Enhancement of alcohol oxidase and alcohol oxidase biosensors using high hydrostatic pressure

摘要

We report the effect of high hydrostatic pressure (HHP) on the activity and stability of alcohol oxidase (AOx) and alcohol oxidase biosensors. The in-situ activity of AOx was measured at 45 "C from 0.1 MPa to 200 MPa using a high pressure optical cellwith sapphire windows interfaced to a fiber optic spectrometer. Alcohol oxidase biosensors were fabricated by immobilization of AOx in poly-o-phenylendiamine by electrochemical polymerization in 5 mM o-phenylenediamine and 0.1 Mphosphate buffer (pH 7.0)at 650 m V vs. AgAgCl at 0.1 MPa to 240 MPa on platinized platinum electrodes. The stability of biosensors was tested by either storing at room temperature for 14 days or heating at 50 "C at 0.1 MPa for 10 min. The activity of both isozymes of AOx increased at high pressure. At 45 "C, the activity of labile and resistant fraction was 1.5, 1.8, 1.4, 2.2, or 1.8 times greater at 40, 80, 120, 160, or 200 MPa compared to 0.1 MPa; the activity of resistant fraction was 1.2, 1.6, 2.0,1.6, or 1.7 times greater at 40, 80, 120, 160, or 200 MPa compared to 0.1 MPa. However, high pressure during electropolymerization did not significantly affect the stability of biosensors upon depressurization regardless of the isozyme. High pressure applied during thermal inactivation of immobilized AOx didn't significantly affect the thermostability, either. The activating effect of HHP has potential to increase the biocatalysis of enzymes and reduce the economic cost in industry.