Optimization of Polysaccharides Extraction from Physalis alkekengi L.Peel and Its Effect on the Expression of Inflammation-Related Proteins in SW620 Cells

Objective:To establish an optimized aqueous extraction process for polysaccharides from Physalis alkekengi L.peel and to preliminarily explore its in vitro anti-inflammatory activity against colorectal cancer SW620 cells.Methods:A single-factor test combined with orthogonal test analysis was used to evaluate the effects of the material-to-liquid ratio,extraction temperature,and extraction time on the yield of polysaccharides from Physalis alkekengi L.peel.The antioxidant activity of the polysaccharides was assessed by analyzing their free radical scavenging ability in vitro,and the anti-inflammatory effect was evaluated using SW620 cells.Results:The optimal extraction conditions were a material-to-liquid ratio of m(g):V(mL)=1:30,an extraction temperature of 100℃,and an extraction time of 40 minutes,with a predicted polysaccharide yield of 25.7%.The polysaccharides from Physalis peruviana peel effectively scavenged DPPH,superoxide anion,and hydroxyl radicals.After treatment with Physalis peruviana polysaccharides,the levels of IL-1β,IL-18,and TNF-αin the cell culture medium were significantly reduced,and the phosphorylation level of P65 protein in SW620 cells was decreased.Conclusion:This extraction method is stable and reliable,and the prepared Physalis alkekengi L.polysaccharides exhibit significant in vitro antioxidant and anti-inflammatory activities.This study provides a theoretical basis for developing drugs for the prevention and treatment of colorectal cancer.

松乳菇Lactarius deliciosus与红汁乳菇L. hatsudake子实体内可培养细菌多样性的研究

分离松乳菇Lactarius deliciosus与红汁乳菇L.hatsudake子实体内可培养的细菌,进行分类鉴定,为后期细菌与乳菇的相互作用研究奠定基础。利用传统平板法分离培养乳菇子实体内的细菌,进行16S r DNA片段的扩增和测序及系统发育树的构建和物种多样性分析。从松乳菇中获得66株细菌,红汁乳菇中获得48株细菌。松乳菇子实体内细菌隶属于3个门6个属10个种,其中变形菌门的γ-变型菌纲为优势类群,占细菌总数的77.28%,β-变形菌纲的Pandoraea和拟杆菌门的金黄杆菌属Chryseobacterium分别占细菌总数的12.12%和10.6%。红汁乳菇子实体内共分离、鉴定出细菌3个门7个属7个种,其中变形菌门中的γ-变型菌纲为优势类群,占细菌总数的77.09%,厚壁菌门的芽胞杆菌属Bacillus和拟杆菌门的金黄杆菌属Chryseobacterium分别占细菌总数的14.58%和8.33%。松乳菇与红汁乳菇子实体内存在一定种类和数量的细菌,其中荧光假单胞菌Psedomonas fluorescens和美洲爱文氏菌Ewingella americana为优势细菌。

Extraction and purification of the polysaccharides in Hippohaere rhamnoides L.

The different extraction technology and purification technology ofHippohpae rhamoides polysaccharides were researched in the paper. The best method of papain extraction were obtained, the ratio of papain 2%, pH at 5.5, temperature at 45℃ and extraction time of 20 min were suitable for papain extraction. The highest content of Hippohpae rhamoides polysaccharides was 44.28 mg·g^-1. The optimum process of ultrasonic extraction were obtained, namely extracted for 55 min at 480 W with the material ratio of 1：20. The highest content of Hippohpae rhamoides polysaccharides was 48.63 mg·g^-1. The results showed that the ultrasonic and papain extraction together was the best method, the content was 54.30 mg·g^-1. After the removing protein, pigment and dialysis. Two fraction were separated from the purified Hippohpae rhamoides by DEAE-cellulose chromatography, the main fraction was collected finally. The fraction was identified by Sepharose CL-4B gel filtration. Ultraviolet spectrometry, freeze-thawing analysis showed that fraction was purified. Its molecular weight was probably 109.4 ku.

Isolation,purification,structural characteristic and antioxidative property of polysaccharides from A.cepa L.var.agrogatum Don

Allium cepa L.var.agrogatum Don(ALAP)is commonly consumed in China as well as other regions and has various beneficial health effects.A novel acidic polysaccharide(named ALAP-21)was obtained from ALAP by ultrasonic and microwave-assisted extraction and purification using DEAE-52 anion exchange and Sephadex G-100 columns.The monosaccharide composition,structural and antioxidative properties of ALAP-21 were investigated by GC–MS chromatography,FT-IR and NMR spectroscopies and three antioxidative activity tests in vitro.The results showed that ALAP-21 was a heteropolysaccharide composed of glucose,galacturonic acid,mannose,galactose,arabinose,rhamnose,xylose,fructose and glucuronic acid with a relative molar ratio of 26.282:27.546:11.400:4.781:2.467:2.445:3.622:1.106:1.753,owning(1→4)-β-d-Glcp,(1→4)-β-d-GalAp6Me3OA,(1→4)-β-d-Galp6OMe,(1→2)-β-l-Rhap,(1→4)-β-d-Manp glycosidic linkages.(1→4)-β-d-Glcp and(1→4)-β-d-GalAp6Me3OA residues might be the main components of the sugar chain backbone of ALAP-21.Furthermore,ALAP-21 exhibited high potential for DPPH radicals(82.02%),hydroxyl radicals(53.33%)and superoxide anion radicals(50.28%).These results provide a reference for further research and rational development of ALAP polysaccharides.

Study on Extraction of Polysaccharides and Antioxidant Activity of Sagittaria sagittifolia L.Polysaccharide

[Objective] This experiment was conducted to investigate the optimal ex- traction process and antioxidant activity of polysaccharide in Sagittaria sagittifo/ia L. [Method] The effects of extraction temperature, extraction time, extraction times and material-to-liquid ratio on content of polysaccharides in S. sagittifo/ia were investi- gated. On the basis of single-factor tests, an orthogonal test was carried out to de- termine the optimal process parameters. In addition, the in-vitro antioxidant activity of S. sagittifo/ia polysaccharides was evaluated on the basis of determination of their scavenging activities against DPPH and hydroxyl free radicals and reducing power. [Result] The results showed that the optimized process parameters were as follows： material-to-liquid ratio at 1：40 （g/ml）, extraction temperature at 90 ℃, extrac- tion time of 4 h, and extraction times of 3 times. Under this condition, the content of S. sagittifo/ia polysaccharides was 29.32%. For 1 mg/ml S. sagittifo/ia polysaccha- rides, the scavenging ratios of DPPH and hydroxyl free radicals were 70.62% and 35.82%, respectively, and in the determination of reducing power, the absorbance at 700 nm was 0.453 1. [Conclusion] It could be concluded that S. sagittifolia polysac- charides have strong antioxidant capacity to have free radicals scavenged in vitro.

Immunoregulatory polysaccharides from Apocynum venetum L.flowers stimulate phagocytosis and cytokine expression via activating the NF-κB/MAPK signaling pathways in RAW264.7 cells

Two immunomodulatory polysaccharides(Vp2a-Ⅱ and Vp3) were isolated and identified from Apocynum venetum L. flowers, and their innate immune-stimulating functions and working mechanisms were evaluated in RAW264.7 cells. Both the level of released nitric oxide(NO) and expression of inducible nitric oxide synthase(iNOS) m RNA were significantly enhanced in the RAW264.7 macrophages cells treated by Vp2a-Ⅱ and Vp3. Vp2a-Ⅱ(100–800 μg/m L) and Vp3(400 μg/mL) could significantly increase the phagocytic activity of RAW264.7 cells and the secretion and m RNA expression of TNF-α and IL-6 in a concentrationdependent manner through affecting mitogen-activated protein kinase(MAPK) activity and nuclear factor κB(NF-κB) nuclear translocation. Vp2a-Ⅱ might activate the MAPK signaling pathways and induce the nuclear translocation of NF-κB p65, whilst Vp3 likely activated the NF-κB and MAPK signaling pathways without influencing the p38 MAPK route.

Determination of the Composition and Content of Lycium barbarum Polysaccharides by Chromatography

Lycium barbarum polysaccharides （LBP） were obtained after Lycium barbarum, the study object, went through chloroform-methanol degreasing, water extraction and alcohol precipitation, acetone and ether wash and dry, hydrogen peroxide bleaching, deproteinization by savage method. Determined LBP molecular weight by HPLC was 73950-138090Da. And the monosaccharides in LBP determined by IC mainly included glucose, arabinose, rhamnose, galactose, xylose and mannose, in which glucose, arabinose and galactose had larger contents, while the contents of other monosaccharides were low.

PURIFICATION AND PARTIAL CHARACTERIZATION OF POLYSACCHARIDES FROM THE POLLEN OF TYPHA ANGUSTIFILIA L.

Three polysaccharides TAA,TAB and TAC were purified from the pollen of Typha An- gustifilia L..TAA is mainly composed of α-L-arabinofuranose,β—D—galactose and α—D—galac- turonic acid.TAB and TAC contain the backbone mainly composed of (1→5)—linked—L—arabi- nosyl residue.

Optimization of extraction process of Amomum longiligulare T.L.Wu polysaccharides by response surface methodology

Objective:To optimize the technological conditions of ultrasonic-assisted extraction of Amomum longiligulare T.L.Wu polysaccharides and increase the yield of polysaccharides.Methods:The polysaccharide was prepared by water extraction and alcohol precipitation method,and the yield of polysaccharide was taken as an index.The effects of ultrasonic time,extraction times,ultrasonic temperature,and water-to-material ratio on polysaccharide yield were investigated through a single factor experiment.Combined experimental design and response surface analysis were used to optimize the extraction process of Amomum longiligulare T.L.Wu polysaccharides.Results:The optimum conditions of ultrasonic-assisted extraction were determined as follows:extraction time of 29 min,three extraction times,water bath extraction temperature of 68℃,water-to-material ratio of 15:1;under these conditions,the polysaccharide yield was 10.69%.Conclusion:The results are close to the predicted values of the model.This optimization test is effective and feasible,and provides a reference for the related research of Amomum longiligulare T.L.Wu.

Effects of Polysaccharides from Dicliptera chinensis(L.)Nees.on TLR4/NF-κB Signaling Pathway in Cell Model of Non-alcoholic Fatty Liver

[Objectives]To observe the effects of polysaccharides from Dicliptera chinensis(L.)Nees.on the expression of TLR/NF-κB pathway related proteins in HepG2 cells induced by oleic acid,and to explore the possible mechanism of polysaccharides from D.chinensis(L.)Nees.in the treatment of non-alcoholic fatty liver disease(NAFLD).[Methods]HepG2 cells were induced with oleic acid to establish a non-alcoholic fatty liver cell model.After intervention with 0.25 and 0.5 mg/mL of D.chinensis(L.)Nees.polysaccharides,the ALT and AST activity and TG and TC contents were detected with kits,and the changes in the expression of CDK5,TLR4,p-NF-κB and NF-κB were analyzed using Western-blotting.[Results]In the HepG2 cells induced with oleic acid,the ALT and AST activity increased significantly,the TG and TC contents increased significantly,and the expression levels of CDK5,TLR4 and p-NF-κB proteins up-regulated significantly.In the HepG2 cells intervened with D.chinensis(L.)Nees.polysaccharides,the activity of ALT and AST,the contents of TG and TC,and the expression levels of CDK5,TLR4 and p-NF-κB proteins all reduced significantly.[Conclusions]Polysaccharides from D.chinensis(L.)Nees.may interfere with NAFLD by inhibiting the TLR4/NF-κB pathway.

Toxicity Experiment of Polysaccharides from Dicliptera chinensis( L. ) Juss.

[Objectives] To study the acute toxicity,mutation,and feeding experiment of polysaccharides from Dicliptera chinensis( L.)Juss.,and assess its safety. [Methods]In accordance with Procedures for toxicological Assessment on Food Safety,acute toxicity to mice,micronucleus experiment for bone marrow cell in mice,sperm shape abnormality test in mice,and Ames experiment were carried out,and 30 d feeding experiment was detected for rats. [Results] when the maximum tolerance dose( MTD) of mice was higher than 20 g/kg,at polysaccharides from Dicliptera chinensis( L.) Juss. ≤10 g/kg,there was no statistical difference between the control group and mouse bone marrow cell micronucleus and sperm shape abnormality test group; Ames experiment indicated that the number of revertant colonies of different dose groups of polysaccharides from Dicliptera chinensis( L.) Juss. with and without S9 did not exceed two times of the blank control group; rat 30 d feeding experiment indicated that when polysaccharides from Dicliptera chinensis( L.) Juss. ≤ 3. 2 g/kg,rats grew and developed well,and there was no statistical difference with the control group in blood,multiple organ biochemical indicators and liver tissue. [Conclusions]Polysaccharides from Dicliptera chinensis( L.) Juss. are safe and non-toxic chemical compounds and can be used as health products or drugs.

UV,HPLC测定红花中黄色素、多糖和腺苷的含量

目的：为红花药材的质量评价提供依据。方法：采用ＵＶ，ＨＰＬＣ分析不同产地红花的化学成分含量（黄色素、多糖、腺苷）。结果：不同产地红花黄色素含量在２４．９０％～４０．３４％的范围内，多糖含量在５．６２％～１０．２２％的范围内，腺苷含量在３８．７～３９２．７μｇ·ｇ－１的范围内。结论：不同产地红花化学成分含量存在差异（Ｐ＜０．０１），巍山红花黄色素含量最高，新乡红花多糖含量最高；吉木萨尔红花腺苷含量最高。

当归多糖诱导L－细胞产生造血生长因子的实验研究

用造血祖细胞体外培养、造血生长因子检测和流式细胞术等方法，研究当归多糖（ＡＰ）诱导Ｌ－细胞（成纤维细胞株）产生造血生长因子及其对血细胞发生的调节作用。结果表明，Ｌ－细胞条件培养液（ＬｃＣＭ）对造血祖细胞（ＣＦＵ－ＧＭ、ＢＦＵ－Ｅ、ＣＦＵ－Ｅ）的体外增殖呈双向调节作用；１０％ＬｃＣＭ能促进骨髓造血细胞进入增殖活跃的Ｓ＋Ｍ／Ｇ２期；经ＡＰ诱导制备的ＬｃＣＭ在有或无外源性造血生长因子（如Ｅｐｏ，ＢＰＡ，ＧＭ－ＣＳＦ）存在时均对造血祖细胞的克隆增殖显示较高刺激活性；ＡＰ或ＩＬ－１与造血生长因子间似无协同作用。本研究提示ＡＰ可能通过诱导造血微环境的成纤维细胞分泌某些造血生长因子，从而促进造血祖细胞增殖分化，这或许是当归＂补血＂的生物学机理之一。

猪苓多糖对毒激素-L诱导大鼠恶病质样表现的抑制作用

目的：探讨肿瘤恶病质的病因及猪苓多糖治疗作用机理。方法：观察猪谷多糖对毒激素－Ｌ诱导的恶病质样表现的作用。结果：从人原发性肝癌患者腹水中提取的毒激素－Ｌ可在体外显著诱导脂解，体内抑制摄入行为，降低血锌及升高血铜（Ｐ＜０．０１）。结论：毒激素－Ｌ与肿瘤恶病质密切相．关，猪苓多糖可抑制毒激素－Ｌ在诱导肿瘤恶病质中的作用。

HPLC衍生化法分析决明子多糖水解产物中单糖组分及其多糖组成特征的研究

分别利用分光光度法和1-苯基-3-甲基-5-吡唑啉酮（PMP）衍生化/HPLC法分析了12批不同批次药材的决明子多糖及其水解产物中的单糖组分,以水解的单糖组分建立了决明子多糖的HPLC指纹图谱,并研究了决明子多糖的组成特征。结果表明,经过纯化的决明子多糖含量在93.41%-98.57%之间,以甘露糖为参照峰建立了12批药材的指纹图谱,相似度在0.963-0.999之间,多糖水解产物中单糖组分的含量在1.617-304.5 mg/g之间,7个共有峰组分含量大小分布均依次为：甘露糖〉木糖〉半乳糖〉葡萄糖〉阿拉伯糖〉葡萄糖醛酸〉氨基半乳糖,显示了决明子多糖的基本组成结构特征,12批药材的决明子多糖中各单糖的平均摩尔比为甘露糖∶木糖∶半乳糖∶葡萄糖∶阿拉伯糖∶葡萄糖醛酸∶氨基半乳糖=43.0∶24.2∶15.4∶5.6∶5.5∶3.4∶2.9。指纹图谱相似度分析和多糖的组成特征分析对市售药材的分析鉴定结果一致,可作为决明子多糖结构分析及其质量控制的依据。

松乳菇多糖对人骨肉瘤MG-63细胞Caspase-3、Caspase-9、Bax和Bcl-2蛋白表达的影响

目的研究松乳菇多糖体外对人骨肉瘤MG-63细胞生长、凋亡及凋亡相关蛋白的影响。方法体外培养MG-63细胞,将不同浓度的松乳菇多糖作用于细胞,MTT法检测不同时间点和不同浓度的松乳菇多糖对细胞生长的抑制作用; Annexin V-FITC/PI双染流式细胞仪检测对细胞凋亡的影响,并分析细胞周期; Western blot法检测Caspase-3、Caspase-9、Bax和Bcl-2蛋白的表达。结果松乳菇多糖可时间和浓度依赖性地抑制MG-63细胞的生长;通过阻滞G2-M期促进MG-63细胞凋亡;促进Caspase-3、Caspase-9和Bax蛋白的表达,抑制Bcl-2蛋白的表达。结论松乳菇多糖体外可抑制人骨肉瘤MG-63细胞的生长,诱导其凋亡,其诱导凋亡的作用机制与调节Caspase-3,Caspase-9,Bax和Bcl-2蛋白的表达有关。

南瓜多糖对糖尿病大鼠胰岛Fas、Fas-L、Bcl-2及Bax表达的影响

目的观察南瓜多糖(PP)对链脲佐菌素(STZ)诱导的糖尿病大鼠胰腺组织中Fas、Fas-L、Bcl-2及Bax表达的影响。方法建立链脲佐菌素性糖尿病模型,PP(150、300、600mg.kg-1)灌胃进行治疗。3wk后检测大鼠空腹血糖值;采用免疫组化的方法探讨PP对链脲佐菌素性糖尿病大鼠Fas和Fas-L蛋白表达的影响;采用RT-PCR的方法观察PP对实验性糖尿病大鼠胰岛凋亡调控基因Bax和Bcl-2表达的影响。结果PP(300、600mg.kg-1)能明显降低STZ诱导的糖尿病大鼠的空腹血糖值;PP(150、300、600mg.kg-1)治疗后大鼠胰腺组织中Fas/Fas-L蛋白的表达率明显低于STZ模型组;PP(150、300、600mg.kg-1)给药3wk后,大鼠Bcl-2基因的表达明显增加,PP(300、600mg.kg-1)治疗后大鼠Bax基因的表达明显下降,Bcl-2与Bax的比值明显增加。结论PP对实验性糖尿病大鼠有降血糖作用,其作用与降低大鼠胰腺组织中Fas、Fas-L蛋白的表达、调节Bcl-2与Bax基因的表达有关。

大粒车前子多糖组分PLP-1的结构表征

研究了由大粒车前子多糖分离纯化所得组分PLP-1的一级精细结构特征.通过单糖组成分析、部分酸水解及甲基化分析,并结合NMR,GC和GC-MS等技术测定了PLP-1的一级结构.结果表明,PLP-1由Ara(33.98%),Xyl(58.23%),Rha(2.36%),Glc(2.23%),Gal(2.36%)和Man(0.83%)组成,糖醛酸以GlcA形式存在.PLP-1中主要糖残基有α-T-linked Araf(9.58%),β-T-linked Xylp(8.71%),α-1,3-linked Araf(18.22%),β-1,3-linked Xylp(8.05%),β-1,4-linked Xylp(5.85%),β-1,2,4-linked Xylp(16.98%)和β-1,3,4-linked Xylp(27.52%),GlcA以α-T-linked GlcAp形式存在;此外还有1,2-linked Rhap,T-linked Glcp,1,6-linked Glcp,T-linked Galp,1,3-linked Galp和1,3,6-linked Galp等少量其它残基.根据所得结果推断出PLP-1可能的一级结构.

HPLC法比较2种银杏外种皮多糖的相对分子质量及含量

目的:采用HPLC法比较2种银杏外种皮多糖的相对分子质量及含量。方法:从银杏外种皮中提取出多糖,分别于常温下贮藏5年和3年后测定并比较其相对分子质量和含量。以右旋糖酐为标样,采用HPLC法,PLaquagel-OHMIXED(300mm×7.5mm,8μm)色谱柱,柱温25℃,示差折光检测器,流动相为超纯水,流速1.0mL·min-1。结果:贮藏5年和3年的2种银杏外种皮多糖的平均相对分子质量分别为10825.3(RSD=0.45%)和11062.5(RSD=0.78%);其多糖含量分别为66.8%(RSD=5·.9%)和68.7%(RSD=3.4%)。结论:在常温下,银杏外种皮多糖随着贮藏时间的延长,其相对分子质量和含量会发生变化。

油菜花粉多糖对荷瘤鼠脾细胞IL-2、TNF-α产生及其mRNA表达的影响

【目的】研究油菜花粉多糖(LBPP)对荷瘤鼠脾细胞IL-2、TNF-α的诱生及其mRNA表达的影响。【方法】通过动物抑制性肿瘤法制备肿瘤模型,采用双抗体夹心ELISA法检测IL-2、TNF-α含量,逆转录-聚合酶链反应(RT-PCR)检测IL-2、TNF-α mRNA的表达。【结果】LBPP有明显抗肿瘤作用,200mg·kg-1高剂量组抑瘤率可达51.26%,明显优于环磷酰胺46.22%;LBPP可显著促进荷瘤鼠脾细胞IL-2、TNF-α分泌(P<0.01),提高荷瘤鼠脾细胞IL-2、TNF-α mRNA的表达(P<0.01),高剂量组提高率分别达56.37%、105.24%、1364%和1289%。【结论】LBPP通过提高机体IL-2、TNF-α mRNA的表达而发挥抗肿瘤作用。