Factors affecting the production and measurement of hydrogen peroxide in honey samples.

Many Australian native honeys possess significant antimicrobial properties due to the production of hydrogen peroxide (H2O2) by glucose oxidase, an enzyme derived from the honeybee. The level of H2O2 produced in different honey samples is highly variable, and factors governing its production and stability are not well understood. In this study, highly active Australian honeys that had been stored for >10 years lost up to 54 % of their antibacterial activity, although almost all retained sufficient activity to be considered potentially therapeutically useful. We used a simple colourimetric assay to quantify H2O2 production. Although we found a significant correlation between H2O2 production and antibacterial activity across diverse honey samples, variation in H2O2 only explained 47 % of the variation observed in activity, limiting the assay as a screening tool and highlighting the complexity of the relationship between H2O2 and the killing power of honey. To further examine this, we tested whether H2O2 detection in honey was being inhibited by pigmented compounds and if H2O2 might be directly degraded in some honey samples. We found no correlation between H2O2 detection and honey colour. Some honey samples rapidly lost endogenous and spiked H2O2, suggesting that components in honey, such as catalase or antioxidant polyphenols, may degrade or quench H2O2. Despite this rapid loss of H2O2, these honeys had significant peroxide-based antibacterial activity, indicating a complex relationship between H2O2 and other honey components that may act synergistically to augment activity.

Inhibition of Dermatophyte Fungi by Australian Jarrah Honey.

Superficial dermatophyte infections, commonly known as tineas, are the most prevalent fungal ailment and are increasing in incidence, leading to an interest in alternative treatments. Many floral honeys possess antimicrobial activity due to high sugar, low pH, and the production of hydrogen peroxide (H2O2) from the activity of the bee-derived enzyme glucose oxidase. Australian jarrah (Eucalyptus marginata) honey produces particularly high levels of H2O2 and has been found to be potently antifungal. This study characterized the activity of jarrah honey on fungal dermatophyte species. Jarrah honey inhibited dermatophytes with minimum inhibitory concentrations (MICs) of 1.5-3.5% (w/v), which increased to ≥25% (w/v) when catalase was added. Microscopic analysis found jarrah honey inhibited the germination of Trichophyton rubrum conidia and scanning electron microscopy of mature T. rubrum hyphae after honey treatment revealed bulging and collapsed regions. When treated hyphae were stained using REDOX fluorophores these did not detect any internal oxidative stress, suggesting jarrah honey acts largely on the hyphal surface. Although H2O2 appears critical for the antifungal activity of jarrah honey and its action on fungal cells, these effects persisted when H2O2 was eliminated and could not be replicated using synthetic honey spiked with H2O2, indicating jarrah honey contains agents that augment antifungal activity.