木棉花黄酮的大孔树脂纯化及其祛痘功效分析

Purification of Kapok Flavonoids by Macroporous Resin and Its Anti-Acne Efficacy

以广东肇庆木棉花为原料,经过超声辅助醇提后,筛选最适大孔树脂类型并确定木棉花黄酮粗提物的纯化工艺,采用体外抑菌圈实验方法测定纯化精制的木棉花黄酮对表皮葡萄球菌和痤疮丙酸杆菌的抑制效果;采用体外美白活性方法测定木棉花黄酮对酪氨酸酶的抑制率,并通过斑马鱼胚胎黑色素抑制测试法对木棉花黄酮的美白活性进行进一步验证;最后采用体外抗氧化活性测定方法考察木棉花黄酮对ABTS·^(+)和DPPH等自由基的清除能力。通过对比8种大孔树脂对木棉花黄酮的吸附率和解析率,筛选出D101为最佳树脂类型,同时考察得出其解析液最佳的乙醇体积分数为60%;其最佳的动态吸附-解析条件为:上样体积2 BV,解析液洗脱体积0.625 BV。纯化后木棉花黄酮的纯度由32.5%提高至98.2%。精制的木棉花黄酮对ABTS·^(+)和DPPH自由基清除率的IC_(50)(半抑制浓度)值分别为(616.90±11.15)mg/L和(11.80±0.62)mg/L。木棉花黄酮对酪氨酸酶抑制率的IC_(50)值为(1.69±0.01)g/L。木棉花黄酮质量浓度在1.0 g/L时对斑马鱼胚胎黑色素的抑制率为12%,具有显著的美白活性。通过抑菌圈实验可得木棉花黄酮质量浓度为12.5 g/L时,对痤疮丙酸杆菌表现为中度敏感,对表皮葡萄球菌表现为高度敏感,且此抑菌效果呈现剂量依赖效应。通过检测木棉花黄酮对RAW264.7分泌细胞炎症因子TNF-α和IL-6的抑制效果,可得当木棉花黄酮质量浓度大于0.5 g/L时能显著抑制炎症因子TNF-α的分泌,当木棉花黄酮的质量浓度为0.25 g/L及以上时即可显著抑制炎症因子IL-6的分泌。

This study use Guangdong Zhaoqing kapok as raw material.After ultrasonic-assisted alcohol extraction,the most suitable macroporous resin type was investigated,and the purification process of the crude kapok flavonoids was determined.The inhibitory effect of purified kapok flavonoids on Staphylococcus and Propionibacterium acnes was measured by tests of antibacterial activity in vitro.The inhibition rate of kapok flavonoids on tyrosinase was determined by in vitro whitening activity method,and the whitening activity of kapok flavonoids was further verified by zebrafish embryo melanin inhibition test method.Finally,the scavenging ability of kapok flavonoids to free radi-cals such as ABTS·^(+) and DPPH was investigated by in vitro antioxidant activity assay.By comparing the adsorption and desorption rates of 8 macroporous resins to kapok flavonoids,D101 was screened out as the best resin type and volumetric concentration of ethanol for the best analytical solution is 60%.The optimal adsorption-analytical conditions are:the sample volume is 2 BV,and the elution volume of the analytical solution is 0.625 BV.After purification,the purity of kapok flavone increased from 32.5%to 98.2%.The IC_(50) values of the purified kapok flavone for the free radical scavenging rate of ABTS·^(+) and DPPH are(616.90±11.15)mg/L and(11.80±0.62)mg/L,respectively.The IC_(50) of kapok flavone on tyrosinase inhibition is(1.69±0.01)g/L.When the concentration of kapok flavone is 1.0 g/L,the inhibition rate of zebrafish embryo melanin is 12%,with significant whitening activity.The inhibition zone experiment shows that when the mass concentration of kapok flavonoids is 12.5 g/L,it is moderately sensitive to Propionibacterium acnes and highly sensitive to Staphylococcus epidermidis,and the antibacterial effect shows a dose-dependent effect.The inhibitory effect of kapok flavonoids on the inflammatory factors TNF-αand IL-6 secreted by RAW264.7 cells is detected by ELISA,which shows when the mass concentration of kapok flavonoids is more than 0.5 g/L,it can significantly inhibitthe secretion of inflammatory cytokines TNF-α;when the mass concentration of kapok flavonoids is 0.25 g/L and above,the secretion of inflammatory factor IL-6 can be significantly inhibited.