巨尾桉树皮总多酚的提取分离及其生物活性研究

Extraction Technology for Total Polyphenols from Eucalyptus grandis × E. urograndis Bark and Their Bioactivities

以巨尾桉树皮为原料提取按树皮中的总多酚,通过正交试验优化了酶处理与超声波辅助联合提取法及超声波辅助提取法的工艺条件,并对抗氧化活性较强的乙酸乙酯萃取物进一步分离,通过大孔吸附树脂分离纯化制备得到11个活性组分(Fr.1~Fr.11)及1个活性单体化合物(Pu1),同时对其进行了体外抗氧化活性实验,并对Pu1进行了结构表征。研究结果表明:超声波辅助提取法的最佳工艺条件为60%乙醇、提取温度60℃、超声波作用时间50 min、料液比为1∶12(g∶mL),总多酚得率为5.12%。酶处理与超声波辅助联合提取法中最佳酶处理工艺条件为:酶溶液质量浓度0.15 g/L、酶解温度50℃、酶解时间50 min、酶溶液pH值5.0,在此基础上再经超声波辅助提取,总多酚得率为6.23%。乙酸乙酯萃取物的分离活性组分中Fr.5、Fr.6、Fr.7和Fr.8在质量浓度为24 mg/L时对DPPH·清除率大于60%,具有较好抗氧化能力,Pu1的半数抑制质量浓度(IC_(50))值为8.99 mg/L,小于对照样Vc的IC_(50)值(10.46 mg/L),说明Pu1具有显著的体外抗氧化活性。活性组分的总多酚含量与其抗氧化活性有很好的相关性(R=0.944 8,P<0.000 1)。通过UV、FT-IR、MS和NMR等分析表明:化合物Pu1的相对分子质量为498,分子式为C21H_22O14,确定化合物Pu1为2'-甲氧基-六羟基联苯二酸-4-O-α-L-鼠李糖苷。

The optimum extraction conditions of total polyphenols from Eucalyptus grandis×E.urograndis bark via enzymolysis treatment and ultrasonic-assisted coupling extraction method and ultrasonic extraction method were investigated by using orthogonal experiment.Vitro antioxidant activities of different solvent extracts by ultrasonic extracts were studied and the ethyl acetate extract exhibited stronger antioxidant activity.Then,11 kinds of fractions(Fr.1-Fr.11)and a purified compound(Pu1)were separated from the ethyl acetate extract by AB-8 macroporous resin.The optimum crafts for ultrasonic extraction method were 60%ethanol,ultrasonic time 50 min,ratio of material-liquid 1∶12(g∶mL),ultrasonic temperature 60℃,and the yield of total polyphenols was 5.12%.The optimum crafts for enzymolysis treatment and ultrasonic-assisted coupling extraction method were 0.15 g/L enzyme concentration,enzymolysis temperature 50℃,enzymolysis time 50 min,enzyme solution pH 5.0,combining with ultrasonic extraction,the yield of total polyphenols was 6.23%.The results of radical scavenging capacity assay showed that the scavenging rates of Fr.5,Fr.6,Fr.7 and Fr.8 active fractions at 50 mg/L were greater than 60%,and they had high antioxidant activity.The IC 50 of the active compound Pu1 was 8.99 mg/L,less than that of Vc(10.46 mg/L),indicating significant antioxidant activities.Moreover,the phenolic content of the active fractions showed positive correlation with their antioxidant activities(R=0.944 8,P<0.000 1).The structure of compound Pu1 was characterized by UV,FT-IR,MS and NMR and the results showed that its molecular formula and molar mass were C 21 H 22 O 14 and 498,respectively,which was named as 2′-methoxy-hexadylic biphenylate-4-O-α-L-rhamnoside.