Extraction of Polysaccharides from Spirulina with Boiling Water

[Objective] The aim of this study was to develop a simple and effective method for the polysaccharide extraction from Spirulina. [Method] The polysaccha- rides were directly extracted from the fresh algae mat or dry powder of Spirulina in boiling water. The contents and quality of the extracted polysaccharides were measured. [Result] On average, 236.06 g of polysaccharide was extracted from 25 kg of fresh Spirulina mat, with a yield rate of 0.94%, while 191.95 g of polysac- charide was extracted from 2.5 kg of dry powder of Spirulina with a yield rate of 0.77%. The polysaccharide content in the extract of fresh Spirulina mat was 12.56% （according to glucose content）, while that of the dry powder was 12.38% （according to glucose content）; the glucose was produced during the hydrolysis of Spirulina polysaccharide. [Conclusion] Extraction polysaccharide from Spirulina with boiling wa- ter greatly reduces the use of ethanol, and the possibility of pollution from the ex- ogenous non-food chemical reagents, so that the extracted polysaccharides can be used as food materials. This method makes it possible to establish the production line for Spirulina polysaccharide.

Protective effects of a polysaccharide from Spirulina platensis on dopaminergic neurons in an MPT-Pinduced Parkinson's disease model in C57BL/6J mice

The present study aimed to determine whether a polysaccharide obtained fromSpirulina platensis shows protective effects on dopaminergic neurons. A Parkinson’s disease model was established through the intraperitoneal injection of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyr-idine （MPTP） in C57BL/6J mice. Prior to the MPTP injection, some mice were pretreated with intraperitoneal injections of a polysaccharide derived fromSpirulina platensis once daily for 10 days. The results showed that the immunoreactive staining and mRNA expression of the dopa-mine transporter and tyrosine hydroxylase, the rate-limiting enzyme in dopamine synthesis, in the substantia nigra, were signiifcantly increased in mice pretreated with 800 mg/kg of the poly-saccharide compared with those in MPTP-treated mice. The activities of superoxide dismutase and glutathione peroxidase in the serum and midbrain were also increased signiifcantly in mice injected with MPTP after pretreatment with the polysaccharide fromSpirulina platensis. By con-trast, the activity of monoamine oxidase B in serum and midbrain maintained unchanged. These experimental ifndings indicate that the polysaccharide obtained fromSpirulina platensis plays a protective role against the MPTP-induced loss of dopaminergic neurons in C57BL/6J mice, and that the antioxidative properties of this polysaccharide likely underlie its neuroprotective effect.

Fractionation and characterization of polysaccharides from cyanobacterium Spirulina (Arthrospira) maxima in nitrogen-limited batch culture

By a sequentially selective extraction procedure, polysaccharides (PS) from cyanobacterium Spirulina (Arthrospira) maxima that can be bio functionally healthy components were fractionated in four parts including the culture medium (PSCM), the external layers of the cell (PSEL), the cell wall (PSCW) and storage granules (PSSG). The four fractionated parts of polysaccharides were characterized by gas liquid chromatography (GLC) equipped with FID and fused silica capillary column (15 m×0.53 mm i. d.). The contents of PSCM, PSEL, PSCW and PSSG were about 2.0%, 2.6%, 10.0% and 52.0% to cell dry matter, respectively. Glucose was almost the only monosaccharide in PSSG and PSCW and most predominant in PSEL, while in PSCM, xylose, rhamnose and glucose were the main ones. Two uronic acids represented by glucuronic acid and galacturoic acid, six neutral monosaccharides including fucose, rhamnose, xylose, mannose, galactose and glucose, and two possible unidentified sugars were found in PSEL and PSCM. These data are valuable for the selective productions of high added value sugars from Spirulina .

Effects of the polysaccharides from Spirulina platensis on the activities of hepatitis B virus

To investigate the effects of the polysaccharides from Spirulina platensis (PSP) on hepatitis B virus (HBV), the cytotoxic effect of PSP was assessed by choosing the maximal concentration of PSP without cytotoxic effect on HepG2 2.2.15 cell line for further experiments. Four concentrations (400-50 μg/ml) of PSP were adopted to treat the cells, and the supernatants or cells were collected after 24, 48, 72 and 144 h respectively. HBsAg and HBeAg in the supernatants of cell cultures were tested with ELISA and copies of HBV DNA in supernatants were measured by real-time fluorescence quantitative PCR (FQ-PCR). Meanwhile, DNA/RNA hybridization was performed to evaluate the expression of IFN-α receptor (IFN-αR) on the cells. The experimental results showed that the secretion of HBV antigens decreased under the influence of PSP in a dose and time-dependent manner. PSP in concentration of 400 μg/ml could significantly decrease the secretion of HBsAg in 24 h. Although no obvious effect was observed on the secretion of HBeAg at that time, the inhibitory effects were observed in a dose-dependent manner from 48 to 144 h. In addition, the copies of HBV DNA were declined under the influence of PSP in the same manner, moreover, the maximal suppressive effect of PSP in concentration of 400 μg/ml was as great as that of lamivudine. The expression of IFN-αR was much higher in PSP-treated cells than that of the un-treated cells also in dose and time-dependent manner. It is concluded that PSP in non-cytotoxic concentration not only significantly decreases the secretion of the HBV antigens and the replication of HBV DNA, but also increases the expression of IFN-αR on HepG2 2.2.15 cell line. The results of the present investigation strongly support the notion that PSP exerts the anti-HBV effect both through enhancing the anti-HBV immunity and acting on HBV directly.

Antiviral effect of polysaccharide from Spirulina platensis(PSP) on HSV-2 in vitro

To explore the antiviral effect and mechanism of polysaccharide from Spirulina platensis （PSP） on herpes simplex virus type 2 （HSV-2）, a standard strain of HSV-2 （333 strain） was used to investigate the antiviral effect of PSP in vitro. PSP in various concentrations was applied to different stages of HSV-2 replication cycle. Finally, the virus infectivity （TCID50）, cytopathic effect （CPE）, and MTT staining method for viable cells （MTT assay） were used as markers to evaluate the effect of PSP on HSV-2. The quantity of HSV-DNA was detected by real-time fluorescence quantitative PCR （FQ-PCR）. The HSV-2 infected Vero cell tdtrastructures were observed by transmission electron microscopy （TEM）. The results showed that PSP had little cytotoxic effect on Vero cells, it could not directly inactivate HSV-2 infectivity. PSP not only interfered in adsorption of HSV-2 to Vero cells but also inhibited HSV-2 biosynthesis in the ceils. FQ-PCR results showed that the inhibitory rate on HSV- DNA also increased in a dose-dependent and time-dependent manner. TEM also confirmed that PSP exhibited pronounced inhibitory effect on HSV-2. In conclusion, the antiviral effect of PSP on HSV-2 may be attributed to the inhibition of virus adsorption, virus replication and synthesis in cells.

螺旋藻(Spirulina Platensis)多糖蛋白对辐射诱发小鼠PCE微核率及白细胞数的影响

本试验通过对NIH小鼠骨髓细胞微核分析及外周血白细胞计数表明,螺旋藻多糖蛋白能显著减轻辐射对小鼠PCE微核率及白细胞数的影响。故螺旋藻多糖蛋白有可能作为一种新的有效的天然辐射防护剂。

Effect of Spirulina Platensis Polysaccharide on Hematopoietic Recovery and Related Cytokines in Mice with Transplanted Tumor Treated by Chemotherapy

Objective:To evaluate the effect of Spirulina platensis polysaccharide (SPP) on hematopoietic recovery and related cytokines in mice with transplanted tumor after chemotherapy. Methods:Mouse model of transplanted solid tumor was established and treated with chemotherapy. Peripheral blood cells, bone marrow nucleated cells, and colony forming unit spleen (CFU S) were counted; the content of DNA in bone marrow was inspected by ultraviolet spectrophotometer; serum content of cytokines, interleukin (IL)1, IL 3, granulocyte macrophage colony stimulating factor (GM CSF) and tumor necrosis factor α (TNF α) were determined by double antibody sandwich ELISA.Results:Cyclophosphamide (CTX) could induce evident myelosuppression, manifested as decrease of peripheral blood cells, bone marrow nucleated cell and DNA, and the CFU S number. SPP could significantly ameliorate the myelosuppression induced by CTX without reducing anti tumor effect of CTX. In addition, it could also increase the contents of IL 1, IL 3, GM CSF, TNF α in serum. Conclusion:SPP can probably accelerate the hematopoietic recovery in mice after chemotherapy through promoting endogenous secretion of cytokines.

Isolation and structural elucidation of a polysaccharide with radiation-protective activity from Spirulina platensis

The structure of a radiation-protective sulfated polysaccharide, SP-APS, isolated from Spirulina platensis, was investigated by methylation analysis, partial acetolysis and H-1-NMR spectroscopy. The repeating-unit of SP-APS as branched structures was elucidated.

响应面法优化螺旋藻多糖提取及脱色工艺

研究螺旋藻多糖热碱液的提取、过氧化氢法脱色的条件并进行工艺优化。在单因素实验结果的基础上,分别以多糖得率和多糖脱色率为指标,通过响应面法优化热碱法提取螺旋藻多糖和过氧化氢法脱色的工艺条件,并分别得到热碱液提取螺旋藻多糖的最优提取工艺条件和过氧化氢法最优脱色条件。结果表明,热碱液提取法最优条件:NaOH质量分数1.5%、提取温度为90℃、提取时间4 h及料液比1:20 g/mL,在此条件下,得到螺旋藻多糖得率为5.18%±0.23%;过氧化氢法的最佳脱色条件:过氧化氢溶液添加量为17%、脱色时间为55 min、脱色温度为50℃,在此条件下得到的实际脱色率为82.54%±0.03%,多糖保留率为83.45%±0.13%。本研究所得结果对于螺旋藻多糖的开发利用具有一定参考价值。

螺旋藻多糖水解工艺研究

为提高螺旋藻多糖的提取率,本文用超声波辅助提取法提取螺旋藻多糖,在单因素实验基础上,用正交试验对提取工艺进行优化,并进行体外抗氧化活性的测定。结果表明最佳提取工艺为pH=11,液固比40∶1,提取温度70℃,提取5 h,此优化条件下螺旋藻多糖的提取率为8.03%。测定螺旋藻多糖的DPPH清除力和还原力,结果显示螺旋藻多糖具有抗氧化活性。

固定化培养中氮磷浓度对钝顶螺旋藻生长及其代谢产物和叶绿素荧光参数的影响

该研究以钝顶螺旋藻为实验对象,探究培养基中不同氮浓度(5、15、30、45 mmol·L^(-1))和磷浓度(0.5、1.5、3.0、4.5 mmol·L^(-1))对固定化生物膜培养模式下螺旋藻生长及其代谢产物和叶绿素荧光参数的影响,为生产实际中规模化高效培养螺旋藻提供理论依据。结果表明:(1)随着氮、磷浓度的升高,螺旋藻的生物量密度和产率先增高后降低,并在30 mmol·L^(-1)氮和3.0 mmol·L^(-1)磷处理时达到最大值;低氮、低磷条件会影响螺旋藻的藻体结构,表现为单细胞藻丝变短,螺旋变少,导致生物膜细胞密度降低。(2)与生物量变化趋势一致,螺旋藻光合色素含量随着培养基中氮、磷浓度的增加呈先升后降的变化趋势,且均在30 mmol·L^(-1)氮或者3.0 mmol·L^(-1)磷条件下达到最大值。(3)培养基中磷浓度固定时,螺旋藻的PSⅡ反应中心最大光能转化效率(F v/F m)和电子传递速率(ETR)随着氮浓度增加而增大,氮的缺乏会影响光合色素的合成,进而削弱螺旋藻光合作用;培养基中氮浓度固定时,螺旋藻的F v/F m和ETR随着磷浓度的增加呈现先升后降的趋势。(4)低氮、低磷条件下虽不利于可溶性蛋白的增加,却可以刺激可溶性多糖的积累,并且磷源限制程度越严重,螺旋藻蛋白质含量越低,多糖含量越高。研究认为,调节营养液中氮、磷浓度可定向诱导螺旋藻蛋白和多糖等高值化产物的合成,生产中可通过调控氮、磷浓度来促进相应目标产物的合成积累;螺旋藻生物膜对氮浓度的耐受程度较磷浓度更强,螺旋藻叶绿素荧光参数在高氮(45 mmol·L^(-1))培养下均较高,同时生物质产量也较高。

菌草灵芝多糖肽对Caco-2细胞MDR1、MRP2基因表达水平的影响

使用CCK-8比色法研究菌草灵芝多糖肽(JCGLPP)对人结肠腺癌上皮细胞(Caco-2)存活率的影响;采用PCR检测JCGLPP作用不同时间后对Caco-2细胞中P-糖蛋白的编码基因MDR1和多药耐药相关蛋白-2的编码基因MRP2表达水平的影响.结果显示:0.1~500μg·mL^(-1)JCGLPP对Caco-2细胞无毒性;10μg·mL^(-1)JCGLPP作用1~48 h能抑制MDR1基因的表达,对MRP2基因的表达呈先抑后扬的趋势.表明JCGLPP可抑制MDR1基因的表达,对MRP2基因表达的影响与其作用时间有关.

一株太空培育螺旋藻诱变株的皮肤修护和保湿研究

利用太空育种方法,培育得到新型的钝顶螺旋藻诱变株H11,超声法提取水溶性胞内多糖,使用斑马鱼胚胎尾鳍检测其皮肤修护能力,使用3D表皮模型检测其对天然保湿因子分泌的影响。实验结果表明,野生株和诱变株螺旋藻多糖得率分别为4.88%和20.32%(干重);野生株和诱变株螺旋藻多糖在0.1 mg/mL浓度下,斑马鱼胚胎尾鳍再生生长率分别提高1.44%和13.12%;在1%浓度下3D表皮模型反式尿刊酸的生成分别增加了16.50%和156.43%。结果表明,螺旋藻诱变株多糖含量远远高于野生株,并且表现出优于野生株螺旋藻的皮肤修护和保湿能力。

极大螺旋藻胞内多糖对体外肿瘤细胞抑制作用的实验研究

研究了极大螺旋藻(Spirulina maxima)胞内多糖对SMMC7721等肿瘤细胞系的生长及凋亡的影响。在体外培养条件下,用半抑制浓度(80,5 mg/L)的极大螺旋藻胞内多糖(IPSⅠ,IPSⅡ)处理肿瘤细胞,并用MTT,SRB,DNA ladder,AnnexinⅤ等方法测定。结果表明,IPSⅠ(80 mg/L)对SMMC7721,A549,HeLa,U251等肿瘤细胞系抑制率分别为64.97%、46.1%、43.3%、27.02%;IPSⅡ(5 mg/L)能诱导SMMC7721细胞凋亡。

醋柴胡多糖对拉米夫定体外抗乙型肝炎病毒的增效作用研究

目的:考察醋柴胡多糖对拉米夫定抗乙型肝炎病毒(HBV)的增效作用,并初步探讨其作用机制。方法:将不同浓度的醋柴胡多糖、拉米夫定及二者联合作用于人肝癌细胞HepG2.2.15,同时设立对照组,孵育48 h,酶联免疫吸附试验试剂盒检测细胞上清液中乙型肝炎表面抗原(HBsAg)、乙型肝炎e抗原(HBeAg)分泌量,荧光探针定量聚合酶链式反应(PCR)检测细胞HBV脱氧核糖核酸(DNA)表达量,金(正均)氏公式定量分析醋柴胡多糖的增效作用。采用高效液相色谱法测定细胞内拉米夫定含量;Western blot法测定有机阳离子转运蛋白(OCT)1、OCT2、P糖蛋白(P-gp)和多药耐药蛋白2(MRP2)的表达量。结果:与拉米夫定单用组相比,醋柴胡多糖增加拉米夫定对HBsAg分泌的抑制作用,表现为相加作用、对HBeAg作用表现为协同增强,Q值达6.55、对HBV-DNA抑制作用表现为相加。醋柴胡多糖低剂量组、醋柴胡多糖低中剂量组、醋柴胡多糖低高剂量组可显著促进拉米夫定的摄取;醋柴胡多糖高剂量联用组可显著降低P-gp的表达;醋柴胡多糖单用及联用组均可显著提高OCT1的表达。结论:醋柴胡多糖可通过增加拉米夫定的摄取发挥协同抗HBV作用,其作用机制可能P-gp、OCT1有关。

螺旋藻糖蛋白的分离纯化及其性质研究

采用新的分离工艺从螺旋藻中提取得到粗多糖，经环流泡沫分离去除蛋白和ＳｅｐｈａｃｒｙｌＳ－１００柱层析纯化后得到一种糖蛋白．经ＳｅｐｈａｄｅｘＧ－２００和凝胶电泳证实所得的糖蛋白为均一组成；由ＳＤＳ－ＰＡＧＥ测定分子量为１３００００；红外色谱呈现出典型的多糖吸收峰，含有α－型糖苷连接键；

螺旋藻多糖提取新工艺的研究

提取螺旋藻多糖的传统工艺存在着流程长，操作复杂，有机溶剂消耗大等问题。采用三氯乙酸（ＴＣＡ）法代替Ｓｅｖａｇ法去除蛋白质，使改进后的提取工艺中蛋白质去除率增加，多糖损失率降低，流程缩短。通过用ＤＥＡＥ－Ｓｐｅｈａｒｏｓｅ和Ｓｅｐｈａｄｅｘ－Ｇ５０柱的分离纯化得到２个多糖组分。经测定，其相对分子质量分别为Ｍｒ１＝１２６００，Ｍｒ２＝１６６００。组分１的单糖组成：Ｄ－葡萄糖、Ｄ－甘露糖、Ｄ－半乳糖和葡萄糖醛酸；组分２的单糖组成：Ｄ－葡萄糖、Ｄ－木糖、Ｌ－鼠李糖和葡萄糖醛酸。

钝顶螺旋藻多糖和藻蓝蛋白对小鼠急性放射病的防护作用

本文报告了钝顶螺旋藻多糖（ＳＰＰ）和藻蓝蛋白（Ｃ－ＰＣ）对小鼠受致死剂量￣（６０）Ｃｏγ射线照射后３０天存活率的影响。在照射前５天每天给小鼠腹腔注射ＳＰＰ１２５ｍｇ／ｋｇ或Ｃ－ＰＣ５０ｍｇ／ｋｇ均能分别比对照组提高小鼠的存活率３３％和２８％同时观察了ＳＰＰ和Ｃ－ＰＣ对经￣（６０）Ｃｏγ射线亚致死剂量照射后小鼠血细胞生成的影响，结果显示，ＳＰＰ和Ｃ－ＰＣ可刺激照射后小鼠粒单系祖细胞和造血干细胞的形成，并增加骨髓有核细胞的数量。ＳＰＰ和Ｃ－ＰＣ还可以增加照射小鼠外周血细胞的总数。ＳＰＰ和Ｃ－ＰＣ对外周血红细胞数与血红蛋白量无显著的影响。结果提示：ＳＰＰ和Ｃ－ＰＣ可能促进￣（６０）Ｃｏγ射线照射小鼠造血功能的恢复。

螺旋藻中藻胆蛋白和多糖的分离纯化及产品特性

提出一种从螺旋藻中提取纯化藻蓝蛋白和糖蛋白的新型分离工艺。等电聚焦分析结果表明所得到的藻蓝蛋白为电泳纯，等电点为４．３，测定了藻蓝蛋白的氨基酸组成及其红外吸收光谱。ＳＤＳ－ＰＡＧＥ分析结果表明所得到的糖蛋白组成均一，其分子量为１３０ｋｄ，测定了螺旋藻糖蛋白的氨基酸组成，红外光谱分析表明螺旋藻糖蛋白含有α－型糖苷连接键。

钝顶螺旋藻糖缀合物SPPA-1的理化性质及活性的研究

目的 研究螺旋藻多糖中的均一组分SPPA 1的分离方法、糖组成、分子量及生物活性。方法 采用丙酮分部沉淀 ,三氯乙酸去蛋白 ,再经SephadexG 75和CM SephadexC 5 0柱色谱得到SPPA 1,用气相色谱法测定单糖组成 ,GPC法测定分子量 ,IR和NMR法确定糖苷键类型。结果 经高压液相色谱和毛细管电泳鉴定为均一性组分。其总糖含量为 91 70 % ,N含量为 0 96 % ,分子量为 6 9 0 0× 10 4 Da。糖组成分析表明其由单一的葡萄糖组成 ,红外光谱、核磁共振1HNMR谱分析表明 ,其单糖类型为α 吡喃型。ESR研究表明SPPA 1具有清除O·2 自由基活性。结论 SPPA