ID: 2203

  • Title:
    Chemistry and cytotoxic properties of cell culture media modified by He/O2 plasma treatment

    Lukes, Petr - Institute of Plasma Physics of the Czech Academy of Sciences
    Jirasek, Vit - Institute of Plasma Physics of the Czech Academy of Sciences
    Kovalova, Zuzana - Institute of Plasma Physics of the Czech Academy of Sciences
    Tarabova, Barbora - Institute of Plasma Physics of the Czech Academy of Sciences

    The application of cold atmospheric plasma (CAP) in medicine is a perspective and rapidly increasing research topic. Besides direct treatment of living tissue or cells with CAP source, properties of plasma-treated biologically relevant liquids such as cell culture media gain significant interest and have been applied for various medical and biological trials. The secondary aqueous-phase chemical reactions induced by plasma-supplied reactive species play an important role. The post-discharge activity can last in these liquids for a significant period after the solution’s exposure to the plasma. Various transient reactive oxygen and nitrogen species such as OH•, O2•-, NO• and NO2• radicals, peroxynitrite may be produced in plasma-treated liquids. These species have highly cytotoxic properties and cause biochemical and antibacterial activity of plasma-treated solutions through post-discharge processes. Nevertheless, the chemical content of treated liquids also plays an important role. Culture media contain a complex mixture of inorganic salts and organic compounds such as amino acids, vitamins, glucose, antibiotics, etc., significantly influencing the properties and activity of these plasma-treated liquids.

    In this work, we studied plasmachemical modifications of Dulbecco’s Modified Eagle Medium (DMEM) treated by He/O2 plasma. We performed a chemical analysis of plasma-treated DMEM and correlated it with its antibacterial properties observed on E. coli. Special attention was paid to the role of amino acids in the plasma-induced biocidal activity of DMEM.

    The COST-Reference plasma jet was used as a plasma source to treat the culture media [1-5]. A sinusoidal voltage with a 230 V root mean square was used to power the electrode of the capacitively coupled 13.56 MHz RF plasma source. The plasma was operated at a helium flow rate of 1.4 slm with a 0.6% oxygen admixture. The jet was aimed at a liquid solution, with a distance of 4 mm between the jet outlet and the liquid surface. The organic chloramines concentrations in plasma-treated DMEM were determined using the 3,3´,5,5´-tetramethylbenzidine colorimetric assay. The presence of aldehydes was examined using a thiobarbituric acid reactive assay. FTIR spectroscopy was used to analyze the media (Bruker Vertex 70v). Bacterial experiments were performed using the strain Escherichia coli K12 (CCM 3880). Bacterial suspensions initially contained 10^9 (CFUs)/ml.

    Plasma-treated cell culture media have shown long-term post-discharge chemical and bactericidal activity. This behavior was attributed to the action of tertiary chemical products formed by the decay of organic dichloramines. By chemical analysis of the plasma-treated media, the initial formation of organic monochloramines was observed, followed by their post-discharge decay to C-1 aldehydes. The monochloramines were formed by the chlorination of amino acids with hypochlorite, produced in the reaction of O atoms with Cl- ions present in culture media. At higher O atoms dose, further chlorination of monochloramines to dichloramines occurred. The thiobarbituric acid assay revealed malondialdehyde formation from glucose oxidation as the major component of the media.

    non-thermal plasma, culture media, amino acid, O atom, bacteria

    [1] M. M. Hefny, C. Pattyn, P. Lukes, J. Benedikt, J. Phys. D.: Appl. Phys., 49, 404002 (2016). [2] V. Jirasek, P. Lukes, Plasma Sources Sci. Technol., 28, 035015 (2019). [3] V. Jirasek, P. Lukes P, J. Phys. D: Appl. Phys., 53, 505206 (2020). [4] V. Jirasek, Z. Kovalova, B. Tarabova, P. Lukes, J. Phys. D: Appl. Phys., 54, 505206 (2021). [5] V. Jirasek, B. Tarabova, P. Lukes, Plasma Proc. Polym., 19, 2200079 (2022).

    Topic 1:
    4. Diagnostics, analytics, experimental techniques

    Topic 2:

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