CN1203083C - 知母提取物及其制备方法 - Google Patents
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Abstract
本发明涉及医药技术领域。具体涉及知母提取物及其制备方法。将知母药材采用含水乙醇热提或冷浸，采用大孔吸附树脂分离，精制知母提取物。知母提取物的主要活性成分是芒果苷及新芒果苷，二者总含量高达50%以上，芒果苷与新芒果苷的含量比为1∶1～5∶1。经动物试验，所得的知母提取物能有效防治糖尿病。
Description
技术领域：
本发明涉及医药技术领域，具体而言，涉及一种知母提取物及其制备方法。
背景技术：
知母为百合科植物知母Anemarrhena asphodeloides Bunge的根茎，具有滋阴降火、润燥滑肠的功效，临床多用于消渴证。目前知母药用时主要为单味知母饮片或与其他中药配伍煎汤后服用。
知母的化学成分主要为皂苷、黄酮类成分，其中知母的黄酮类成分主要为芒果苷(mangiferin)、新芒果苷(neomangiferin)。有报道，自发性糖尿病高血清胰岛素动物模型KK-Ay小鼠(II型糖尿病的一种有效的动物模型)口服芒果苷或新芒果苷3周后降糖作用明显，能降低胰岛素抵抗，而对正常小鼠无明显作用(1、Planta Med，1997，63(4)：320-2；2、Biol Pharm Bull，1998，21(12)：1389-90；3、Phytomedicine，2001，8(2)：85-7)。
发明内容：
本发明提供一种从知母中提取得到的知母提取物及其制备方法，经动物试验，口服制得的知母提取物，可显著降低高血糖模型动物的血糖值。因此，本发明所制备的知母提取物可用于制备防治糖尿病。
本发明知母提取物制备方法为：
将知母粉碎后，用含水乙醇热提或冷浸，提取液浓缩后离心或过滤，取上清液或过滤液进行大孔树脂吸附，先用水淋洗除去杂质，再用含水低级醇洗脱，洗脱液浓缩至一定体积，静置，析出沉淀，过滤，将沉淀干燥粉碎得提取物1；再在过滤液中加入乙醇醇沉，弃沉淀，再将上清液浓缩至干后干燥粉碎，或将上清液浓缩后喷雾干燥，得提取物2；合并提取物1与提取物2并混合均匀，即为本发明的知母提取物。
获得的知母提取物成分分析：经硅胶、sephadexLH-20、ODS等反相色谱担体柱色谱获得了2个化合物，即芒果苷和新芒果苷，用化学、波谱法测定结构，化学结构通式如下：
其中：芒果苷的R基团为H
新芒果苷的R基团为Glu(葡萄糖)
所以，知母提取物主要是芒果苷与新芒果苷的混合物，用HPLC定量分析表明，芒果苷与新芒果苷的含量总和占知母提取物的50％以上，且芒果苷与新芒果苷的含量比为1∶1～5∶1。
生物活性测定：
制作糖尿病模型小鼠：ICR种小鼠，雌雄兼用，体重20g左右，先测空腹血糖值，选用血糖值正常者，然后一次性腹腔注射链佐菌素柠檬酸溶液260mg/kg，之后按胰岛素维持量，每天皮下注射精蛋白锌胰岛素注射液0.2U/只，连续注射21天。末次注射精蛋白锌胰岛素后7小时禁食12小时，再次测定空腹血糖值，取血糖值16.0mmol/L以上者作为糖尿病模型小鼠。
选75只糖尿病模型小鼠，按血糖值分为5组：溶剂对照组、阳性药(吡格列酮30mg/kg)对照组以及本发明知母提取物小、中、大剂量组(20，60，180mg/kg)。每组各15只。
每天固定时间，灌胃给药，同时每只小鼠继续皮下注射精蛋白锌胰岛素(0.2U/只)，每日1次，连续7天，末次给药前禁食10小时，给药后2小时(继续禁食)测定血糖，比较各组给药前后血糖降值(见表1)。继续灌胃给药7天，但各组均停止注射精蛋白锌胰岛素，同法测定血糖，比较各组血糖降值(见表2)。
表1 知母提取物并同时注射小量胰岛素对糖尿病模型小鼠血糖影响(x±s)
药物剂量
血糖(mmol/L) 血糖降值
组别
(mg/kg) 给药前 给药后 (mmol/L)
模型对照组 - 20.45±2.75 19.01±2.96 1.44±2.57
阳性药物对照组 30 20.33±2.55 6.68±1.51** 13.65±3.16**
知母提取物组 20 20.26±2.76 16.53±2.76* 3.73±2.40*
知母提取物组 60 20.44±2.62 10.23±1.78** 10.21±2.76**
知母提取物组 180 20.64±2.79 7.85±2.66** 12.79±2.31**
P＜0.05，**P＜0.01与模型对照比较
表2 知母提取物对不同时注射小量胰岛素的糖尿病模型小鼠血糖影响(x±s)
剂量
血糖(mmol/L) 血糖降值
组别
(mg/kg) 给药前 给药后 (mmol/L)
模型对照组 - 20.45±2.75 17.37±2.77 3.08±3.94
阳性药物对照组 30 20.33±2.55 16.99±2.35 3.34±2.62
知母提取物组 20 20.26±2.76 17.56±2.63 2.70±2.47
知母提取物组 60 20.44±2.62 17.81±2.74 2.63±3.16
知母提取物组 180 20.64±2.79 15.75±3.67 4.89±3.87
从表1可看出，联合使用小剂量胰岛素，知母提取物3个剂量均能够显著降低模型小鼠的血糖水平。从表2可看出，不使用小剂量胰岛素，知母提取物3个剂量对模型小鼠的血糖水平均无显著影响。结果表明，本发明所获得的知母提取物具有确切的降血糖作用，其作用机制可能与提高胰岛素敏感指数有关。故本发明所获得的知母提取物可用于制备防治糖尿病药物。
具体实施方式：
将粉碎的知母药材用20～95％乙醇热提，首次溶媒用量约为生药量的4～8倍；或将粉碎的知母药材用20～95％乙醇渗漉提取，溶媒用量约为生药量的4～12倍；提取液浓缩，浓缩液离心或过滤；浓缩液的离心上清液或过滤液通过大孔树脂吸附(大孔树脂为以苯乙烯为架桥材料的聚苯乙烯型多孔性吸附树脂，如D101、D201、ZTC-1、AB-8、1300-1型大孔树脂等)，用水淋洗除去杂质；再用含水低级醇洗脱，低级醇为甲醇、乙醇或丙醇等C1～C5醇类，其浓度为15～25％，用量为生药量的5～15倍；收集低级醇洗脱液，浓缩至无醇，静置后有沉淀析出，离心或过滤，得沉淀，为提取物1；其离心上清液或过滤液再用乙醇醇沉，乙醇终浓度为70～95％，弃沉淀，上清液或浓缩至干，干燥粉碎得提取物2，或浓缩后喷雾干燥，也可得提取物2；合并提取物1与提取物2并混合均匀，即为本发明知母提取物，成品收率为生药量的1.0％以上。
下面给出最佳实施例进一步说明本发明但并不对本发明限制。
实施例1
100公斤粉碎后的知母药材，投入提取罐，在90～100℃依次用70％乙醇400升热提3次，每次提取时间为90分钟，合并3次提取液，浓缩至80升，过滤，将滤液用蒸馏水调节至100升。再缓缓通过装有350升ZTC-1型大孔树脂(天津正天成澄清技术有限公司生产)的吸附柱，用500升蒸馏水淋洗除去杂质；再用1000升20％乙醇洗脱，收集洗脱液，浓缩至10升，静置24小时，沉淀析出，过滤，干燥粉碎后得浅黄色沉淀0.4公斤，为提取物1；再将滤液在搅拌下缓缓加入100升95％乙醇，静置12小时，离心，并沉淀上清液浓缩至干，干燥粉碎后得黄色粉末1.6公斤，为提取物2；合并提取物1与提取物2并混合均匀，即为本发明知母提取物，共重2.0公斤，按生药量计得率为2.0％。
Claims (5)
1.一种治疗糖尿病的知母提取物，其特征在于知母提取物中芒果苷和新芒果苷的含量总和大于50％，它是通过如下方法制备而成的：
(1)提取
知母粉碎后，用20-95％含水乙醇热提或冷浸得提取液，提取液浓缩后离心或过滤，弃沉淀，上清液或过滤液备用；
(2)吸附洗脱
将上述上清液或过滤液进行聚苯乙烯型多孔性吸附树脂吸附、水淋洗除去杂质后，用上样生药质量的5～15倍量体积的15～25％含水的C1～C5低级醇洗脱；洗脱液浓缩至无醇，静置，析出沉淀，离心或过滤，得沉淀提取物1，上清液或过滤液备用；
(3)纯化
在上清液或过滤液中加入乙醇醇沉，乙醇终浓度为70-95％，弃沉淀，将醇沉后的上清液浓缩至干，得固体物为提取物2；或将上清液浓缩后，喷雾干燥，也可得提取物2；
(4)成品制备
合并提取物1与提取物2并混合均匀，即得知母提取物，该提取物中含芒果苷和新芒果苷。
2.根据权利要求1所述的知母提取物，其特征在于：步骤(1)中可以用20-95％含水乙醇渗漉提取得到提取液。
3.根据权利要求1所述的知母提取物，其特征在于：步骤(1)中的乙醇体积用量为生药质量的4-8倍。
4.根据权利要求2所述的知母提取物，其特征在于：乙醇的体积用量为生药质量的4-12倍。
5.根据权利要求1～4任意一项所述的知母提取物，其特征在于：芒果苷与新芒果苷的含量比为1∶1～5∶1。
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Families Citing this family (15)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN100366588C (zh) * | 2003-09-10 | 2008-02-06 | 广州中一药业有限公司 | 知母总酚的提取物及其制备方法和用途 |
| CN1326459C (zh) * | 2005-01-31 | 2007-07-18 | 王喜军 | 知母制剂作为防治植物龙胆斑枯病的新用途 |
| CN100335493C (zh) * | 2005-05-24 | 2007-09-05 | 浙江大学 | 从中药材中提取菝葜皂甙元的方法 |
| CN100336822C (zh) * | 2005-09-01 | 2007-09-12 | 桂林莱茵生物科技股份有限公司 | 一种提取芒果甙的方法 |
| CN101062249B (zh) * | 2007-05-22 | 2010-05-19 | 中国人民解放军第二军医大学 | 知母提取物与黄柏提取物的组合物及其用途 |
| AT505539B1 (de) * | 2007-09-03 | 2009-02-15 | Peter Dr Laszloffy | Extraktionsverfahren für die herstellung eines kräuterextraktes sowie eine kosmetische pflegesalbe |
| CN101229316B (zh) * | 2008-01-31 | 2010-08-25 | 广东药学院 | 一种知母提取物 |
| CN103288898B (zh) * | 2013-06-06 | 2016-05-04 | 广州莱泰制药有限公司 | 新芒果苷对照品提取纯化分离方法 |
| CN103349724A (zh) * | 2013-08-02 | 2013-10-16 | 石任兵 | 一种知母总黄酮提取物及其同时制备知母总黄酮提取物及总皂苷提取物的方法 |
| CN103479856A (zh) * | 2013-09-22 | 2014-01-01 | 山东省医药工业研究所 | 知母化学成分群提取物及其制备工艺 |
| CN103908567B (zh) * | 2014-03-14 | 2017-04-05 | 中国人民解放军第二军医大学 | 一种用于治疗糖尿病痛性神经病变的复方制剂及其应用 |
| CN105031178A (zh) * | 2015-08-03 | 2015-11-11 | 南京中医药大学 | 一种高效利用知母的提取精制方法 |
| CN105582104A (zh) * | 2015-12-30 | 2016-05-18 | 中国中医科学院中医临床基础医学研究所 | 一种治疗类风湿关节炎的中药组合物 |
| CN105974014A (zh) * | 2016-05-04 | 2016-09-28 | 广州市香雪制药股份有限公司 | 知母的一测多评检测方法 |
| CN111166823A (zh) * | 2020-03-17 | 2020-05-19 | 皖南医学院 | 一种知母总提取物用于制备抗肺纤维化的药物的医药用途 |
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