S-8大孔树脂-C18柱联用分离牡丹籽壳中短叶松素及抗氧化研究

Study on separation of pinobanksin from peony seed shell by S-8 macroporous resin-C18 column and its antioxidation

以牡丹籽壳为原料,利用大孔树脂和C18反相键合硅胶柱(C18柱)联合分离短叶松素,回收率为(98.33±0.64)%。用70%(体积分数)乙醇提取牡丹籽壳粗黄酮(Mu Dan Ke Flavonoids,MDKF),得率最高为(10.54±0.13)%;比较了6种大孔树脂(AB-8、S-8、DM301、HPD600、HPD100和D101)的吸附率和解析率,发现S-8大孔树脂的吸附率和解析率最佳,分别为83.47%和84.46%;优化S-8大孔树脂分离MDKF的最佳条件为:上样液质量浓度1.6 mg/mL,上样液流速2.0 mL/min,洗脱剂乙醇体积分数60%,洗脱液流速1.5 mL/min,洗脱液体积100 mL;对经过C18柱分离后的组分进行LC-MS分析得到短叶松素,回收率为(98.33±0.64)%;分离前后抗氧化能力比较:C18纯化物>S-8大孔树脂纯化物>MDKF粗提物;分子对接实验表明,短叶松素与超氧化物歧化酶、过氧化氢酶、过氧化物酶、谷胱甘肽过氧化物酶均有结合能力,过氧化氢酶的结合能力最强为-9.1 kcal/mol。实验表明,短叶松素具有较好的抗氧化能力,可作为食品、药品或化妆品等抗氧化剂的添加,应用前景非常广泛,对牡丹籽壳的进一步开发利用提供理论支持。

Pinobanksin was isolated from a peony seed shell using macroporous resin and C18 reversed bonded silica gel column(C18 column)with a recovery of(98.33±0.64)%.The maximum yield of MDKF from peony seed shell was(10.54±0.13)%with 70%ethanol(volume fraction).The adsorption and analytical rates of six macroporous resins(AB-8,S-8,DM301,HPD600,HPD100,and D101)were compared,and the adsorption and analytical rates of S-8 macroporous resins were the best,which were 83.47%and 84.46%,respectively.The optimal conditions for the separation of MDKF by S-8 macroporous resin were that the mass concentration of loading liquid was 1.6 mg/mL,the flow rate of loading liquid was 2.0 mL/min,the volume fraction of elution agent ethanol was 60%,the flow rate of eluent was 1.5 mL/min,and the eluent volume was 100 mL.The fractions separated by the C18 column were analyzed by LC-MS and the recovery was(98.33±0.64)%.Comparing antioxidant activities before and after separation,results showed that C18 purified substance>S-8 macroporous resin purified substance>MDKF crude extract.Molecular docking experiments showed that pinobanksin had a binding ability with super oxide dimutese,catalase from Micrococcus lysodeiktic,peroxidase,and glutathione peroxidase enzymes,and the binding ability of catalase was-9.1 kcal/mol.Results showed that pinobanksin had a good antioxidant capacity and could be used as an antioxidant supplement for food,medicine,or cosmetics.It has a very broad application prospect and provides theoretical support for further development and utilization of peony seed shells.