ORIGINAL RESEARCH article
Front. Microbiol.
Sec. Antimicrobials, Resistance and Chemotherapy
This article is part of the Research TopicExpanding Multidisciplinary Potential of Cold Atmospheric Plasma Beyond Antimicrobial ResistanceView all 3 articles
Antimicrobial Efficacy and Cytotoxic Assessment of Plasma-Activated Water Generated by a Dielectric Barrier Discharge Microbubble System
Provisionally accepted
Raquel Oliveira Ferreira1*
Nilson Cristino da Cruz1*
Danielli C.F.S. Spigarollo2Iolanda Cristina Silveira Duarte3Elaine C. de Oliveira4- 1São Paulo State University, São Paulo, Brazil
- 2Universidade Estadual Paulista Julio de Mesquita Filho, São Paulo, Brazil
- 3Universidade Federal de Sao Carlos, São Carlos, Brazil
- 4Universidade Estadual de Campinas, Campinas, Brazil
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The growing threat of antimicrobial resistance has intensified the search for alternative disinfection strategies. Plasma-activated water (PAW), enriched with reactive oxygen and nitrogen species (RONS), has emerged as a promising non-antibiotic antimicrobial approach. In this study, PAW was generated using a bench-scale dielectric barrier discharge (DBD) reactor coupled with a microbubble diffusion system, operating with compressed ambient air at 8-9 kV, frequencies of 40 kHz, 1.25 W, air flow rate of 5 L/min with activation times of 7 and 14 minutes. In the generated PAW, ROS were produced in higher abundance than RNS, with hydrogen peroxide (H₂O₂) and ozone (O₃) representing the dominant species contributing to the antimicrobial activity, alongside detectable levels of nitrite (NO₂⁻), nitrate (NO₃⁻), and nitrous acid (HNO₂). Physicochemical characterization included measurements of pH, oxidation-reduction potential (ORP), conductivity, and total dissolved solids (TDS), as well as qualitative analysis of RONS using UV–Vis spectrophotometry (190–400 nm), for detection of diagnostic wavelengths, with quantitative assessment based on colorimetric methods for H₂O₂, NO₂⁻, and NO₃⁻, and a photometric assay for O₃. Antimicrobial activity was evaluated against Staphylococcus aureus (ATCC 6538), Escherichia coli (ATCC 25922), and Candida albicans (ATCC 10231) using PAW generated at 7 and 14 minutes of activation. In addition, cytotoxicity was evaluated in L929 fibroblasts following ISO 10993-5 criteria, and cell viability remained above the 70% threshold after exposure to PAW. Morphological assessments were performed in both L929 and B16F10 cells using fluorescence microscopy to examine potential differential cellular responses. These findings indicate that longer activation times enhanced antimicrobial activity while preserving fibroblast viability, supporting the potential of PAW as an effective and safe alternative for biomedical applications.
Keywords: Antimicrobial activity, cold atmospheric plasma, Cytotoxicity, DBD plasma, microbubble diffusion, Physicochemical characterization, Plasma-activated water, reactive species
Received: 14 Nov 2025; Accepted: 20 Jan 2026.
Copyright: © 2026 Ferreira, da Cruz, Spigarollo, Silveira Duarte and de Oliveira. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
* Correspondence:
Raquel Oliveira Ferreira
Nilson Cristino da Cruz
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