超声提取的麦冬多糖POJ-U1b的结构分析及AFM观测

本文利用超声提取麦冬多糖,经DEAE-52纤维素柱层析进行分离纯化,利用Sephadex G-150凝胶柱层析对得到的多糖组分POJ-U1b进行纯度鉴定,结果显示POJ-U1b为单一组分。利用气相色谱法、红外光谱法及核磁共振对POJ-U1b的结构进行了研究,利用原子力显微镜对不同浓度的POJ-U1b表面形貌进行了观测。结果显示,麦冬多糖POJ-U1b是由葡萄糖组成的葡聚糖,具有糖类物质的特征吸收峰,其糖环构型既有吡喃型又有呋喃型,糖链主要由→6)-α-D-Glcp(1→连接方式构成。AFM观测结果表明POJ-U1b具有高度分支的化学结构,糖链间通过糖单元间不同的链接方式衍生出许多环状和分支结构。随着多糖浓度的增大,POJ-U1b呈现出链状及片状粘连结构,这种现象可能与糖链间的范德瓦尔斯相互作用及糖链间氢键缔合有关。     

滑菇多糖wPNP-a1的制备及结构分析

利用传统热水浸提法提取滑菇粗多糖(wPNP),进而用季铵盐沉淀法和Sephadex G-150凝胶色谱柱层析对wPNP进行纯化,得滑菇多糖wPNP-a1.以高效液相色谱(HPLC)、傅里叶变换红外光谱(FIR)、气相色谱(GC)、核磁共振(NMR)、环境扫描电子显微镜(ESEM)和原子力显微镜(AMF)等方法分析wPNP-a1的结构,结果表明,滑菇多糖wPNP-a1的分子量为419310 Da,具有一般类多糖的特征吸收峰,糖苷键为β-型,不含乙酰基,由木糖、甘露糖、半乳糖和葡萄糖组成,其摩尔比为3.91∶2.77∶1.91∶1;wPNP-a1的分子间排斥力较大,分子间吸引力较小,在云母表面呈直链状分布,链宽23 nm左右,链高0.7～0.8 nm.     

旱莲草多糖的提取及其抗氧化活性研究

采用水/甲醇混合溶剂浸泡提取旱莲草的多糖组分,利用硅胶柱层析和中压液相色谱,对多糖进行分离纯化,纯化后的多糖组分经薄层层析分析发现,这些多糖主要组成单糖均为葡萄糖和果糖。旱莲草多糖在总还原能力、总抗氧化能力和FRAP抗氧化能力测试中均显示较好的活性,能够有效清除羟自由基和DPPH自由基,在1.5 mg/m L的浓度下,对羟自由基的清除效率为(12.33~56.81)%,粗多糖组分在1 mg/m L的浓度下对DPPH自由基的清除效率为(17.88~84.47)%,精分多糖组分在5 mg/m L的浓度下,对DPPH自由基的清除效率为(18.64~91.35)%;极性小的多糖抗氧化活性显著优于极性大的多糖,抗氧化活性最高可达后者的11倍之多;粗多糖组分的抗氧化活性一般优于纯化后的相应精多糖组分,这说明旱莲草的不同多糖分子之间可能存在协同抗氧化作用。     

海洋红细菌Lentibacter algarum胞外多糖的分离纯化及结构解析

从青岛潮间带的海水中分离得到的红细菌科细菌Lentibacter algarum的发酵液中提取胞外多糖,对其进行离子交换色谱分离纯化,得到水洗和0.1、0.5、1.0 mol/L NaCl溶液4个洗脱组分。对含量最高的0.1 mol/L NaCl洗脱组分La0.1进行进一步的凝胶排阻柱层析纯化,得到组分La0.1-1。通过化学测定和高效液相色谱(HPLC)、高效凝胶渗透色谱法(HPGPC)等分析方法对其理化性质、分子量、单糖组成、连接方式及初步结构进行研究。结果表明,La0.1-1总糖含量为66%,平均分子量为12.0 kDa。其单糖组成主要为半乳糖、甘露糖和氨基葡萄糖,比例为Gal∶Man∶GlcN=1.35∶1.1∶1.0。对La0.1-1进行气相色谱质谱联用(GC-MS)和一维核磁(1-D NMR)分析结果显示,La0.1-1的连接方式是以β构型为主,主要存在→2)-Manp(1→,→3)-Galp(1→连接,还存在少量的→4)-Galp(1→和→4)-Manp(1→等连接方式,表明该多糖以直链为主,还存在一定的分支,分支发生在→2)-Manp(1→的O-6位和→3)-Galp(1→的O-4或O-6位。氨基葡萄糖主要为→4)GlcN(1→和末端连接方式。核磁分析还显示La0.1-1存在一定的乙酰基取代,初步判断主要取代在氨基葡萄糖的N-2位上,也可能存在于甘露糖和半乳糖上。本研究是首次对Lentibacter属细菌的胞外多糖进行测定,获得了结构较为新颖的胞外多糖资源,为开发海洋多糖资源提供物质基础。     

桦褶孔菌多糖的分离纯化及清除自由基活性

利用水提醇沉法对桦褶孔菌Lenzites betulina粗多糖HZKJc进行提取,随后经Sevage法和酶法联合脱蛋白,再经DEAE‐Sephadex A‐25,G‐100柱层析纯化后得到多糖纯品HZKJv。通过高效液相色谱(HPLC)测定上述纯化多糖的纯度和相对分子量分布;经纸层析(PC)、气相色谱(GC)、红外光谱(IR)进行单糖组成分析;并研究其清除自由基活性。分离纯化后得到的HZKJv为纯多糖,相对分子质量约为11 687。当HZKJv溶液浓度为3.0mg/m L时,其DPPH自由基清除率可达82%;浓度为2.0mg/m L时,对?OH自由基的清除率可达85%。表明HZKJv对?OH与DPPH自由基存在很好的清除作用。     

苦楝果多糖的分离纯化及组成分析

利用水提醇沉法提取苦楝果实中的多糖,经DEAE-52柱层析分离,得到MP1、MP2和MP3三个多糖组分,用Sephadex G-100凝胶色谱柱对MP1进行纯化鉴定,结果显示为单一峰。借助气质联用仪,对苦楝粗多糖和组分MP1进行了成分分析。红外光谱分析表明苦楝多糖的单糖残基以吡喃环和呋喃环的形式存在。     

碱液提取箬叶多糖的纯化及其结构性质的研究

通过高碘酸氧化、Smith降解、部分酸水解分析、NMR分析等多种方法对以不同浓度的NaOH溶液箬叶中提取的两种多糖FⅢ-a及FⅣ-a进行了研究,结果表明二者均具有多分枝结构,FⅢ-a主链以α(1→3)连接的木糖为主,分子侧链由半乳糖、阿拉伯糖、葡萄糖醛酸构成,葡萄糖醛酸主要位于分子的末端;FⅣ-a主链由α(1→3)木糖和β(1→6)半乳糖构成,以阿拉伯糖、葡萄糖醛酸组成侧链,葡萄糖醛酸主要位于分子的末端.     

皂荚半乳甘露聚糖胶分级醇沉及表征

使用不同浓度乙醇和异丙醇分别对皂荚半乳甘露聚糖胶水溶液进行分级沉降,沉淀物用凝胶渗透色谱(GPC)和高效液相色谱(HPLC)等进行表征.结果表明,异丙醇可在较小浓度下更快沉降皂荚多糖胶,当异丙醇溶液浓度为28.6％ (V/V)时,沉淀物中半乳甘露聚糖浓度达到12.50％(w/w);随着醇浓度上升,沉降组分半乳甘露聚糖得率呈增加趋势,且在后期增加幅度最大,多糖最高得率可达80％,纯化后皂荚多糖胶(GSG)表现出较高的甘露糖/半乳糖(M/G)之比值,在异丙醇沉降中表现更加明显(低浓度的异丙醇达到最高的M/G =4.1);低浓度醇沉主要得到大分子组分,随乙醇浓度增加组分分子量明显降低,多糖胶更加均匀,而在异丙醇沉降后期均一性有所下降.     

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。     

野木瓜水溶性多糖的分离纯化及抗氧化活性研究(英文)

采用水提醇沉法获得了水溶性野木瓜粗多糖(CCCPs),经过脱蛋白、脱色及透析,得到了野木瓜精多糖(FCCPs),再经DEAE-纤维素和Sepharose CL-6B柱层析得到均多糖组分(CCP1)。高效液相色谱法测定CCP1的单糖组成为:鼠李糖、阿拉伯糖、果糖、甘露糖、葡萄糖,其摩尔比为0.034∶0.228∶0.045∶0.055∶0.638。体外抗氧化实验结果表明:不同纯度的野木瓜多糖都具有一定的抗氧化活性,并且随着多糖浓度增加,抗氧化活性增强,野木瓜粗多糖(CCCPs)对·OH和O-·2的清除能力比精多糖(FCCPs)和均多糖(CCP1)强。     

Net charge affects morphology and visual properties of ovalbumin aggregates

The effect of ovalbumin net charge on aggregate morphology and visual properties was investigated using chromatography, electrophoresis, electron microscopy, and turbidity measurements. A range of differently charged ovalbumin variants (net charge ranging from -1 to -26 at pH 7) was produced using chemical engineering. With increasing net charge, the degree of branching and flexibility of the aggregates decreased. The turbidity of the solutions reflected the aggregate morphology that was observed with transmission electron microscopy. Increasing the stiffness of the aggregates transformed the solutions from turbid to transparent. Artificially shielding the introduced net charge by introducing salt in the solution resulted in an aggregate morphology that was similar to that for low-net-charge variants. The morphology of heat-induced aggregates and the visual appearance of the solutions were significantly affected by net charge. We also found that the morphology of ovalbumin aggregates can be rapidly probed by high-throughput turbidity experiments.     

Trichomes in Camptotheca acuminata Decaisne (Nyssaceae): Morphology,distribution, structure,and secretion

Camptotheca acuminata is a main source of the anti-cancer drug camptothecin (CPT). In this species, several studies have observed non-glandular trichomes (NGTs) and glandular trichomes (GTs). It has been assumed that GTs contain CPT, yet this has not been proven and no information is available on the accumulation of other secondary metabolites. The objective of this study was to describe the morphology, distribution and structure of C. acuminata trichomes and to investigate the chemical nature of the substances secreted by GTs. Light and fluorescence microscope, scanning electron microscope (SEM) and transmission electron microscope (TEM) were used to determine the morphology, distribution and structure of GTs and NGTs. Thin layer chromatography (TLC) and high-performance liquid chromatography (HPLC) analyses were carried out to confirm the presence of CPT in GTs, and histochemical tests were performed to investigate the presence of other secondary metabolites. C. acuminata possesses two types of GTs (GT1 and GT2), which differ in terms of their morphology, pattern of distribution and accumulated substances. The chemical analyses demonstrated that both GT1 and GT2 accumulate CPT. Histochemical analysis showed that phenols accumulate in the vacuole of GT2s. No isoprenoids were detected in GTs.     

Low molecular weight polyethylenimine grafted N-maleated chitosan for gene delivery: properties and in vitro transfection studies

To develop chitosan-based efficient gene vectors, chitosans with different molecular weights were chemically modified with low molecular weight polyethylenimine. The molecular weight and composition of polyethylenimine grafted N-maleated chitosan (NMC-g-PEI) copolymers were characterized using gel permeation chromatography (GPC) and (1)H NMR, respectively. Agarose gel electrophoresis assay showed that NMC-g-PEI had good binding ability with DNA, and the particle size of the NMC-g-PEI/DNA complexes was 200-400 nm, as determined by a Zeta sizer. The nanosized complexes observed by scanning electron microscopy (SEM) exhibited a compact and spherical morphology. The NMC-g-PEI copolymers showed low cytotoxicity and good transfection activity, comparable to PEI (25 KDa) in both 293T and HeLa cell lines, except for NMC 50K-g-PEI. The results indicated that the molecular weight of NMC-g-PEI has an important effect on cytotoxicity and transfection activity, and low molecular weight NMC-g-PEI has a good potential as efficient nonviral gene vectors.     

Purification of a factor from the granules of a rat natural killer cell line (RNK) that reduces tumor cell growth and changes tumor morphology. Molecular identity with a granule serine protease (RNKP-1).

We have purified a protein from the granules of the rat NK leukemia cell line (RNK) that is cytostatic to a variety of tumor cells. This protein shows no species specificity because certain tumor cell lines of mouse, rat, and human origin were equally sensitive to its growth inhibitory effects. Treatment of sensitive cells resulted in a rounding of the cells followed by homotypic aggregation into large aggregates. The granule protein was distinct from cytolysin, Na-Cbz-Lys-thiobenzylester-esterase, or leukolexin. It had a molecular mass of 29 to 31 kDa, bound strongly to heparin, was inactivated by heating at 70 degrees C for 5 min or reduction, but was stable to trypsin treatment. By using molecular sieve chromatography, heparin agarose chromatography, and reverse phase HPLC, this protein was purified to homogeneity. The first 33 amino acids of the N-terminal amino acid sequence showed complete identity to the sequence predicted from a rat serine protease gene recently cloned and designated RNKP-1. Therefore we have purified a novel serine protease and demonstrated that it has effects on the growth and morphology of certain tumor cells. Other serine proteases that were structurally related and have substantial homology with RNKP-1 at the amino acid level showed neither growth inhibitory properties nor affected the morphology of the tumor target cells we used.     

Galactosyl conjugated N-succinyl-chitosan-graft-polyethylenimine for targeting gene transfer

Through incorporating lactobionic acid (LA) bearing a galactose group to N-succinyl-chitosan-graft-polyethylenimine (NSC-g-PEI), NSC-g-PEI-LA copolymers were synthesized as gene vectors with hepatocyte targeting properties. The molecular weight and composition of NSC-g-PEI-LA copolymers were characterized using gel permeation chromatography (GPC) and (1)H nuclear magnetic resonance spectroscopy ((1)H NMR) respectively. Agarose gel electrophoresis assays showed good DNA binding ability of NSC-g-PEI-LA, and the particle size of the NSC-g-PEI-LA/DNA complexes were between 150 and 400 nm as determined by a Zeta sizer. The NSC-g-PEI-LA/DNA complexes observed by scanning electron microscopy (SEM) exhibited a compact and spherical morphology. The zeta potentials of these complexes were increased with the weight ratio of NSC-g-PEI-LA/DNA. NSC-g-PEI-LA has a lower cytotoxicity than PEI (25 kDa) and the toxicity decreased with increasing substitution of LA. The transfection efficiency of different complexes was evaluated by luciferase assay. Compared with PEI (25 kDa) and NSC-g-PEI/DNA, NSC-g-PEI-LA showed good transfection activity and cell specificity to HepG2 cells. The results suggested that NSC-g-PEI-LA has the potential to be used as a safe and effective targeting gene vector.     

Isolation and characterization of α-N-acetylβ-endorphin(1–26) from the rat posterior/intermediate pituitary lobe

An extract from 50 rat posterior intermediate pituitaries was fractionated by gel filtration followed by cation exchange chromatography. α-N-Acetylated derivatives of β-endorphin-like molecules were detected with a specific radioimmunoassay for α-N-acetylβ-endorphins. Six peaks of α-N-acetylβ-endorphin-like immunoreactivity were observed in the cation exchange chromatography fractions. One of these peaks was purified to homogeneity using reverse phase high performance liquid chromatography (RP-HPLC). The isolated peptide was characterized by tryptic digestion followed by RP-HPLC and by amino acid analysis. The results showed that the isolated peptide was α-N-acetylβ-endorphin(1–26) with an oxidized methionine residue at position 5. Two previously unrecognized α-N-acetylβ-endorphin derivatives were also observed during the isolation procedure.     

菰黑粉菌的分离鉴定及其发酵液中植物激素的检测

在显微镜下观察孢子形态,并观察单菌落的形态和菌株的微观形态,PCR扩增其ITS-5.8S rDNA序列,测序并在NCBI中进行同源比较,确定其种属。液体培养该菌株,通过高效液相色谱法检测发酵液中植物激素。结果表明：用菌落形态与孢子形态鉴定和分子生物学鉴定的方法,对茭白中分离的一个菌株鉴定为菰黑粉菌,且在其发酵液中检测到植物激素IAA、ABA和GA3,其中IAA含量为0.1306mg/L,ABA含量为0.01367mg/L。     

Purification and characterization of a trypsin inhibitor from seeds of Murraya koenigii

A protein with trypsin inhibitory activity was purified to homogeneity from the seeds of Murraya koenigii (curry leaf tree) by ion exchange chromatography and gel filtration chromatography on HPLC. The molecular mass of the protein was determined to be 27 kDa by SDS-PAGE analysis under reducing conditions. The solubility studies at different pH conditions showed that it is completely soluble at and above pH 7.5 and slowly precipitates below this pH at a protein concentration of 1 mg/ml. The purified protein inhibited bovine pancreatic trypsin completely in a molar ratio of 1:1.1. Maximum inhibition was observed at pH 8.0. Kinetic studies showed that Murraya koenigii trypsin inhibitor is a competitive inhibitor with an equilibrium dissociation constant of 7 x 10(-9) M. The N-terminal sequence of the first 15 amino acids showed no similarity with any of the known trypsin inhibitors, however, a short sequence search showed significant homology to a Kunitz-type chymotrypsin inhibitor from Erythrina variegata.     

Monocytes treated with ciprofloxacin and oxyLDL express myristate,priming atherosclerosis

Antibiotics are essential in many life‐threatening diseases. On the other hand, improper use of antibiotics can be disastrous. Cell morphological changes were observed in the ciprofloxacin‐treated cells starting at 48 hours. Changes in cell morphology were continuously observed up to 14 days, which showed gradual morphological changes from monocyte to plaque‐like cells at day 12, and foam cell, which is an intermediate stage in atherosclerosis was observed at day 8, which was confirmed with Oil Red O staining. Flow cytometry data revealed that oxidized LDL (oxyLDL)‐induced cells showed 60.16% of CD64 (proinflammatory macrophage markers) and no expression of CD23 (anti‐inflammatory macrophage markers), whereas ciprofloxacin‐treated cells expressed 67.97% of CD64 and 13.78% of CD23. Chemokine antibody array analysis revealed that ciprofloxacin exposed cells showed a proinflammatory role (ENA78, Eotaxin1, Eotaxin2, IP‐10, MIG, MIP‐3β, SDF‐1β, TECK, CXCL16, and Fractalkine). Liquid chromatography with tandem mass spectrometry (LC‐MS/MS) revealed that myristic acid was incorporated into a protein with 68 kDa molecular mass in exposing oxyLDL‐induced monocytes with ciprofloxacin, which could be a reason for the observed foam cells and in vitro plaque formation. As myristic acid primes atherosclerosis, it is better to limit the intake of antibiotics like ciprofloxacin for common illness, specifically the high‐risk patients, which may contribute to atherosclerosis.     

产黄芩苷内生真菌的筛选与鉴定

从药用植物黄芩根、茎、叶和花中分离得到17株内生真菌,其发酵液对10种指示菌进行抑菌活性测定,并通过薄层色谱(TLC)和高效液相色谱(HPLC)对所分离到的内生真菌代谢产物进行分析。结果显示,所分离到的内生真菌中12株至少对一种指示菌有抑菌活性,其中3株(G2、J4和J5)具有较广的抑菌作用。从黄芩茎和花中分离得到的2株内生真菌J1、H3可以在人工培养基中产生黄芩活性成分——黄芩苷,结合菌落形态特征、显微观察及ITS序列分析,初步鉴定这2株菌株均属于青霉菌属。