Plasma-treated water (PTW), i.e. distilled water (DW) exposed to low-temperature atmospheric pressure helium plasma, exhibited strong bactericidal activity against Escherichia coli in suspension even within a few minutes of preparation. This effect was enhanced under acidic conditions. The bactericidal activity of PTW was attenuated according to first-order kinetics and the half-life was highly temperature dependent. The electron spin resonance (ESR) signal of an adduct of the superoxide anion radical ( O 2 − • ) was detected in an aqueous solution using a spin-trapping reagent mixed with PTW, and adding superoxide dismutase to the PTW resulted in a loss of the bactericidal activity and weakening of the ESR adduct signal of O 2 − • in the spin-trapping. These results suggest that O 2 − • plays an important role in imparting bactericidal activity to PTW. Moreover, molecular nitrogen was required both in the ambient gas and in the DW used to prepare the PTW. We, therefore, suggest that the reactive molecule in PTW with bactericidal effects is not a free reactive oxygen species but nitrogen atom(s)-containing molecules that release O 2 − • , such as peroxynitrous acid (ONOOH) or peroxynitric acid (O ₂ NOOH). Considering the activation energy for degradation of these species, we conclude that peroxynitric acid stored in PTW induces the bactericidal effect.

To understand plasma-induced chemical processing in liquids, we investigated the formation of free radicals in aqueous solution exposed to different types of non-contact atmospheric-pressure helium plasma using the spin-trapping technique. Both hydroxyl radical (OH·) and superoxide anion radical (O₂⁻·) adducts were observed when neutral oxygen gas was additionally supplied to the plasma. In particular, O₂⁻· can be dominantly induced in the solution via oxygen flow into the afterglow gas of helium plasma. This type of plasma treatment can potentially be used in medical applications to control infectious diseases, because the O₂⁻· is crucial for sterilization of liquids via atmospheric-pressure plasma.