A hydrophobin-based-biosensor layered by an immobilized lactate dehydrogenase enzyme for electrochemical determination of pyruvate

Appropriate enzyme immobilization on the electrode surface in order to access its active site has always been an important strategy for electrode modification. In this report, lactate dehydrogenase enzyme was appropriately immobilized on the glassy carbon electrode via hydrophobin (HFB1) and graphen...

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Published in:Bioelectrochemistry (Amsterdam, Netherlands) Netherlands), 2019-12, Vol.130, p.107323-107323, Article 107323
Main Authors: Mirzaei, Fatemeh, Mirzaei, Mohammad, Torkzadeh-Mahani, Masoud
Format: Article
Language:English
Subjects:
Biosensors
Dehydrogenase
Dehydrogenases
Diffusion coefficient
Electrical measurement
Electrochemical impedance spectroscopy
Electrochemistry
Electrodes
Enzymes
Glassy carbon
Graphene
Hydrophobins
Immobilization
L-Lactate dehydrogenase
Lactate dehydrogenase
Lactic acid
NADH
Nanocomposites
Nicotinamide adenine dinucleotide
Oxidation
Pyruvic acid
Reproducibility
Spectroscopy
Voltammetry
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Summary:Appropriate enzyme immobilization on the electrode surface in order to access its active site has always been an important strategy for electrode modification. In this report, lactate dehydrogenase enzyme was appropriately immobilized on the glassy carbon electrode via hydrophobin (HFB1) and graphene oxide nanocomposite. The step-by-step modification was successfully confirmed by water contact analysis, cyclic voltammetry, and electrochemical impedance spectroscopy. Under optimum conditions, this biosensor demonstrated a detection limit of 8.69 nM and RSD of 4.3% and 3.6% (n = 5) for reproducibility and repeatability. The effect of scan rate on the oxidation behavior of NADH was investigated by cyclic voltammetry; and diffusion coefficient for NADH was estimated at 6.27 × 10  cm .s . The apparent Michaelis-Menten constant (K ) was amperometrically determined and it was lower than K for the free enzyme. Also, the modified electrode represented good stability after nine days with 6% decrease in current. The proposed assay was successfully used in real sample-serum-analysis and the obtained recoveries were between 93% and 104.0%.
ISSN:1567-5394
1878-562X
DOI:10.1016/j.bioelechem.2019.06.008