Generation of superoxide anions by a glycation reaction in conventional laboratory media

We reported that generation of superoxide anion (O2−) was detected from conventional laboratory media. The generated O2− is non-enzymatic converted to hydroxyl radicals, which cause damage to lipids, proteins, and nucleic acids. However, the O2− generating mechanism from culture media is unclear. We considered that the O2− generation was implicated in a glycation reaction between reducing sugar and proteins, which is the early stage of Maillard reaction. It has been suggested that the glycated proteins, such as Schiff base and Amadori compounds, undergo a spontaneous autoxidation reaction, catalyzed by transition metal ions, involving the O2− generation. Therefore, we investigated the effect of Chelex 100 on the O2− generation from brain-heart-infusion (BHI) medium, which is a nutritional culture medium for bacteria. However, the O2− generation from the BHI medium treated with Chelex 100 was significantly increased in comparison to it treated without Chelex 100. The quantity of O2− generation from BHI medium was significantly increased by addition of glucose, and in alkaline environment as well as a glycation reaction model system that autoclaved a mixture solution of glucose and tryptophan. In addition, the O2− generation from BHI medium was significantly inhibited by pyridoxamine that is a Maillard reaction inhibitor. Therefore, it was suggested that the O2− generation from BHI medium is closely related to the glycation reaction of amide compounds such as proteins containing in the medium without the transition metals.