scholarly journals Employment of 1-Methoxy-5-Ethyl Phenazinium Ethyl Sulfate as a Stable Electron Mediator in Flavin Oxidoreductases-Based Sensors

Sensors ◽  
2020 ◽  
Vol 20 (10) ◽  
pp. 2825
Author(s):  
Maya Fitriana ◽  
Noya Loew ◽  
Arief Budi Witarto ◽  
Kazunori Ikebukuro ◽  
Koji Sode ◽  
...  
Keyword(s):  
Glucose Dehydrogenase ◽  
High Sensitivity ◽  
Lactate Oxidase ◽  
Electron Mediator ◽  
Effective Electron ◽  
Ethyl Sulfate ◽  
Enzyme Sensor ◽  
Aerococcus Viridans ◽  
Enzyme Sensors ◽  
Lactate Sensor

In this paper, a novel electron mediator, 1-methoxy-5-ethyl phenazinium ethyl sulfate (mPES), was introduced as a versatile mediator for disposable enzyme sensor strips, employing representative flavin oxidoreductases, lactate oxidase (LOx), glucose dehydrogenase (GDH), and fructosyl peptide oxidase (FPOx). A disposable lactate enzyme sensor with oxygen insensitive Aerococcus viridans-derived engineered LOx (AvLOx), with A96L mutant as the enzyme, was constructed. The constructed lactate sensor exhibited a high sensitivity (0.73 ± 0.12 μA/mM) and wide linear range (0–50 mM lactate), showings that mPES functions as an effective mediator for AvLOx. Employing mPES as mediator allowed this amperometric lactate sensor to be operated at a relatively low potential of +0.2 V to 0 V vs. Ag/AgCl, thus avoiding interference from uric acid and acetaminophen. The lactate sensors were adequately stable for at least 48 days of storage at 25 °C. These results indicated that mPES can be replaced with 1-methoxy-5-methyl phenazinium methyl sulfate (mPMS), which we previously reported as the best mediator for AvLOx-based lactate sensors. Furthermore, this study revealed that mPES can be used as an effective electron mediator for the enzyme sensors employing representative flavin oxidoreductases, GDH-based glucose sensors, and FPOx-based hemoglobin A1c (HbA1c) sensors.

scholarly journals Ratiometric Fluorescent Biosensors for Glucose and Lactate Using an Oxygen-Sensing Membrane

Biosensors ◽  
2021 ◽  
Vol 11 (7) ◽  
pp. 208
Author(s):  
Hong Dinh Duong ◽  
Jong Il Rhee
Keyword(s):  
Limit Of Detection ◽  
Oxygen Sensing ◽  
High Sensitivity ◽  
Glucose Sensing ◽  
Oxidation Reactions ◽  
Lactate Oxidase ◽  
Lactate Oxidation ◽  
Detection Range ◽  
Quenching Effect ◽  
Sensing Membrane

In this study, ratiometric fluorescent glucose and lactate biosensors were developed using a ratiometric fluorescent oxygen-sensing membrane immobilized with glucose oxidase (GOD) or lactate oxidase (LOX). Herein, the ratiometric fluorescent oxygen-sensing membrane was fabricated with the ratio of two emission wavelengths of platinum meso-tetra (pentafluorophenyl) porphyrin (PtP) doped in polystyrene particles and coumarin 6 (C6) captured into silica particles. The operation mechanism of the sensing membranes was based on (i) the fluorescence quenching effect of the PtP dye by oxygen molecules, and (ii) the consumption of oxygen levels in the glucose or lactate oxidation reactions under the catalysis of GOD or LOX. The ratiometric fluorescent glucose-sensing membrane showed high sensitivity to glucose in the range of 0.1–2 mM, with a limit of detection (LOD) of 0.031 mM, whereas the ratiometric fluorescent lactate-sensing membrane showed the linear detection range of 0.1–0.8 mM, with an LOD of 0.06 mM. These sensing membranes also showed good selectivity, fast reversibility, and stability over long-term use. They were applied to detect glucose and lactate in artificial human serum, and they provided reliable measurement results.

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.

X-ray Structures of Aerococcus viridans Lactate Oxidase and Its Complex with d-Lactate at pH 4.5 Show an α-Hydroxyacid Oxidation Mechanism

2008 ◽  
Vol 378 (2) ◽  
pp. 436-446 ◽  
Author(s):  
Makio Furuichi ◽  
Nobuhiro Suzuki ◽  
Balasundaresan Dhakshnamoorhty ◽  
Hirotaka Minagawa ◽  
Ryosuke Yamagishi ◽  
...  
Keyword(s):  
Oxidation Mechanism ◽  
Lactate Oxidase ◽  
X Ray ◽  
Aerococcus Viridans

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 FMN-dependent oligomerization of putative lactate oxidase from Pediococcus acidilactici

2019 ◽  
Author(s):  
Yashwanth Ashok ◽  
Mirko M. Maksimainen ◽  
Tuija Kallio ◽  
Pekka Kilpeläinen ◽  
Lari Lehtiö
Keyword(s):  
Hydrogen Peroxide ◽  
Active Site ◽  
Pediococcus Acidilactici ◽  
Active Site Residues ◽  
Lactate Oxidase ◽  
Monooxygenase Activity ◽  
Aerococcus Viridans ◽  
Lactate Levels

AbstractLactate oxidases belong to a group of FMN-dependent enzymes and they catalyze a conversion of lactate to pyruvate with a release of hydrogen peroxide. Hydrogen peroxide is also utilized as a read out in biosensors to quantitate lactate levels in biological samples. Aerococcus viridans lactate oxidase is the best characterized lactate oxidase and our knowledge of lactate oxidases relies largely to studies conducted with that particular enzyme. Pediococcus acidilactici lactate oxidase is also commercially available for e.g. lactate measurements, but this enzyme has not been characterized before in detail. Here we report structural characterization of the recombinant enzyme and its co-factor dependent oligomerization. The crystal structures revealed two distinct conformations in the loop closing the active site, consistent with previous biochemical studies implicating the role of loop in catalysis. Despite the structural conservation of active site residues when compared to Aerococcus viridans lactate oxidase we were not able to detect either oxidase or monooxygenase activity when L-lactate or other potential alpha hydroxyl acids were used as a substrate. Pediococcus acidilactici lactate oxidase is therefore an example of a misannotation of an FMN-dependent enzyme, which catalyzes likely a so far unknown oxidation reaction.

scholarly journals A Disposable Electrochemical Glucose Sensor Using Catalytic Subunit of Novel Thermostable Glucose Dehydrogenase

2007 ◽  
Vol 1 (1) ◽  
pp. 26-30 ◽  
Author(s):  
Hideaki Yamaoka ◽  
Koji Sode
Keyword(s):  
Blood Glucose ◽  
Whole Blood ◽  
Glucose Sensor ◽  
Glucose Dehydrogenase ◽  
Blood Glucose Measurement ◽  
Catalytic Subunit ◽  
Glucose Measurement ◽  
Carbon Electrode ◽  
Electron Mediator ◽  
Screen Printed Carbon Electrode

We report here the development of a novel disposable-type glucose enzyme sensor based on a screen-printed carbon electrode (SPCE) employing a catalytic subunit of FAD-dependent glucose dehydrogenase (FADGDH) and a Rucomplex (hexaammineruthenium (III) chloride) as the electron mediator. The whole blood glucose measurement was achieved in 2 seconds using a 500 nl sample of the range of 100-800 mg/ml.

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