Bactericidal effect of plasma-activated water generated by a novel super-potable plasma device as a novel antibacterial method: an in vitro study.

Abstract

Background: A continuous risk from bacterial infection poses a major environmental and public health challenge. As an emerging strategy for inhibiting bacterial infections, plasma-activated water (PAW) has proved highly effective, environment-friendly, and non-drug resistant to many bacteria. However, the plasma device is too big and requires specialists to operate, which inevitably limits its real-life application. It has been confirmed that the bactericidal effect of traditional PAW would be significantly reduced in the organic-rich environments. To better resolve the limitations, a novel super-portable plasma device was applied, and the low concentration of H₂O₂ was added to enhance the bactericidal effect of PAW in the organ-rich environments.

Methods: A super-portable plasma device, weighing only 1 kg, was applied to generate PAW. Staphylococcus aureus (S. aureus), Escherichia coli (E. coli) and Porphyromonas gingivalis (P. gingivalis) were selected as the test bacteria. Colony-forming units (CFU) counting and bactericidal rate were employed to evaluate the bactericidal effect of PAW. Optical emission spectroscopy (OES) was used to identify the major reactive species in PAW. Different concentration of H₂O₂ was added to PAW (H₂O₂/PAW), and the pH, oxidation-reduction potential (ORP), conductivity, H₂O₂ and NO content of HP/PAW were measured to explore the bactericidal mechanisms. CCK-8 assay was used to assess the cytotoxicity of 0.1% H₂O₂/PAW on human periodontal ligament fibroblasts (hPDLFs).

Results: The 20 mL of PAW generated by the novel super-portable plasma device achieved the bactericidal rate of 98.94% against S. aureus (t = 10 min) and 99.66% (t = 20 min). When the PAW was prepared with a volume of 20 mL, activation times of 6 min and 8 min, and the treatment time was 30 min, the bactericidal rates all exceeded 98% against P. gingivalis. The addition of 0.1% H₂O₂ significantly enhanced the antibacterial effect of PAW in the presence of Bovine Serum Albumin (BSA) (p<0.05). The addition of 0.1% H₂O₂ lowered the pH, increased the ORP and conductivity, and raised the content of H₂O₂ and NO in the solution. Cytotoxicity evaluation indicated that 0.1% H₂O₂/PAW had moderate cytotoxicity only after 20 min of preparation and 48 h of treatment, while other preparation and treatment times showed mild or no cytotoxicity.

Conclusions: The PAW generated by the novel super-portable plasma device exhibited an excellent bactericidal effect. The addition of low concentrations of H₂O₂ to PAW, specifically 0.1% H₂O₂/PAW, maintained an effective antibacterial effect in complex environments in the presence of BSA with non-cytotoxicity.