CN100413869C - 白果内酯配位色谱分离纯化方法 - Google Patents
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- 239000003795 chemical substances by application Substances 0.000 claims abstract description 22
- 239000000284 extract Substances 0.000 claims abstract description 12
- JIAARYAFYJHUJI-UHFFFAOYSA-L zinc dichloride Chemical compound [Cl-].[Cl-].[Zn+2] JIAARYAFYJHUJI-UHFFFAOYSA-L 0.000 claims abstract description 10
- SQGYOTSLMSWVJD-UHFFFAOYSA-N silver(1+) nitrate Chemical compound [Ag+].[O-]N(=O)=O SQGYOTSLMSWVJD-UHFFFAOYSA-N 0.000 claims abstract description 6
- 235000005074 zinc chloride Nutrition 0.000 claims abstract description 5
- 239000011592 zinc chloride Substances 0.000 claims abstract description 5
- VMHLLURERBWHNL-UHFFFAOYSA-M Sodium acetate Chemical compound [Na+].CC([O-])=O VMHLLURERBWHNL-UHFFFAOYSA-M 0.000 claims abstract description 4
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- 229910001961 silver nitrate Inorganic materials 0.000 claims abstract description 3
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- 229910000365 copper sulfate Inorganic materials 0.000 claims description 4
- ARUVKPQLZAKDPS-UHFFFAOYSA-L copper(II) sulfate Chemical compound [Cu+2].[O-][S+2]([O-])([O-])[O-] ARUVKPQLZAKDPS-UHFFFAOYSA-L 0.000 claims description 4
- 229910052742 iron Inorganic materials 0.000 claims description 4
- CNFDGXZLMLFIJV-UHFFFAOYSA-L manganese(II) chloride tetrahydrate Chemical compound O.O.O.O.[Cl-].[Cl-].[Mn+2] CNFDGXZLMLFIJV-UHFFFAOYSA-L 0.000 claims description 4
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- 230000005526 G1 to G0 transition Effects 0.000 claims description 3
- UQGFMSUEHSUPRD-UHFFFAOYSA-N disodium;3,7-dioxido-2,4,6,8,9-pentaoxa-1,3,5,7-tetraborabicyclo[3.3.1]nonane Chemical compound [Na+].[Na+].O1B([O-])OB2OB([O-])OB1O2 UQGFMSUEHSUPRD-UHFFFAOYSA-N 0.000 claims description 3
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- 229920000858 Cyclodextrin Polymers 0.000 abstract description 2
- 235000010338 boric acid Nutrition 0.000 abstract description 2
- VTJUKNSKBAOEHE-UHFFFAOYSA-N calixarene Chemical compound COC(=O)COC1=C(CC=2C(=C(CC=3C(=C(C4)C=C(C=3)C(C)(C)C)OCC(=O)OC)C=C(C=2)C(C)(C)C)OCC(=O)OC)C=C(C(C)(C)C)C=C1CC1=C(OCC(=O)OC)C4=CC(C(C)(C)C)=C1 VTJUKNSKBAOEHE-UHFFFAOYSA-N 0.000 abstract description 2
- 150000002500 ions Chemical class 0.000 abstract description 2
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- HFHDHCJBZVLPGP-UHFFFAOYSA-N schardinger α-dextrin Chemical compound O1C(C(C2O)O)C(CO)OC2OC(C(C2O)O)C(CO)OC2OC(C(C2O)O)C(CO)OC2OC(C(O)C2O)C(CO)OC2OC(C(C2O)O)C(CO)OC2OC2C(O)C(O)C1OC2CO HFHDHCJBZVLPGP-UHFFFAOYSA-N 0.000 abstract description 2
- 229910021380 Manganese Chloride Inorganic materials 0.000 abstract 1
- GLFNIEUTAYBVOC-UHFFFAOYSA-L Manganese chloride Chemical compound Cl[Mn]Cl GLFNIEUTAYBVOC-UHFFFAOYSA-L 0.000 abstract 1
- PMZURENOXWZQFD-UHFFFAOYSA-L Sodium Sulfate Chemical compound [Na+].[Na+].[O-]S([O-])(=O)=O PMZURENOXWZQFD-UHFFFAOYSA-L 0.000 abstract 1
- 239000012530 fluid Substances 0.000 abstract 1
- FBAFATDZDUQKNH-UHFFFAOYSA-M iron chloride Chemical compound [Cl-].[Fe] FBAFATDZDUQKNH-UHFFFAOYSA-M 0.000 abstract 1
- 235000002867 manganese chloride Nutrition 0.000 abstract 1
- 239000011565 manganese chloride Substances 0.000 abstract 1
- 229940099607 manganese chloride Drugs 0.000 abstract 1
- 238000004519 manufacturing process Methods 0.000 abstract 1
- 229910052938 sodium sulfate Inorganic materials 0.000 abstract 1
- 235000011152 sodium sulphate Nutrition 0.000 abstract 1
- BSVBQGMMJUBVOD-UHFFFAOYSA-N trisodium borate Chemical compound [Na+].[Na+].[Na+].[O-]B([O-])[O-] BSVBQGMMJUBVOD-UHFFFAOYSA-N 0.000 abstract 1
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 description 9
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- 235000008100 Ginkgo biloba Nutrition 0.000 description 3
- 238000006243 chemical reaction Methods 0.000 description 3
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- TWNQGVIAIRXVLR-UHFFFAOYSA-N oxo(oxoalumanyloxy)alumane Chemical compound O=[Al]O[Al]=O TWNQGVIAIRXVLR-UHFFFAOYSA-N 0.000 description 3
- 150000008442 polyphenolic compounds Chemical class 0.000 description 3
- HIXDQWDOVZUNNA-UHFFFAOYSA-N 2-(3,4-dimethoxyphenyl)-5-hydroxy-7-methoxychromen-4-one Chemical compound C=1C(OC)=CC(O)=C(C(C=2)=O)C=1OC=2C1=CC=C(OC)C(OC)=C1 HIXDQWDOVZUNNA-UHFFFAOYSA-N 0.000 description 2
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- IDGUHHHQCWSQLU-UHFFFAOYSA-N ethanol;hydrate Chemical compound O.CCO IDGUHHHQCWSQLU-UHFFFAOYSA-N 0.000 description 2
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Abstract
一种白果内酯配位色谱分离纯化方法,包括白果内酯初提物在流动相作用下流经固定相以及流份收集、浓缩和干燥,其特征在于在含白果内酯初提物中或柱填料添加配位剂。所述的配位剂为能形成配离子的无机物或者包合物等,如硝酸银、硼酸、硼钠、醋酸钠、硫酸钠、氯化锌、氯化铁、氯化锰、杯芳烃、环糊精等。本方法得到的白果内酯纯度95%以上,回收率超过70%,而且工艺简单,经济适用,对环境友好。
Description
一、技术领域
本发明涉及一种植物活性成分的分离纯化方法,确切地说是白果内酯配位色谱分离纯化方法。
二、背景技术
白果内酯是银杏叶药用活性成分的重要组份之一。也是保护脑细胞、神经元细胞免遭缺血、缺氧性损伤作用的关键成分之一。实现白果内酯的高效分离纯化具有现实意义。
银杏叶粗提物(含白果内酯15%左右,又称白果内酯初提物)中白果内酯通常与其他银杏内酯、银杏黄酮以及糖类、胶体等混为一体呈粘膏状,分离、纯化白果内酯极为困难。
色谱是一种广为使用的现代分离技术。但传统的色谱柱用于分离纯化白果内酯或达不到预期的分离纯化效果,或分离柱成本过高(如使用反相色谱填料)。
三、发明内容
色谱分离又称柱层析。其包括目标物和杂质的原料在流动相(洗脱液)的作用下流经固定相(色谱填料)以及流份收集、浓缩和干燥。本发明所称的配位色谱分离就是在上述色谱分离介质中引入配位剂,利用配位结合的高选择性,即配位剂与白果内酯或其他杂质的非共价键合形成配合物,扩大白果内酯与其他杂质的物性差异,这种差异可以是空间结构的、可以是溶解性的、可以是电特性的、也可以是与分离介质间的结合力的,如此等等,最终表现为扩大了白果内酯与杂质之间在色谱分离特性上的差异。从而提高色谱分离的效果和产品的纯度。
所称的引入配位剂有以下两种方法。
一是柱前配位衍生化法。就是将配位剂与白果内酯初提物(目标物)的乙醇溶液混合均匀发生配位反应,经真空脱溶至干上色谱柱。初提物中含多酚类化合物和有机酸,先用弱碱性的盐,如醋酸钠、硼酸钠等中和至弱酸性或中性,然后再加入配位剂进行配位反应。
二是固定相添加法。就是将配位剂与柱填料混合均匀后再装柱,形成配位色谱柱。配位剂含量2~15%。当银杏叶提取物在流动相作用下通过该配位色谱柱时,在配位剂的作用下,白果内酯与其他产生较大的选择性差异,从而提高分离效率。所述的柱填料可以硅胶、氧化铝、葡聚糖凝胶、聚酰亚胺等常规色谱填料。
本发明所称的配位剂可以是能形成配离子的无机物或者包合物或者是其混合配位剂。
所述的无机物如硝酸银、硼酸、硼酸钠、醋酸钠、硫酸铜、氯化锌、氯化铁、氯化锰等中的一种或两种以上混合配位剂。
所述的包合物如杯芳烃、环糊精等中的一种或两种以上混合配位剂或者其与无机物的混合配位剂。
本方法得到的白果内酯纯度95%以上,回收率超过70%,比普通色谱法分别提高20%和15%以上。而且工艺简单,经济适用,对环境友好。
四、具体实施方式
1、将15%白果内酯初提物用70%乙醇水溶解成溶液,依其多酚类化合物和有机酸的含量,加入NaAc的饱和水溶液,混合均匀使溶液成中性,反应完全后按摩尔比1∶1~3加入H3BO3的饱和水溶液,室温放置过夜;将过夜溶液真空减压浓缩至干,上硅胶柱,采用石油醚——乙酸乙酯的混合溶液梯度洗脱,混合溶剂中石油醚从100%变化到0%的同时,乙酸乙酯从0%变化到100%。分段收集含较纯白果内酯的流份,浓缩、结晶、干燥,可得95%以上纯度白果内酯,分离效率提高15%。
2、将15%白果内酯初提物用70%乙醇水溶解成溶液,依其多酚类化合物和有机酸的含量,加入Na2B4O7的饱和水溶液,混合均匀使溶液呈中性,反应结束后按摩尔比1∶1~3加入H3BO3饱和水溶液,室温放置过夜;将过夜溶液真空减压浓缩至干,上硅胶柱,采用石油醚——甲酸乙酯的混合溶液梯度洗脱,混合溶剂中石油醚从100%变化到0%的同时,甲酸乙酯从0%变化到100%。分段收集含较纯白果内酯的流份,浓缩、结晶、干燥,可得95%以上纯度白果内酯,分离效率提高15%。
3、将200~400目硅胶与氯化锌混合,氯化锌含量2%,制成配位色谱柱,用石油醚-甲酸乙酯作流动相,梯度洗脱,分离白果内酯,浓缩、结晶、干燥,得到95%以上纯度白果内酯,分离效率提高10%。
4、将200~400目硅胶与氯化铁混合,氯化铁含量5%,制成配位色谱柱,用石油醚-乙酸乙酯作流动相,梯度洗脱,分离白果内酯,浓缩、结晶、干燥,得到95%以上纯度白果内酯,分离效率提高15%。
5、将氧化铝与氯化锰混合,氯化锰含量15%,制成配位色谱柱,用石油醚——甲酸乙酯流动相,梯度洗脱,分离白果内酯,浓缩、结晶、干燥,得到95%以上纯度白果内酯,分离效率提高10%。
6、将氧化铝与硫酸铜混合均匀,使硫酸铜含量10%,制成配位色谱柱,用石油醚——乙酸乙酯梯度洗脱,分离白果内酯,浓缩、结晶、干燥,得到95%以上纯度白果内酯,分离效率提高15%。
Claims (1)
1. 一种白果内酯配位色谱分离纯化方法,包括白果内酯初提取物在流动相作用下流经固定相以及流份收集、浓缩和干燥,其特征在于:在白果内酯初提取物中添加配位剂,添加配位剂的方法是先用醋酸钠或硼酸钠中和白果内酯初提物乙醇溶液至中性,再按摩尔比1∶1~3加入硼酸饱和水溶液,室温静置过夜,脱溶浓缩至干;或者在柱填料中添加配位剂,添加配位剂的方法是在常规色谱填料中添加2~15%的配位剂,所述的配位剂为硝酸银、硫酸铜、氯化锌、氯化铁、氯化锰中的一种或两种以上混合配位剂。
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Citations (3)
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US5389370A (en) * | 1989-12-04 | 1995-02-14 | Montana Limited | Active component concentrates and new active component combinations from ginkgo biloba leaves, their method of preparation and pharmaceuticals containing the active component concentrates or the active component combinations |
CN1287121A (zh) * | 2000-06-02 | 2001-03-14 | 天津大学 | 由银杏叶或银杏叶浸膏制备药物银杏内酯a、b的方法 |
CN1374311A (zh) * | 2002-03-26 | 2002-10-16 | 贵州省生化工程中心 | 采用柱层析法提取银杏萜内脂的工艺 |
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Patent Citations (3)
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---|---|---|---|---|
US5389370A (en) * | 1989-12-04 | 1995-02-14 | Montana Limited | Active component concentrates and new active component combinations from ginkgo biloba leaves, their method of preparation and pharmaceuticals containing the active component concentrates or the active component combinations |
CN1287121A (zh) * | 2000-06-02 | 2001-03-14 | 天津大学 | 由银杏叶或银杏叶浸膏制备药物银杏内酯a、b的方法 |
CN1374311A (zh) * | 2002-03-26 | 2002-10-16 | 贵州省生化工程中心 | 采用柱层析法提取银杏萜内脂的工艺 |
Non-Patent Citations (2)
Title |
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银杏萜内酯提纯新工艺研究. 何照范等人.西南农业大学学报,第21卷第6期. 1999 |
银杏萜内酯提纯新工艺研究. 何照范等人.西南农业大学学报,第21卷第6期. 1999 * |
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