# Water Modification by Cold Plasma Jet with Respect to Physical and Chemical Properties
## Appendix A
Optical emission spectroscopy of the plasma phase revealed the formation in the He:1%N\({}_{2}\) plasma jet of nitric oxide NO and hydroxyl HO\({}^{\bullet}\) derivatives which can lead to numerous RONS after dissolution in the exposed phosphate buffered saline. Experiments showed that an evaporation of the solution occurred under gas exposition and was amplified by the plasma, being mostly related to the interaction between the ionization waves and the gas flow. Further, UV-visible absorption spectroscopy was used to identify and quantify long-lived reactive oxygen and nitrogen species, namely nitrite (NO\({}_{2}^{-}\)), nitrate (NO\({}_{3}^{-}\)), as well as a short-lived species, i.e., peroxynitrite anion (ONOO\({}^{-}\)). [PERSON] et al. [63] used a micro-plasma jet produced in helium and demonstrated that the concentration of H\({}_{2}\)O\({}_{2}\), NO\({}_{2}^{-}\) and NO\({}_{3}^{-}\) can fully account for the majority of RONS produced in the plasma activated buffer. The role of these species on the viability of normal and tumor cell lines was investigated. Although the degree of sensitivity to H\({}_{2}\)O\({}_{2}\) is cell type dependent, it was showed that H\({}_{2}\)O\({}_{2}\) alone cannot account for the toxicity of helium plasma. Indeed, NO\({}_{2}^{-}\), but not NO\({}_{3}^{-}\), acts in synergy with H\({}_{2}\)O\({}_{2}\) to enhance cell death in normal and tumor cell lines to a level similar to that observed after plasma treatment. These findings suggested that the efficiency of plasma treatment strongly depends on the combination of H\({}_{2}\)O\({}_{2}\) and NO\({}_{2}^{-}\) in determined concentrations. It was also showed that the interaction of the helium plasma jet with the ambient air is required to generate NO\({}_{2}^{-}\) and NO\({}_{3}^{-}\) in solution. [PERSON] et al. [64] addressed the analysis of the reactive species generated by non-thermal atmospheric pressure plasmas, which are widely employed in industrial and biomedical research, as well as first clinical applications. They summarized the progress in the detection of plasma-generated, short-lived, reactive oxygen, and nitrogen species in aqueous solutions, discussed the potential and limitations of various analytical methods in plasma-liquid systems, and provided an outlook on the possible future research goals in the development of short-lived, reactive species analysis methods. [PERSON] et al. [65] stated that nitric oxide is a relatively stable free radical and an important signal molecule in plants, animals, and humans with high relevance for biological processes involving inflammatory processes, e.g., wound healing or cancer. The molecule can be detected in the gas phase of non-thermal plasma jets, making it a valuable tool for clinical intervention, but transport efficiency from the gas phase into the liquid phase or tissue remains to be clarified. To elucidate this fact, the nitric oxide concentration in buffered solutions was determined using electron paramagnetic resonance spectroscopy. The origin of the nitric oxide in the liquid could be excluded; therefore, potential precursors such as hydroxyl radicals, superoxide anions, atomic hydrogen, and stable species (nitrite, nitrate and hydrogen peroxide) were detected and the potential formation pathway, as well as ways of enhancing the production of nitric oxide by alteration of the feed gas and the surrounding gas composition during plasma treatment of the liquid, were pointed out. [PERSON] et al. [54] presented a 2D fluid dynamics model for the interaction between a plasma jet and liquid water. The results indicated that the formed reactive species originate either from the gas phase (with further solvation) or are formed at the liquid interface. A clear increase in the aqueous density of H\({}_{2}\)O\({}_{2}\), HNO\({}_{2}\)/NO\({}_{2}^{-}\) and NO\({}_{3}^{-}\) was observed as a function of time, while the densities of O\({}_{3}\), HO\({}_{2}\)/O\({}_{2}^{-}\) and ONOOH/ONOO\({}^{-}\) were found to quickly reach a maximum due to chemical reactions in solution.