CN109364119B - 从青钱柳叶中制备具有降血糖作用的总三萜的方法及应用 - Google Patents
从青钱柳叶中制备具有降血糖作用的总三萜的方法及应用 Download PDFInfo
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- CN109364119B CN109364119B CN201811598696.7A CN201811598696A CN109364119B CN 109364119 B CN109364119 B CN 109364119B CN 201811598696 A CN201811598696 A CN 201811598696A CN 109364119 B CN109364119 B CN 109364119B
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Abstract
本发明提供一种从青钱柳叶中制备具有降血糖作用的总三萜的方法,以青钱柳叶为原料,经乙醇水混合溶剂提取,有机溶剂液液萃取后,通过聚酰胺柱色谱有机溶剂洗脱和大孔树脂柱色谱乙醇水洗脱进行分离纯化,可得到高纯度的总三萜。经研究表明,该总三萜及其中含有的主要成分阿江榄仁酸、积雪草酸、青钱柳酸B、化合物4,能有效增加3T3‑L1脂肪细胞的葡萄糖摄取,具有抗糖尿病活性,可将总三萜及其主要成分用于制备降血糖的抗糖尿病的药物、保健食品和功能食品等产品。本发明开发了一种青钱柳叶中总三萜的制备方法及其和化合物4在制备抗糖尿病药物中的应用,为推广利用和深加工中药材提供依据,并提供了新的应用方向。
Description
技术领域
本发明属医药食品技术领域,具体涉及一种从青钱柳叶中制备具有降血糖作用的高纯度总三萜的方法,及其总三萜在制备抗糖尿病药物、保健食品和功能食品等产品中的应用。
背景技术
青钱柳叶为胡桃科青钱柳属植物青钱柳Cyclocarya paliurus(Batalin)Iljinsk.的干燥叶。青钱柳是我国特有的单种属植物,广泛分布于江西、浙江、安徽、湖南、广东等长江以南地区。民间将其叶作为茶饮,该茶味甜,具有生津止渴、清热解暑、降血糖、降血压和延年益寿的功效。2013年底,经国家卫计委批准青钱柳叶成为新食品原料。现代药理实验证实青钱柳叶具有治疗糖尿病、高脂血症、高血压及肥胖的作用,目前大部分研究都是针对青钱柳叶中的黄酮类、多糖类物质,对青钱柳叶中所含有的三萜类化合物的药理学研究则少有报道。
Zhu等人(Zhu KN,Jiang CH,Tian YS,et al.Two triterpeniods fromCyclocarya paliurus(Batal)Iljinsk(Juglandaceae)promote glucose uptake in 3T3-L1adipocytes:The relationship to AMPK activation.Phytomedicine,2015,22(9):837-846.)研究了青钱柳叶80%乙醇提取物的氯仿部位以及从中分离得到的两个三萜类主要成分cyclocaric acid B和cyclocarioside H的降糖作用,结果表明对于3T3-L1脂肪细胞,该部位和两个三萜类成分均能显著促进其葡萄糖消耗。Wu等人(Wu ZF,Meng FC,CaoLJ,et al.Triterpenoids from Cyclocarya paliurus and their inhibitory effecton the secretion of apoliprotein B48in Caco-2cells.Phytochemistry,2017,142,76-84.)从青钱柳叶80%乙醇提取物的氯仿部位分离得到了22个三萜类成分,发现三萜类成分能有效抑制Caco-2细胞apoB48的分泌,降低高脂饲料喂养的高血脂症大鼠血脂水平。中国专利CN101792479A提供了一种青钱柳中常春藤皂苷H的提取工艺,主要特征为90%乙醇提取,石油醚、乙醚脱脂,取其正丁醇萃取液,HPD400型大孔树脂纯化获得。中国专利CN103242422A公开了一种从青钱柳叶中提取青钱柳酸A的方法,主要特征为50~90%甲醇溶液超声提取,大孔树脂纯化,高速逆流色谱再纯化获得。中国专利CN103232515A公开了一种制备青钱柳苷Ⅰ的方法,其特征为50~80%甲醇浸泡提取,大孔树脂40~70%乙醇溶液洗脱纯化,之后再用大孔树脂吸附并以乙酸乙酯、甲醇洗脱获得。尹忠平等人(尹忠平,上官新晨,黎冬明,等.超声辅助提取青钱柳叶总三萜化合物研究.江西农业大学学报,2010,32(2):0373-0377.)优化了青钱柳叶中总三萜的提取工艺,即在65%的乙醇、料液比1:12,超声30min的条件下,总三萜得率为9.19%。随后,尹忠平等人(尹忠平,上官新晨,张月红,等.大孔树脂吸附纯化青钱柳叶三萜化合物.食品科学,2011,32(6):61-65.)采用AB-8大孔树脂对青钱柳叶65%乙醇超声三萜粗提物进行吸附纯化,三萜类物质纯度达到44.30%。尹忠平等人(尹忠平,上官新晨,张月红,等.高纯度青钱柳叶总三萜化合物精制研究.食品科技,2011,36(2):161-165.)之后又对经大孔树脂纯化的总三萜,进行乙酸乙酯固液萃取和活性炭脱色处理,获得精制总三萜,纯度可达94.61%。未见有关本申请所公开的以聚酰胺材料正相洗脱体系为核心的制备总三萜的制备工艺的研究或报道。
发明内容
本发明的目的是提供一种从青钱柳叶中制备总三萜的方法,是制备青钱柳总三萜的方法,通过以下步骤实现:将青钱柳植物的叶子按药材/溶剂(重量/体积比)加入5-12倍量的乙醇水混合溶液,室温浸提、或回流提取2-3次,合并提取液减压浓缩至无醇味,先用石油醚脱色,然后用中极性有机溶剂等体积萃取,回收溶剂得到浸膏,所得浸膏直接与1至2倍(重量比)的聚酰胺材料拌样吸附,再将3至10倍的聚酰胺装入柱色谱,经混合有机溶剂洗脱,浓缩获得浸膏,然后用蒸馏水分散溶解浸膏,上大孔树脂柱,先用20%~50%乙醇水混合溶液(体积比)洗去杂质,收集50%~95%乙醇水洗脱液,干燥得到高纯度总三萜。
上述提取用的乙醇水混合溶液的用量优选原料的8-10倍,浓度为50%~95%乙醇,更优选60%~80%乙醇水混合溶液(体积比);萃取所用的中极性有机溶剂包括三氯甲烷、乙酸乙酯、正丁醇或其中的两种的混合物;洗脱用的混合有机溶剂为石油醚、己烷、二氯甲烷、三氯甲烷、乙酸乙酯、丙酮、甲醇等之中的两种按20:1至1:1的比例混合而成;去除杂质所用的乙醇水浓度优选40%(体积比);收集洗脱液所用的乙醇水浓度优选70%~80%。
所述提取方法为室温浸提或回流提取,提取2-3次。
所述的聚酰胺材料为60~200目,优选100~200目;大孔树脂类型选自DM130、AB-8、D101型号树脂材料。
用紫外-可见分光光度仪测定总三萜成品中三萜类成分的含量,以人参皂苷Re为对照品,5%香草醛-冰醋酸和高氯酸为显色剂,检测波长为560nm,测得总三萜含量为80%以上。经HPLC检测确定该总三萜中含有多种三萜类单体,主要包括阿江榄仁酸(化合物1)、积雪草酸(化合物2)、青钱柳酸B(化合物3)和(20S,24R)-20,24-epoxy-25-hydroxy-12β-(α-L-arabinopyranosyloxy)-3,4-seco-dammara-4(28)-en-3-oic acid(化合物4)。
与已知的文献对比,Zhu等人报道了青钱柳叶80%乙醇提取物的氯仿部位和两个三萜类成分cyclocaric acid B和cyclocarioside H的降糖活性,但并未研究总三萜的纯化工艺,且其主要成分与本发明所述总三萜主要成分不同。Wu等人报道的青钱柳叶80%乙醇提取物氯仿部位的三萜类成分能抑制Caco-2细胞apoB48的分泌,降低高血脂症大鼠血脂水平,同样也并未提供高纯度总三萜的制备方法和降糖活性研究。中国专利CN101792479A提供的一种青钱柳中常春藤皂苷H的提取工艺,与本发明所公开的研究对象以及制备方法都截然不同。中国专利CN103242422A公开的一种从青钱柳叶中提取青钱柳酸A的方法,同样也与本发明所涉及的研究对象以及提取纯化工艺明显不同。中国专利CN103232515A公开的一种制备青钱柳苷Ⅰ的方法,与本发明公开的以聚酰胺材料正相洗脱体系为核心的高纯度总三萜制备工艺以及主要成分明显不同。尹忠平等人报道的青钱柳叶中总三萜的提取优化工艺,即采用超声提取的方法,与本发明公开的总三萜制备工艺截然不同,总三萜纯度也不高。之后,尹忠平等人采用AB-8大孔树脂对青钱柳叶65%乙醇超声三萜粗提物进行吸附纯化,三萜类物质纯度达到44.30%,而本发明所制备的总三萜纯度能达到80%及以上,且其纯化方法与本发明公开的总三萜制备工艺有所不同。随后,尹忠平等人又将青钱柳叶依次通过石油醚脱脂,超声辅助提取2次,大孔树脂吸附纯化,乙酸乙酯超声辅助固液萃取,活性炭脱色步骤,获得精制总三萜,纯度可达94.61%,与本发明公开的高纯度总三萜制备工艺相比,超声辅助提取与超声辅助固液萃取工业化生产相对较难,而且提取纯化步骤多,工艺相对复杂,活性炭反复利用率低,成本高。由于尹忠平等人文章中缺少青钱柳叶经石油醚脱脂后的总三萜提取得率的关键数据,所以无法计算出具体总三萜得率,未能与本发明公开的高纯度总三萜得率相比较。另外,其总三萜测定方法与本发明所使用的方法中的标准品不同,因此含量指标也不具备完全的可比性。从成分上来看,尹忠平等人所述的精制总三萜特征为“石油醚:乙酸乙酯:甲醇=30:10:7展开后仅显示出紫色点,基本上无杂色点(非紫色点),而粗提总三萜展开显色后还有黄色、绿色、棕色和黑褐色等杂色点”,而本发明所述的总三萜主要TLC(二氯甲烷:甲醇=10:1)特征为紫色点和蓝色点(附图2),主要成分包括阿江榄仁酸(1,蓝色点)、积雪草酸(2,蓝色点)、青钱柳酸B(3,紫色点)和(20S,24R)-20,24-epoxy-25-hydroxy-12β-(α-L-arabinopyranosyloxy)-3,4-seco-dammara-4(28)-en-3-oic acid(4,紫色点)等,所以化学成分明显不同。
本发明提供的总三萜制备工艺步骤简单易行,实用性强;大孔树脂和聚酰胺价格便宜,可再生和反复利用,成本低;并且提高了总三萜的含量,使三萜类成分含量在80%以上,总三萜的总提取率在2%以上。
本发明的另一个目的是提供所述的总三萜和化合物(4)在制备抗糖尿病药物、保健食品和功能食品中的应用。所述化合物(4)为(20S,24R)-20,24-epoxy-25-hydroxy-12β-(α-L-arabinopyranosyloxy)-3,4-seco-dammara-4(28)-en-3-oic acid。本发明选取了3T3-L1脂肪细胞对总三萜、阿江榄仁酸(1)、积雪草酸(2)、青钱柳酸B(3)和(20S,24R)-20,24-epoxy-25-hydroxy-12β-(α-L-arabinopyranosyloxy)-3,4-seco-dammara-4(28)-en-3-oic acid(4)的抗糖尿病活性进行了研究,结果显示总三萜及化合物(1)-(4)能有效增加3T3-L1脂肪细胞的葡萄糖摄取。其中总三萜(10μg/mL)的葡萄糖摄取百分率为124.27%,阿江榄仁酸(10μM)、积雪草酸(10μM)、青钱柳酸B(10μM)和(20S,24R)-20,24-epoxy-25-hydroxy-12β-(α-L-arabinopyranosyloxy)-3,4-seco-dammara-4(28)-en-3-oic acid(10μM)的葡萄糖摄取百分率为120.12%,122.02%,125.03%,145.89%,表明它们具有明显的抗糖尿病活性,其中化合物4的效果达到了极显著。
本发明开发了青钱柳叶中总三萜的制备方法及其抗糖尿病的用途,为推广利用和深加工中药材提供依据,并提供了新的应用方向。
以青钱柳叶为原料,经乙醇水混合溶剂提取,有机溶剂液液萃取后,通过聚酰胺柱色谱有机溶剂洗脱和大孔树脂柱色谱乙醇水洗脱进行分离纯化,获得高纯度的总三萜。该方法
经研究表明,该总三萜及其中含有的主要成分阿江榄仁酸、积雪草酸、青钱柳酸B、化合物4,能有效增加3T3-L1脂肪细胞的葡萄糖摄取,具有抗糖尿病活性,可将总三萜及其主要成分用于制备降血糖的抗糖尿病的药物、保健食品和功能食品等产品。本发明开发了一种青钱柳叶中总三萜的制备方法及其和化合物4在制备抗糖尿病药物中的应用,为推广利用和深加工中药材提供依据,并提供了新的应用方向。
附图说明
图1为人参皂苷Re的标准曲线。
图2为本发明所得产品的薄层色谱图(展开剂:二氯甲烷/甲醇=10/1)。
图3为本发明所得产品的HPLC色谱图;图中峰标:(1)阿江榄仁酸;(2)积雪草酸;(3)青钱柳酸B;(4)(20S,24R)-20,24-epoxy-25-hydroxy-12β-(α-L-arabinopyranosyloxy)-3,4-seco-dammara-4(28)-en-3-oic acid。
具体实施方式
本发明结合附图和具体实施例作进一步的说明。
实施例1
干燥的青钱柳叶5kg,打粉,用50L的80%乙醇溶液室温浸提两次,每次5天。合并提取液,减压浓缩至干得总浸膏750g。总浸膏用1L的水分散,先后分别用石油醚、三氯甲烷等体积萃取,得到三氯甲烷萃取部分280g。三氯甲烷萃取部分经60-80目聚酰胺色谱柱(二氯甲烷:甲醇=15:1)洗脱,收集洗脱部分,获得浸膏。然后用水溶解浸膏,吸附于AB-8大孔树脂柱上,先用30%乙醇洗去杂质,后用60%乙醇洗脱。收集60%乙醇洗脱液,减压浓缩至干,得到总三萜干燥粉末105g,总提取率为2.1%。测得总三萜含量为80.3%。
实施例2
干燥的青钱柳叶10kg,打粉,用80L的70%乙醇溶液回流提取两次,每次2h。合并提取液,减压浓缩至干得总浸膏1600g。总浸膏用2L的水分散,先后分别用石油醚、乙酸乙酯等体积萃取,得到乙酸乙酯萃取部分560g。乙酸乙酯萃取部分经100-200目聚酰胺色谱柱(石油醚:丙酮=1:1)洗脱,收集洗脱部分,获得浸膏。然后用水溶解浸膏,吸附于D101大孔树脂柱上,先用40%乙醇洗去杂质,后用70%乙醇洗脱。收集70%乙醇洗脱液,减压浓缩至干,得到总三萜干燥粉末280g,总提取率为2.8%。测得三萜含量为85.4%。
实施例3
干燥的青钱柳叶1kg,打粉,用10L的90%乙醇回流提取两次,每次100min。合并提取液,减压浓缩至干得总浸膏182g。总浸膏用500mL的水分散,先后分别用石油醚、乙酸乙酯:正丁醇(10:1)等体积萃取,得到乙酸乙酯正丁醇萃取部分65g。萃取部分经100-200目聚酰胺色谱柱(二氯甲烷:乙酸乙酯=2:1)洗脱,收集洗脱部分,获到总三萜提取物。然后用水溶解浸膏,吸附于DM130大孔树脂柱上,先用50%乙醇洗去杂质,后用90%乙醇洗脱。收集90%乙醇洗脱液,减压浓缩至干,得到总三萜干燥粉末25g,总提取率为2.5%。测得三萜含量为81.1%。
实施例4对比实验
对比实验采用的是以大孔树脂材料反相洗脱体系为主的总三萜制备方法。
干燥的青钱柳叶3kg,打粉,用30L的70%乙醇溶液回流提取两次,每次2h。合并提取液,减压浓缩至干得总浸膏658g。总浸膏用1L的水分散,先后分别用石油醚、乙酸乙酯等体积萃取,得到乙酸乙酯萃取部分250g。乙酸乙酯萃取部分加水溶解,过D101大孔树脂柱,先用40%乙醇洗去杂质,后用95%乙醇洗脱。收集95%乙醇洗脱液,减压浓缩至干,得到总三萜干燥粉末180g,总提取率为6.0%。测得三萜含量为47.8%。
实施例5抗糖尿病活性研究
总三萜、阿江榄仁酸(1)、积雪草酸(2)、青钱柳酸B(3)和(20S,24R)-20,24-epoxy-25-hydroxy-12β-(α-L-arabinopyranosyloxy)-3,4-seco-dammara-4(28)-en-3-oic acid(4)的抗糖尿病活性研究。
方法:3T3-L1前脂肪细胞用含10%胎牛血清和1%双抗的高糖型DMEM培养基于37℃、5%CO2细胞培养箱中孵育,2天后再转移至含10%胎牛血清和DMI(1μM地塞米松,0.5mM3-异丁基-1-甲基黄嘌呤和5μg/mL胰岛素)的DMEM培养基中诱导分化2天,隔天更换新鲜培养液,第8天采集分化好的脂肪细胞。
为评价3T3-L1脂肪细胞的受试物安全给药浓度,以每孔10000个细胞密度接种3T3-L1细胞到96孔板,37℃、5%CO2培养箱中培养24h,弃去培养液,加入用培养基配制一定浓度受试物干预24h,弃去培养液,每孔加入1mg/mL MTT的DMEM溶液,37℃孵育4h,弃去上清液,每孔加入100μL的DMSO,震荡10min,在酶标仪570nm波长处测定其吸光值,计算细胞活力,确定无细胞毒性的受试物浓度做为3T3-L1细胞葡萄糖摄取实验的浓度。细胞活力=(实验组吸光值/空白对照组吸光值)×100%
实验时,完全分化的3T3-L1脂肪细胞接种于96孔板,并设无细胞空白对照孔。待细胞生长至80-90%融合,弃去原培养基,用KRP缓冲液洗2遍,换上含0.2%BSA的KRP培养液,分组加药。为刺激葡萄糖消耗,用含0.1μM胰岛素的KRP缓冲液孵育30min。设DMSO对照组,RSV(白藜芦醇)对照组(终浓度为5μM)和不同受试组。作用24h后,更换培养液,用含100μM2-NBDG的KRP培养液培养30min,立即于荧光酶标仪475nm和550nm波长处测定细胞内2-NBDG的含量,每组设3个以上复孔。相同实验重复6次。
结果:阳性药RSV和受试物作用于3T3-L1脂肪细胞24h后,与溶剂对照组比较,5μMRSV的葡萄糖摄取百分率为152.82%(P<0.01);10μM化合物1、2、3和4以及总三萜10μg/mL的葡萄糖摄取百分率分别为120.12%(P<0.05),122.02%(P<0.05),125.03%(P<0.05),145.89%(P<0.01)以及124.27%(P<0.05),具体数据见表1。
实施例6总三萜片剂的制备
总三萜20.0g与淀粉500g混匀,加10%淀粉浆10g制成软材,加入硬脂酸镁0.1g,干淀粉8g混匀后压制成2000片,即得。每片含总三萜10mg。
实施例7总三萜滴丸剂的制备
精密称取1.0g的总三萜,加适量无水乙醇,微热溶解,加入到3.75g的PEG4000熔融液中,搅拌混合均匀,直至乙醇挥尽,静置于90℃水浴上保温30min。待气泡除尽,在保温的条件下,用1.6mm的注射器吸取熔融物,控制滴距在6~8cm范围内,冷却高度为15cm,滴入5℃冷凝液液体石蜡中待冷凝完全,倾去冷凝液,收集滴丸,沥净和用滤纸除去滴丸上的冷凝液,即得。每粒滴丸含总三萜10mg。
实施例8本发明所述的总三萜含量,由紫外可见分光光度法以人参皂苷Re为对照品进行测定,方法如下:
(1)标准曲线的确定(附图1)
精密称定人参皂苷Re对照品5.47mg,加甲醇定容成25mL制成总皂苷标准溶液。精密量取标准溶液0,0.2,0.4,0.5,0.6,0.8mL(相当于标准品0,40,80,100,120,160μg),分别加入50mL小烧杯中,50℃水浴蒸干。在已挥干的小烧杯中加入0.2mL的5%香草醛-冰醋酸溶液,转动小烧杯使残渣溶解,再加入0.8mL高氯酸,摇匀,保鲜膜封口,于60℃水浴加热15min,冷却后加入冰醋酸5.0mL,摇匀后在紫外-可见分光光度计560nm处测定吸光度。所得人参皂苷Re标准曲线如图1所示。
(2)总三萜的含量测定方法
取总三萜粉末适量,加入5%香草醛-冰醋酸溶液0.2mL、高氯酸0.8mL,60℃水浴15min,冷却后加冰醋酸5.0mL,摇匀,以相应的溶液为空白。在560nm处用紫外分光光度仪测定吸光度,计算得到总三萜含量。
实施例9本发明通过HPLC方法分析本发明所述总三萜中的主要成分,方法如下:
(1)供试品溶液的制备
精密称取10mg总三萜粉末,加甲醇溶解定容至10mL备用;
(2)色谱条件
色谱柱:XSelect TM HSS T3柱(250mm×4.6mm,5μm);流动相:乙腈(A)-0.1%磷酸水(B),梯度洗脱(0~20min,20%~50%A;20~30min,50%~55%A;30~42min,55%~60%A;42~60min,60%~85%A);流速:1.0mL/min;检测波长:210nm;柱温:30℃;进样量:10μl。通过化学成分分离鉴定,将4种三萜混合对照品和总三萜样品的HPLC色谱图对比,确定本发明所述总三萜中含有阿江榄仁酸(1)、积雪草酸(2)、青钱柳酸B(3)和(20S,24R)-20,24-epoxy-25-hydroxy-12β-(α-L-arabinopyranosyloxy)-3,4-seco-dammara-4(28)-en-3-oic acid(4)等三萜类成分,参见图3。
化合物(1):阿江榄仁酸arjunolic acid的结构如下:
化合物(2):积雪草酸asiatic acid的结构如下:
化合物(3):青钱柳酸B cyclocaric acid B的结构如下:
化合物(4):(20S,24R)-20,24-epoxy-25-hydroxy-12β-(α-L-arabinopyranosyloxy)-3,4-seco-dammara-4(28)-en-3-oic acid的结构如下:
化合物4为新化合物,其核磁共振波谱学数据如表2所示。
表2.化合物(4)(CD3OD)的1H(500MHz)和13C NMR(125MHz)NMR数据及信号归属
Claims (8)
1.一种从青钱柳叶中制备具有降血糖作用的总三萜的方法,其特征在于,通过以下步骤实现:将青钱柳植物的叶子按重量/体积比的药材/溶剂加入5-12倍量的乙醇水混合溶液,室温浸提或回流提取2-3次,合并提取液减压浓缩至无醇味,然后先用石油醚脱色,用中极性有机溶剂萃取,回收溶剂得到浸膏,所得浸膏直接用重量比1至2倍的聚酰胺材料吸附,再将3至10倍的聚酰胺材料装入柱色谱,经有机溶剂洗脱,浓缩获得浸膏,再用蒸馏水分散溶解浸膏,上大孔树脂柱,先用体积比为20%~50%乙醇水混合溶液洗去杂质,收集50%~95%乙醇水洗脱液,干燥得到高纯度的总三萜;其中提取用的乙醇水混合溶液的浓度中含50%~95%乙醇;萃取所用的中极性有机溶剂为三氯甲烷、乙酸乙酯、正丁醇或其中的两种的混合物;洗脱用的有机溶剂为石油醚、二氯甲烷、乙酸乙酯、丙酮、甲醇之中的两种按20:1至1:1的比例混合而成;去除杂质选用浓度体积比为40%的乙醇水溶液;收集洗脱液所用的乙醇水浓度选用70%~80%,大孔树脂柱选用DM130、AB-8、D101型号树脂材料。
2.根据权利要求1所述的一种从青钱柳叶中制备具有降血糖作用的总三萜的方法,其特征在于,所述提取方法为室温浸提或回流提取,提取2-3次。
3.根据权利要求1所述的一种从青钱柳叶中制备具有降血糖作用的总三萜的方法,其特征在于,所述的聚酰胺材料选用60~200目。
4.根据权利要求1所述的一种从青钱柳叶中制备具有降血糖作用的总三萜的方法,其特征在于,提取用的乙醇水混合溶液的浓度中含60%~80%乙醇。
5.根据权利要求3所述的一种从青钱柳叶中制备具有降血糖作用的总三萜的方法,其特征在于,所述的聚酰胺材料选用100~200目。
7.权利要求1方法制备的总三萜和权利要求6所述的化合物(4)在制备抗糖尿病药物中的应用。
8.权利要求1方法制备的总三萜和权利要求6所述的化合物(4)在制备辅助降血糖的保健品中的应用。
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