Phytochemical Contents and Antioxidant Properties of Ganoderma lucidum Polysaccharides in Relation to Growth Stage and Culture Substrate

[Objectives]To determine physicochemical characteristics and antioxidant activity of Ganoderma lucidum polysaccharides(GLP)in different growth stages and culture media.[Methods]Five polysaccharides(GLP1,GLP2,GLP3,GLPW and GLPB)were extracted and purified from the fruiting body of G.lucidum at three growth stages and culture substrates(wood culture and bag culture).The chemical components and antioxidant activity of the five polysaccharides were determined.[Results]GLP1 contained the highest content of neutral sugar(87.65%)and GLP3 had the highest uronic acid content.All the samples mainly comprised glucose,galactose,mannose,xylose and arabose with different ratios.Moreover,their antioxidant activities were investigated on the basis of DPPH radical,ABTS radical,SOD-like activity,and antihaemolytic activity.Results indicated in all three growth stages GLP2 had the greatest antioxidant properties.In addition,the antioxidant activity of GLPW was significantly higher than that of GLPB.[Conclusions]Overall,by comparison,G.lucidum in growth stage two may have potential health benefits,and wood culture may be a superior choice of artificial cultivation due to their abundance of active polysaccharides.

The Effect of Culture Condition on Type 5 Capsular Polysaccharide Production of Staphylococcus aureus from Diary Cattle

[Objective] The effect of different culture conditions on type 5 capsular polysaccharide production of Staphylococcus aureus from diary cattle was studied to provide simple way for CP production and preparation and laid foundation for carrying out new polysaccharide vaccine research. [Method] Staph-ylococcus aureus was isolated from milk sample of sick dairy cattle and capsular polysaccharide serotypes were identified. Type 5 capsular polysaccharide was cultured on BHI,solid columbia and mod110 culture media. Glucose and lactose were taken as carbon sources for every culture media in solid and liquid state. Therefore 9 different culture conditions were taken to study the effect of culture conditions on capsular polysaccharide production. [Result] Different culture conditions indicated that compared with columbia culture media, BHI culture media could decline capsular polysaccharide production and mod110 culture media could increase capsular polysaccharide production. While for same culture media, solid culture media was better for capsular polysaccharide production,meanwhile,taken lactose as carbon source could increase capsular polysaccharide production.

Structural Analysis of Water-soluble Polysaccharide PIP_1 Extracted from the Cultured Mycelium of Phellinus igniarius

Water-soluble crude polyseccharide（PIP） was extracted from cultured mycelium of the fungus Phellinus igniarius. After ethanol precipitation and sepharose CL-6B gel filtration, the fraction of PIP1 was obtained, which was shown to be a homogeneous polysaccharide by means of high-performance liquid chromatography. The structure of PIPt was determined by using several methods. C.,C analysis indicates that PIP1 is composed of the monosaccharides of glucose, galactose, and mannose. Their malar ratio is 3. 70： 4. 06： 1.00. The molar weight was estimated to be 17 kd via HPLC. IR, GC, partial hydrolysis with acid, pefiedate oxidation, Smith degradation, methylation, and GC-MS analysis were used for the structural analyses of PIP1. The results show that PIP1 has a small quantity of branch structure, The main glycosidic linkage of PIP1 has a β-configurafion. The main chain is made up of a large mass of glucose （ 1→3 ） and few mannose （ 1→4 ） ; the side chain is composed of glucose （ 1 →3 ） and galactose （ 1→6 ） ; the nonreduced end is composed of galactose and glucose. The side chains are branched at 6-0 of glucose（ 1→3,6） and mannose（1→4,6）. On an average, there are three branches among 20 residues. It is presumable that the existence of 1,3-linked Glc in the main and side chains is the main reason for its higher antitumor activity.

COMPARISON OF POLYSACCHARIDES ISOLATED FROM THE MYCELIA OF A CULTIVATED STRAIN OF PORIA COCOS GROWN IN DIFFERENT LIQUID CULTURE MEDIA

Mycelium of a cultivated strain of Poria cocos was grown by submerged fermentation in a liquid mediumcontaining corn steep liquor with orbital shaking. Six polysaccharides coded as ac-PCM1, ac-PCM2, ac-PCM3-Ⅰ andⅡ, ac-PCM4-Ⅰand Ⅱ were isolated from the myelium by extracting with 0.9% NaCl aqueous solution, hot water, 0.5 mol/L NaOHaqueous solution and 88% formic acid. Exo-polysaccharide was obtained from the culture medium and coded as ac-PCM0.The monosaccharide composition and molecular weights of these polysaccharides were characterized by using infraredspectroscopy, gas chromaography, elemental analysis, ^(13)C-NMR, viscometry and light scattering. The results indicated thatac-PCM0, ac-PCM1 and ac-PCM2 are heteropolysaccharides containing glucose, galactose, mannose and fucose, and ac-PCM3-Ⅰ and ac-PCM3-Ⅱ mainly consist of D-glucose. The content of the glucose in the polysaccharides increased with theisolation progress. Remarkably, α-glucan and β-glucan coexisted in the extract by NaOH aqueous solution (ac-PCM3), andcould be separated by chemical methods. The protein in the ac-PCM polysaccharides cultured from the medium containingcorn steep liquor was higher than that in the ab-PCM from the medium with bran extract. Therefore, the polysaccharidesfrom Poria cocos mycelia cultured in different media have different chemical composition, molecular weights and conformations.

Inhibitory Effect of Extracellular Polysaccharide EPS-II from Pseudoalteromonas on Candida adhesion to Cornea in vitro

Objective Fungal keratitis（FK） is a vision-threatening infection,whose treatment requires more effective and safer anti-fungal agent exploitation urgently.With this aim,we focused on the effect of an extracellular polysaccharide on fungal adhesion to human corneal epithelial cells.Methods We performed the cytotoxicity assays of the extracellular polysaccharide EPS-II from an antarctic bacterium Pseudoaltermonas and evaluated its inhibitory effect on Candida albicans cells＇ adherence to human corneal epithelial cells（HCECs）.Results EPS-II,which displayed minor cytotoxicity but also promoted proliferation of HCECs,could inhibit the adherence of yeast cells to HCECs in a dose-dependent manner.EPS-II could also suppress the subsequent PI3K/AKT signaling pathway,and thereby decrease the expression of early inflammatory cytokines.Conclusions ExtracellularpolysaccharideEPS-IIwassuggestedasanewnaturalagentforattenuatingFK.

Dynamic culture of a thermosensitive collagen hydrogel as an extracellular matrix improves the construction of tissue-engineered peripheral nerve

Tissue engineering technologies offer new treatment strategies for the repair of peripheral nerve injury, hut cell loss between seeding and adhesion to the scaffold remains inevitable. A thermosensitive collagen hydrogel was used as an extracellular matrix in this study and combined with bone marrow mesenchymal stem cells to construct tissue-engineered peripheral nerve composites in vitro. Dynamic culture was performed at an oscillating frequency of 0.5 Hz and 35° swing angle above and below the horizontal plane. The results demonstrated that bone marrow mesenchymal stem cells formed membrane-like structures around the poly-L-lactic acid scaffolds and exhibited regular alignment on the composite surface. Collagen was used to fill in the pores, and seeded cells adhered onto the poly-L-lactic acid fibers. The DNA content of the bone marrow mesenchymal stem cells was higher in the composites constructed with a thermosensitive collagen hydrogel compared with that in collagen I scaffold controls. The cellular DNA content was also higher in the thermosensitive collagen hydrogel composites constructed with the thermosensitive collagen hydrogel in dynamic culture than that in static culture. These results indicate that tissue-engineered composites formed with thermosensitive collagen hydrogel in dynamic culture can maintain larger numbers of seeded cells by avoiding cell loss during the initial adhe-sion stage. Moreover, seeded cells were distributed throughout the material.

Production and characterization of an extracellular polysaccharide of antarctic marine bacteria Pseudoalteromonas sp. S-15-13

Twenty-seven antarctic bacteria producing extracellular polysaccharide （EPS） were selected from 57 strains by staining technology. The effects of major environmental factors on the growth and EPS production of Pseudoalteromonas sp. S - 15 - 13 were investigated, and the EPS was separated and purified for characterization analysis. The results showed that the optimal conditions for the EPS production were culture period, 56 h; growth temperature, 8 ℃ ; carbon source, 1.0% glucose; NaCI concentration, 3.0% ; pH 6.0 - 7.0. The EPS was purified by cold ethanol precipitation, proteins removal, ion exchange chromatography and gel chromatography technology. The molecular mass of EPS - H was 62 kDa as determined by the high performance gel permeation chromatography. Its sugar composition was a homopolymer of marmose analyzed by gas chromatograph spectroscopy. After repeated freezing and thawing of the bacteria hiomass in the presence of EPS, the bacterial growth was much higher than that observed after freezing in the absence of EPS and the difference augmented with the increase of freeze-thaw cycles. It is hypothesized that the adaptation of Pseudoalteromonas sp. S- 15 - 13 to the antarctic marine conditions, characterized by low temperature, high NaCl concentration and repeated freeze-thaw cycles, might be related to the EPS production ability.

Optimization for Production of Intracellular Polysaccharide from Cordyceps ophioglossoides L2 in Submerged Culture and Its Antioxidant Activities in vitro

Cordyceps ophioglossoides is a valuable traditional medicinal material.We have found that intracellular polysaccharide(IPS) is the major biologically active ingredient in Cordyceps ophioglossoides.This study is the first time to optimize the yield of IPS from Cordyceps ophioglossoides.The optimal medium for IPS production consists of glucose 54.50 g·L·1,yeast powder 25.50 g·L·1,NaH2PO4 0.4 g·L·1 and K2HPO4 0.4 g·L·1.The suggested culture conditions are 24 ℃,initial pH 4.5 with a rotary speed of 120 r·min·1 for 168 h.The yield of IPS is 737.93 mg·L·1,which is 50% higher than the yield under the conditions prior to optimization.The anti-oxidative activities of IPS in Cordyceps ophioglossoides L2 are also characterized using various in vitro assay.The anti-oxidative activity may explain the reason why IPS from Cordyceps ophioglossoides can be used to fight against neurodegenerative dis-eases and menopausal symptoms.

A Polysaccharide-Degrading Marine Bacterium Flammeovirga sp.MY04 and Its Extracellular Agarase System

Bacteria of the genus Flammeovirga can digest complex polysaccharides (CPs), but no details have been reported regarding the CP depolymerases of these bacteria. MY04, an agarolytic marine bacterium isolated from coastal sediments, has been identified as a new member of the genus Flammeovirga. The MY04 strain is able to utilize multiple CPs as a sole carbon source and grows well on agarose, mannan, or xylan. This strain produces high concentrations of extracellular proteins (490 mg L-1 ± 18.2 mg L-1 liquid culture) that exhibit efficient and extensive degradation activities on various polysaccharides, especially agarose. These proteins have an activity of 310 U mg-1 ± 9.6 U mg-1 proteins. The extracellular agarase system (EAS) in the crude extracellular enzymes contains at least four agarose depolymerases, which are with molecular masses of approximately 30-70 kDa. The EAS is stable at a wide range of pH values (6.0-11.0), temperatures (0-50℃), and sodium chloride (NaCl) concentrations (0- 0.9 mol L-1). Two major degradation products generated from agarose by the EAS are identified to be neoagarotetraose and neoagarohexaose, suggesting that β-agarases are the major constituents of the MY04 EAS. These results suggest that the Flammeovirga strain MY04 and its polysac-charide-degradation system hold great promise in industrial applications.

Extracellular Polysaccharides Matrix—An Often Forgotten Virulence Factor in Oral Biofilm Research

Oral diseases related to dental biofilms continue to afflict the majority of the world＇s population. Among them, dental caries continues to be the single most prevalent and costly oral infectious disease （Marsh, 2003; Dye et al., 2007）. Dental caries results from the interaction of specific bacteria with constituents of the diet within a dental biofilm known as plaque （Bowen, 2002）. Sucrose is considered to be the ＂arch criminal＂ from the dietary aspect because it serves as a substrate for synthesis of extracellular （EPS） and intracellular （IPS） polysaccharides in dental biofilm and is also fermentable （Bowen, 2002）.

Isolation, chemical characteristics and immunity activity of an extracellular polysaccharide EPS I isolated from Antarctic bacterium Pseudoalteromonas sp.S-15-13

A new extracelluar polysaccharide （EPS） was isolated and purified from Antarctic bacterium S-15-13, identified as Pseudoalteromonas sp. After being separated and purified by DEAE-Sephadex A-50 ionexchange and Sephadex G-100 gel chromatography, two mains fractions （EPS I and EPS Ⅱ ） were ob-tained. EPS I was composed of mannose, glucose and galactose with a molecular weight of 23kDa and EPS Ⅱ was composed of mannose only with a molecular weight of 62kDa. The effect of the polysaccharide EPS I on the cellular immune response of mice was investigated. Results demonstrated that EPS I could markedly facilitate lymphocyte proliferation, and might be a strong immunomodulator.

In Vitro Antioxidant Function of Extracellular Polysaccharides from Tremella aurantialba

Extracellular polysaccharide from Tremella aurantialba (TEP) significantly inhibited H2O2-induced haemolysis of RBCs, significantly decreased the amount of malondialdehyde (MDA) produced in mice liver homogenates as a result of autoxidation, and inhibited VitC-Fe2+-induced swelling of liver mitochondria.

Ultrastructural and Extracellular Protein Changes in Cell Suspension Cultures of Populus euphratica Associated with Low Temperature-induced Cold Acclimation

Populus euphratica Olive is the only tree species that can grow in the saline land and also survive cold winters in northwest China, and it plays a very important role in stabilizing the vulnerable ecosystem there. A cell suspension culture was initiated from callus derived from plantlets of Populus euphratica. Cold acclimation was induced (LT50 of 17.5 ℃) in cell suspension at 45 ℃ in the dark for 30 days and the freezing tolerance increased from LT50 of 12.5 ℃ in nonacclimated cells to LT50 of 17.5 ℃ in cold-acclimated cells. Microvacuolation, cytoplasmic augmentation and accumulation of starch granules were observed in cells that were cold-acclimated by exposure to low temperatures. Several qualitative and quantitative changes in proteins were noted during cold acclimation. Antibodies to carrot extracellular (apoplastic) 36 kD antifreeze protein did not cross react on immunoelectroblots with extracellular proteins in cell suspension culture medium of Populus euphratica, indicating no common epitopes in the carrot 36 kD antifreeze protein and P. euphratica extracellular proteins. The relationship of these changes to cold acclimation in Populus euphratica cell cultures was discussed.

Effect of polygonatum polysaccharide on the hypoxia-induced apoptosis and necrosis in in vitro cultured cerebral cortical neurons from neonatal rats

BACKGROUND： Cardiocerebrovascular diseases induced cerebral circulation insufficiency and senile vascular dementia can result in ischemic/hypoxic apoptosis of central neurons, which we should pay more attention to and prevent and treat as early as possible. Traditional Chinese medicine possesses the unique advantage in this field. Polygonatum, a Chinese herb for invigorating qi, may play a role against the hypoxic apoptosis of brain neurons. OBJECTIVE ： To observe the protective effect of polygonatum polysaccharide on hypoxia-induced apoptosis and necrosis in cerebral cortical neurons cultured in vitro. DESIGN： A comparative experiment.SETTING： Laboratory of Cell Biology, Institute of Basic Medical Sciences, Jiangxi Provincial Academy of Traditional Chinese Medicine. MATERIALS： The experiment was carried out in the Laboratory of Cell Biology, Institute of Basic Medical Sciences, Jiangxi Provincial Academy of Traditional Chinese Medicine from November 2003 to April 2005. Totally 218 Wistar rats （male or female） of clean degree within 24 hours after birth were purchased from the animal center of Jiangxi Medical College （certification number was 021-97-03）. METHODS：① Preparation of cerebral cortical neurons of rats： The cerebral cortical tissues were isolated from the Wistar rats within 24 hours after birth, and prepared to single cell suspension, and the cerebral cortical neurons of neonatal rats were in vitro cultured in serum free medium with Neurobasal plus B27 Supplement. ② Observation on the non-toxic dosage of polygonatum polysaccharide on neurons： After the neurons were cultured for 4 days, polygonatum polysaccharide of different dosages （1-20 g/L） was added for continuous culture for 48 hours, the toxicity and non-toxic dosage of polygonatum polysaccharide on neurons were observed and detected with trypan blue staining. ③Grouping： After hypoxia/reoxygenation, the cultured neurons were divided into normal control group, positive apoptotic group and polygonatum polysaccharide group. In the normal control group, the neurons were cultured at 37℃ in CO2 with the volume fraction of 0.05 under saturated humidity for 6 days. In the apoptotic positive group, the neurons were cultured with hypoxia for 12 hours after 4-day culture, and followed by reoxygenation for 48 hours. In the polygonatum polysaccharide group, polygonatum polysaccharide with the terminal concentration of 0.5, 1 and 1.5 g/L was added to some neurons at 10 hours before the hypoxia culture, and then the neurons were cultured with hypoxia for 12 hours, followed by reoxygenation for 48 hours; polygonatum polysaccharide with the terminal concentration of 0.5, 1 and 1.5 g/L was added to the other neurons at 12 hours after hypoxia followed by reoxygenation for 48 hours.④ The Hoechst33342 fluorescence staining, Annexin V/PI flow cytometer, appearance of DNA agarose gel electrophoresis gradient strap and immunohistochemical staining were used to observe the expressions of Bcl-2, Bax and Caspase-3 apoptotic and anti-apoptotic proteins and the ratio of Bcl-2/Bax, and observe the effect of polygonatum polysaccharide against the hypoxic apoptosis of cerebral cortical neurons of neonatal rats. MAIN OUTCOME MEASURES： ① Toxicity and non-toxic dosage of polygonatum polysaccharide on neurons;② Apoptotic rate of neurons detected with Hoechst33342 fluorescence staining;③ Early apoptotic rate and necrotic rate of neurons detected with Annexin V/PI flow cytometer; ④DNA agarose gel electrophoresis ladder-like strap appeared or not;⑤ Expressions of Bcl-2, Bax and Caspase-3 apoptotic and anti-apoptotic proteins and the ratio of Bcl-2/Bax. RESULTS：① Polygonatum polysaccharide within 6 g/L had no cytotoxicity on the normal cultured cerebral cortical neurons （P 〉 0.05）. ②The apoptotic rates of neurons detected with Hoechst33342 fluorescence staining had significant differences between the polygonatum polysaccharide groups and positive apoptosis group added to neurons at 10 hours before the hypoxia culture [（13.00±4.52）%,（12.72±2.15）%, （11.80±1.18）%,（38.03±1.05）%, P 〈 0.01], and had no significant differences between the polygonatum polysaccharide groups and positive apoptosis group added to neurons at 12 hours after the hypoxia culture （36.77±1.45）%, （36.60±1.61）%, （36.37±2.02）%, （38.03±1.05）%, P 〉 0.05].③ Annexin V/PI flow cytometer detected that the anti-necrotic effect was enhanced with the increased concentration of polygonatum polysaccharide within 0.5-1.5 g/L （P 〈 0.01）. Polygonatum polysaccharide of 0.5-1.5 g/L added before hypoxia could significantly decrease the apoptotic rate of neurons induced by hypoxia/reoxygenation （P 〈 0.01）. ④ No DNA agarose gel electrophoresis ladder-like strap appeared in the groups with polygonatum polysaccharide of 0.5-1.5 g/L added at 10 hours before hypoxia;⑤ After Polygonatum polysaccharide of 0.5-1.5 g/L was added before hypoxia, the expression of Bcl-2 protein of hypoxic neurons was increased （P 〈 0.01）, and those of Bax protein and Caspase-3 protein were reduced （P 〈 0.01）, and the ratio of Bcl-2/Bax was increased （P 〈 0.01）. CONCLUSION： Polygonatum polysaccharide within 6 g/L has no cytotoxicity on the normal cultured cerebral cortical neurons. Polygonatum polysaccharide of 0.5-1.5 g/L added before hypoxia plays a role agains necrosis of neurons induced by hypoxia. Polygonatum polysaccharide of 0.5-1.5 g/L can significantly reduce the apoptosis of neurons induced by hypoxia through up-regulating the expression of Bcl-2 protein, down-regulating the expressions of Bax protein and Caspase-3 protein, and increasing the ratio of Bcl-2/Bax.

Quantitative detection and comparison of sulfate glycosaminoglycans content in extracellular matrix of in vitro cultured epiphyseal, articular and rib chondrocytes

Objective To establish a method for quantitative detection of the sulfate glycosaminoglycans ( GAG) content in extracellular matrix of in vitro cultured chondrocytes so as to evaluate the biological characteristics of epiphyseal, articular and rib chondrocytes. Methods Sulfate GAG content in extracellular matrix of three chondrocytes was measured by the modified dimethylmethylene blue (DMB) method. The changes of the toluidine blue (TB) stain of chondrocytes were observed by light microscope. Results Primary chondrocytes had the highest content of sulfate GAG in the extracellular matrix, ie, epiphyseal chondrocytes reached ( 70. 12 ± 7. 72 )μg/cm2, articular chondrocytes (92.00 ± 10.15) μg/cm2 and rib chondrocytes (80.61 ± 11. 40) μg/cm2, respectively. On the third pasage chondrocytes, epiphyceal chondrocytes decreased to (53.27 ± 9. 50 ) μg/cm2, articular chondrocytes to (63.88 ± 11.92) μg/cm2 and rib chondrocytes to (58.94 ±8.21) μg/cm2, respectively. The change of TB in every passage

Protective Effect and Autophagy Mechanism of Lycium barbarum Polysaccharides on Retinal Pigment Epithelial Cells Under High-Glucose Conditions

Objective:To study the effects of Lycium barbarum polysaccharide(LBP)on the proliferation,apoptosis,and autophagy of retinal pigment epithelial(RPE)cells cultured under high-glucose conditions.Methods:The ARPE-19 cell line was randomly divided into a control group(normally cultured in Dulbecco's Modified Eagle Medium/Nutrient Mixture F-12[DMEM/F-12]medium),a high-glucose group(HG;50 mmol/L glucose added to DMEM/F-12 medium),and a HG+LBP group(incubated in DMEM/F-12 medium containing 1 mg/mL LBP for 24 h,and then treated with 50 mmol/L glucose for 24 h).Following Ad-mCherry-GFP-LC3B infection,cell proliferation,apoptosis,mammalian target of rapamy-cin(mTOR)expression,and autophagic flux were determined by Cell Counting Kit-8(CCK-8),AnnexinV-APC/7-AAD Apoptosis Detection Kit,Western blot,and laser confocal microscopy,respectively.Results:The proliferation rate of ARPE-19 cells in the HG group was significantly lower than that in the control group(P<0.05),while the proliferation rate of ARPE-19 cells in the HG+LBP group was significantly higher than that in the HG group(P<0.05).The apoptosis rate of ARPE-19 cells in the HG group was significantly higher than that in the control group(P<0.05),while the apoptosis rate of ARPE-19 cells in the HG+LBP group was significantly lower than that in the HG group(P<0.05).The relative expression of phosphorylated mTOR(p-mTOR)of ARPE-19 cells in the HG group was significantly lower than that in the control group(P<0.05),with enhanced autophagic flux;when compared with the HG group,the HG+LBP group had significantly higher expression of p-mTOR(P<0.05),with diminished autophagic flux.Conclusion:LBP has a protective effect on RPE cells with high glucose-induced injury,and its mechanism may be related to LBP inhibition of high glucose-induced abnormal autophagy.

基于MAPK/ERK通路探讨茯苓多糖对卵巢早衰模型大鼠的作用及其分子机制

目的基于丝裂原活化蛋白激酶/细胞外调节蛋白激酶(MAPK/ERK)通路探讨茯苓多糖对卵巢早衰(POF)模型大鼠的作用及其分子机制。方法取50只SD雌性大鼠腹腔注射环磷酰胺诱导复制POF模型,随机分为5组:模型组、茯苓多糖低剂量组(100 mg/kg)、茯苓多糖高剂量组(200 mg/kg)、茴香霉素组(MAPK激活剂,10 mg/kg)、茯苓多糖高剂量+茴香霉素组,每组10只;另取10只SD雌性大鼠腹腔注射等剂量生理盐水作为对照组。分组处理后分别测定各组大鼠子宫指数及卵巢指数;酶联免疫吸附试验(ELISA)测定各组大鼠血清性激素[雌二醇(E2)、促黄体生成素(LH)、卵泡刺激素(FSH)]水平;HE染色观察各组大鼠卵巢形态;酶联免疫吸附试验检测各组大鼠血清及卵巢组织肿瘤坏死因子-α(TNF-α)、白细胞介素-4(IL-4)、白细胞介素-18(IL-18)水平;Western blotting检测各组大鼠卵巢组织MAPK/ERK通路相关蛋白表达。结果与对照组比较,模型组大鼠卵巢间质结构有一定程度的出血和纤维化损伤,卵泡发育失常,血清FSH和LH水平、血清和卵巢组织TNF-α、IL-4、IL-18水平、卵巢组织p-p38 MAPK/p38 MAPK及p-ERK1/2/ERK1/2相对表达量均升高(P<0.05),子宫指数和卵巢指数、E_(2)水平均降低(P<0.05)。与模型组比较,茯苓多糖低剂量组、茯苓多糖高剂量组大鼠卵巢形态损伤及卵泡发育失常均减轻,血清FSH及LH水平、血清和卵巢组织TNF-α、IL-4、IL-18水平、卵巢组织p-p38 MAPK/p38 MAPK及p-ERK1/2/ERK1/2相对表达量均降低(P<0.05),子宫指数和卵巢指数、E_(2)水平均升高(P<0.05),且茯苓多糖高剂量组效果更优(P<0.05)。茴香霉素组大鼠卵巢形态损伤及卵泡发育失常加重,血清FSH及LH水平、血清和卵巢组织TNF-α、IL-4、IL-18水平、卵巢组织p-p38 MAPK/p38 MAPK及p-ERK1/2/ERK1/2相对表达量均升高(P<0.05),子宫指数及卵巢指数、E_(2)水平均降低(P<0.05)。茯苓多糖高剂量+茴香霉素组逆转茯苓多糖作用,呈相反趋势(P<0.05)。结论茯苓多糖可通过抑制MAPK/ERK通路的激活减轻POF大鼠炎症,从而改善其卵巢功能和性激素分泌,缓解大鼠POF症状。

细菌根际黏附的界面作用机制研究

界面作用是细菌在环境中黏附、定殖、形成生物膜和发挥生态功能的基础和前提,对植物吸收养分、病原微生物拮抗具有重要意义。微生物-植物根际相互作用研究大多数从生态学和分子生物学角度出发,应用多种组学手段研究根系分泌物对于根际微生物定殖数量、群落组成和生理功能的影响,忽略了细菌定殖过程中物理、化学的界面作用机制及其对黏附的贡献。本文从界面作用机制出发,探讨了不同类型根系分泌物对细菌表面性质、细菌胞外聚合物(EPS)分子组成以及黏附功能的调控作用;梳理了黏附过程中细菌-植物生物大分子相互作用的主要模式及其微观机制;归纳了根际定殖过程可视化研究方法和微生物-植物生物大分子相互作用的分析手段;提出了根际生物分子组成分析、黏附蛋白质功能预测、根际定殖原位观测方法等亟需加强的研究方向。

巴马地区产胞外多糖益生乳酸菌的筛选及鉴定

采用含碳酸钙的快速初筛培养基,结合形态学观察、革兰氏染色、过氧化氢酶接触实验、菌落拉丝法和硫酸-苯酚法,从巴马自然发酵食品中筛选产胞外多糖乳酸菌,对其耐酸、耐胆盐、细胞表面疏水性和生物安全性进行研究,并采用分子生物学技术对筛选益生菌株进行鉴定。结果表明,从巴马自然发酵食品中筛选到6株产胞外多糖乳酸菌,编号分别为R2、R3、R4、R5、R6和R9;其中菌株R6多糖产量最高(106.73 mg/L),p H 3时耐酸性最佳,0.05%胆盐时延滞期最短(1.50 h),细胞表面疏水性最突出(在十六烷中的细胞表面疏水性达到了93.28%),对沙门氏菌、大肠杆菌和金黄色葡萄球菌的抑制作用最显著,且对多种常见抗生素敏感和不具有溶血活性,菌株R6被鉴定为植物乳杆菌(Lactiplantibacillus plantarum),可作为潜在益生菌株。

葡聚糖蔗糖酶在食品级乳酸菌中重组表达

为研究葡聚糖在乳酸菌中的合成情况以及结构分析,本研究将葡聚糖蔗糖酶DsrB与带有SP45分泌信号肽的乳酸乳球菌表达载体pNZ8148质粒载体进行连接,电转入乳酸乳球菌NZ9000中进行异源表达。经由一系列醇沉、三氯乙酸去除蛋白质、二次醇沉、透析以及纯化等步骤得到多糖溶液。采用高效液相色谱法检测单糖组成;利用HPSEC测定葡聚糖的分子量;通过傅里叶红外技术、核磁共振技术以及扫描电镜分析多糖的结构并观察多糖表面特征。结果表明,将葡聚糖蔗糖酶在乳酸菌中进行表达,在蔗糖浓度为10%的条件下,葡聚糖产量为10.54 g/L;单糖组成显示仅含有葡萄糖一种单糖;胞外多糖分子量测定结果为2.4×10^(6) Da;由傅里叶红外和核磁联合表明体外合成的胞外多糖仅含有α-(1,6)糖苷键;扫描电镜表示胞外多糖表面呈现为多孔状结构。在乳酸菌中异源表达,实现了葡聚糖的食品级生产,为葡聚糖于食品中应用提供了理论基础。