紫娟茶化学成分及其抗炎抗氧化活性研究

本文对紫娟茶的化学成分进行分离纯化及其抗炎、抗氧化活性进行考察。实验采用现代分离纯化方法,从紫娟茶中共分离得到12个化合物,根据波谱学数据进行结构鉴定为:山柰酚(1)、(+)-表儿茶素(2)、(-)-表儿茶素-3-O-没食子酸酯(3)、杨梅素(4)、表没食子儿茶素-3-O-(3″-O-甲基)没食子酸酯(5)、槲皮素(6)、没食子酸(7)、表没食子儿茶素没食子酸酯(8)、3-O-没食子酰基奎宁(9)、小木麻黄素(10)、1,4,6-三-O-没食子酰基-β-D-葡萄糖(11)、咖啡因(12),其中化合物9为首次从紫娟茶中分离得到。化合物11表现出良好的抗氧化活性,化合物1、3、4、6、8～11均具有显著的抗炎活性。     

高速逆流色谱制备玫瑰红景天中的三种酚性化合物

采用高速逆流色谱方法(HSCCC,High-speed Counter-current Chromatography)同时分离三种玫瑰红景天酚性化合物。玫瑰红景天提取物经聚酰胺吸附多酚后经硅胶柱分级得预分离样品,采用正己烷∶乙酸乙酯∶甲醇∶水(4∶5∶4∶5,v/v/v/v)组成的两相溶剂系统对预分离样品进行分离纯化,一次进样150 mg,一次色谱分离得到化合物1:68.5 mg、化合物2:8.5 mg、化合物3:45.5 mg,纯度都超过98%。通过ESI-MS、1H NMR对其结构进行鉴定化合物1为没食子酸(Gallic acid),化合物2为没食子酸甲酯(Methyl gallate),化合物3为山奈酚(Kaempferol)。结果表明利用HSCCC可以成功分离三种酚性化合物,分离效果好,产品纯度高。     

高速逆流色谱分离纯化雷公藤中的雷公藤红素

以雷公藤植物粗提物为原料,建立了高速逆流色谱分离纯化雷公藤红素的分离纯化方法。优化了两相溶剂体系的组成及配比。优化后的分离纯化溶剂体系为正己烷-乙酸乙酯-甲醇-水,其体积之比为2∶3∶3∶2(上相为固定相,下相为流动相),实验温度为室温,主机转速为800 rpm,正向洗脱,流动相流速为2.0 m L/min。目标产物的分离时间较短、产品纯度高(97.5%)、分离过程稳定。     

HSCCC-ELSD法分离纯化青葙子中的皂苷

连接有蒸发光散射检测器的高速逆流色谱仪首次成功的应用于制备和分离青葙子中的皂苷celosins A和B.二氯甲烷∶正丁醇∶甲醇∶水(4∶0.3∶3∶2)+0.5％冰醋酸作为洗脱溶剂系统.从半制备型HSCCC收集到的组分进行HPLC分析,可以得到:celosin A纯度为98.9％,celosin B的纯度为98.1％.这是高速逆流色谱仪首次被用于纯化青葙子中的皂苷,两个化合物的结构通过碳谱和质谱来确定.     

展毛野牡丹酚类化学成分的研究

为了明确展毛野牡丹的化学成分,该研究采用Diaion HP20SS、MCI gel、Sephadex LH-20柱层析和反相高效液相色谱等方法,对展毛野牡丹根和茎的醇提物分别进行了分离纯化。结果表明:从展毛野牡丹中分离得到11个化合物,它们的结构经波谱数据分析和鉴定。它们分别是4-羟基-3-甲氧基苯酚1-O-β-D-(6'-O-没食子酰)-吡喃葡萄糖苷(1)、3,4-二羟基苯乙醇4-O-β-D-(6'-O-没食子酰基)-吡喃葡萄糖苷(2)、龙胆酸5-O-β-D-(6'-O-没食子酰基)-吡喃葡萄糖苷(3)、2,4,6-三甲氧基苯酚1-O-β-D-(6'-O-没食子酰)-吡喃葡萄糖苷(4)、甲基6-O-没食子酰基-β-D-吡喃葡萄糖苷(5)、乙基6-O-没食子酰基-β-D-吡喃葡萄糖苷(6)、6'-O-没食子酰基黑樱苷(7)、没食子酸甲酯(8)、没食子酸乙酯(9)、2,6-二甲氧基对苯二酚4-O-β-D-吡喃葡萄糖苷(10)、2-甲氧基对苯二酚4-O-β-D-吡喃葡萄糖苷(11)。所有化合物均为首次从展毛野牡丹中分离得到,化合物2-7、10和11为首次从该属植物中分离得到。     

高速逆流色谱分离制备蛹虫草中虫草素和N6-(2-羟乙基)-腺苷

采用高速逆流色谱（HSCCC）技术从蛹虫草子实体粗提物中分离制备高纯度虫草素和N⁶-(2-羟乙基)-腺苷。利用高效液相色谱（HPLC）测定目标产物在溶剂体系中的分配系数,优化HSCCC分离虫草素和N⁶-(2-羟乙基)-腺苷的溶剂体系,确定了以乙酸乙酯-正丁醇-1.5%氨水（1:4:5,V/V/V）为HSCCC的两相溶剂体系,并运用此溶剂体系,上相为固定相,下相为流动相,主机转速850r/min,流动相流速为1.5mL/min,检测波长为254nm条件下进行分离制备,在250min内从200mg蛹虫草子实体粗提物中一步分离得到10.8mg纯度99%的虫草素和6.1mg 纯度98%的N⁶-(2-羟乙基)-腺苷。该方法简便、快速,为虫草素和N⁶-(2-羟乙基)-腺苷的大量制备建立了基础。     

高速逆流色谱分离雨生红球藻中虾青素的工艺优化

应用高速逆流色谱(HSCCC)进行了雨生红球藻中虾青素的分离制备工艺优化,结果最优条件为正己烷∶乙酸乙酯∶乙醇∶水(6. 5∶5∶6. 5∶3,v/v/v/v)作为两相溶剂系统,以下相为固定相,上相为流动相,转速850 r/min,流速3 mL/min,温度25℃,上样浓度10 mg/mL,上样量10 mL。进一步应用高效液相色谱、质谱并与标准品比对,对所得虾青素进行鉴定。本文的研究结果为应用HSCCC高效制备雨生红球藻虾青素提供了技术支持。     

鬼灯擎中丹宁类化合物的分离及结构的核磁共振分析（Ⅱ）

利用Toyopearl HW-40(C)凝胶反复柱层析,从鬼灯擎(Rodsersia aesculifolia Ba-tal.)根茎的70％Me_2CO提取物中分离得到六种丹宁类化合物,其中四种经化学及核磁共振分析分别确定其结构为3-0-没食子酰基-(-)-表儿茶素[3-0-galloyl-(-)-epicatechin]RA-11,1,3-0-没食子酰基-表儿茶素-(4β-8)-(3-0-没食子酰基)-表儿茶素[(3-0-galloyl)-epieatechin-(4β-8)-(3-0-Salloyl)-epicatechin]RA-13,2;1,2,4,6-四-0-没食子酰基-β-D-葡萄糖[1,2,4,6-tetra-0-galloyl-β-D-glucose]RA-14,3和没食子酸甲酯(methyl gallate)RA-16,4。RA-14为首次获得的结晶性没食子丹宁,这些成分均为首次从该植物中得到。     

溶剂系统极性在高速逆流色谱分离油橄榄叶多酚类化合物过程中的影响

为探讨目标化合物极性和溶剂系统极性在HSCCC分离过程中的相关性,本文以油橄榄叶中的2种主要有效成分-橄榄苦苷和木犀草素-7'-O-β-D-葡萄糖苷为目标化合物,系统考察正己烷-乙酸乙酯-甲醇-水系统[(1∶19∶1∶19,v/v)、(1∶9∶1∶9,v/v)、(1∶9∶2∶8,v/v)、(1∶9∶3∶7,v/v)、(1∶6∶1∶6,v/v)、(1∶5∶1∶5,v/v)]不同比例组成的两相溶剂系统对目标化合物分离的影响。结果表明:橄榄苦苷、木犀草素-7'-O-β-D-葡萄糖苷随着溶剂系统的极性的减小,其分离效果呈现先增强后减弱的趋势,在最佳的分离条件下,分离制备所得橄榄苦苷、木犀草素-7'-O-β-D-葡萄糖苷的纯度按照HPLC归一化法检测,分别为87.8%和77.5%。本实验结果为HSCCC溶剂系统快速筛选及提高HSCCC在复杂体系中分离制备目标化合物的效率提供技术支持。     

大花红景天多元酚类化学成分的研究

从大花红景天Rhodiola crenulata (HK.f.et thoms)H.Ohba的乙酸乙酯提取物中分离出12个多元酚类化合物。其中六个经光谱分析和化学方法鉴定为没食子酸(gallic acid,1),酪醇(p-tyrosol,2),6-氧-没食子酰基红景天甙(6-O-galloylsalidroside,3),1,2,3,4,6-五氧-没食子酰基-β-D-吡喃葡萄糖(1,2,3,4,6-penta-O-galloyl-β-D-gluco pyranoside,4),草质素-7-氧-α-L-吡喃鼠李糖甙(rhodionin,5),草质素-7-氧-(3-氧-β-D-吡喃葡萄糖基)-α-L-吡喃鼠李糖甙(rhodiosin,6),1和4具有抗毒肝素的活性,4还有抗脂质过氧化的活性,2,5,6具有提高机体抗逆境能力的活性。     

HPLC特征图谱结合含量测定的白芍及其炮制品的质量对比研究

建立白芍、炒白芍、酒白芍、硫熏白芍HPLC特征图谱,并结合多成分含量测定,为白芍、炒白芍、酒白芍和硫熏白芍的质量控制提供参考。采用Intersustain C₁₈(250 mm×4.6 mm,5μm)色谱柱,流动相为乙腈-0.1%醋酸水溶液,流速为每分钟1 mL,梯度洗脱,检测波长为230 nm,柱温为30℃,进样量为10μL。14批白芍、炒白芍、酒白芍和硫熏白芍的特征图谱,标定了6个共有峰,并均被指认,分别为没食子酸、儿茶素、芍药内酯苷、芍药苷、1,2,3,4,6-五没食子酰葡萄糖和苯甲酰芍药苷,而硫熏白芍标定7个共有峰,峰7为白芍硫熏后产生;且各色谱谱峰有较好的分离,但不同炮制品特征图谱存在一定差异;含量测定结果显示,白芍炒制、酒制及硫熏后,6种成分均有不同程度的变化;借助中药色谱指纹图谱相似度评价系统和SIMCA-P13.0软件对14批白芍、炒白芍、酒白芍和硫熏白芍进行相似度和正交偏最小二乘判别(OPLS-DA)分析,所建立的白芍和炮制品及硫熏品的质量评价方法稳定性、重复性好,可用于白芍、炒白芍、酒白芍和硫熏白芍的质量控制和评价。     

Application of high-speed counter-current chromatography for preparative separation of cyclic peptides from Vaccaria segetalis

Following an initial clean-up step on silica gel, high-speed counter-current chromatography (HSCCC) was used to separate cyclic peptides from an extract of the seeds of Vaccaria segetalis. The two-phase solvent system used for HSCCC separation was composed of petroleum ether-ethyl acetate-methanol-water at an optimized volume ratio of 0.5:3.5:1:5. From 190 mg of crude extract, 38.0 mg of segetalin B and 28.5 mg of segetalin A were obtained with purities of 98.1% and 95.6% as determined by HPLC, respectively. The chemical structures of the target compounds were confirmed by high resolution electrospray ionization time of flight MS (HRESI-TOF-MS) and (1)H NMR analyses.     

应用高速逆流色谱分离桑枝酚类成分

建立了高速逆流色谱(HsCCC)分离制备高纯度的桑枝酚类成分的新方法.分离条件如下:溶剂系统为正己烷-乙酸乙酯-甲醇冰(1∶1∶1∶2,v/v),上相为固定相,下相为流动相;流速2.0 mL/min;转速900rpm;进样量75 mg.收集得到三个高纯度化合物,经HPLC、MS、1H和13C NMR等分别鉴定为反式氧化白藜芦醇(25.2mg),反式白藜芦醇(7.4 mg)和桑辛素M(29.1 mg).高速逆流色谱可以高效分离桑枝成分,方法简便,技术可行,优于传统的柱色谱法.     

Preparative isolation and purification of theaflavins and catechins by high-speed countercurrent chromatography

High-speed countercurrent chromatography (HSCCC) has been applied for the separation of theaflavins and catechins. The HSCCC run was carried out with a two-phase solvent system composed of hexane-ethyl acetate-methanol-water-acetic acid (1:5:1:5:0.25, v/v) by eluting the lower aqueous phase at 2 ml/min at 700 rpm. The results indicated that pure theaflavin, theaflavins-3-gallate, theaflavins-3'-gallate and theaflavin-3,3'-digallate could be obtained from crude theaflavins sample and black tea. The structures of the isolated compounds were positively confirmed by (1)H NMR and (13)C NMR, MS analysis, HPLC data and TLC data. Meanwhile, catechins including epigallocatechin gallate, gallocatechin gallate, epicatechin gallate and epigallocatechin were isolated from the aqueous extract of green tea by using the same solvent system. This study developed a modified method combined with enrichment theaflavins method by using HSCCC for separation of four individual theaflavins, especially for better separation of theaflavins monogallates.     

高速逆流色谱法分离制备丹酚酸B

采用高速逆流色谱法分离纯化丹参水溶性成分丹酚酸类物质,制备丹酚酸B化学对照品。分离采用的溶剂系统为正己烷-乙酸乙酯-水-甲醇(1.5:5:5:1.5),上相做固定相,下相做流动相,流速为1.7 mL/min,仪器转速850 rpm,进样量80 mg,纯度用HPLC方法测定。结果表明:一次分离可制备63.4 mg丹酚酸B,其纯度为98.6%。该方法操作简单,可作为高纯度丹酚酸B化学对照品的制备分离方法。     

Preparative separation of helvolic acid from the endophytic fungus <Emphasis Type="Italic">Pichia guilliermondii</Emphasis> Ppf9 by high-speed counter-current chromatography

High-speed counter-current chromatography (HSCCC) was applied for preparative separation of helvolic acid from the crude extract of the endophytic fungus Pichia guilliermondii Ppf9, associated with the medicinal plant Paris polyphylla var. yunnanensis for the first time. The two-phase solvent system consisted of n-hexane-ethyl acetate-methanol-water (4.5:4.5:5.0:5.0, v/v) appending with phosphoric acid (0.2%, v/v) was employed. The revolution speed of the separation column, flow rate of the mobile phase and separation temperature of the apparatus were 800 rpm, 3 ml min(-1) and 25°C, respectively. About 6.8 mg of helvolic acid was successfully obtained from 450 mg of the crude extract by HSCCC within 4 h separation procedure, and its purity reached to 93.2% according to the HPLC analysis. The product was further characterized by MS, (1)H-NMR and (13)C-NMR spectra.     

Combinative application of pH-zone-refining and conventional high-speed counter-current chromatography for preparative separation of alkaloids from Stephania kwangsiensis

A method which involves the combination of pH-zone-refining counter-current chromatography (pH-zone-refining CCC) and conventional high-speed counter-current chromatography (HSCCC) was established for the preparative separation of alkaloids from the crude extracts of Stephania kwangsiensis. pH-zone-refining CCC was first performed with the solvent system composed of n-hexane-ethyl acetate-methanol-water (3:7:1:9, v/v), where triethylamine (10 mM) was added to the upper organic stationary phase as a retainer and hydrochloric acid (5 mM) to the aqueous mobile phase as an eluter. From 2.0 g of crude extract, 370 mg of sinoacutine and 600 mg of a mixture of three other alkaloids were obtained. Then, the mixture was further separated by conventional HSCCC with the solvent system composed of n-hexane-ethyl acetate-methanol-water (7:3:6:4, v/v), yielding 42 mg of (-)-crebanine, 50 mg of (-)-stephanine and 30 mg of l-romerine from 150 mg mixture of three other alkaloids, respectively. The purities of the four compounds were all over 98% as determined by HPLC, and the chemical structures of the four compounds were confirmed by positive ESI-MS and (1)H NMR data. Results of the present study successfully indicated that this method was efficient for the preparative separation of alkaloids from natural plants.     

Preparative separation of alkaloids from Nelumbo nucifera leaves by conventional and pH-zone-refining counter-current chromatography

Two modes of high-speed counter-current chromatography (HSCCC) were successfully applied to the separation of alkaloids from crude extract of Nelumbo nucifera leaves. The conventional HSCCC separations were performed with a two-phase solvent system composed of tetrachloromethane–CHCl₃–methanol–0.1 M HCl at a volume ratio of 1:3:3:2 (v/v/v/v), and 120 mg crude extract could be successfully separated. pH-Zone-refining CCC was performed with a two-phase solvent system composed of petroleum ether (60–90 °C)–ethyl acetate–methanol–water (5:5:2:8, v/v/v/v) where triethylamine (10 mM) was added to the upper organic stationary phase as a retainer and hydrochloric acid (5 mM) to the aqueous mobile phase as an eluent. From 4.0 g of the crude extract, 120 mg N-nornuciferine, 1020 mg nuciferine and 96 mg roemerine were obtained in a single run each with a purity of over 98% as determined by HPLC. The structures of the isolated compounds were identified by ESI-MS, ¹H NMR and ¹³C NMR.     

Application of microwave‐assisted extraction coupled with high‐speed counter‐current chromatography for separation and purification of Dehydrocavidine from Corydalis saxicola Bunting

Introduction – Dehydrocavidine is a major component of Corydalis saxicola Bunting with sedative, analgesic, anticonvulsive and antibacterial activities. Conventional methods have disadvantages in extracting, separating and purifying dehydrocavidine from C. saxicola. Hence, an efficient method should be established. Objective – To develop a suitable preparative method in order to isolate dehydrocavidine from a complex C. saxicola extract by preparative HSCCC. Methodology – The methanol extract of C. saxicola was prepared by optimised microwave‐assisted extraction (MAE). The analytical HSCCC was used for the exploration of suitable solvent systems and the preparative HSCCC was used for larger scale separation and purification. Dehydrocavidine was analysed by high‐performance liquid chromatography (HPLC) and further identified by ESI‐MS and 1H NMR. Results – The optimised MAE experimental conditions were as follows: extraction temperature, 60°C; ratio of liquid to solid, 20; extraction time, 15 min; and microwave power, 700 W. In less than 4 h, 42.1 mg of dehydrocavidine (98.9% purity) was obtained from 900 mg crude extract in a one‐step separation, using a two‐phase solvent system composed of chloroform–methanol–0.3 m hydrochloric acid (4 : 0.5 : 2, v/v/v). Conclusion – Microwave‐assisted extraction coupled with high‐speed counter‐current chromatography is a powerful tool for extraction, separation and purification of dehydrocavidine from C. saxicola. Copyright © 2009 John Wiley & Sons, Ltd.     

Application of analytical and preparative high-speed counter-current chromatography for the separation of Z-ligustilide from a crude extract of Angelica sinensis

Z-Ligustilide was separated and purified from the traditional Chinese medicinal plant Angelica sinensis by high-speed counter-current chromatography (HSCCC). Analytical HSCCC was first used for the systematic selection of the two-phase solvent system. Preparative HSCCC separation was performed with a two-phase solvent system composed of petroleum ether (60-90 degrees C)-ethanol-water at an optimum volume ratio of 10:17:10 (v/v). A total of 38 mg Z-ligustilide at 98.8% purity was obtained in one step from 200 mg crude extract as determined by HPLC analysis. The structure of the target compound was identified by electron impact ionisation mass spectrometry.