electrode ◽  
The Body ◽  
In Vitro Tests ◽  
In Vivo Evaluation ◽  
Heat Curing ◽  
Layer Membrane ◽  
Outer Coating

We have developed an implantable glucose sensor based on a new tri-layer membrane configuration. The needle-type sensor integrates a Pt working electrode and a Ag/AgCI reference electrode. Its size is equivalent to a 25 gauge needle (0.5 mm in diamater). Poly (o-phenylenediamine) was used as an inner coating to reduce interference by small compounds present in the body fluids, and the perfluorinated ionomer, Nation as a biocompatible, protective, outer coating. Glucose oxidase trapped in an albumin/glutaraldehyde matrix was sandwiched between these coatings. In vitro tests in buffer showed the sensors had a good selectively, a sensitivity of about 25 nA/mM, and a 90% response time of 33 s. Stabilization of the current following polarization required 10 to 30 min in vitro and 30 to 40 in vivo. Although these sensors remained stable for many weeks in saline solution, their implantation in animals resulted in the degradation of the protective Nation outer coating, which in turn, led to the failure of the incorporated reference electrode. We demonstrated that if unprotected, the AgCI layer of the reference electrode rapidly dissolves in the biological environment. However, we later showed that in vivo degradation of Nation can be prevented by heat curing. When heat cured sensors were subcutaneously implanted in dogs, the sensors' signal closely followed the plasma glucose level during glucose tolerance tests. The response of the sensors implanted in dogs was retained for 10 days.

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