扁藻多糖的分离分析及其生物活性的初步研究

目的:初步确定亚心型扁藻多糖的理化性质、结构特征及抗肿瘤活性。方法:从培养至平衡期后的亚心型扁藻细胞内分离出乙醇沉淀的多糖级分,经过Sephadex G-150凝胶层析纯化后,利用化学分析及仪器分析进行理化性质和结构特征分析,并通过此多糖对小鼠体内黑色素瘤的抑制作用确定其抗肿瘤活性。结果:扁藻多糖含有氨基和硫酸基,并且含有以葡萄糖为主的多种已知和未知的单糖糖基和糖醛酸,其对小鼠体内黑色素瘤的抑制率为34.2%。结论:此实验结果为扁藻多糖在医疗方面作为抗肿瘤药物的可能性提供了基础依据。     

米氏凯伦藻多糖的酶法制备及其体外抑制肿瘤血管生成活性

研究碱性蛋白酶酶解制备米氏凯伦藻多糖及其抑制肿瘤血管活性。采用闪式萃取结合酶法制备米氏凯伦藻多糖,其适宜提取条件为碱性蛋白酶酶解温度40℃、酶解p H为8.5、酶用量为2.5%、酶解时间为2 h。其提取液经三氯乙酸除蛋白、乙醇沉淀获得米氏凯伦藻多糖。制备的米氏凯伦藻多糖在一定浓度时可抑制肿瘤细胞培养液诱导的内皮细胞增殖和迁移作用,具有一定的抑制肿瘤血管生成活性。     

海边月见草叶提取物乙醇洗脱级分的抗氧化作用

海边月见草(Oenothera drummondii Hook.)叶提取物经X-5大孔树脂吸附及体积分数10%、30%、50%和70%乙醇溶液梯度洗脱获得4个级分,研究了这4个级分对DPPH、·OH和O2-的清除作用及其在卵黄脂蛋白过氧化体系中的抗氧化作用。4个洗脱级分均表现出较强的清除DPPH和·OH的能力,清除率随各级分浓度的升高而提高,其中,30%乙醇洗脱级分的清除作用最强,对DPPH和·OH的50%抑制浓度(IC50)分别为0.069和0.741g·L-1。浓度达1.0g·L-1,4个洗脱级分对O2.均有一定的清除能力,且对卵黄脂蛋白过氧化作用的抑制率均在50%以上。结果表明,海边月见草叶提取物的不同浓度乙醇洗脱级分具有较强的清除自由基的能力,并有一定的抗卵黄脂蛋白过氧化的作用,但总体效果较槲皮素略差。     

亚心形扁藻多糖的提取分离纯化及组成研究

以实验室培养的亚心型扁藻为原料,热水提取扁藻多糖。多糖经加热除蛋白、乙醇分级、Sephadex G-50柱层析等一系列纯化步骤后,得到一扁藻多糖(简称PPSc-1)。PPSc-1经醋酸纤维素薄膜电泳和SephadexG-100柱层析的纯度鉴定,组分均一。将多糖PPSc-1用三氟乙酸全水解,然后进行HPAEC-PAD分析,结果表明组成该多糖的单糖为D-半乳糖、D-2-氨基葡萄糖、D-阿拉伯糖、D-鼠李糖和3种未知单糖。各种已知单糖的摩尔比是D-半乳糖∶D-2-氨基葡萄糖∶D-阿拉伯糖∶D-鼠李糖=59.64∶40.9∶18∶1。     

网地藻多糖清除DPPH·自由基活性的动力学研究

该研究通过超声辅助并采用醇沉、脱蛋白、脱色、干燥的方法,分别检测低(0.1 mg·mL~(-1))、中(0.25mg·mL~(-1))、高(0.5 mg·mL~(-1))三种浓度下的网地藻多糖对DPPH·自由基的清除能力,探讨质量浓度和反应时间对网地藻多糖清除DPPH·自由基活性的变化规律。按照一级反应动力学方程和二级反应动力学方程分别建立反应动力学模型。结果表明:不同的质量浓度和反应时间对网地藻多糖清除DPPH·自由基活性均有影响,网地藻多糖质量浓度提高,其清除DPPH·自由基的能力逐渐加强,当网地藻多糖浓度为0.5 mg·mL~(-1)时,反应20 min,网地藻多糖清除DPPH·自由基的清除率最高为86.06%,其清除DPPH·自由基活性半数清除率(IC_(50))为0.25 mg·mL~(-1)。准一级动力学模型拟合的线性相关性较差,相关系数R~2的范围分别为0.848~0.891;准二级动力学模型拟合的相关系数R~2的范围为0.902~0.967,因此采用二级动力学拟合方程能较好地描述网地藻多糖对DPPH·自由基的清除能力。网地藻多糖在低(0.1 mg·mL~(-1))、中(0.25 mg·mL~(-1))、高(0.5 mg·mL~(-1))三种浓度时对DPPH·的二级反应的清除速率常数(k_2)分别为0.011、0.054、0.421。这说明网地藻多糖随着反应浓度逐渐升高其清除DPPH·自由基的速度越来越快,清除自由基能力也越来越强,结合IC_(50)值来共同评价抗氧化能力,IC_(50)值越小,反应速率值越大,表明其抗氧化活性越好,这与实验得出的数据一致。     

当归不同提取液中阿魏酸、咖啡酸含量及抗氧化作用的比较研究北大核心CSCD

为考察不同提取条件对当归提取液中阿魏酸和咖啡酸的含量及抗氧化作用的影响,本研究通过HPLC外标一点法检测当归提取液中阿魏酸和咖啡酸的含量,通过DPPH法和ABTS法检测当归提取液的抗氧化作用。研究结果表明,不同提取条件下当归提取液中阿魏酸的含量为0. 574～0. 707 mg/g,咖啡酸为0. 012～0. 082 mg/g;碱水溶液提取时阿魏酸与咖啡酸的含量最高,其抗氧化活性也最强,而95%乙醇提取时的含量最低,抗氧化活性最弱;提取溶剂随醇浓度的增加,咖啡酸的含量逐渐减少,阿魏酸的含量呈现先增加后减少的趋势,30%～50%乙醇提取时各组分有良好的协同抗氧化作用。     

当归不同提取液中阿魏酸、咖啡酸含量及抗氧化作用的比较研究

为考察不同提取条件对当归提取液中阿魏酸和咖啡酸的含量及抗氧化作用的影响,本研究通过HPLC外标一点法检测当归提取液中阿魏酸和咖啡酸的含量,通过DPPH法和ABTS法检测当归提取液的抗氧化作用。研究结果表明,不同提取条件下当归提取液中阿魏酸的含量为0. 574～0. 707 mg/g,咖啡酸为0. 012～0. 082 mg/g;碱水溶液提取时阿魏酸与咖啡酸的含量最高,其抗氧化活性也最强,而95%乙醇提取时的含量最低,抗氧化活性最弱;提取溶剂随醇浓度的增加,咖啡酸的含量逐渐减少,阿魏酸的含量呈现先增加后减少的趋势,30%～50%乙醇提取时各组分有良好的协同抗氧化作用。     

仙人掌多糖对蛋白质糖基化终产物和醛糖还原酶的抑制效应

以仙人掌为原料提取多糖产品,通过超滤浓缩和不同浓度乙醇分级沉淀,分别得到55%乙醇沉淀的多糖(OPMC I),进一步乙醇浓度增至80%沉淀的多糖(OPMCⅡ),以及一次性80%乙醇沉淀的多糖(OPMCⅢ)。比较了三种多糖对蛋白质非酶糖基化反应的终端产物(AGEs)和醛糖还原酶(AR)形成的抑制作用。结果表明,在对AGEs形成的抑制过程中,第四周时OPMC II的抑制作用较强,且强于同剂量的氨基胍;在对AR活性的抑制作用中,OPMC II在三种多糖样品中抑制作用最强,但明显低于阳性对照物依帕司他。     

沙棘果皮多糖清除氧自由基的活性研究

沙棘(Hippophae rhamnosides)果皮经80℃恒温水浴提取, 乙醇沉淀得粗多糖。Sevag法去蛋白,经50%和70%乙醇分级, 得3种级分沙棘多糖H1、H2和H3; 以Fenton 反应, 即H2O2/Fe2+/水杨酸为.OH产生和检测体系; 以邻苯三酚/EDTA/Tris-HCl为O2 -. 产生体系, 对沙棘多糖H1、H2和H3进行抗氧自由基活 性研究。结果表明, 沙棘多糖对.OH和O2 -. 有较显著的清除能力。不同级分多糖H1、H2和H3浓度达200mg.mL-1时, 对.OH的清除率分别为44.9%、49.0%和26.4%, 抗O2-. 活性分别为36.9%,15.4%和23.1%。多糖质量浓度增大时,两种自由基清除率增加, 且呈量效关系。     

沙棘果皮多糖清除氧自由基的活性研究

沙棘(Hippophae rhamnosides)果皮经80℃恒温水浴提取,乙醇沉淀得粗多糖.Sevag法去蛋白,经50%和70%乙醇分级,得3种级分沙棘多糖H1、H2和H3;以Fenton反应,即H2O2/Fe2 /水杨酸为·OH产生和检测体系;以邻苯三酚/EDTA/Tris-HCl为O2-.产生体系,对沙棘多糖H1、H2和H3进行抗氧自由基活性研究.结果表明,沙棘多糖对·OH和O2-.有较显著的清除能力.不同级分多糖H1、H2和H3浓度达200μg·mL-1时,对·OH的清除率分别为44.9%、49.0%和26.4%,抗O2-.活性分别为36.9%,15.4%和23.1%.多糖质量浓度增大时,两种自由基清除率增加,且呈量效关系.     

灵芝液态发酵高产胞外多糖菌株筛选及多糖特性分析

对10株灵芝菌株发酵菌丝体生物量、胞内多糖含量、胞外粗多糖得率及其多糖含量和单糖组成特征进行了系统分析。结果表明,随着培养时间的延长,10株菌株的生物量均有不同程度增加,但不同菌株胞内多糖含量和胞外粗多糖得率变化趋势有所不同。在培养至7d时,G0023和G0160菌株胞外粗多糖得率均较高,分别为3.02g/L和3.14g/L,其多糖含量分别达到了84.11%和91.03%,可作为发酵高产胞外多糖的优良菌株。分级醇沉胞外多糖分析结果表明,各菌株胞外液20%乙醇沉淀所得20E组分的得率和多糖含量均高于50%乙醇沉淀所得的50E组分,说明胞外液中主要以大分子量多糖为主。20E主要由葡萄糖组成,含有少量木糖和甘露糖;50E主要由葡萄糖和甘露糖组成,含有少量木糖和半乳糖。胞外液表观粘度随剪切速率变化曲线分析结果显示,各菌株胞外液均呈现剪切变稀非牛顿流体特性,其表观粘度大小与菌株对应20E组分的得率及多糖含量呈正相关。     

蝶蛹金小蜂毒液蛋白不同提取分离方法的比较

为了建立蝶蛹金小蜂Pteromalus puparum毒液抑制寄主血细胞免疫活性组分合适的分离纯化方法,就等电点沉淀法、乙醇沉淀法、75%硫酸铵沉淀法、75%硫酸铵沉淀法+40℃加热处理法,以及75%硫酸铵沉淀法分别与3种不同滤膜的分子大小截留法的组合等7种方法对毒液蛋白分离效果及活性的影响进行了比较。结果表明：等电点沉淀法获得的组分抑制寄主菜粉蝶Pieris rapae离体血细胞延展和包囊的活性最强,乙醇沉淀法次之,75%硫酸铵沉淀法最弱。从蛋白组分的SDS-PAGE图谱来看,等电点沉淀法获得毒液组分相对最纯,仅有3条主要谱带,分子量大小在45～116.2 kDa范围内;乙醇沉淀法次之,有5条主要谱带,分子量大小在24～116.2 kDa范围内;硫酸铵沉淀法的谱带组成与毒液蛋白粗提液相似。3种分子大小截留法获得的毒液组分的活性分析表明,强活性组分分子量大小可能都大于100 kDa。综合认为,7种方法中以等电点沉淀法提取分离蝶蛹金小蜂毒液蛋白相对为最适。     

Extraction of DNA,RNA, and glycogen from oysters

The macromolecules DNA, RNA, and glycogen (CHO)_N were extracted with phenol from eggs, sperm, and the combined gill, mantle, and digestive gland tissues of oysters, using methods effective for the HeLa cells. From the eggs, most of the (CHO)_N and RNA coprecipitated in 20% ethanol whereas the DNA precipitated from 50% ethanol solutions. From the combined tissues all three macromolecules precipitated in 20% ethanol whereas from sperm they precipitated from 50% ethanol solutions. The DNA preparation from sperm was viscous and white but contained mostly (CHO)_N and some RNA. However, the DNA-(CHO)_N-RNA preparation was rendered insoluble in saline by CaPO₄ treatment and was not toxic for mammalian kidney cells in tissue culture. Thus, the methods do not yield pure nucleic acids but appear suitable for attempts to extract infectious DNA from oysters.     

Isolation and Characterization of A Temperature Sensitive Protein from dog Colostrum And Milk

A temperature sensitive protein (one of several) that was soluble at 2°C but reversibly precipitated upon warming to room temperature was isolated from dog milk by precipitation with 50% saturated ammonium sulfate, ion exchange chromatography, centrifugation at 30°C and preparative isoelectric focusing in polyacrylamide gel. The molecular weight by sedimentation equilibrium was 13, 400; however, by thin layer gel filtration it appeared to be much larger. Ultracentrifu-gation studies at conditions near those at which the protein precipitated revealed no evidence of aggregation. The protein had a fairly high content of nonpolar amino acids with proline being in the highest amount. The effects of pH, ionic strength, heat and ethanol on the solubility of the protein were studied. A circular dichroism spectrum of the temperature sensitive protein indicated that the protein is quite unordered. An antiserum against the temperature sensitive protein reacted with each of the other temperature sensitive proteins from milk but not with dog serum or saliva.     

大孔吸附树脂分离纯化人参二醇类和三醇类皂甙

本实验研究大孔吸附树脂从人参根提取物中富集、分离人参二醇类和人参三醇类皂甙的工艺条件及参数。用不同浓度的乙醇洗脱,使人参二醇类和三醇类皂甙实现富集分离,人参二醇类皂甙富集在80％乙醇洗脱液部分,人参三醇类皂甙富集在40％洗脱液部分。得到含量大于25％的人参三醇类皂甙,含量大于50％的人参二醇类皂甙,总皂甙洗脱率在91％以上。此法能够较好地分离、纯化人参二醇类和三醇类皂甙。     

Investigation of the pharmaceutical and pharmacological equivalence of different Hawthorn extracts

Seven Hawthorn extracts were tested in isolated guinea pig aorta rings. The effect on noradrenaline- (10 microM) induced contraction was investigated. The extracts were prepared using ethanol (40 to 70% v/v), methanol (40 to 70% v/v), and water as the extraction solvents. The aqueous-alcoholic extracts displayed similar spectra of constituents. They were characterised by similar procyanidin, flavonoid, total vitexin and total phenols content and by similar TLC fingerprint chromatograms. The aqueous extract, however, showed a different fingerprint and a noticeably lower concentration of procyanidins, flavonoids and total phenols but a similar total vitexin content. All 7 extracts had a relaxant effect on the aorta precontracted by noradrenaline and led to relaxations to 44 until 29% of the initial values. The EC50 values of the aqueous-alcoholic extracts varied between 4.16 and 9.8 mg/l. The aqueous extract produced a similarly strong maximal relaxation as the other extracts, but the EC50, at 22.39 mg/l, was markedly higher. The results show that Hawthorn extracts with comparable quality profiles were obtained by using aqueous-alcoholic extraction solvents (40 to 70% ethanol or methanol). The extracts exerted comparable pharmacological effects. When using water as the extraction solvent, both, the spectrum of constituents and the pharmacological effect, deviated remarkably. It is thus possible to obtain bioequivalent extracts with comparable effect profiles by using 40 to 70% ethanol or methanol as the extraction solvent.     

Role of phosphatidylinositol (PI) in ethanol production and ethanol tolerance by a high ethanol producing yeast

The effect of inositol addition on phospholipids, cell growth, ethanol production and ethanol tolerance in a high ethanol producing Saccharomyces sp were studied. Addition of inositol greatly influenced major phospholipid synthesis. With inositol in the fermentation medium, phosphatidylinositol (PI) content was increased, while phosphatidylcholine (PC) and phosphatidylethanolamine (PE) were decreased. However, without inositol in the fermentation medium, PI content dropped down within 24 h, then increased, but was lower than in the presence of inositol. When yeast cells had a higher content of PI, they produced ethanol much more rapidly and tolerated higher concentrations of ethanol. During ethanol shock treatment at 18% (v/v) ethanol, yeast cells with a higher concentration of PI lost their viability much more slowly than those with a lower concentration of PI, indicating that the PI content in these yeast cells can play an important role in ethanol production and ethanol tolerance. Fatty acids and ergosterol were not responsible for high ethanol tolerance and high ethanol production in this yeast strain. Received 22 September 1998/ Accepted in revised form 20 December 1998     

Fixation-associated quantitative variations of DNA fluorescence observed in flow cytometric analysis of hemopoietic cells from adult diploid frogs.

We have examined, by flow cytometry, the apparent DNA content of frog blood cells that had been fixed with either 50% ethanol, 70% ethanol, or 66% methanol, before being stained with either mithramycin, propidium iodide, or Hoechst 33258. After 50% ethanol fixation, regardless of the dye used, the DNA content of the hemopoietic cells appeared unimodal, but after either 70% ethanol or 66% methanol fixation, it appeared bimodal. Cell sorting revealed that the lower and upper modes are represented by erythrocytes (RBCs) and leukocytes (WBCs), respectively. In amphibians, the chromatin of metabolically inactive RBCs is highly condensed relative to the chromatin of metabolically active WBCs. The bimodal distribution of DNA contents seen with 66% methanol and 70% ethanol, but not 50% ethanol, seems to reflect this disparity in the degree of chromatin condensation existing between the RBCs and WBCs. This, in turn, implies that the accessibility of fluorescent DNA dyes to the chromatin of fixed frog hemopoietic cells, especially of RBCs, can be affected by the concentration of alcohol used for their fixation.     

Physiological diversity and trehalose accumulation in Schizosaccharomyces pombe strains isolated from spontaneous fermentations during the production of the artisanal Brazilian cachaça

Twenty-seven Schizosaccharomyces pombe isolates from seven cacha?a distilleries were tested for maximum temperature of growth and fermentation, osmotolerance, ethanol resistance, invertase production, and trehalose accumulation. Two isolates were selected for studies of trehalose accumulation under heat shock and ethanol stress. The S. pombe isolates were also characterized by RAPD-PCR. The isolates were able to grow and ferment at 41 degrees C, resisted concentrations of 10% ethanol, and grew on 50% glucose medium. Four isolates yielded invertase activity of more than 100 micromol of reducing sugar x mg(-1) x min(-1). The S. pombe isolates were able to accumulate trehalose during stationary phase. Two isolates, strains UFMG-A533 and UFMG-A1000, submitted to a 15 min heat shock, were able to accumulate high trehalose levels. Strain UFMG-A533 had a marked reduction in viability during heat shock, but strain UFMG-A1000 preserved a viability rate of almost 20% after 15 min at 48 degrees C. No clear correlation was observed between trehalose accumulation and cell survival during ethanol stress. Strain UFMG-A1000 had higher trehalose accumulation levels than strain UFMG-A533 under conditions of combined heat treatment and ethanol stress. Molecular analysis showed that some strains are maintained during the whole cacha?a production period; using the RAPD-PCR profiles, it was possible to group the isolates according to their isolation sites.     

乙醇浓度对提取灵芝三萜含量的影响及提取物抗前列腺癌细胞LNCaP的活性

本研究考察乙醇浓度对灵芝子实体中三萜成分提取的影响及提取物抗前列腺癌细胞LNCaP的增殖活性和迁移能力,为提取灵芝三萜时乙醇浓度的选择和灵芝抑制前列腺相关疾病产品开发提供依据。研究采用高效液相（HPLC）色谱法测定了灵芝醇提物中目前具有代表性的9种三萜的含量;并用抑制前列腺癌细胞LNCaP的细胞活性和细胞迁移能力的实验研究灵芝提取物的抗前列腺癌的作用。HPLC测定结果显示,当乙醇浓度为50%-80%时,9种灵芝三萜的含量随着乙醇浓度的升高而增大;当乙醇浓度继续升高至95%时,提取的三萜含量反而有所降低。细胞实验结果显示,4个浓度乙醇的灵芝提取物都具有良好的抑制LNCaP肿瘤细胞增殖作用和阻滞LNCaP细胞迁移能力。因此在使用乙醇提取灵芝三萜时,乙醇浓度升高有利于三萜的提取;在生产灵芝抗前列腺相关产品时,提取灵芝子实体中抗前列腺增生的活性成分时选用80%的乙醇较为适宜。