A Critical Factor in Reactive Oxygen Species (ROS) Studies: The Need to Understand the Chemistry of the Solvent Used

Reactive oxygen species (ROS) play critical roles in normal physiological processes including cellular signaling and immune responses. Various pathological conditions including infections of various types, inflammation, cancer, and respiratory conditions are associated with elevated levels of ROS. Therefore, there is widespread interest in understanding ROS concentrations under various pathophysiological conditions for diagnostic and therapeutic applications including ROS-triggered drug delivery. However, in determining ROS concentration, there are major concerns of inappropriate use of various methods that lead to erroneous results; this has prompted the publication of a consensus paper in Nature Metabolism by a group of ROS experts stating “Unfortunately, the application and interpretation of these measurements are fraught with challenges and limitations. This can lead to misleading claims.” Along this line, we have identified an overlooked factor, which can significantly skew the results and results interpretation: the organic co-solvent. DMSO is one of the most widely used organic co-solvent to dissolve a reagent for bioassays. Herein, we describe the rapid oxidation of DMSO by hypochlorite and how this oxidation impacts results of ROS determination in buffer, cell culture media, cell culture, and cell lysates. We hope to use this one example to draw attention to the convoluted roles that DMSO and possibly other organic co-solvents can play and skew experimental results. We also hope to stimulate additional studies to bring more rigor to studying ROS concentration and biology.