Re-Examining the Role of Hydrogen Peroxide in Bacteriostatic and Bactericidal Activities of Honey

The aim of this study was to critically analyze the effects of hydrogen peroxide on growth and survival of bacterial cells in order to prove or disprove its purported role as a main component responsible for the antibacterial activity of honey. Using the sensitive peroxide/peroxidase assay, broth microdilution assay and DNA degradation assays, the quantitative relationships between the content of H(2)O(2) and honey's antibacterial activity was established(.) The results showed that: (A) the average H(2)O(2) content in honey was over 900-fold lower than that observed in disinfectants that kills bacteria on contact. (B) A supplementation of bacterial cultures with H(2)O(2) inhibited E. coli and B. subtilis growth in a concentration-dependent manner, with minimal inhibitory concentrations (MIC(90)) values of 1.25 mM/10(7) cfu/ml and 2.5 mM/10(7) cfu/ml for E. coli and B. subtilis, respectively. In contrast, the MIC(90) of honey against E. coli correlated with honey H(2)O(2) content of 2.5 mM, and growth inhibition of B. subtilis by honey did not correlate with honey H(2)O(2) levels at all. (C) A supplementation of bacterial cultures with H(2)O(2) caused a concentration-dependent degradation of bacterial DNA, with the minimum DNA degrading concentration occurring at 2.5 mM H(2)O(2). DNA degradation by honey occurred at lower than ≤2.5 mM concentration of honey H(2)O(2) suggested an enhancing effect of other honey components. (D) Honeys with low H(2)O(2) content were unable to cleave DNA but the addition of H(2)O(2) restored this activity. The DNase-like activity was heat-resistant but catalase-sensitive indicating that H(2)O(2) participated in the oxidative DNA damage. We concluded that the honey H(2)O(2) was involved in oxidative damage causing bacterial growth inhibition and DNA degradation, but these effects were modulated by other honey components.