Leucine modifications by He/O₂ plasma treatment in phosphate-buffered saline: bactericidal effects and chemical characterization

Abstract

We studied the chemical and bactericidal properties of plasma-modified amino acid leucine (Leu) treated in phosphate-buffered saline (PBS) using an atmospheric-pressure plasma jet operating with He/O2 gas. We determined the initial formation of hypochlorite by the reaction of plasma-supplied O atoms with chloride ions in PBS, followed by the subsequent chlorination of Leu. The principal chemical process involved the formation of Leu monochloramine and Leu dichloramine. The products of plasma-modified Leu further possessed post-discharge reactivity characterized by the decay of Leu chloramines to the secondary products, i.e. isovaleraldehyde, isovaleronitrile, and N-chloromethyl-3-butaldimine. Our study of the effects of Leu on the viability of Escherichia coli bacteria in the plasma-treated PBS solution demonstrated time-and pH-dependent cytotoxic behavior, which were associated with the formation of different reaction products. At shorter treatment times, the addition of Leu hindered the bactericidal properties of plasma-treated PBS caused by the consumption of bactericidal hypochlorite in reaction with Leu and formation of non-bactericidal Leu monochloramine. Acidification to pH = 3 and longer plasma treatment substantially enhanced the bactericidal effects of Leu solutions, which were tentatively assigned to the formation of Leu dichloramine.

Treatment of phenylalanine and tyrosine in phosphate‐buffered saline by plasma‐supplied oxygen atoms: Chemical characterization and bactericidal effects

“…We can conclude from the obtained data that the bactericidal effect of acidified solutions is related to the chloramine postdischarge reaction pathways. As with our suggestion for Leu‐chloramine, [ 14 ] we hypothesize that acidification promotes the disproportionation of monochloramine to dichloramine as described in reaction (), and that the dichloramine then decays as illustrated in Scheme 3. This hypothesis was indirectly confirmed by the presence of 4‐hydroxybenzyl cyanide following acidification, albeit at a relatively low concentration.…”

“…We recently reported on plasma‐treated leucine [ 14 ] and discovered that acidifying the treated solution to pH 3 enhanced the antibacterial activity of the solution, which was previously inactive. Although the toxic compound was not identified directly, it was speculated that the effect may be related to the amino acid dichloramine or its degradation products (compounds V , VI , and VII in Scheme 3).…”

“…We determined the identical absorbance of TMBD also for chloramines of other amino acids such as taurine, valine, and leucine prepared with the same NaOCl concentrations. [ 14 ] As a result, the concentration of chloramine was determined in this study by diluting the plasma‐treated sample with water (in a ratio of 1:5–1:30) and adding the developing reagent. The developing reagent was prepared with 140 µl of TMB (20 mM in N,N‐dimethylformamide), and 10 µl of 20 mM NaI in 1230 µl of 300 mM acetate buffer (pH 5.4).…”

“…The chlorination of phenol [ 8 ] and leucine (Leu) [ 14 ] was observed using the He/O 2 plasma jet, yielding a variety of primary and secondary products. Both oxidation and chlorination were observed in a review of the degradation of amino acids by two types of plasma jets.…”

“…We have recently published on the chemistry of the interaction between the He/O 2 plasma jet and solutions containing Cl − ions. [ 8,13,14 ] The He plasma jet with a small (≤1%) admixture of oxygen produces a large number of O atoms, as well as other reactive oxygen species (ROS) such as ozone and singlet oxygen. [ 15 ] Recent studies have confirmed that O atoms are metastable in both the gas and liquid phases, that is, they can dissolve, diffuse, and react with bulk liquid species despite partial homogeneous and heterogeneous recombination.…”