Sugar constituents of fucose-containing polysaccharides from various Japanese brown algae

Sugar constituents of the fucose-containing polysaccharides (FCPs) from 21 species of brown algae were analyzed. FCPs were extracted with hot water (100 °C, 4 h), separated by precipitation with 20% (v:v) ethanol in the presence of 0.05 M MgCl₂ to remove contaminating soluble alginate, and purified by DEAE-Sephadex column chromatography. The samples were hydrolyzed with HCI, and neutral sugar and uronic acid were separated by anion exchange chromatography. Their amounts were determined by gas-liquid chromatography. The neutral sugars in the FCPs from Ishige okamurae, Laminaria ochotensis, Myelophycus simplex, Padina arborescens and Sargassum thunbergii all contained arabinose, fucose, galactose, glucose, mannose, rhamnose and xylose residues. The FCPs from Ishige okamurae, Padina arborescens, Sargassum hemiphyllum, S. patents and S. sagamianum contained the four uronic acids, galacturonic acid, glucuronic acid, guluronic acid and mannuronic acid.     

Recherches Cytochimiques sur la Microsporidie Nosema bomycis au cours de son Développement chez le Ver à Soie (Bombyx mori)*

RESUME. La Microsporidie Nosema bombycis, Protozoaire parasite agent de la pébrine du ver à soie, a étéétudiée cytochimiquement à la fois en microscopie photonique et électronique. Les examens ont porté sur la détection et la localisation des acides nucléiques (ADN et ARN), des polysaccharides, de la phosphatase acide, au cours des différents stades du développement dans les cellules de I'hôte (du schizonte à la spore). Les principaux résultats concernent les observations relatives aux polysaccharides et à la phosphatase qui ne sont détectés qu'au stade de la spore et ne sont pas observés au stade du schizonte. Les polysaccharides sont présents au niveau du sac polaire, du filament polaire et sur la membrane cytoplasmique; la phosphatase acide est localisée au niveau du sac polaire, du filament polaire et dans la vacuole postérieure. SYNOPSIS. Nosema bombycis, agent of pebrine disease of silkworm, was studied cytochemically, using both light and electron microscopy. Presence of nucleic acids (DNA and RNA), polysaccharides, and acid phosphatases was demonstrated and localization of these substances was determined in various stages of the parasite (from the schizont to the spore). DNA and RNA were detected in all these stages. Polysaccharides and acid phosphatase were found in the spore but not in the schizogonic stages. Polysaccharides were detected in the polar cap, the polar filament, and the limiting membrane of the cytoplasm of the spore. Acid phosphatase was found in the polar cap, the polar filament, and the posterior vacuole.     

Observations of a set of isomeric anti-lactose antibodies directed against a group D streptococcal polysaccharide

Sets of isomeric anti-lactose antibodies with specificity for the lactose units of a cell wall polysaccharide fromStreptococcus faecalis strain N were induced in rabbits immunized with a vaccine of nonviable cells of the organism. Such sets of anti-lactose antibodies were isolated from the serum of immunized animals by affinity chromatography on lactosyl-Sepharose. Gel electrofocusing experiments showed that the preparations consisted of multiprotein components. One preparation of antibodies of 13 isomers was separated into homogeneous components by liquid isoelectrofocusing. The individual isomeric antibodies exhibit specificity for the lactose units of the antigenic polysaccharide, possess isoelectric points in the range of 5.9–8.0, and belong to the IgG class of immunoglobulins, and each member yields one light chain and one heavy chain on dissociation in sodium dodecyl sulfate (SDS) and mercaptoethanol. These results have been interpreted as evidence for the assembly of the chains of isomeric antibodies by a single-chain pairing mechanism.     

灵芝线粒体及其对菌丝生长、主要活性成分影响的分析

线粒体是为细胞提供能量（ATP）的细胞器,携带着自己的DNA——mtDNA,已有多种灵芝属菌株的线粒体基因组相继报道,但对于种内的线粒体基因组的分析较少,对核相同、线粒体不同的菌株间差异的研究也鲜有报道。本研究对两株灵芝线粒体基因组进行组装注释,根据差异片段构建分子标记进行种间分类。结果显示：两株灵芝线粒体基因组大小分别为49 233bp、61 563bp的闭环结构,含有15个常见蛋白编码基因,rRNA大小亚基基因及26个携带氨基酸的tRNA基因,其差异区段主要为内含子序列、大亚基及基因间区。根据cob、cox2基因序列能够进行灵芝种间区分,用于灵芝种间鉴定。本研究还根据灵芝119、灵芝无孢的单核菌株构建同核异质体（TY-119、TY-W）,分析线粒体对菌落形态、菌丝生长速度及多糖、三萜成分的影响,结果显示：同核异质体TY-W与TY-119菌落形态上有一定差异,同核异质体TY-W菌丝生长速度为4.77mm/d,是TY-119菌丝生长速度4.50mm/d的1.06倍,同核异质体TY-119菌丝、子实体阶段多糖含量分别为4.45mg/g、12.14mg/g是TY-W菌丝体多糖含量（3.23mg/g）的1.38倍、子实体多糖含量（10.24mg/g）的1.19倍;同核异质体TY-W菌丝、子实体阶段的三萜含量分别为6.82mg/g、11.45mg/g是同核异质体TY-119菌丝体三萜含量（9.26mg/g）的0.74倍,子实体三萜含量（9.10mg/g）的1.26倍。利用高效液相色谱分析同核异质体中灵芝酸含量显示同核异质体TY-W灵芝酸A、灵芝酸E、灵芝酸F含量分别为3.77μg/mL、14.29μg/mL、12.91μg/mL;是TY-119灵芝酸A含量（2.59μg/mL）的1.46倍、灵芝酸E含量（13.65μg/mL）的1.17倍、灵芝酸F含量（12.72μg/mL）的1.06倍。对同核异质体菌丝、子实体阶段多糖、三萜合成通路关键基因（pgm、ugp、gls、hmgs、hmgr、mvd、fps、sqs）表达量进行检测,显示两菌株间多数基因具有显著性差异。结果表明线粒体的不同会影响灵芝菌落形态、菌丝生长速度及多糖、三萜的含量,有助于我们进一步研究线粒体基因组。     

Antibacterial potential of Forsythia suspensa polysaccharide against resistant Enterobacter cloacae with SHV‐12 extended‐spectrum β‐lactamase (ESBL)

In this study, a homogenous polysaccharide (FSP), with an average molecular weight of 9.08 × 10⁴ Da, was isolated from Forsythia suspense and its antibacterial potential against Enterobacter cloacae producing SHV‐12 ESBL was investigated. Growth kinetics, in vitro competition and biofilm formation experiments demonstrated that SHV‐12 ESBL contributed to a fitness benefit to E cloacae strain. The antibacterial activity of FSP (2.5, 5.0 and 10.0 μg/mL) was tested against E cloacae bearing SHV‐12 ESBL gene using bacterial sensitivity, agar bioassay and agar well diffusion assays. It was found that the addition of FSP demonstrated potent antibacterial activities against this bacterial as showed by the decrease of bacterial growth and the increase of the inhibition zone diameter. Furthermore, SHV‐12 ESBL gene expression was decreased in E cloacae strain following different FSP treatment in a concentration‐dependent manner. In conclusion, these data showed that FSP exhibited potent good antibacterial activity against E cloacae producing SHV‐12 ESBL via inhibition of SHV‐12 ESBL gene expression, which may promote the development of novel natural antibacterial agents to treat infections caused by this drug‐resistant bacterial pathogen.     

Protective effect of Agrimonia pilosa polysaccharides on dexamethasone‐treated MC3T3‐E1 cells via Wnt/β‐Catenin pathway

A water‐soluble polysaccharide (APP‐AW) was isolated from Agrimonia pilosa and prepared to three sulphated derivatives (S1, S2 and S3). The results showed that pre‐treatment with APP‐AW, S1, S2 and S3 each at the concentration of 50 μg/mL for 48 hours was able to prevent cytotoxicity induced by 1 μmol/L dexamethasone (Dex) in MC3T3‐E1 cells via inhibition of apoptosis, which is in line with the findings in flow cytometry analysis. Meanwhile, the decreased ALP activity, collagen content, mineralization, BMP2, Runx2, OSX and OCN protein expression in DEX‐treated MC3T3‐E1 cells were reversed by the addition of APP‐AW, S1, S2 and S3. Moreover, APP‐AW, S1, S2 and S3 rescued DEX‐induced increase of Bax, cytochrome c and caspase‐3 and decrease of Bcl‐2, Wnt3, β‐catenin and c‐Myc protein expression in MC3T3‐E1 cells. Our findings suggest that pre‐treatment with APP‐AW, S1, S2 and S3 could significantly protect MC3T3‐E1 cells against Dex‐induced cell injury via inhibiting apoptosis and activating Wnt/β‐Catenin signalling pathway, thus application of these polysaccharides may be a promising alternative strategy for steroid‐induced avascular necrosis of the femoral head (SANFH) therapy.     

Development and validation of an experimental and theoretical method for the chiral discrimination of dinotefuran

Dinotefuran is a low-cost agrochemical considered a highly toxic product. In this sense, there is a need for its constant environmental, biological, and food control, aiming to ensure its use to humans as well as to preserve biodiversity and ecosystems. In the present work, we developed an experimental and theoretical method for dinotefuran chiral discrimination. According to the main results, the dinotefuran enantioselective separation was efficiently optimized by high-performance liquid chromatography evaluating the influence of different percentage compositions in the mobile phase to improve the resolution of the peaks in the chromatogram. The novelty of this work was the proposition of a reduced molecular model for the chiral selector amylose-Tris-(3,5-dimethylphenylcarbamate) polysaccharide that was able to adequately describe at the molecular level its interaction with the dinotefuran enantiomers. Besides, the thermodynamic and structural parameters obtained via density functional theory calculations pointed out the chiral discrimination as well as the enantiomeric elution order of the analyte studied, confirming the experimental data, thus validating our proposed method. Finally, hydrogen bonds and repulsive interactions played a key role in the discrimination between the diastereomeric complexes, and consequently, for the dinotefuran enantioselective separation.