山慈菇水层部分抗菌活性筛选和分离纯化研究
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摘要
山慈菇为常用抗癌中药,主要分布于四川,云南,贵州等长江以南各省区。用于治疗瘰疬结核、咽喉肿痛、跌打损伤、蛇虫咬伤、黄疸及癫痫等症。虽然在临床上得到广泛的应用,但目前对山慈菇的化学成分及药理作用的研究报道并不多。因此,本课题针对山慈菇化学成分及其抗菌活性进行研究。
首先对山慈菇采用85%乙醇提取后,经系统溶剂萃取法进行极性划分。得到的水层部分经大孔树脂柱色谱、硅胶柱色谱、制备薄层色谱和重结晶等手段从中分离得到5个化合物,并根据化合物的性质,利用~1H-NMR、~(13)C-NMR、EI-MS等方法及相关文献报道对比鉴定了其中4个化合物的化学结构,分别为:酪氨酸(Ⅰ),邻苯二甲酸二(2-乙基)己酯(Ⅱ),十六烷酸(Ⅲ),蔗糖(Ⅴ)。其中,酪氨酸,邻苯二甲酸二(2-乙基)己酯,十六烷酸为首次从该植物中分离得到。
对山慈菇水层部分进行抑菌实验结果显示:山慈菇水层部分的25%和75%乙醇洗脱物对金黄色葡萄球菌、白色念珠菌具有良好的抑菌活性,而对大肠杆菌抑制效果稍弱。该3种供试菌对两种洗脱物的敏感性依次为:金黄色葡萄球菌>白色念珠菌>大肠杆菌。
从山慈菇中分离得到一个具有激活酪氨酸酶活性的化合物——酪氨酸,并初步探讨山慈菇及伪品激活酪氨酸酶活性的影响因素。结果表明山慈菇的激活作用最高,丽江山慈菇和金果榄作用不明显,白芨呈抑制作用;薄层检测显示山慈菇、丽江山慈菇和金果榄中含有酪氨酸。初步认为酪氨酸为山慈菇等激活酪氨酸酶活性作用的主要成分之一。
本论文通过对山慈菇的化学成分及活性研究,对山慈菇的深入研究与二次开发,提供了初步的理论和实验依据。
Cremastra appendiculata is general anti-cancer Taditional Chinese Medicine, and mainly grow in Sichuan,Yunnan,Guizhou province and the southern part of Yangtze River.Cremastra appendiculata are used to cure the crewels and concretion, swelling of throat,injury from fall,snake and insect bite,aurigo and epilepsy,et al. However,the compouds and pharmacologic action of Cremastra appendiculata (D.Don) Makino were reported poorly.Cremastra appendiculata,Pleione bulbocodioides(Franch.)Rolfe and P.yunnanensis Rolfe are the Pseudobulb of appendiculate Cremastra.So,we will study on the activity of inhibiting bacteria and the chemical constiuents about water-solubility components of Cremastra appendiculata.
Extracts were obtained with the methanol and ethanol from Cremastra appendiculata, and the polarity was separated by the means of systematic solvent extraction. The chemical constituents of the water layer were studied and 5 compounds were isolated with macroporous resin and silica gel columns,preparative TLCs,recrystallization etc.On the basis of ~1H-NMR,~(13)C-NMR,EI-MS data,physicochemical characters and reported in literature,4 chemical structures of them were elucidated as:Tyrosine (Ⅰ),Bis(2-ethylhexyl) phthalate(Ⅱ),Hexadecanoic acid(Ⅲ),sucrose(Ⅴ).They are all isolated firstly from water of Cremastra appendiculata.
The antibacterial activity of water-solubility components of Cremastra appendiculata of Staphylococcus aureus,Candida albicans and Escherichia coli was investigated. The results showed that 25%and 75%ethanol eluates had obvious inhibitory effect on the growth of Staphylococcus aureus and Candida albicans.however,the antibacterial activity on Escherichia coli was relatively smaller and the sensitive strain is Staphylococcus aureus>Candida albicans>Escherichia coli.
Tyrosine activity of tyrosinase which was isolated with Cremastra appendculata, and to study the effects of Cremastra appendculatai and spurious breed on activity of tyrosinase.The results showed that the Cremastra appendiculata significantly increased the activity of tyrosinase,but the Rhizoma Bletillae inhibited.The Cremastra appendiculata,Iphigenia indica Kunthet Benth and Tinospora capilipes Gagnep have tyrosine that was detected by TLCs.Tyrosine is one of the major components to effect the Cremast- ra appendiculata,Iphigenia indica Kunthet Benth and Tinospora capilipes Gagnep on activity of tyrosinase.
The chemical constituents,antibacterial activityand tyrosine activity of Cremasttra appendiculata was studied and provid theoretical and experimental bases for the more research and development of Cremastra appendiculata.
首先对山慈菇采用85%乙醇提取后,经系统溶剂萃取法进行极性划分。得到的水层部分经大孔树脂柱色谱、硅胶柱色谱、制备薄层色谱和重结晶等手段从中分离得到5个化合物,并根据化合物的性质,利用~1H-NMR、~(13)C-NMR、EI-MS等方法及相关文献报道对比鉴定了其中4个化合物的化学结构,分别为:酪氨酸(Ⅰ),邻苯二甲酸二(2-乙基)己酯(Ⅱ),十六烷酸(Ⅲ),蔗糖(Ⅴ)。其中,酪氨酸,邻苯二甲酸二(2-乙基)己酯,十六烷酸为首次从该植物中分离得到。
对山慈菇水层部分进行抑菌实验结果显示:山慈菇水层部分的25%和75%乙醇洗脱物对金黄色葡萄球菌、白色念珠菌具有良好的抑菌活性,而对大肠杆菌抑制效果稍弱。该3种供试菌对两种洗脱物的敏感性依次为:金黄色葡萄球菌>白色念珠菌>大肠杆菌。
从山慈菇中分离得到一个具有激活酪氨酸酶活性的化合物——酪氨酸,并初步探讨山慈菇及伪品激活酪氨酸酶活性的影响因素。结果表明山慈菇的激活作用最高,丽江山慈菇和金果榄作用不明显,白芨呈抑制作用;薄层检测显示山慈菇、丽江山慈菇和金果榄中含有酪氨酸。初步认为酪氨酸为山慈菇等激活酪氨酸酶活性作用的主要成分之一。
本论文通过对山慈菇的化学成分及活性研究,对山慈菇的深入研究与二次开发,提供了初步的理论和实验依据。
Cremastra appendiculata is general anti-cancer Taditional Chinese Medicine, and mainly grow in Sichuan,Yunnan,Guizhou province and the southern part of Yangtze River.Cremastra appendiculata are used to cure the crewels and concretion, swelling of throat,injury from fall,snake and insect bite,aurigo and epilepsy,et al. However,the compouds and pharmacologic action of Cremastra appendiculata (D.Don) Makino were reported poorly.Cremastra appendiculata,Pleione bulbocodioides(Franch.)Rolfe and P.yunnanensis Rolfe are the Pseudobulb of appendiculate Cremastra.So,we will study on the activity of inhibiting bacteria and the chemical constiuents about water-solubility components of Cremastra appendiculata.
Extracts were obtained with the methanol and ethanol from Cremastra appendiculata, and the polarity was separated by the means of systematic solvent extraction. The chemical constituents of the water layer were studied and 5 compounds were isolated with macroporous resin and silica gel columns,preparative TLCs,recrystallization etc.On the basis of ~1H-NMR,~(13)C-NMR,EI-MS data,physicochemical characters and reported in literature,4 chemical structures of them were elucidated as:Tyrosine (Ⅰ),Bis(2-ethylhexyl) phthalate(Ⅱ),Hexadecanoic acid(Ⅲ),sucrose(Ⅴ).They are all isolated firstly from water of Cremastra appendiculata.
The antibacterial activity of water-solubility components of Cremastra appendiculata of Staphylococcus aureus,Candida albicans and Escherichia coli was investigated. The results showed that 25%and 75%ethanol eluates had obvious inhibitory effect on the growth of Staphylococcus aureus and Candida albicans.however,the antibacterial activity on Escherichia coli was relatively smaller and the sensitive strain is Staphylococcus aureus>Candida albicans>Escherichia coli.
Tyrosine activity of tyrosinase which was isolated with Cremastra appendculata, and to study the effects of Cremastra appendculatai and spurious breed on activity of tyrosinase.The results showed that the Cremastra appendiculata significantly increased the activity of tyrosinase,but the Rhizoma Bletillae inhibited.The Cremastra appendiculata,Iphigenia indica Kunthet Benth and Tinospora capilipes Gagnep have tyrosine that was detected by TLCs.Tyrosine is one of the major components to effect the Cremast- ra appendiculata,Iphigenia indica Kunthet Benth and Tinospora capilipes Gagnep on activity of tyrosinase.
The chemical constituents,antibacterial activityand tyrosine activity of Cremasttra appendiculata was studied and provid theoretical and experimental bases for the more research and development of Cremastra appendiculata.
引文
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