核桃楸叶降血糖和抗氧化有效部位的筛选及其成分分析
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摘要
目的:筛选核桃楸叶乙醇提取物降血糖和抗氧化有效部位并确定其活性成分。方法:制备核桃楸叶乙醇提取物的不同溶剂萃取部位:二氯甲烷萃取部位、乙酸乙酯萃取部位、水饱和正丁醇萃取部位以及剩余水溶性成分部位,采用体外α-葡萄糖苷酶和α-淀粉酶抑制活性筛选模型,测定核桃楸叶乙醇提取物各萃取部位降血糖活性,以清除DPPH自由基能力研究其抗氧化活性,进而应用紫外分光光度法考察各萃取部位总黄酮含量,综合筛选并确定核桃楸叶乙醇提取物降血糖和抗氧化的有效部位,最终通过超高效液相色谱法(UPLC)确定其活性成分。结果:体外酶活性抑制试验显示,核桃楸叶乙醇提取物的乙酸乙酯萃取部位在体外有明显的抑制α-葡萄糖苷酶和α-淀粉酶作用,其最大半数抑制浓度(IC_(50))分别为0.014、0.13 mg·mL~(-1),均强于阳性药阿卡波糖(IC_(50)分别为0.044、0.158 mg·mL~(-1));DPPH自由基清除与黄酮含量测定实验结果表明,其乙酸乙酯萃取部位清除DPPH自由基能力亦强于其他萃取部位,其IC_(50)为6.89 mg·mL~(-1);乙酸乙酯活性部位中总黄酮质量分数为86.11%,为该部位的主要成分,经标品比对较好的3个活性成分为金丝桃苷、异槲皮素和紫云英苷。结论:核桃楸叶乙醇提取物乙酸乙酯萃取部位应为核桃楸叶降血糖和抗氧化有效部位,主要活性成分为金丝桃苷、异槲皮素和紫云英苷。
Objective:To screen and chemical analysis hypoglycemic and antioxidant effective fractions from the leaves of Juglans mandshurica.Methods:Activities of dichloromethane extract,ethyl acetate extract,butyl alcohol extract,water extract after extraction and ethanol extraction from the leaves of J.mandshurica were compared by using α-Glucosidase inhibition,α-Amylase inhibition and DPPH radical scavenging assays in vitro.The content of total flavonoids of effective fraction was measured by UV.The active components were determined by UPLC.Results:Enzyme inhibition kinetics in vitro showed the ethyl acetate part of ethanol extract of the leaves of J.mandshurica had strong α-glucosidase and α-Amylase inhibitory activity,maximum 50% inhibition concentration(IC_(50))were 0.014 and 0.13 mg·mL~(-1),respectively;all were stronger than the positive drug acarbose(IC_(50) were 0.044 and 0.158 mg·mL~(-1),respectively).The experimental results of DPPH free radical scavenging and flavone content determination showed that the DPPH free radical scavenging capacity of its extraction site of ethyl acetate was also better than that of other extraction sites,and its IC_(50) was 6.89 mg·mL~(-1).The total flavonoids content in the active site of ethyl acetate was 86.11%,which was the main component of the site.The three active ingredients which are better than the standard were hypericin,isoquercetin and astragalin.Conclusion:Ethyl acetate extract of ethanol extract of the leaves of J.mandshurica was hypoglycemic and antioxidant effective fraction,the major active ingredient was hypericin,isoquercetin and astragalin.
Objective:To screen and chemical analysis hypoglycemic and antioxidant effective fractions from the leaves of Juglans mandshurica.Methods:Activities of dichloromethane extract,ethyl acetate extract,butyl alcohol extract,water extract after extraction and ethanol extraction from the leaves of J.mandshurica were compared by using α-Glucosidase inhibition,α-Amylase inhibition and DPPH radical scavenging assays in vitro.The content of total flavonoids of effective fraction was measured by UV.The active components were determined by UPLC.Results:Enzyme inhibition kinetics in vitro showed the ethyl acetate part of ethanol extract of the leaves of J.mandshurica had strong α-glucosidase and α-Amylase inhibitory activity,maximum 50% inhibition concentration(IC_(50))were 0.014 and 0.13 mg·mL~(-1),respectively;all were stronger than the positive drug acarbose(IC_(50) were 0.044 and 0.158 mg·mL~(-1),respectively).The experimental results of DPPH free radical scavenging and flavone content determination showed that the DPPH free radical scavenging capacity of its extraction site of ethyl acetate was also better than that of other extraction sites,and its IC_(50) was 6.89 mg·mL~(-1).The total flavonoids content in the active site of ethyl acetate was 86.11%,which was the main component of the site.The three active ingredients which are better than the standard were hypericin,isoquercetin and astragalin.Conclusion:Ethyl acetate extract of ethanol extract of the leaves of J.mandshurica was hypoglycemic and antioxidant effective fraction,the major active ingredient was hypericin,isoquercetin and astragalin.
引文
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[11] 杨楠,贾晓斌,张振海,等.黄酮类化合物抗肿瘤活性及机制研究进展[J].中国中药杂志,2015,40(3):373-381.
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[13] 徐婷,褚江洪,丁萍英,等.葛根素对α-葡萄糖苷酶抑制作用机理的探讨[J].中国生化药物杂志,2015,35(1):43-45,48.
[14]白冰,李玉萍,叶军,等.马齿苋活性成分中α-葡萄糖苷酶抑制剂的筛选[J].安徽农业科学,2009,37(2):657-658,660.
[15]孙佳明,杜延佳,宗颖,等.白芍降血糖和抗氧化的有效部位筛选研究[J].时珍国医国药,2014,25(9):2113-2114.
[16]赵蓉,李多伟,任涛,等.DNS比色法测定白芸豆中α-淀粉酶抑制剂活性的方法研究[J].中成药,2013,35(3):573-576.
[17]恽祥惠,邢丽娜,李云,等.建立基于96孔板检测的FRAP法和DPPH法及其在橘核抗氧化活性研究中的应用[J].时珍国医国药,2014,25(5):1275-1278.
[18]冯改利,宋小妹,邓羽,等.DPPH法筛选大血藤抗氧化活性有效部位[J].陕西中医,2011,32(9):1233-1235.
[19]孙佳明,宗颖,雷岱虹,等.甘草乙醇提取物的体外抑制α-葡萄糖苷酶、抗氧化活性及HPLC-MS分析[J].中成药,2013,35(9):1943-1946.
[20]罗海青,曾子盈,邢奕丹,等.黄皮叶总黄酮的含量测定及其抗氧化活性研究[J].广东药科大学学报,2018,34(4):438-441.
[21]钱慧琴,秦晶晶,赵媛,等.月季不同部位总黄酮含量分析及其抗氧化活性研究[J].食品研究与开发,2018,39(20):19-22.
[2] 常仁龙,孙佳明,张博,等.核桃楸叶化学成分研究[J].中成药,2009,31(7):1082-1085.
[3] 赵娜娜,刘汇,孙道磊,等.核桃楸不同药用部位的抗氧化活性比较[J].长春中医药大学学报,2018,34(1):26-28.
[4] 任晓蕾,曹贵阳,初东君,等.核桃楸不同药用部位总黄酮含量测定及变化规律[J].中国实验方剂学杂志,2012,18(24):104-106.
[5] 陈栏心.广西1996～2012年艾滋病流行特征及流行地区数理判别模型研究[D].南宁:广西医科大学,2014.
[6] 孙佳明,张林林,宗颖,等.甘草降血糖和抗氧化的有效部位筛选研究[J].时珍国医国药,2013,24(12):2879-2880.
[7] 冯长根,陈凌,刘霞.以中草药为来源的α-葡萄糖苷酶抑制剂筛选研究进展[J].中国新药杂志,2005,14(6):669-672.
[8] 吕凤霞,陆兆新.α-淀粉酶抑制剂的研究进展[J].食品科学,2002,23(3):152-155.
[9] 洪佳敏,林宝妹,张朝坤,等.番石榴醇提物抗氧化能力及α-葡萄糖苷酶抑制活性研究[J].西北农林科技大学学报(自然科学版),2019,47(1):1-7.
[10] 胡绪乔,原菲,严春艳,等.银杏多糖的分离鉴定和体外抗氧化活性测试[J].中药材,2011,34(12):1950-1953.
[11] 杨楠,贾晓斌,张振海,等.黄酮类化合物抗肿瘤活性及机制研究进展[J].中国中药杂志,2015,40(3):373-381.
[12] 许有瑞,伊辉,张可锋,等.瑶药葫芦钻中α-葡萄糖苷酶抑制活性部位的筛选[J].中国药房,2017,28(16):2235-2237.
[13] 徐婷,褚江洪,丁萍英,等.葛根素对α-葡萄糖苷酶抑制作用机理的探讨[J].中国生化药物杂志,2015,35(1):43-45,48.
[14]白冰,李玉萍,叶军,等.马齿苋活性成分中α-葡萄糖苷酶抑制剂的筛选[J].安徽农业科学,2009,37(2):657-658,660.
[15]孙佳明,杜延佳,宗颖,等.白芍降血糖和抗氧化的有效部位筛选研究[J].时珍国医国药,2014,25(9):2113-2114.
[16]赵蓉,李多伟,任涛,等.DNS比色法测定白芸豆中α-淀粉酶抑制剂活性的方法研究[J].中成药,2013,35(3):573-576.
[17]恽祥惠,邢丽娜,李云,等.建立基于96孔板检测的FRAP法和DPPH法及其在橘核抗氧化活性研究中的应用[J].时珍国医国药,2014,25(5):1275-1278.
[18]冯改利,宋小妹,邓羽,等.DPPH法筛选大血藤抗氧化活性有效部位[J].陕西中医,2011,32(9):1233-1235.
[19]孙佳明,宗颖,雷岱虹,等.甘草乙醇提取物的体外抑制α-葡萄糖苷酶、抗氧化活性及HPLC-MS分析[J].中成药,2013,35(9):1943-1946.
[20]罗海青,曾子盈,邢奕丹,等.黄皮叶总黄酮的含量测定及其抗氧化活性研究[J].广东药科大学学报,2018,34(4):438-441.
[21]钱慧琴,秦晶晶,赵媛,等.月季不同部位总黄酮含量分析及其抗氧化活性研究[J].食品研究与开发,2018,39(20):19-22.