黄绿蜜环菌菌丝体多糖分离纯化与含量测定

分离纯化黄绿蜜环菌菌丝体粗多糖,提高多糖含量,以进一步应用于工业化生产。采用DEAE-52纤维素层析柱分离纯化黄绿蜜环菌菌丝体多糖,依次用水、0.1 mol/L Na Cl、0.2 mol/L Na Cl、0.4 mol/L Na Cl、0.8 mol/L Na Cl进行洗脱;苯酚-硫酸法测定纯化后多糖含量,测定波长490nm,葡萄糖浓度与吸光度的回归方程为A=6.3137X+0.2156R2=0.9909。黄绿蜜环菌菌丝体水体粗多糖经DEAE-52纤维素层析柱分离纯化后,多糖含量达到99%以上。经过中试试验验证提取及分离纯化工艺适用于工业化生产。     

聚酰胺树脂分离纯化芦荟多糖的研究

文章采用聚酰胺吸附树脂,采用水、0．2M／LNaCl溶液、5％NaOH水溶液为洗脱剂梯度洗脱分离纯化,得到了三个芦荟多糖组分,通过IR、UV、GC等手段对其进行了分析。以甘露糖为标准,采用硫酸．苯酚法,测得水洗和NaOH水溶液洗脱所得多糖含量分别为90．10％和93．28％;以木糖、甘露糖、葡萄糖、半乳糖为单糖标准的GC分析结果显示：水洗所得多糖主要以甘露糖为主,同时含有少量的葡萄糖。     

地木耳多糖的提取与纯化研究

对地木耳采用水提醇沉法获得的地木耳多糖粗提取物,采用Sevage法脱蛋白质、醇沉,干燥得粗多糖,进一步用DEAE-52纤维素柱层析分离纯化,用纸色谱和琼脂糖凝胶电泳对洗脱组分进行纯度鉴定。结果表明：Sevage法脱蛋白7次可脱除94%的蛋白质,多糖得率为13.75%。DEAE-52纤维素柱层析后得到10种组分,浓缩干燥后得到白色粉末状多糖组分,每个组分经过纯度鉴定后均为单一的多糖。选择水和NaC l溶液为洗脱剂的温和条件分离纯化多糖效果较好。     

长白楤木根、茎、叶水溶性多糖的纯化及组成分析

长白楤木是一种具有药用价值的植物。本实验用热水浸提、醇析、Severge法除蛋白,水相透析得到粗多糖。粗多糖经过DEAE-纤维素柱层析洗脱,再经SephadexG-200葡聚糖凝胶柱层析后,得到4种纯多糖,命名为根(gNaCl)茎(JH2O)茎(JNaCl)叶(YNaCl)。红外光谱和其完全水解的高效液相色谱分析表明,4种多糖不含硫酸基团和乙酰基,具备β-糖苷键以及糖类特征吸收峰。其中,根的NaCl洗脱多糖组分为D-半乳糖醛酸、D-果糖、D-葡萄糖,茎的水洗脱液多糖组分为D-甘露糖、D-葡萄糖,茎的NaCl洗脱多糖组分D-葡萄糖、D-半乳糖,叶的洗脱多糖组分为D-半乳糖醛酸和D-山梨糖。     

复方多糖口服液的制备及质量研究

目的制备复方多糖口服液,并进行定量分析,对各项指标进行检测。方法采用水提醇沉法制备复方多糖口服液,薄层色谱法进行定性鉴别,苯酚-硫酸法测定含量。结果检测波长为490 nm,在4.86～24.7mg范围内多糖呈线性关系,加样回收率为99.84%。结论本法简便易行,结果稳定,可做复方多糖口服液中多糖的含量测定方法。     

皱皮木瓜多糖的分离、纯化及抗氧化活性研究

为了研究木瓜多糖的提取、分离、纯化与抗氧化活性,采用水提醇沉法提取皱皮木瓜中的多糖,得多糖Ⅰ;利用Sevag法除去多糖中的蛋白质后得多糖Ⅱ;以30%H_2O_2脱除色素后再次醇沉得到精制多糖Ⅲ;透析除去小分子后利用AB-8大孔树脂进行分离以水、30%、50%、70%和95%乙醇洗脱,其中水洗脱部分多糖为Ⅳ。用苯酚-硫酸法测定多糖含量。多糖Ⅰ得率为9.83%,多糖含量(纯度,下同)为64.45%;脱蛋白后多糖Ⅱ中多糖含量为78.23%;经脱色后多糖Ⅲ含量达88.39%;大孔树脂水洗脱部分多糖Ⅳ含量为89.74%。以DPPH(2,2-二苯基-1-苦肼基)清除率和Fe~(3+)还原力方法测定木瓜多糖的抗氧化活性,木瓜多糖均体现出一定的抗氧化作用,呈浓度依赖性增强,其中多糖Ⅰ、Ⅱ表现出更好的作用。     

白花蛇舌草多糖的分离提取及含量测定

采用热水浸提法提取白花蛇舌草水溶性多糖,薄层层析法鉴定其多糖的单糖组成。通过正交试验优选浸提显色条件,以苯酚-硫酸法制得有色糖醛衍生物,用分光光度法在490nm波长处测定吸光度,其曲线方程为Y=0．01361x-0．08161,相关系数r=0．9997。结果表明,白花蛇舌草多糖由鼠李糖、葡萄糖、半乳糖及甘露糖等组成,含量为15．10％,回收率达95．68％。     

猴头多糖HEP-2化学成分研究

猴头菌(Hericium erinaceus (Bull.) Pers．)的菌丝体培养物经水溶、乙醇沉淀、除蛋白等步骤得到猴头多糖(HEP),利用柱层析从HEP中分离得到两个组分HEP-1和HEP-2。用高效液相凝胶法(HPGPC)鉴定了HEP-2的纯度及分子量,通过有机元素分析、IR、ICP等多种分析方法确定HEP-2为一种硫酸化复合多糖,首次确定了猴头多糖的基本化学组成。     

茯苓中多糖的提取及含量测定

采用水煎煮、稀碱浸提茯苓中的多糖,通过正交试验优选工艺条件。结果表明,稀碱浸提茯苓中的多糖的工艺较为合理,其操作简单,提取时间短,收率较高．为测定提取液中的多糖含量,以苯酚—硫酸法,制得有色糖醛衍生物,用分光光度法,在490nm波长处测定吸光度,回归方程线性关系好。方法简单易行、稳定、快速。     

超滤法提取裂褶菌胞外多糖几个主要参数的研究

对超滤法提取裂褶菌胞外多糖进行了研究,采用“二次回归旋转正交组合设计”得出超滤法提取裂褶菌胞外多糖的最优条件为:压力0.11MPa,温度55.20℃,pH4.01。在上述条件下超滤速度达31.92ml/min,制得的裂褶菌胞外多糖是以D-葡聚糖为主的混合多糖,其纯度为75.13%,收率可达82.06%。     

Purification and partial characterization of cellular retinoic acid-binding protein from human placenta

Cellular retinoic acid-binding protein (CRABP) has been purified to homogeneity from human placenta by a series of procedures, including acetone powder extraction, gel filtration on Sephadex G-50, and ion-exchange chromatography on DEAE-cellulose and on SP-Sephadex. Cellular retinol-binding protein (CRBP) was isolated concurrently. CRABP was purified 75,400-fold, based on total soluble acetone powder extract of placenta. The protein is a single polypeptide chain with a molecular mass of 14,600 Da, estimated by sodium dodecyl sulfate (SDS) gel electrophoresis or gel filtration, and has an isoelectric point of 4.78 (apo-CRABP, 4.82). On analysis of absorption and fluorescence spectra, the protein was seen to exhibit an absorption peak at 350 nm, fluorescence excitation maxima at 350 and 370 nm, and a fluorescence emission maximum at 475 nm. Human CRABP was immunologically distinct from human CRBP and serum retinol-binding protein.     

Does ionic strength affect the configuration of aquatic humic substances,as indicated by gel filtration?

SUMMARY. 1. The effect of ionic strength on the configuration of aquatic humic substances was studied by gel filtration and dialysis of water from small. Finnish forest lakes of varying colour.
2. Sephadex gel filtration of water from the most humic lake gave similar elution profiles of UV absorbance and dissolved organic carbon (DOC). Gel filtration of unconcentrated samples from all three lakes under natural conditions of ionic strength (I = 1.7 × 10⁻⁴) and pH (5.5–6.0) gave similar fractionation of humic substances, despite their widely differing colour (30–350 mg Pt l⁻¹) and DOC (5–25mg C l⁻¹).
3. Increasing the ionic strength by two orders of magnitude caused considerable retardation on the Sephadex columns of the humic substances, suggesting a decrease in their molecular size and/or shape.
4. Dialysis experiments strongly indicated that ionic strength-induced changes in the configuration of the aquatic humic substances are indeed real. Hence it is probable that the elution behaviour of aquatic humic substances on Sephadex gels has previously been wrongly attributed to ionic strength-dependent interactions between the gel and the humic substances.     

The use of fluorescence spectroscopy for identification of A- and B-thrombin chains during their chromatographic separation

The separation of A- and B-chains of human thrombin has been performed by gel filtration on Sephadex G-100 under the reduction of disulphide bonds with dithiothreitol. Identification of A- and B-chains has been provided by measurements of the fluorescence intensity of fractions at 310 nm and 350 nm which are near the maximum positions of tyrosine and tryptophan fluorescence, respectively. The appearance of A-chain was monitored by an increase of the ratio of Ifl310/Ifl350 greater than 2. The fluorescence spectrum of A-chain has maximum position at 304 nm, which is characteristic of tyrosine fluorescence. The fluorescence spectrum of B-chain has maximum position at 347.5 nm which corresponds to fluorescence of tryptophan residues. The identification of A- and B-chains has been confirmed by the gel electrophoresis data.     

Comparative biochemistry of acid proteinase from animal origins

Acid proteinases from 17 tissues of 12 animal species were compared with respect to molecular weight, inhibition by pepstatin and activation by tripolyphosphate. Gel filtration of acid proteinases from protochordates and vertebrates showed a common elution profile and three peaks with mol. wts of -20,000, -45,000 and above 150,000 were detected with acid-denatured hemoglobin as substrate at pH 3.6. The main component of vertebrate acid proteinases was identified as cathepsin D. In the invertebrate acid proteinases, the elution profiles through gel filtration were characteristic to the tissues examined, and were not so distinct as those of vertebrates. Through a biochemical survey, the animal acid proteinase was discussed from a comparative point of view.     

Purification and characterization of a 28-kDa major protein from ginseng root

A major protein was isolated from ginseng root (Panax ginseng C.A. Meyer) using a combination of ammonium sulfate fractionation, gel filtration chromatography, ion-exchange FPLC, and fast performance liquid chromatofocusing. Electrophoretic and gel permeation chromatographic studies revealed that the major protein, GMP, is composed of two subunits of approximately 28 kDa. During purification, it was found that the elution profiles of GMP from gel filtration chromatography were significantly different, depending on the ionic strength of buffers used. GMP in a buffer of low ionic strength was isolated as a complex with carbohydrate, which could be only dissociated at high ionic strength. Carbohydrate composition in GMP detected by gas chromatography varied, depending on the isolation method of the protein from ginseng roots. These results suggest that carbohydrates are bound non-covalently to GMP whose amino acid composition analysis showed high amounts of acidic amino acids.     

两步串联层析法纯化鼠抗人CD80单克隆抗体4E5

采用阴离子交换与凝胶过滤两步串联层析法,纯化了小鼠腹水来源的CD80阻断型单克隆抗体4E5。腹水样品经离心、过滤预处理后,在Tris-HCl缓冲溶液(pH8.0, 50mmol/L)条件下上阴离子交换柱对目的单抗进行捕集,采用0-0.5 mol/L NaCl浓度分步洗脱;含目的单抗的洗脱馏分再上凝胶过滤柱纯化,用PB缓冲溶液（pH7.2, 20mmol/L）洗脱,获得目的单抗4E5,其生物学活性高、纯度大于95%,抗体总回收率达61%。     

Cryptomonad biliprotein: phycocyanin-645 from a Chroomonas species.

The properties of phycocyanin-645 from the fresh water cryptomonad Chroomonas spec. were investigated after the pigment was isolated and purified by a combination of differential ammonium sulphate fractionation, gel filtration chromatography and ammonium sulphate gradient elution. Phycocyanin-645 is characterized by absorption maxima at 645 nm, 584 nm, 369 nm, 275 nm and shoulders at 340 nm and 620 nm. The CD spectrum has a negative maximum at 645 nm and a positive maximum at 584 nm with a shoulder at 610 nm. The fluorescence emission spectrum is asymmetrical and shows a maximum at 660 nm and a shoulder at approximately 715 nm. The molecular weight of the native phycocyanin-645, estimated by gel filtration, is 45000 for all multiple pigment forms below. Phycocyanin-645 is heterogeneous as revealed by isoelectric focusing with pIs at 7.03, 6.17, 5.75, 5.25 and 4.88, respectively, the main bands lying at pI 7.03 and pI 6.17. This was confirmed by polyacrylamide gel electrophoresis; five pigment compoents differing in mobility were found. We propose the term "multiple pigment forms" for these five phycocyanin-645 modifications. Calibrated SDS gel electrophoresis shows phycocyanin-645 to consist of three subunits, two light chains (alpha1, alpha2), having molecular weights of 9200 and 10400, respectively, and one heavy chain (beta), having a molecular weight of 15 500. Suggesting a 1:1:2 ratio between the subunits, the quaternary structure of the pigment molecule is alpha1beta--alpha2beta1.     

Characterization of vertebrate collagenase activity by high-performance liquid chromatography using a synthetic substrate

The hydrolysis of the model collagenase substrate, 2,4-dinitrophenyl-Pro-Gln-Gly-Ile-Ala-Gly-Gln- -Arg by partially purified tadpole back-skin collagenase was monitored by separation of the substrate peptide from the product peptides 2,4-dinitrophenyl-Pro-Gln-Gly and Ile-Ala-Gly-Gln- -Arg by reverse-phase high-performance liquid chromatography. The method provides a sensitive, relatively rapid means of determination of collagenase activity using purified enzyme samples. It is not by itself, however, suitable for use with impure systems since the tissue culture medium from tadpole back skin was found to contain at least three peptidases which could be separated by gel filtration and which showed identical high-performance liquid chromatographic elution profiles using the octapeptide model substrate, but only one of which cleaved triple helical collagen.     

alpha-D-Fucosidase activity of human and pig kidney: a property of alpha-D-galactosidase.

α-d-Fucosidase activity was demonstrated in human and pig kidneys. Two forms of α-d-fucosidase were separated by gel filtration on Sephadex G-200. The elution profiles of α-d-fucosidases and α-d-galactosidases were identical upon gel filtration and isoelectric focusing. A comparison of the properties of α-d-fucosidase and α-d-galactosidase from human and pig kidney showed that both activities had similar pH optima and similar thermostability and were inhibited to the same extent by d-fucose, d-galactose, and d-galactono-(1→4)-lactone. These data suggest that the hydrolysis of both α-d-fucoside and α-d-galactoside are catalyzed by the same enzyme, α-d-galactoside (fucoside) hydrolase.     

An improved treatment of data for estimation of molecular weights by sephadex gel filtration

A method for analysis of elution data of proteins, obtained from Sephadex gel filtration experiments, is described. The relevant elution data from seven different proteins, with known molecular weights and Stoke's radii, were fitted into various equations relating elution parameters and molecular size parameters. It was observed that polynomial relationships represented elution data for proteins with a much greater degree of precision than linear equations. The validity of this procedure was also checked by analysing gel filtration data available in the literature and it was concluded that a better fit was obtained using polynomial relationships, provided a sufficiently large number of experimental points were available for numerical analysis. Using this method, values of 320,000 ± 7000 for the molecular weight, and (60 ± 0.4) × 10^?8 cm for the Stoke's radius of Neurospora NAD-specific glutamate dehydrogenase were calculated.