杜仲叶多糖、杜仲精粉、杜仲胶的提取分离及其性能分析
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摘要
为实现杜仲叶中生物活性成分和杜仲胶的高效提取分离,采用连续热水浸提、碱提取、酶水解和石油醚提取杜仲叶,分别得到了杜仲精粉、杜仲多糖(水溶性多糖和碱提多糖)及杜仲胶。考察了料液比和温度对活性成分提取得率的影响,探讨了酶解时间和酶种类对杜仲叶残渣水解的影响,最后对杜仲精粉、水溶性多糖和碱提多糖的铁离子还原能力和DPPH自由基(DPPH·)清除活性进行了测定,并对杜仲胶相对分子质量进行了测定。研究结果表明:料液比为1∶10(g∶mL),温度为80℃时,总黄酮和绿原酸的提取得率最大分别为50.08和6.83 mg/g。杜仲水溶性多糖、碱提多糖、杜仲精粉均具有较好的抗氧化能力,其中杜仲精粉抗氧化活性最高, 5 g/L时杜仲精粉的铁离子还原能力为4.01 mmol/L, 300 mg/L时DPPH·的清除率为94.46%,与Vc相当。杜仲叶水提残渣酶解48 h时,葡聚糖和木聚糖水解率最大,分别为86.77%和15.62%。杜仲叶中水溶性多糖的得率为13.2%,杜仲胶得率为1.86%,碱提多糖得率为9.8%,杜仲精粉得率为8.0%。杜仲胶的M_w和M_n分别为330 398和84 526,远大于水溶性多糖和碱提多糖的相对分子质量,但相对分子质量分布指数为3.91,分散程度明显低于杜仲多糖。
In order to extract bioactive components and gutta-percha effectively from leaves of Eucommia ulmoides Oliver, the successive extractions of the leaves by hot water, alkali, and enzyme-petroleum ether were used to obtain the extractum, polysaccharides(water-soluble polysaccharides and alkali-extracted polysaccharides) and gutta-percha, respectively. Effects of solid to liquid ratio and temperature on yields of bioactive components were investigated. Meanwhile, the effects of time and type of enzymes on enzymatic hydrolysis of the residues were discussed. Finally, the ferric reducing/antioxidant power and the scavenging DPPH radical activity of bioactive constituents, water-soluble and alkali-extracted polysaccharides were determined, and the molecular weights of gutta-percha were determined. The results showed that the highest yields of flavonoids and chlorogenic acid were 50.08 mg/g and 6.83 mg/g with solid to liquid ratio of 1∶10(g∶mL)at 80 ℃. The water-soluble and alkali-extracted polysaccharides and extractum from E. ulmoides Oliver exhibited good antioxidant activity, in which the antioxidant activity of extractum was the highest. The total antioxidant activity of 5 g/L extractum was 4.01 mmol/L, and the scavenging rate of DPPH radical at 300 mg/L was 94.46%, which was the same as that of Vc. The hydrolysis rates of glucan and xylan were the highest when water-extracted residues of E. ulmoides leaves were hydrolysed for 48 h, which were 86.77% and 15.62%, respectively. The yields of extractum, water-soluble polysaccharides, alkali-extracted polysaccharides and gutta-pereha were 8.0%, 13.2%, 9.8%, 1.86%, respectively. The weight average relative molecular mass(M_w) and the number average relative molecular mass(M_n) of gutta-pereha were 330 398 and 84 526, respectively, which were higher than the molecular weights of polysaccharides. Meanwhile, M_w/M_n=3.91,and the dispersity of extractum was lower than that of polysaccharides.
In order to extract bioactive components and gutta-percha effectively from leaves of Eucommia ulmoides Oliver, the successive extractions of the leaves by hot water, alkali, and enzyme-petroleum ether were used to obtain the extractum, polysaccharides(water-soluble polysaccharides and alkali-extracted polysaccharides) and gutta-percha, respectively. Effects of solid to liquid ratio and temperature on yields of bioactive components were investigated. Meanwhile, the effects of time and type of enzymes on enzymatic hydrolysis of the residues were discussed. Finally, the ferric reducing/antioxidant power and the scavenging DPPH radical activity of bioactive constituents, water-soluble and alkali-extracted polysaccharides were determined, and the molecular weights of gutta-percha were determined. The results showed that the highest yields of flavonoids and chlorogenic acid were 50.08 mg/g and 6.83 mg/g with solid to liquid ratio of 1∶10(g∶mL)at 80 ℃. The water-soluble and alkali-extracted polysaccharides and extractum from E. ulmoides Oliver exhibited good antioxidant activity, in which the antioxidant activity of extractum was the highest. The total antioxidant activity of 5 g/L extractum was 4.01 mmol/L, and the scavenging rate of DPPH radical at 300 mg/L was 94.46%, which was the same as that of Vc. The hydrolysis rates of glucan and xylan were the highest when water-extracted residues of E. ulmoides leaves were hydrolysed for 48 h, which were 86.77% and 15.62%, respectively. The yields of extractum, water-soluble polysaccharides, alkali-extracted polysaccharides and gutta-pereha were 8.0%, 13.2%, 9.8%, 1.86%, respectively. The weight average relative molecular mass(M_w) and the number average relative molecular mass(M_n) of gutta-pereha were 330 398 and 84 526, respectively, which were higher than the molecular weights of polysaccharides. Meanwhile, M_w/M_n=3.91,and the dispersity of extractum was lower than that of polysaccharides.
引文
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[6]张学俊,伊廷金,孙黔云,等.杜仲叶多糖的提取分离、抗补体活性及结构研究[J].天然产物研究与开发,2011,23(4):606-611,637.ZHANG X J,YI T J,SUN Q Y,et al.Isolation,anti-complement activities,and chemical structures of polysaccharide from Eucommia ulmoides leaves[J].Natural Product Research and Development,2011,23(4):606-611,637.
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[8]张学俊,王庆辉,宋磊,等.不同温度条件下溶剂循环溶解析出提取杜仲胶[J].天然产物研究与开发,2007,19(6):1062-1066.ZAHNG X J,WANG Q H,SONG L,et al.The solution-precipitation of Eucommia ulmoids gum in petroleum ether at different temperatures to extract the gum [J].Natural Product Research and Development,2007,19(6):1062-1066.
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[10]刘景煜,李晨,高锦明,等.枸杞子总黄酮含量检测方法优化[J].食品工业科技,2016,37(13):304-308.LIU J Y,LI C,GAO J M,et al.Optimization on test method of total flavonoids from Lycium barbarum L[J].Science and Technology of Food Industry,2016,37(13):304-308.
[11]DONG J E,MA B L,JIA E H.Study on measuring method of aucubin in Eucommia ulmoids leaf[J].Journal of Northwest Foresty University,2001,16(1):53-55.
[12]申迎宾,张友维,黄才欢,等.提取溶剂对青稞提取物总酚、黄酮含量及其抗氧化活性的影响[J].食品与机械,2016,32(11):133-136.SHEN Y B,ZHANG Y W,HUANG C H,et al.Effect of different solvents on total phenolic content,total flavonoid content and antioxidant activity of highland barley[J].Food & Machinery,2016,32(11):133-136.
[13]彭密军,周春山,钟世安,等.杜仲中桃叶珊瑚苷的提取工艺研究[J].林产化学与工业,2003,23(3):65-68.PENG M J,ZHOU C S,ZHONG S A,et al.Study on technology of extracting aucubin from Eucommia [J].Chemistry and Industry of Forest Products,2003,23(3):65-68.
[14]张学俊,宫本红,王庆辉,等.酶水解杜仲纤维素细胞壁及长丝杜仲胶的提取[J].天然产物研究与开发,2009,21(1):115-121.ZHANG X J,GONG B H,WANG Q H,et al.Enzymatic hydrolysis of cellulose cell wall and filum Eucommia ulmoides extract[J].Natural Product Research and Development,2009,21(1):115-121.
[15]刘贵华,张永康,肖美凤,等.纤维素酶解预处理法提取杜仲胶的工艺研究[J].林产化学与工业,2010,30(2):77-82.LIU G H,ZHANG Y K,XIAO M F,et al.Study on the process of extracting gutta-percha by cellulase hydrolysis pretreatment[J].Chemistry and Industry of Forest Products,2010,30(2):77-82.
[16]曾维才,石碧.天然产物抗氧化活性的常见评价方法[J].化工进展,2013,32(6):1205-1213.ZEN W C,SHI B.Common evaluation method for antioxidant activity of natural products[J].Chemical Industry and Engineering Progress,2013,32(6):1205-1213.
[17]杨娜,王鸿飞,许凤,等.蕨麻多糖提取及抗氧化活性研究[J].中国食品学报,2014,14(2):60-65.YANG N,WANG H F,XU F,et al.Study on the extraction and antioxidant activity of polysaccharides from fern ramus[J].Journal of Chinese Institute of Food Science and Technology,2014,14(2):60-65.
[2]TAKESHI D,SANSEI N,YOSHIHISA N.Constituents and pharmacological effects of Eucommia and Siberian ginseng[J].Acta Pharmacologica Sinica,2001,22:1057-1070.
[3]DONG J E,MA X H,WEI Q,et al.Effects of growing location on the contents of secondary metabolites in the leaves of four selected superior clones of Eucommia ulmoides Oliver[J].Industrial Crops and Products,2011,34(3):1607-1614.
[4]董娟娥,马柏林,刘丽,等.超声波提取杜仲叶中有效成分工艺研究[J].西北林学院学报,2003,18(3):66-68.DONG J E,MA B L,LIU L,et al.Ultrasonic extraction of active ingredients from Eucommia ulmoides[J].Journal of Northwest Forestry University,2003,18(3):66-68.
[5]肖卓炳,郭满满,郭瑞轲,等.杜仲胶的热稳定性、分解动力学及热老化贮存期[J].林产化学与工业,2013,33(6):7-13.XIAO Z B,GUO M M,GUO R K,et al.Thermal stability,decomposition kinetics and storage time of gutta-percha[J].Chemistry and Industry of Forest Products,2013,33(6):7-13.
[6]张学俊,伊廷金,孙黔云,等.杜仲叶多糖的提取分离、抗补体活性及结构研究[J].天然产物研究与开发,2011,23(4):606-611,637.ZHANG X J,YI T J,SUN Q Y,et al.Isolation,anti-complement activities,and chemical structures of polysaccharide from Eucommia ulmoides leaves[J].Natural Product Research and Development,2011,23(4):606-611,637.
[7]ZHU M Q,WEN J L,ZHU Y H,et al.Isolation and analysis of four constituents from barks and leaves of Eucommia ulmoides Oliver by a multi-step process[J].Industrial Crops and Products,2016,83:124-132.
[8]张学俊,王庆辉,宋磊,等.不同温度条件下溶剂循环溶解析出提取杜仲胶[J].天然产物研究与开发,2007,19(6):1062-1066.ZAHNG X J,WANG Q H,SONG L,et al.The solution-precipitation of Eucommia ulmoids gum in petroleum ether at different temperatures to extract the gum [J].Natural Product Research and Development,2007,19(6):1062-1066.
[9]SLUITER A,HAMES B ,RUIZ R ,et al.Determination of structural carbohydrates and lignin in biomass:Laboratory Analytical Procedure(LAP):NREL/TP-510-42618[G].Golden:National Renewable Energy Laboratory,2012-08-03.
[10]刘景煜,李晨,高锦明,等.枸杞子总黄酮含量检测方法优化[J].食品工业科技,2016,37(13):304-308.LIU J Y,LI C,GAO J M,et al.Optimization on test method of total flavonoids from Lycium barbarum L[J].Science and Technology of Food Industry,2016,37(13):304-308.
[11]DONG J E,MA B L,JIA E H.Study on measuring method of aucubin in Eucommia ulmoids leaf[J].Journal of Northwest Foresty University,2001,16(1):53-55.
[12]申迎宾,张友维,黄才欢,等.提取溶剂对青稞提取物总酚、黄酮含量及其抗氧化活性的影响[J].食品与机械,2016,32(11):133-136.SHEN Y B,ZHANG Y W,HUANG C H,et al.Effect of different solvents on total phenolic content,total flavonoid content and antioxidant activity of highland barley[J].Food & Machinery,2016,32(11):133-136.
[13]彭密军,周春山,钟世安,等.杜仲中桃叶珊瑚苷的提取工艺研究[J].林产化学与工业,2003,23(3):65-68.PENG M J,ZHOU C S,ZHONG S A,et al.Study on technology of extracting aucubin from Eucommia [J].Chemistry and Industry of Forest Products,2003,23(3):65-68.
[14]张学俊,宫本红,王庆辉,等.酶水解杜仲纤维素细胞壁及长丝杜仲胶的提取[J].天然产物研究与开发,2009,21(1):115-121.ZHANG X J,GONG B H,WANG Q H,et al.Enzymatic hydrolysis of cellulose cell wall and filum Eucommia ulmoides extract[J].Natural Product Research and Development,2009,21(1):115-121.
[15]刘贵华,张永康,肖美凤,等.纤维素酶解预处理法提取杜仲胶的工艺研究[J].林产化学与工业,2010,30(2):77-82.LIU G H,ZHANG Y K,XIAO M F,et al.Study on the process of extracting gutta-percha by cellulase hydrolysis pretreatment[J].Chemistry and Industry of Forest Products,2010,30(2):77-82.
[16]曾维才,石碧.天然产物抗氧化活性的常见评价方法[J].化工进展,2013,32(6):1205-1213.ZEN W C,SHI B.Common evaluation method for antioxidant activity of natural products[J].Chemical Industry and Engineering Progress,2013,32(6):1205-1213.
[17]杨娜,王鸿飞,许凤,等.蕨麻多糖提取及抗氧化活性研究[J].中国食品学报,2014,14(2):60-65.YANG N,WANG H F,XU F,et al.Study on the extraction and antioxidant activity of polysaccharides from fern ramus[J].Journal of Chinese Institute of Food Science and Technology,2014,14(2):60-65.