engleski

Hydrogen peroxide content and glucose oxidase activity of honeys

Hydrogen peroxide (H2O2) accumulation in honey occurs upon honey dilution due to the action of reactivated glucose oxidase (GOX) which oxidize glucose to gluconic acid, simultaneously liberating H2O2 as by-product. Although the presence of GOX in honey seems necessary for H2O2 evolution, there is no report on the correlation between GOX activity and H2O2 content in diluted honeys, so far. Besides numerous factors that may affect GOX activity and H2O2 content, one of the possible reason for the lack of correlation might be a difference in methods and conditions used for their determination. Namely, H2O2 content is measured in diluted honeys (realistic conditions) by various methods, while GOX activity of honey mostly in the model solution of 1.5 M glucose as substrate by standard o-dianisidine/HRP method (artificial conditions). In order to test whether the difference in determination methods might be a reason for the lack of correlation, we have determined GOX activity and H2O2 content in 15 different samples of honey at five different mass to volume ratios (1:1 ; 1:2 ; 1:4 ; 1:8 ; 1:16) using o-dianisidine/HRP method. For measurement of GOX activity, HRP and o-dianisidine were added to diluted honey prior to 30 minute incubation, while for H2O2 content at the end of incubation period. In addition, sugar profile of honeys, GOX activity in model solutions of glucose or mixture of glucose and fructose equal to their concentrations in diluted honeys, as well as water activity of diluted honeys and model solutions were determined. GOX activity curves followed the curves of H2O2 accumulation in diluted honeys, indicating applicability of o-dianisidine/HRP method for simultaneous determination of GOX activity and H2O2 content. Similarity of curves was confirmed by relatively high coefficients of correlation ranging from 0.853 for the lowest, up to 0.971 for highest dilution. However, GOX activities in model solutions of glucose or glucose and fructose at concentrations equal to those in diluted honeys were found quite higher. On the basis of water activity determination, it seems that both, GOX activity and H2O2 accumulation in honeys are affected by macromolecular crowding milieu of honey biomolecules.

honey ; hydrogen peroxide ; glucose oxidase

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