Chemical and bactericidal effects induced by plasma in water upon electro‐spraying through DC‐driven positive transient spark discharge in air were investigated. Inactivation of E. coli was determined in dependence on pH (controlled by buffers) and correlated with chemical changes induced in water. Productions of hydrogen peroxide, nitrites, nitrates, peroxynitrites, and pH changes were determined, and the extent of oxidative stress induced in bacteria was evaluated. The degree of inactivation and oxidative damage of bacteria increased with the increasing acidity of the solution. Acidified nitrites interacting with hydrogen peroxide were determined as the most important bactericidal ROS/RNS agents in plasma‐treated water. A possible role of peroxynitrites, ozone, and metal nanoparticles is discussed.

Air transient spark (TS) discharge generates cold plasma, which is a rich source of reactive oxygen and nitrogen species (RONS). The gaseous products both in ambient air and air humidified by water electrospray (ES) are nitrogen oxides (NO and NO2). The rotational and vibrational temperatures determined by optical emission spectroscopy (OES) are lower with water ES through the discharge than in ambient air, which reduces the formation of NOx. The investigation of the specificity of Griess colorimetric assay for the detection of long‐lived nitrites in plasma activated solutions confirms its accuracy by comparison with ion chromatography (IC) and excludes possible interferences with hydrogen peroxide by using the enzyme catalase. Examination of the specificity of the Indigo blue assay for ozone detection shows strong interferences with the peroxynitrite chemistry. Phenol as the chemical probe confirms that the TS air discharge produces no aqueous ozone.

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