瘢痕消复方提取工艺及其对增生性瘢痕的抑制作用研究

Study on extraction technology of Banhenxiao Compound and its inhibitory effect on hypertrophic scar

目的:在单因素试验的基础上,以乙醇体积分数、料液比、提取时间为影响因素,研究瘢痕消复方的提取工艺,并考察提取物对增生性瘢痕的抑制作用。方法:采用加热回流法提取瘢痕消复方,以丹参酮ⅡA、丹酚酸B、芦荟苷、没食子酸为主成分进行含量综合评分为评价指标,Box-Behnken响应面法研究提取工艺。采用CCK-8法检测不同浓度的提取液对人增生性瘢痕成纤维细胞(hypertrophic scar fibroblasts,HSFb)细胞活力,流式细胞术检测细胞凋亡率,Western blot检测凋亡蛋白Bax、Bcl-2水平。结果:最佳提取工艺为13倍量73%乙醇提取2次,每次2.1 h。低、中、高剂量提取液均能使HSFb细胞生长抑制率、细胞凋亡率、Bax水平明显升高,Bcl-2水平降低,而且较高剂量比较低剂量更明显。结论:瘢痕消复方提取工艺稳定、可行。复方提取物能有效抑制HSFb增殖,促进细胞凋亡,影响细胞凋亡蛋白的表达。

OBJECTIVE On the basis of a single factor test,the extraction process of Banhenxiao Compound was studied with ethanol volume fraction,solid-liquid ratio and extraction time as the influencing factors,and the inhibitory effect of the extract on hypertrophic scar was investigated.METHODS The heating reflux method was used to extract Banhenxiao Compound.TanshinoneⅡA,salvianolic acid B,aloin and gallic acid were used as the evaluation index.The extraction process was studied by Box-Behnken response surface method.CCK-8 method was used to detect the activity of human hypertrophic scar fibroblasts(HSFb)with different concentrations of extracts.Flow cytometry was used to detect the apoptosis rate,and Western blot was used to detect the levels of apoptotic proteins Bax and Bcl-2.RESULTS The optimal extraction process was 13 times the amount of 73%ethanol extraction twice,2.1 h each time.The growth inhibition rate,apoptosis rate and Bax level of HSFb were significantly increased and the Bcl-2 level was decreased by low,medium and high doses of extracts.And the higher dose was more pronounced than the lower dose.CONCLUSION The extraction process of Banhenxiao Compound is stable and feasible.The compound extract can effectively inhibit HSFb proliferation,promote cell apoptosis and affect the expression of apoptosis protein.