中药黄芪有效成份的研究 Ⅰ.多糖体的分离、性质及其生理活性

从内蒙黄芪(Astragalus mongholicus Bunge)根的水提液中分得三种多糖:黄芪多糖Ⅰ,Ⅱ及Ⅲ。经玻璃纤维纸电泳及Sephadex G-150凝胶过滤表明为均一体。黄芪多糖Ⅰ是杂多糖,由D-葡萄糖、D-半乳糖和L-阿拉伯糖组成。其克分子比为1.75:1.63:1此外还含痕迹量的五碳糖,经已知分子量的标准葡聚糖在Sephadex G-75凝胶层析所标定的曲线上求得其平均分子量约为36300,比旋度为正,[α]_D~(30.5)+101.5(H _2O)。黄芪多糖Ⅱ及Ⅲ均为D-葡聚糖,其平均分子量分别约为12300和34600,比旋度亦为正,分别为[α]_D~(30.5)+170.8(H_2O)和[α]_D~(26)+177.6(H_2O)。黄芪多糖Ⅱ及Ⅲ经过碘酸氧化及Smith降解结果,除了产生大量的赤藓醇外,还产生丙三醇,这个结果表明两者的组成除含少量α-(1→6)-D-葡萄糖缩合键外。主要是α-(1→4)-D-葡萄糖的缩合键。 APS是水提取液中所分得的均一多糖部位,主要由黄芪多糖Ⅰ及Ⅱ组成,在小鼠上具有广泛的各种负疫作用,能增加小鼠脾脏的重量及细胞数,增加小鼠脾脏对绵羊红细胞的免疫应答反应和促进腹腔巨噬细胞的吞噬功能。若将黄芪多糖Ⅰ及Ⅱ分别试验,初步结果表明多糖Ⅰ能增加脾脏重量及细胞数。但抑制脾细胞对绵羊红细胞的免疫应答反应;多糖Ⅱ作用与多糖Ⅰ相似,但较弱;而多糖Ⅲ则没有作用。     

气相色谱-质谱联用同时分析新型细菌多糖中的单糖和糖醛酸

对纯化的新型细菌多糖进行酸水解,用乙硫醇-三氟乙酸和醋酐-吡啶体系先后对酸水解物进行衍生,与之前报道不同的是糖醛酸得到有效衍生化。以木糖为内标,采用气相色谱-质谱联用(GC-MS)定量分析该多糖酸水解物中单糖和糖醛酸衍生物发现,该多糖的糖链由岩藻糖、葡萄糖、葡萄糖醛酸和半乳糖组成,其相对物质的量比为1.50:1.0:0.79:2.06;中性糖比例与糖醇乙酸酯化分析岩藻糖、葡萄糖和半乳糖的相对物质的量比(1.76:1.0:1.98)接近;糖醛酸咔唑法与该方法分析葡萄糖醛酸的含量分别为16.19%和14.85%。以上结果表明所建立的衍生化方法及GC-MS同时定量分析多糖酸水解物中单糖和糖醛酸的方法可行。此外还对葡萄糖醛酸的质谱裂解机理进行了阐述。     

当归多糖的水解特征及其水解产物分析

对当归多糖ASP3的水解特征及其水解产物的组成和红外光谱特征进行了研究。分别采用0.05、0.2和0.5mol/L三氟乙酸(trifluoroacetic acid,TFA)和内切-α-(1→4)-聚半乳糖醛酸酶对ASP3进行部分酸水解和酶水解,并研究其水解特征;结合GC、FT-IR等方法对其水解产物的组成和红外光谱特征进行分析。实验结果表明,ASP3是一种果胶多糖,主要由光滑区(半乳糖醛酸聚糖)和毛发区(富含中性糖侧链的鼠李半乳糖醛酸聚糖)两部分组成:GalA和Rha位于多糖分子的主链,由于Rha含量较低,大部分GalA相连形成半乳糖醛酸聚糖光滑区;Gal、Ara、Man及Glc位于多糖分子的支链,其中Gal以较高的聚合度(半乳聚糖)与主链相连,Ara以还原性末端或低聚寡糖的形式与主链相连或与半乳聚糖末端相连形成阿拉伯半乳聚糖。     

白骨壤酸性果胶多糖HAM-3-Ⅱb-Ⅱ的结构分析

以分离纯化得到的白骨壤酸性多糖HAM-3-Ⅱb-Ⅱ为研究对象, 采用高碘酸氧化-Smith降解、部分酸水解以及甲基化分析等技术对该多糖的结构进行分析. 结果表明, HAM-3-Ⅱb-Ⅱ为典型的鼠李半乳糖醛酸聚糖I 型酸性果胶类多糖, 主链骨架包括: 1,4-连接的α-D-GalpA构成的无分支的半乳糖醛酸聚糖(光滑区)和通过α-D-GalpA的O4位与1,2-和1,2,4-L-Rhap的O2交替相连构成的含有较多分支的鼠李半乳糖醛酸聚糖(毛发区). 由T-、1,6-、1,3,6-、1,4-、1,4,6-D-Galp和T-、1,2-、1,3-、1,5-、1,2,5-、1,3,5-Aaraf聚合成的AGⅠ型阿拉伯半乳聚糖、AGⅡ型阿拉伯半乳聚糖、半乳聚糖以及阿拉伯聚糖, 构成了HAM-3-Ⅱb-Ⅱ的侧链部分, 通过Rha残基的O4位与主链相连.     

海水小球藻中多糖的提取及其单糖组成的气相色谱-质谱分析

在温度为70℃、功率为600 W、提取时间为30 m in的条件下,海水小球藻中的多糖可以得到较好的提取。经过优化,当超声波作用时间为90 m in、超声波功率为300 W、水解液为1 mol/L乙酸时,小球藻多糖的超声水解效果最好。碱提、水提、酸提海水小球藻多糖中的单糖组成,经过GC-MS分析,它们的单糖组成各不相同。碱提粗多糖中所含单糖为鼠李糖、L-岩藻糖、D-阿拉伯糖、D( )-木糖、D-甘露糖、D-葡萄糖、D-半乳糖,各单糖百分含量分别为20.68%、0.32%、1.0%、2.20%、4.58%、50.28%、20.91%(W/W);水提海水小球藻粗多糖中含鼠李糖、核糖、岩藻糖、阿拉伯糖、木糖、甘露糖、葡萄糖、半乳糖8种单糖,它们百分比分别为2.32、2.36、0.24、0.89、0.56、3.44、78.21、11.99%(W/W);酸提粗多糖中所含单糖为鼠李糖、核糖、岩藻糖、阿拉伯糖、木糖、甘露糖、葡萄糖、半乳糖,各单糖百分含量分别为2.75、1.75、0.23、1.04、0.61、5.99、74.33、13.30%(W/W)。本实验所建立的微波辅助提取、超声水解多糖、GC-MS分析多糖中的单糖组成等方法,具有简便、快速、准确的特点。可用于海藻和其它植物中的多糖研究。     

铁皮石斛原球茎多糖DCPP3c-1的分离纯化及结构初步分析

铁皮石斛原球茎粗多糖(DCPP)经阴离子交换纤维素柱(DEAE-2)和凝胶柱(Sephadex G-200)依次层析,分离纯化得灰色多糖DCPP3c-1.其纯度经比旋光度法、柱层析、紫外扫描检测,组分和结构经薄层层析、高效液相色谱、红外光谱及高碘酸钠氧化等试验分析.结果显示DCPP3c-1为均一组分,相对分子质量为72.4 ku.由甘露糖、鼠李糖、半乳糖醛酸、葡萄糖、半乳糖和阿拉伯糖组成,其分子物质的量之比为1.120 4:1:1.046:23.354:3.828:1.046.分子中1→6残基占14%,1→2或1→4残基占40.7%,1→3键占45.3%.红外光谱显示其具有多糖特征吸收峰,并存在吡喃糖苷键.DCPP3c-1是首次从液体悬浮培养的原球茎中分离得到的新型酸性杂多糖组分.     

黄芪药渣中阿拉伯木聚糖(AX-Ⅰ-1)的提取纯化、结构分析及体外抗氧化作用

黄芪药渣经分级提取,再通过二乙氨乙基(DEAE)-纤维素52阴离子交换柱层析和Sephacryl S-400HR凝胶柱层析分离纯化,得到4个多糖组分AX-Ⅰ-1~AX-Ⅰ-4.对多糖AX-Ⅰ-1的理化性质和结构进行研究发现,AX-Ⅰ-1含有鼠李糖、阿拉伯糖、木糖、甘露糖、葡萄糖和半乳糖(摩尔比为0.006∶14.113∶8.284∶0.116∶0.468∶1),主链由阿拉伯糖和木糖通过β-(1→2),(1→3)和(1→4)苷键组成,支链由(1→4)βArap,(1→3)βGalp和(1→2)βMan组成,非还原末端由αRhap,βGclp和βGalp组成.对多糖AX-Ⅰ-1~AX-Ⅰ-4的抗氧化研究结果表明,这4个组分对羟基自由基、超氧阴离子自由基和1,1-二苯基-2-三硝基苯肼(DPPH)自由基均有一定的清除作用;当浓度为0.1 mg/m L时,多糖AX-Ⅰ-2~AX-Ⅰ-4对超氧阴离子的清除率是阳性对照维生素C(Vc)的2倍.     

巴戟天中一种多糖的分离与结构表征

以巴戟天的根为原料, 经热水浸提、Sevag法除蛋白、乙醇沉淀和DEAE-Sepharose CL-6B离子交换柱层析, 得到一种水溶性的巴戟天多糖(MOPI-3). 通过UV、IR、NMR、GC-MS、高碘酸氧化、Smith降解和甲基化等物理化学方法对MOPI-3的纯度、理化性质和组成结构进行表征. 结果表明, MOPI-3分子量为36061, 是一种由阿拉伯糖、半乳糖和葡萄糖组成的杂多糖, 以α-1,3-吡喃葡萄糖和α-1,4-吡喃半乳糖为主链, 平均每5个葡萄糖连接一个半乳糖, 每个重复单元具有一个支链, 支链由3个呋喃阿拉伯糖以α-1,3-键型组成, 连接在主链葡萄糖的6位碳上, MOPI-3含有乙酰基, 连接在主链半乳糖的2位碳原子上.     

红栓菌多糖的分级、表征、分子链构型参数和抗癌活性研究

本文研究了以水为溶剂,乙醇为沉淀剂,获得6个级分的红栓菌多糖,并运用动态膜渗透压法、VPO法、超速离心沉降法、小角激光光散射和高效液相色谱等,对其进行了表征。使用分光光度计,以浊点滴定法与修正的Elias公式确定红栓菌多糖溶液的θ组成为:V_(H_2O):V_(CH_3COCH_3)=89.82:10.18(体积比)。再在θ条件下建立红栓菌多糖的Mark-Houwink方程:[η]=5.01×10~(-4)M~(0.502)计算出它的无扰尺寸。由色谱法和董履和函数适应法解得红栓菌多糖的分子量分布。 通过抗癌药理活性实验,发现该多糖对小白鼠肉瘤S_(180)有较强的生理活性,能抑制肿瘤生长。同时还发现多糖Ⅱ活性高于多糖Ⅰ活性。     

金钗石斛免疫活性多糖DNP-W1B的结构特征

从金钗石斛茎中获得了1个均一多糖组分DNP-W1B, 采用化学与光谱学结合的手段对其一级结构进行了解析. 结果表明, DNP-W1B分子量为7.7×10⁵, 比旋度²⁰_D=+81.3°(c0.3, H₂O), 由葡萄糖、 阿拉伯糖和半乳糖3种单糖组成, 摩尔比为6.2:3.1:0.9. DNP-W1B主链由1,4-和1,6-连接的葡萄糖残基组成, 在主链1,6-连接葡萄糖残基的4位形成分支, 支链由末端半乳糖残基和阿拉伯糖残基组成. 初步的免疫活性实验结果表明, DNP-W1B在体外能促进免疫细胞的增殖.     

Determination of the compositions of polysaccharides from Chinese herbs by capillary zone electrophoresis with amperometric detection

In this paper, capillary zone electrophoresis with amperometric detection (CZE-AD) was applied to determine the compositions of hetero-polysaccharides from Chinese herbs, Angelica sinensis and flax by analyzing their hydrolyzed monosaccharides: fucose, galactose, glucose, arabinose, rhamnose and xylose. Under the selected optimum conditions, the six monosaccharides could be perfectly separated within 25 min and showed significant current responses at copper electrodes. The linear ranges of the six monosaccharides were all from 5.0 x 10(-6) to 2.0 x 10(-4) mol L(-1) and their detection limits were lower or near 1.0 x 10(-6) mol L(-1) (S/N = 3). Experiments showed that the Angelica sinensis polysaccharide was composed of fucose, galactose, glucose, arabinose, rhamnose and xylose (mole ratio 1.0:13.6:15.0:8.7:21.3:3.7), and the flax polysaccharide was composed of galactose, glucose and arabinose (mole ratio 1.0:4.98:1.1). The purity of these polysaccharides leached by the introduced leaching method was 98.3 and 97.6%, respectively. Analyzing polysaccharides by this method has some merits of speed, simple instrumentation and operation, high sensitivity and high reproducibility.     

Novel antiviral fucoidan from sporophyll of Undaria pinnatifida (Mekabu)

Structural characterization and antiviral activities of fucoidan from sporophyll of Undaria pinnatifida (Mekabu) was examined. The fucoidan was composed of fucose and galactose with an approximately ratio of 1.0:1.1. Degree of substitution of sulfate was 0.72 and its apparent molecular weight was 9,000. Methylation analyses showed that fucoidan had various sugar linkages, and revealed that the fucoidan might have complicated structure. This fucoidan showed potent antiviral activities against herpes simplex virus type 1 (HSV-1), HSV-2, and human cytomegalovirus.     

Purification of polysaccharide from artificially cultivated Anoectochilus roxburghii (wall.) Lindl. by high‐speed counter current chromatography and its antitumor activity

To establish a systematic method for the extraction, purification, characterization and antitumor activity study of polysaccharide from artificially cultivated Anoectochilus roxburghii (wall.) Lindl. (AC‐ARPS). High‐speed counter current chromatography with two‐phase aqueous systems was successfully applied to purify AC‐ARPS after one‐step separation. The purity of the AC‐ARPS obtained by phenol/sulfuric acid method was 95.01%. The chemical structures of AC‐ARPS were identified by a series of analytical methods including high‐performance liquid chromatography and liquid chromatography with mass spectrometry. High‐performance liquid chromatography and liquid chromatography with mass spectrometry indicated that AC‐ARPS was mainly composed of mannose, ribose, glucose, galactose and arabinose with a molar ratio of 1.00:8.47:47.30:1.17:1.19. AC‐ARPS is a homogeneous polysaccharide with a molecular weight of 25 681 Da. The antitumor effect of AC‐ARPS was evaluated on lung cancer A549, osteosarcoma 143B, rat adrenal pheochromocytoma PC 12, breast cancer MCF‐7, acute leukemia HL 60, chronic leukemia K562, colon cancer SW620, esophageal cancer OE 19, liver cancer HepG2, and neuroglioma U251 cells in vitro. AC‐ARPS showed the best inhibitory effect on OE 19 cells, and the IC₅₀ value was 5.67 ± 0.831 μmol/L. Fluorescence analysis and flow cytometry results showed that AC‐ARPS induced apoptosis and G2/M phase arrest in OE 19 cells.     

An Acidic Polysaccharide from Tribulus terrestris

An aqueous acidic polysaccharide, named rhamnogalacturonan (designated as TIP-D2) was isolated from Tribulus terrestris L by means of DEAE-cellulose chromatography and gel filtration. The molecular mass of TTP-D2 was estimated to be 26 KDa by gel filtration.TTP-D2 is composed of galacturonic acid, rhamnose, arabinose, galactose,fucose,mannose,xylose and glucose in a ratio of 71.4:13.5:5.6:4.9:3.1:1.9:1.9:1.0. The main chain structure of TTP-D2 was elucidated as an acidic hetero-polysaccaride with the connection of α-(1-4) galacturonic acid with α-（1-3) rhamnose by GC analysis of partially hydrolyzed products and determination of ^1H,^13C-NMR spectra.     

Characterization of polysaccharides purified from Lacquer tree seed cakes and its antioxidant activity

Three polysaccharides, LTPS-1, LTPS-21 and LTPS-31 were isolated and purified from the seed cakes of lacquer tree using DEAE-52 cellulose and Sephadex G-200 column chromatography. The total sugar contents of LTPS-1, LTPS-21 and LTPS-31 were 931.8, 958.2 and 895.1 g kg^?1, respectively. LTPS-1 (3.48 kDa) was mainly composed of rhamnose, arabinose, glucose and galactose in a ratio of 35.36:5.06:1:2. LTPS-21 (11.4 kDa) was mainly composed of rhamnose, arabinose, mannose and galactose in a ratio of 41.93:21.8:1.01:9.24. LTPS-31 (19.49 kDa) was mainly composed of rhamnose, arabinose and mannose in a ratio of 38.31:16.44:1.1. IR analysis suggested they contained lower sulphuric acids, the LTPS-21 and LTPS-31 belonged to β-type polysaccharide. Among the three polysaccharides, LTPS-21 exhibited the strongest reducing power, scavenging activity on ABTS and hydroxyl radicals. These findings suggested that polysaccharides from the seed cakes could be potentially developed as natural functional ingredients in the food and cosmetic industry.     

Development of SPE for recovery of polysaccharides and its application to the determination of monosaccharides composition of the polysaccharide sample of a lactobacillus KLB 58

A new SPE cartridge has been prepared in this study to purify polysaccharides of high molecular weights. A porous nonpolar styrene-divinylbenzene copolymer phase (Hamilton PRP-1) was used to make the new cartridge. The cartridge was conditioned with methanol, water, and ACN in sequence, and the sample dissolved in a small amount of water was loaded. Impurities of low molecular weights were removed first by elution of 80:20 or 90:10 v/v% ACN/water, and polysaccharides were quantitatively recovered by elution of 50:50 v/v% ACN/water or pure water. The recovery of pure dextran 10000 was 90-95%. The SPE method was applied to purification of the polysaccharide sample of KLB58, a new lactobacillus discovered in Korea. The purified KLB 58 sample (weight recovery after SPE purification; 60%) was hydrolyzed for analysis of composition of monosaccharides. The hydrolysate was found to be composed primarily of fructose, glucose, galactose, rhamnose, mannose with small amounts of fucose and ribose.     

The true Johari-Goldstein beta-relaxation of monosaccharides

Broadband isothermal dielectric relaxation measurements of anhydrous fructose, glucose, galactose, sorbose, and ribose were made at ambient pressure in their liquidus and glassy states. We found a new secondary relaxation in fructose and glucose that is slower than those seen before by others. This new secondary relaxation also appears in the dielectric spectra of galactose, sorbose, and ribose, and hence it is a general feature of the relaxation dynamics of the monosaccharides. Dielectric measurements at elevated pressure of fructose and ribose show that the new secondary relaxation shifts to lower frequencies with applied pressures, mimicking the behavior of the alpha-relaxation. In contrast, the faster secondary relaxation remains stationary on applying pressure. These results together with other inferences indicate that the slower secondary relaxation bears relations to the alpha-relaxation, and hence, it is the true Johari-Goldstein secondary relaxation of the monosaccharides.     

Structural Elucidation and Antitumor Activity of Polysaccharide AMP-1 from Atractyloids macrocephalae K.

A polysaccharide named as AMP‐1 was isolated from the root of Atractylodis macrocephalae Koidz, and purified by ethanol fractionation and gel filtration. The homogeneity of AMP‐1 was determined by HPLC and capillary electrophoresis that gave a single peak. AMP‐1 is composed of galactose and mannose in a molar ratio of 1.0:1.9. Its molecular weight is 3.8 × 10³. The structure of ghycan was elucidated by IR, ¹H NMR, ¹³C NMR and methylation analysis, and on the basis of the results was suggested that AMP‐1 contain a backbone of β. (1–2)‐D‐galactose residues with branches of single β‐D‐mannose residue being substituted at the O‐6. Bioactivity assay of AMP‐1 showed that it could inhibit growth of Sarcoma 180 and Lewis pulmonary carcinoma implanted in mice.     

Study of the separation and determination of monosaccharides in soluble coffee by capillary zone electrophoresis with electrochemical detection

A simple, fast and reliable method, based on capillary electrophoresis with electrochemical detection, for the separation and determination of six monosaccharides, namely glucose, galactose, arabinose, fructose, xylose and ribose, in soluble coffees was developed. A copper disk electrode was used as the working electrode. The optimum conditions for separation and detection were 50 mmol L-1 sodium hydroxide buffer (pH 12.7), separation voltage 5 kV and detection potential 0.65 V (vs. Ag/AgCl). The linear ranges were from 5.0 x 10(-3) to 0.5 mmol L-1 for all six sugars. All regression coefficients were > 0.99. The detection limits for all the sugars were 1.0 x 10(-3) mmol L-1. The RSD of the peak current was < 4.2% (n = 5). The proposed method was applied directly to the separation and determination of the six sugars without prior derivatization, and the assay results were satisfactory.