Amperometric assessment of glucose electrode behaviour in mixed solvents and determination of glucose in dairy products

An amperometric biosensor based on a ruthenium(III), nickel(II) and iron(II) hexacyanometallate (HCM)-modified graphite electrode and immobilized glucose oxidase has been used for the determination of glucose in water-miscible organic solvent/aqueous buffer mixtures. Although the specific activity of biochemically active molecule such as enzyme is reduced in organic environment, it was established that the presence of water soluble organic solvents such as methanol, ethanol and acetonitrile (φ = 10%) enhance the biosensor response. Hydrogen peroxide, produced by enzyme-catalysed reduction of glucose, was measured in phosphate buffer solution (pH = 6.86) at −50 mV against a reference Hg|Hg₂Cl₂|3 M KCl electrode to determine the concentration of glucose. The influence of the addition of different volume fractions (φ = 10–60%) of methanol, ethanol, acetone, acetonitrile and isopropanol on biosensor response was investigated. The obtained amperometric signals were fast, reproducible and linearly proportional to glucose concentrations in the range of 0.1–0.8 mM, with a squared correlation coefficient of 0.9994 for buffer solution. With the addition of ethanol (φ = 10% and 40%) the plateau on I/c curve was obtained for concentrations of glucose higher than 0.8 and 1.1 mM, respectively. The biosensor proved to be stable for several months. The recoveries of added glucose (0.200 and 0.300 mM) from aqueous solution and from solution with ethanol φ = 10% ranged from 96.0% to 108.0%. The biosensor was used for the determination of glucose in some food samples of dairy industry, and the results were consistent with those obtained with the commercially available glucose enzyme photometric kit.