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目的评价微量过氧乙酸的细胞毒性,为过氧乙酸灭菌时残余量限值的制定提供实验依据。方法采用体外细胞培养技术,观察体积分数(V/V)为1×10-6、2×10-6、3×10-6、4×10-6、5×10-6和10×10-6的过氧乙酸对第3代L929小鼠成纤维细胞生长的影响,将不含过氧乙酸的培养基作为对照组。分别于培养后2、4和7d经MTT比色法测定各组吸光度(A)值,绘制细胞生长曲线,计算各组细胞相对增殖率,对不同浓度过氧乙酸进行细胞毒性评价。结果培养后2、4和7d,1×10-6组与对照组细胞形态及生长状况相似,其余各组细胞生长较差。MTT法检测显示,各时间点对照组A值最大,除1×10-6组与对照组比较差异无统计学意义(P>0.05)外,其余各浓度组与对照组比较差异均有统计学意义(P<0.05)。相关分析显示,各时间点随着过氧乙酸浓度增加A值逐渐下降,细胞增殖与过氧乙酸浓度成负相关;培养后2、4和7d,相关系数分别为—1.000、—1.000、—0.964(P<0.001),相关程度随时间延长逐渐减小。1×10-6组与对照组生长曲线近似,具有明显的指数增长期,其余组生长曲线较平缓。1×10-6组过氧乙酸的细胞毒性分级为1级,2×10-6组为2级,3×10-6组为3级,4×10-6组为3～4级,5×10-6、10×10-6组均为4级。结论 1×10-6过氧乙酸无明显细胞毒性,浓度超过1×10-6的过氧乙酸具有不同程度细胞毒性。
Objective To evaluate the cytotoxicity of trace peracetic acid and provide an experimental basis for the formulation of the limit of residual quantity for peracetic acid sterilization. Methods The in vitro cell culture technique was used to observe the effects of volume fraction (V / V) of 1 × 10-6, 2 × 10-6, 3 × 10-6, 4 × 10-6, 5 × 10-6 and 10 × 10-6, 6 of peracetic acid on the 3rd generation L929 mouse fibroblasts growth, the medium without peracetic acid as a control group. The absorbance (A) value of each group was determined by MTT colorimetric method at 2, 4 and 7 days after culture respectively. The cell growth curve was plotted, the relative proliferation rate of each group was calculated, and the cytotoxicity of different concentrations of peracetic acid was evaluated. Results After 2, 4 and 7 days, the cell morphology and growth of 1 × 10-6 group were similar to that of the control group, while the other groups showed poor cell growth. MTT assay showed that the value of A in control group was the highest at all time points except for 1 × 10-6 group (P> 0.05), but the difference between the other concentration groups and the control group was statistically significant Significance (P <0.05). Correlation analysis showed that A decreased gradually with the increase of peracetic acid concentration at each time point, and the cell proliferation was negatively correlated with peracetic acid concentration. The correlation coefficients at 2,4 and 7 days after culture were -1.000, -1.000 and -0.964 (P <0.001), the degree of correlation decreased gradually with time. The growth curve of 1 × 10-6 group was similar to that of the control group, with a significant exponential growth phase, while the growth curve of the other groups was gentle. 1 × 10-6 group of peracetic acid cytotoxicity grade 1, 2 × 10-6 group 2, 3 × 10-6 group 3, 4 × 10-6 group 3 to 4, 5 × 10-6, 10 × 10-6 group are 4 levels. Conclusion 1 × 10-6 peracetic acid has no obvious cytotoxicity. Peracetic acid with a concentration higher than 1 × 10-6 has different degrees of cytotoxicity.