## 3 Results and Discussion
### Changes in Lipids Associated with Cell Membranes
While SM levels continued to rise sharply in A4K1H cells, Hex1 Cer levels remained relatively stable (Figure 2d). Given the inconsistency between our hypothesis and the observed lipid changes in A4K1H cells, we conducted a secondary analysis involving samples collected up to 12 h post-irradiation (B4K12H cells). Consistent with our initial findings (i.e., the results of set A), total cell membrane lipids were significantly (\(p<0.05\)) elevated in B4K0H cells compared to the controls in set B. As anticipated, there were no significant differences in lipid levels between B4K12H cells and the controls, indicating a return to normal levels 12 h post-irradiation (Figure 3a,b). \begin{table}
\begin{tabular}{c c c c c} \hline \hline \multirow{2}{*}{**Lipid Categories**} & \multirow{2}{*}{**Classes**} & \multicolumn{4}{c}{**Composition (\%)**} \\  & & **(Calculated from Mean Area Values)** \\ \cline{3-5}  & & **Control** & **A4K0H** & **A4K1H** \\ \hline \multirow{4}{*}{Glycerophospholipid (GP)} & Phosphatidylcholine (PC) & 82.76 & 82.21 & 80.52 \\ \cline{2-5}  & Phosphatidylethanolamine (PE) & 9.70 & 9.74 & 9.69 \\ \cline{2-5}  & Phosphatidylserine (PS) & 2.00 & 2.29 & 2.48 \\ \cline{2-5}  & Phosphatidylinositol (PI) & 0.56 & 0.49 & 0.45 \\ \hline \multirow{4}{*}{Sphingolipid} & Sphingomyelin (SM) & 4.84 & 5.14 & 6.73 \\ \cline{2-5}  & Ganglioside (GM3) & 0.06 & 0.05 & 0.05 \\ \cline{2-5}  & Monohexosylceramide (Hex1 Cer) & 0.05 & 0.05 & 0.06 \\ \cline{2-5}  & Dihexosylceramide (Hex2 Cer) & 0.03 & 0.03 & 0.02 \\ \hline \multirow{4}{*}{Sum} & Sum & 100 & 100 & 100 \\ \cline{2-5}  & & & & \\ \hline \hline \end{tabular}
\end{table} 
Table 2: Composition of cell membrane lipids in set A cell samples. \begin{table}
\begin{tabular}{c c c c c} \hline \hline \multirow{2}{*}{**Lipid Categories**} & \multirow{2}{*}{**Classes**} & \multicolumn{4}{c}{**Composition (\%)**} \\  & & **(Calculated from Mean Area Values)** \\ \cline{3-5}  & & **Control** & **B4K0H** & **B4.5K0H** & **B4K12H** \\ \hline \multirow{4}{*}{Glycerophospholipids (GP)} & Phosphatidylcholine (PC) & 79.73 & 79.51 & 79.81 & 78.98 \\ \cline{2-5}  & Phosphatidylethanolamine (PE) & 10.29 & 10.09 & 9.84 & 10.75 \\ \cline{2-5}  & Phosphatidylserine (PS) & 1.38 & 1.50 & 1.17 & 1.40 \\ \cline{2-5}  & Phosphatidylinositol (PI) & 0.90 & 0.86 & 0.75 & 1.00 \\ \hline \multirow{4}{*}{Sphingolipids (SP)} & Sphingomyelin (SM) & 7.57 & 7.93 & 8.33 & 7.70 \\ \cline{2-5}  & Ganglioside (GM3) & 0.05 & 0.03 & 0.03 & 0.04 \\ \cline{2-5}  & Monohexosylceramide (Hex1 Cer) & 0.04 & 0.03 & 0.03 & 0.06 \\ \cline{2-5}  & Dihexosylceramide (Hex2 Cer) & 0.04 & 0.04 & 0.04 & 0.07 \\ \hline \multirow{4}{*}{Sum} & Sum & 100 & 100 & 100 & 100 \\ \cline{2-5}  & & & & & \\ \hline \hline \end{tabular}
\end{table} 
Table 3: Composition of cell membrane lipids in set B samples. Figure 2: Changes in cell membrane lipids until 1 h following microplasma irradiation at 4.0 kV condition. These data correspond to set #1 samples. (**a**) Total cell membrane-associated lipid alterations were determined from the absolute area values derived from LC/MS analysis. Statistical analysis compared the A4K0H group (n = 5) and A4K1H (n = 5) groups with the control group (n = 5). (**b**) Percentage changes in total cell membrane-associated lipids in the A4K0H and A4K1H groups relative to the control. (**c**) Relative changes in glycerophospholipids, including phosphatidylcholinecholine (PC), phosphatidylcholinetanolamine (PE), phosphatidylserine (PS), and phosphatidylcholinetanol (PI) species in A4K0H and A4K1H groups compared to the control. (**d**) Relative changes in sphingomyelin (SM), ganglioside (GM3), monohexosylceramide (Hex1 Cer), and dihexosylceramide (Hex2 Cer) species in the A4K0H and A4K1H groups compared to the control. Figure 3: Changes in cell membrane lipids until 12 h after microplasma irradiation at 4.0 kV condition. These data belong to set #2 samples. (**a**) Total cell membrane-associated lipid alterations were calculated based on the absolute area values obtained from LC/MS analysis.