金顶侧耳多糖PC-4的结构确定与抗肿瘤活性的研究

从3％三氯乙酸浸提过的金顶侧耳(Pleurotus citrinopileatus)子实体中分离纯化另一水溶性多糖PC-4。该多糖分子量约为189Kd。纸层析与气相层析分析表明其为单一葡聚糖。经高碘酸氧化,Smith降解,甲基化,气相层析,气质联机分析,核磁共振(~1H-NMR,~(13)C-NMR)谱及红外光谱测定等,可确定PC-4的主链结构由β(1→3)糖苷键相连的葡萄糖构成,部份残基C_6上带有分支。约每5个糖残基有两个侧链,侧链仅为1个葡萄糖残基。 给ICR小鼠腹腔注射PC-4,对移植性肿瘤S-180有一定的抑制作用,抑瘤率为67％。离体条件下,PC-4与肿瘤细胞S-180共同培养,则未显示抑瘤作用。     

Anti-tumor polysaccharide from the mycelium of liquid-cultured Agaricus blazei mill.

Anti-tumor active polysaccharide against Sarcoma 180 was isolated by DEAE-Sepharose CL-6B and Sepharose 4B column chromatography from the hot-water soluble fraction of the mycelium of liquid-cultured Agaricus blazei mill. This polysaccharide did not react with antibodies of anti-tumor polysaccharides such as lentinan, gliforan, and FIII-2-b which is one of anti-tumor polysaccharides from Agaricus blazei. Moreover, the analyses of 13C-NMR and GC-MS suggested that this polysaccharide was preliminarily glucomannan with a main chain of beta-1,2-linked D-mannopyranosyl residues and beta-D-glucopyranosyl-3-O-beta-D-glucopyranosyl residues as a side chain. This polysaccharide was completely different from the anti-tumor polysaccharide from fruiting body of Agaricus blazei, beta-1,6-glucan.     

金针菇(Flammulina Velutipes(Curt.ex Fr.)Sing.)子实体多糖PA_(5)DE的提取及性质研究

 从金针菇Flammulina velutipes(Curt.ex Fr.)Sing.子实体中提取水溶性多糖,经乙醇分级,DEAE-Sephadex A-25纯化,得PA_5DE。以聚丙烯酰胺凝胶电泳和凝胶柱Sepharose 4B层析证明是化学均一性多糖,分子量是47.1万。用GLC、I.R、~13C-N.M.R.分析表明含有D-葡萄糖、D-甘露糖、D-岩藻糖。PA_5DE分子可能具有分支结构,含β-型糖苷键,存在β(1→3)和β(1→6)型糖苷键连接,并有抑制肿瘤S-180的活性。     

Effect of immunoglobulin G on membrane-bound enzyme activity of sarcoma 180 cells.

Changes in the activity of membrane bound ATPase of Sarcoma 180 cells caused by immunoglobulin G (IgG) of anti-Sarcoma 180 was investigated in relation to the incorporation of amino acid by the cells. Enzymatic activity of ATPase was increased up to 160% of the original activity upon incubation of the cell with IgG. Kinetic studies showed that IgG did not change the affinity of this enzyme for the substrate, but exerted influence upon catalytic efficiency of the enzyme. The rate of incorporation of leucine into Sarcoma 180 cells was also affected by IgG, as observed in the effect of IgG on the enzymatic reaction of the cells.     

Adenosine phosphyorylase activity as distinct from inosine-guanosine phosphorylase activity in Sarcoma 180 cells and rat liver.

Adenosine phosphorylase (EC 2.4.2.-) activity present in Sarcoma 180 cells grown in culture and in rat liver, is shown to be distinct from inosine-guanosine phosphorylase by several criteria: (a) treatment of Sarcoma 180 cell extract with p-chloromercuribenzoate inhibited the two activities to a different extent, (b) adenine selectively protected the adenosine phosphorylase activity of Sarcoma 180 and rat liver extract against heat inactivation, while hypoxanthine selectively protected inosine-guanosine phosphorylase activity, (c) at nearly saturating substrate concentrations and using Sarcoma 180 extract, the rates of ribosylation of a mixture of adenine + hypoxanthine or adenine + guanine, but not of hypoxanthine + guanine, were found to be almost equal to the sum of their individual rates as measured separately, (d) inosine selectively inhibited the ribosylation of hypoxanthine and guanine catalysed by Sarcoma 180 and rat liver extract while 2-chloroadenosine selectively inhibited the ribosylation of adenine and N6-furfuryladenine, (e) pH vs. activity curves were similar with hypoxanthine or guanine as the substrate but they were markedly different from the curve with adenine as the substrate. The potential role of adenosine phosphorylase activity in vivo is discussed.     

Comparison of antitumor activities of different polysaccharide fractions from the stems of Dendrobium nobile Lindl

The antitumor activities of extracted polysaccharide fractions from the stems of Dendrobium nobile Lindl were investigated. Polysaccharides were sequentially extracted from the stems of D. nobile to obtain three fractions, i.e. water extract fraction (DNP-W), 5% NaOH extract fraction (DNP-OH) and 5% HCl extract fraction (DNP-H). Further the DNP-W was isolated to give six sub-fractions (DNP-W1, DNP-W2, DNP-W3, DNP-W4, DNP-W5 and DNP-W6) by anion-exchange chromatography. The monosaccharide profile, protein content, uronic acid content, total carbohydrate content, viscosity and molecular weight of nine polysaccharide fractions were analyzed. Both the in vivo and in vitro antitumor activities of nine polysaccharide fractions were evaluated and compared. Results indicated that DNP-W1 and DNP-W3 exhibited high antitumor activities against Sarcoma 180 in vivo and HL-60 in vitro. The results suggested that DNP-W1 and DNP-W3 could be considered as an effective natural antitumor source.     

短裙竹荪多糖Dd-S3P的分离纯化及其性质研究

短裙竹荪子实体经2％Na2CO3溶液提取,用蛋白酶法和Sevag法相结合除去蛋白,乙醇分级沉淀,级分3经DEAE－SephadexA－25柱层析纯化得到短裙竹蒸蒸荪多糖DdS3P．经测定该多糖为均一组分,分子量约为3．8×105,红外光谱呈现出典型的多糖吸收峰,含有α-型糖苷连接键,紫外扫描无核酸和蛋白质的特征吸收峰．纸层析和气相层析分析得知Dd－S3P含有D-葡萄糖（D－Glc）、D-甘露糖（D－Man）和D-木糖（D－Xyl）,其摩尔比为1．83：1．00：1．21．经动物试验表明此多糖对小鼠S-180有一定的抑制作用,抑瘤率约31．3％．     

毛木耳子实体中活性多糖APPⅡA的分离纯化与结构初探

毛木耳Auricularia polytricha子实体500g经热水浸提、乙醇沉淀、Sevage法脱蛋白后,得到粗多糖3.94g,得率为0.79%。经小鼠S180载体实验证实,50mg/kg该粗多糖对肿瘤的抑制率达47.89%,呈极显著差异。利用离子交换柱对其进行分离,共得到三个部分,分别命名为APPⅠ、APPⅡ、APPⅢ。其中,APPⅡ对小鼠S180的抑制率达到56.36%,明显优于其他两部分。通过SephacrylS300凝胶过滤层析柱后,APPⅡ被进一步分离为A和B两个部分,APPⅡA的抑瘤率为53.61%,远高于APPⅡB,呈极显著差异。通过紫外光谱、红外光谱等方法对该单一多糖组分进行结构分析,表明APPⅡA为一相对分子质量约为110,000Da、可能同时具有α和β构象的杂多糖,主要有木糖和甘露糖组成,且纯度较高,含有硫酸基。     

Location of F1 ATPase-like Genes on the Physical Map of the Bacillus subtilis 168 Chromosome

A water-soluble polysaccharide, FI, extracted from the mycelium of Granoderma tsugae, was fractionated and purified by ion-exchange chromatography, gel filtration, and affinity chromatography. Sixteen polysaccharides obtained were examined for antitumor effects on Sarcoma 180 in mice.

The three active polysaccharides obtained were as follows:

FI₀-a: A glycan-protein complex containing 9.3% protein, and having a hetero-glyco-chain of mannose and xylose.

FI₀-b-α: Molecular weight 10,000, glucan-protein complex containing 25.8% protein. The inhibition ratio was 61.8% against the solid cancer Sarcoma 180/mice; the survival ratio was more than 194% of the control group (100).

FA-1-b-α: Molcular weight 16,000, a complex of glycan: protein = 42:58 w/w, consisting of glucose as a main component, and associated with arabinose, mannose, xylose, and galactose. This had a tumor inhibition ratio of 56% and a survival ratio of more than 182%.

Comparison of active glycan with the fruiting body and mycelium: Among water-soluble polysaccharides of fruiting body, FI₀-a and FA-1, with antitumor activity, were both glucogalactan-protein complexes of molecular weight 10,000, but that of mycelium was a homoglucan protein complex in FI₀-b-α and heteroglucan protein in both FA-l-a and FA-l-b-α. The heteroglucan had a low tumor inhibition ratio, but caused a high survival ratio in mice.     

Effect of immunoglobulin G on membrane-bound enzyme activity of sarcoma 180 cells

Changes in the activity of membrane bound ATPase of Sarcoma 180 cells caused by immunoglobulin G (IgG) of anti-Sarcoma 180 was investigated in relation to the incorporation of amino acid by the cells. Enzymatic activity of ATPase was increased up to 160% of the original activity upon incubation of the cell Igg. Kinetic studies showed that IgG did not change the affinity of this enzyme for the substrate but exerted influence upon catalytic efficiency of the enzyme. The rate of incorporation of leucine into Sarcoma 180 cells was also affected by IgG, as observed in the effect of IgG on the enzymatic reaction of the cells.     

Antioxidant and antitumor activities of the polysaccharide from seed cake of Camellia oleifera Abel

To explore biomedical potential of the polysaccharide from seed cake of Camellia oleifera Abel, we investigated antioxidant and antitumor capacities of the polymer. The results showed that the polysaccharide is capable of scavenging both superoxide anion and hydroxyl radicals in vitro. The highest scavenging rate of superoxide anion and hydroxyl radicals is 85% and 76%, respectively. Using the model animal, Caenorhabditis elegans, we further show that the polysaccharide can increase antioxidant enzyme activity, decrease lipid peroxidation level, and reduce paraquat-induced oxidative damage at a polysaccharide concentration more than 50mg/l. We also revealed that the polysaccharide has some ferric chelating ability and strong in vivo antitumor activity. The antitumor rate against Sarcoma180 solid tumor grown in BALB/C mice reached 85.6% at the highest dose of 40×20mg/kgdays.     

Water-soluble polysaccharides from Salvia officinalis L. possessing immunomodulatory activity

A water-soluble polysaccharide complex (A) composed of galactose (17.9%), 3-O-methyl-galactose (3.0%), glucose (15.5%), mannose (8.3%), arabinose (30.4%), xylose (7.6%), fucose (2.6%), rhamnose (6.7%), and uronic acids (8.0%) has been isolated from the aerial parts of sage (Salvia officinalis L.) by cold water extraction. It showed a broad molecular-mass distribution pattern (Mw approximately 2000-93,000) with a predominance of polymers with Mw< 10,000. Ion-exchange chromatography of A afforded six polymeric fractions (A1-A6) in which arabinogalactans associated with galacturonan and/or rhamnogalacturonan backbones prevail. Sage polysaccharides were examined for their immunomodulatory activity in the comitogenic thymocyte test which is interpreted as being an in vitro correlate of adjuvant activity. The acidic polysaccharide fractions A2, A3 and A4 exhibited the highest mitogenic and comitogenic activities of all fractions tested, and relatively high SI(comit)/SI(mit) ratios approximately 3 indicate potential adjuvant properties of these polysaccharides.     

Lipoprotein lipase-like activity in the liver of mice with Sarcoma 180

The triglyceride lipase (TGL) activity of liver homogenates of mice with Sarcoma 180 was measured. The liver homogenate of normal or tumor-bearing mice was treated with 0.25% Triton X-100 and centrifuged at 100,000 g for 60 min, and the supernatant was applied to a heparin-Sepharose column. In normal mice, most of the TGL activities in the supernatant was eluted with 0.75 M NaCl from the column. In mice with Sarcoma 180, the TGL gave two peaks on heparin-Sepharose column chromatography, which were eluted with 0.75 M and 1.5 M NaCl, respectively. The activity in the first peak (0.75 M NaCl eluate) decreased; that in the second peak (1.5 M NaCl eluate) increased, and the ratio of the second peak to the first peak increased during tumor development. The livers of normal mice and mice on day 10 after tumor inoculation were perfused with heparin. The highest rate of the TGL release occurred within 1 min of heparin perfusion, and the bulk of heparin-releasable activity appeared within 2 min of perfusion in both normal and tumor-bearing mice. The TGL activity in liver perfusate of tumor-bearing mice, as well as that of liver homogenate, was resolved on a heparin-Sepharose column into two peaks, which were eluted with 0.75 M and 1.5 M NaCl, and most of the activity was eluted with 1.5 M NaCl. The nature of the TGL activity eluted from a heparin-Sepharose column was investigated. In both liver homogenates and liver perfusates, the first peak did not require serum for maximal activity and was relatively resistant to a high concentration of NaCl or protamine sulfate.(ABSTRACT TRUNCATED AT 250 WORDS)     

CYTOCHEMICAL LOCALIZATION OF ADENOSINE TRIPHOSPHATASE IN THE MITOTIC APPARATUS OF HELA AND SARCOMA 180 TISSUE CULTURE CELLS

ATPase was localized in distinct regions of the mitotic apparatus of HeLa and Sarcoma 180 tissue culture cells. ATPase was demonstrated in the metaphase spindle of HeLa and Sarcoma 180 cells fixed in cold buffered 2 per cent formalin (pH 6.5 to 6.8) containing 2 x 10^-3 M CaCl₂. A high concentration of ATPase was frequently observed at the poles of the spindle. ATPase was also demonstrated in the interzonal region of both cell types during anaphase. The narrowing of the band of ATPase activity localized in the interzonal region during telophase indicates that ATPase activity is associated with the central spindle. In polar views of Sarcoma 180 cells fixed in cold, unbuffered, 2 per cent formalin, ATPase was frequently localized in granules in the region of the inner circumference of the ring of chromosomes formed at metaphase. ATPase in the mitotic apparatus of HeLa and Sarcoma 180 cells was shown not to be due to non-specific alkaline phosphatase. Mitotic apparatus ATPase in Sarcoma 180 cells was suppressed by an —SH inhibitor.     

姬松茸菌丝体中各组分多糖的抗肿瘤免疫活性

对深层发酵培养的姬松茸菌丝体依次采用热水、冷碱、热碱提取,得到了四种主要的多糖组分。其中冷碱提取的水不溶性多糖(CAASP)得率最高,达8.5 8g/ 10 0g干菌,且不同的多糖组分具有不同的单糖组成。其中热水提取的水溶性多糖(HWSP)及冷碱提取的水溶性多糖(CAWSP)在2 0mg/kg·d的剂量下对小鼠S180 移植瘤显示出较高的抗肿瘤活性,抑瘤率分别高达5 9.84 %和6 4 .6 6 %。同时发现在四种多糖在发挥抗肿瘤作用过程中,小鼠的脾脏指数和胸腺指数有不同程度的提高。并且各多糖活性组分在体外均具有显著的刺激脾淋巴细胞增殖的作用     

Polysaccharide isolated from Passiflora edulis: Characterization and antitumor properties

A hot water-extracted polysaccharide fraction (PFCM) of Passiflora edulis was characterized by microanalysis, infrared spectroscopy, NMR and high performance size-exclusion chromatography. The major component in PFCM is (1 → 4) linked galacturonic acid (esterified and unesterified). Neutral sugars such as arabinose, glucose, rhamnose, mannose, and fucose were also present. Traces of xylose and ribose were detected. The PFCM sample had a similar molar mass to that of pectin extracted from P. edulis under acidic conditions. Sarcoma 180 tumors treated with PFCM showed a growth inhibition ratio ranging from 40.59% to 48.73% depending on the dosage and type of PFCM administration (oral or intraperitoneal). Toxicological analysis shows that PFCM increases the cell types involved in primary defense mechanisms and no significant changes in the biochemical parameters and organs (e.g., kidney and liver) were observed. However, the use of PFCM treatment increased the spleen weight when compared with the use of 5-fluorouracil.     

Correlation of structure to antitumor activities of five derivatives of a beta-glucan from Poria cocos sclerotium

A water-insoluble (1-->3)-beta-D-glucan isolated from fresh sclerotium of Poria cocos was, respectively, sulfated, carboxymethylated, methylated, hydroxyethylated, and hydroxypropylated, to afford five water-soluble derivatives. Their weight-average molecular masses (Mw) and intrinsic viscosities ([eta]) were determined by size-exclusion chromatography combined with laser light scattering (SEC-LLS), LLS, and viscometry in phosphate buffer solution (PBS) at 37 degrees C. The antitumor activities, against Sarcoma 180 tumor cell (S-180) and gastric carcinoma cell strain (MKN-45 and SGC-7901) of the native beta-glucan and the five derivatives, were tested in vitro and in vivo. The Mw values of the five derivatives in PBS were determined to be 3.8 x 10(4), 18.9 x 10(4), 16.0 x 10(4), 76.8 x 10(4), and 224.3 x 10(4), respectively. The high Mw values of the hydroxyethylated and hydroxypropylated derivatives in aqueous solution resulted from aggregation, and their true Mw values obtained in dimethyl sulfoxide were 20.1 x 10(4) and 19.1 x 10(4). The sulfated and carboxymethylated derivatives having DS of 1.0-1.3 show good water solubility, and exist as relatively expanded chains in aqueous solution. Interestingly, the native beta-glucan did not show antitumor activity, whereas the sulfated and carboxymethylated derivatives exhibit significant antitumor activities against S-180 and gastric carcinoma tumor cells. This work showed that good water solubility, relatively high chain stiffness, and moderate molecular mass of the derivatives in aqueous solution contribute beneficial to enhancement of antitumor activity.     

巴戟天及其糖提取物对于体外培养成骨细胞OPG/RANKL基因系统表达的影响

目的:探讨巴戟天及多糖提取物对成骨细胞骨保护素(OPG)/核因子κB受体活化因子配体(RANKL)基因系统表达的影响。方法:取2~3天的SD大鼠5只分离原代成骨细胞,再取8周龄SD大鼠35只随机分为七组,对照组不进行处理,三组给予10 g/L、50 g/L、100 g/L巴戟天水灌胃,其余三组分别给予10 g/L、50 g/L、100 g/L巴戟天多糖灌胃,72 h后采用采用ELISA法测定培养液中OPG、RANKL及骨钙素的含量,采用MTT法检测不同浓度巴戟天水及多糖提取物对大鼠成骨细胞增殖的影响,采用荧光定量PCR检测OPG和RANKL mRNA表达情况;通过Westernblot检测OPG和RANKL蛋白表达水平。结果:巴戟天水及多糖提取物组A570nm、ALP活性、骨钙素含量、OPG/RANKL mRNA表达量、OPG和RANKL蛋白表达阳性密度均高于对照组(P0.05);A 570 nm、ALP活性、骨钙素含量、OPG/RANKL mRNA表达量、OPG和RANKL蛋白表达阳性密度均高于同等剂量的水提取物各组(P0.05);巴戟天多糖组中随着多糖剂量的升高A 570 nm、ALP活性、骨钙素含量、OPG/RANKL mRNA表达量、OPG和RANKL蛋白表达阳性密度,差异比较有统计学意义(P0.05)。结论:巴戟天水及多糖提取物均能促进体外培养成骨细胞的增殖,提高成骨细胞活性。     

Correlation between the antitumor activity of a polysaccharide schizophyllan and its triple-helical conformation in dilute aqueous solution

Eight samples of a polysaccharide schizophyllan ranging in weight-average molecular weight Mw (in water) from 5 x 10(3) to 1.3 x 10(5) were prepared and their antitumor activity (expressed in terms of the tumor inhibition ratio) against Sarcoma 180 ascites, intrinsic viscosities [eta], and gel-filtration chromatograms in aqueous solution were determined. The tumor inhibition ratio was essentially unity for samples with Mw higher than 9 x 10(4), but reduced to zero or even to a negative value when Mw was lower than 10(4). The [eta] data combined with the chromatographic data showed that above Mw approximately 9 x 10(4) the predominant species of schizophyllan in aqueous solution is the previously found rigid triple helix, whereas below Mw approximately 9 x 10(4) both triple helices and single chains coexist in the solution and the fraction of triple helices decreases monotonically to zero as Mw is decreased to 5 x 10(3). From these findings it was concluded that the antitumor potency of schizophyllan in water is related to the amount of triple helices relative to that of single chains.     

Bioactivities of water-soluble polysaccharides from fruit shell of Camellia oleifera Abel: Antitumor and antioxidant activities

Polysaccharides were extracted from fruit shell of Camellia oleifera Abel. Fruit shell of Camellia oleifera Abel polysaccharide (WEP2) was a water-soluble compound. Its molecular weight was about 362 kDa. HPLC analysis showed that this polysaccharide was composed of rhamnose, fucose, arabinose, mannose, galactose and glucose in the molecular ratio of 4.05, 11.62, 1.78, 3.91, 8.76 and 27.06, respectively. The broad intense characteristic peak around 3463 cm⁻¹ due to the hydroxyl stretching vibration of the polysaccharide was observed in the polysaccharide. The characteristic absorption bands at 852 cm⁻¹ and 893 cm⁻¹ indicated that WEP2 contained both α-glycosidic and β-glycosidic linkages. WEP2 exhibited remarkable antitumor activity against Sarcoma180 cell compared to the negative control group. At the highest dose 40 mg/kg days, the tumor inhibition rate reached 65.2%. The scavenging effects of WEP2 to hydroxyl radical and superoxide radical anion were 72.5% and 86.3% at a concentration of 1.0 mg/ml, respectively.