A strategy for high cell density culture of heterotrophic microalgae with inhibitory substrates

Substrate inhibition is one of the major problems preventing high cell densities of microalgae in heterotrophic culture, so the possibility of overcoming the problem by various culture techniques was examined. It was found that perfusion culture may be the most appropriate technique for high cell densities in heterotrophic culture using inhibitory substrates. An experimental example in which a hollow fibre cell recycle system (HFCRS) was employed to achieve high cell densities of Chlamydomonas reinhardtii on acetate under heterotrophic conditions of growth was demonstrated. The cell density in the HFCRS was much higher than that reported in the literature for this species.     

高胆固醇高脂肪膳食对家兔β-VLDL组成及其与巨噬细胞相互作用的影响的初步研究

给家兔喂以1％胆固醇及10％菜油(A组)或猪油(B组)50多天后A组血胆固醇水平(824.2±265.1mg/dl)明显低于B组(1666±693.8mg/dl);A组甘油三酯水平(51.9±19.1mg/dl)亦低于B组(104±40.2mg/dl)。二组家兔的β—VLDL的脂类组成无差别,但A组β—VLDL的apoE高于B组,分别为45.2％及37.5％。高分子量apoB(apoB_h)为33.6％,低于B组β-VLDL(47.3％)。A组β-VLDL促进小鼠腹腔巨噬细胞胆固醇堆积的程度大于B组,可能与apoE含量高有关。我们认为多不饱和脂酸减轻动脉粥样硬化(As)的作用不在于改变脂蛋白构成后阻碍泡沫细胞的形成而是促进β—VLDL从体内清除。     

Cloning, sequence analysis and expression of gene alyVI encoding alginate lyase from marine bacterium Vibrio sp. QY101.

The gene (alyVI) encoding an alginate lyase of marine bacterium Vibrio sp. QY101, which was isolated from a decaying thallus of Laminaria, was cloned using a strategy of combined degenerate PCR and long range-inverse PCR (LR-IPCR), then sequenced and expressed in Escherichia coli. Gene alyVI was composed of a 1014 bp open reading frame (ORF) encoding 338 amino acid residues. The calculated molecular mass of alyVI product is 38.4 kDa, but a signal peptide is cleaved off, leaving a mature protein of 34 kDa. AlyVI was purified from culture supernatants to electrophoretic homogeneity using affinity chromatography. AlyVI was most active at pH 7.5 and 40 degrees C in the presence of 1 mM ZnCl2. A nine-amino-acid consensus region (YXRESLREM), which was only found in polyguluronate lyases, was also observed in the amino-terminal region of AlyVI. However, AlyVI could degrade both M block and G block. These results indicate that a novel alginate lyase-encoding gene has been cloned.     

Chiral resolution of alpha,alpha'-bis[3-(N,N-diethylcarbamoyl)piperidino]-p-xylene, a novel antiplatelet compound.

alpha,alpha'-Bis[3-(N,N-diethylcarbamoyl)piperidino]-p-xylene dihydrobromide, a novel antiplatelet agent, was resolved into three isomers A, B, and C, on a chiral alpha 1-acid glycoprotein analytical column using a mobile phase of 0.025 M phosphate buffer containing 0.025 M tetrabutylammonium hydrogen sulfate, at a pH of 6.5. The effect of molarity, temperature, pH, flow rate, and organic modifiers on the enantioselectivity was examined. Based on circular dichroic spectra at 220 nm, A and C appear to be the (-)- and (+)-enantiomers, respectively, and B the meso diastereomer. Attempts at resolution using Pirkle type columns gave unsatisfactory results. It appears that both hydrophobic and polar interactions between the compound and the stationary phase are important determinants of resolution.     

Detection of covalent enzyme-substrate complexes of nitrilase by ion-spray mass spectroscopy

Nitrilase from Rhodococcus ATCC 39484 was found to consist of two species of Mr 40,258 +/- 2 and 40,388 +/- 2 Da. When the enzyme was incubated with nitrile substrates and the reaction quenched with acid, higher Mr species were observed. The mass differences were consistent with addition of a substrate molecule to each species. These results represent the first reported demonstration that this, or any other nitrilase forms a covalent intermediate with its substrates. The observation that the intermediate, suggested to be either a thioimidate or an acylenzyme, can be trapped by acidification indicates that the rate of breakdown of the intermediate is rate-limiting.     

Participation of lipopolysaccharide in hyperplasic adipose expansion: Involvement of NADPH oxidase/ROS/p42/p44 MAPK‐dependent Cyclooxygenase‐2

Obesity is a world‐wide problem, especially the child obesity, with the complication of various metabolic diseases. Child obesity can be developed as early as the age between 2 and 6. The expansion of fat mass in child age includes both hyperplasia and hypertrophy of adipose tissue, suggesting the importance of proliferation and adipogenesis of preadipocytes. The changed composition of gut microbiota is associated with obesity, revealing the roles of lipopolysaccharide (LPS) on manipulating adipose tissue development. Studies suggest that LPS enters the circulation and acts as a pro‐inflammatory regulator to facilitate pathologies. Nevertheless, the underlying mechanisms behind LPS‐modulated obesity are yet clearly elucidated. This study showed that LPS enhanced the expression of cyclooxygenase‐2 (COX‐2), an inflammatory regulator of obesity, in preadipocytes. Pretreating preadipocytes with the scavenger of reactive oxygen species (ROS) or the inhibitors of NADPH oxidase or p42/p44 MAPK markedly decreased LPS‐stimulated gene expression of COX‐2 together with the phosphorylation of p47^phox and p42/p44 MAPK, separately. LPS activated p42/p44 MAPK via NADPH oxidase‐dependent ROS accumulation in preadipocytes. Reduction of intracellular ROS or attenuation of p42/p44 MAPK activation both reduced LPS‐mediated COX‐2 expression and preadipocyte proliferation. Moreover, LPS‐induced preadipocyte proliferation and adipogenesis were abolished by the inhibition of COX‐2 or PEG₂ receptors. Taken together, our results suggested that LPS enhanced the proliferation and adipogenesis of preadipocytes via NADPH oxidase/ROS/p42/p44 MAPK‐dependent COX‐2 expression.     

成熟促进因子对克隆重构胚核重编程的调控

成熟促进因子(maturation promoting factor,MPF)由催化亚单位P34cdc2和调节亚单位cyclin组成,对细胞周期的调控起着重要作用。目前,在核移植研究中发现：供体核在MPF的作用下发生核膜破裂(nuclear envelop breakdown．NEBD)和早熟染色体凝集(premature chromosome condensation,PCC),促进了核、质蛋白质因子的交换,有利于核重编程的进行。PCC还会对供体核的倍性及形态产生影响。     

Linking seasonal inorganic nitrogen shift to the dynamics of microbial communities in the Chesapeake Bay

Seasonal shifts of dissolved inorganic nitrogen (DIN) and the dynamics of microbial communities for nitrogen transformation were investigated in the water column of Chesapeake Bay. The relative abundance of nitrogen over phosphorus (N*) showed a strong seasonal and spatial pattern: gradually decreased from upstream to downstream; high in winter and low in summer. Because the phosphorus concentration remained relatively stable, the spatiotemporal pattern of N* implied that a substantial fraction of DIN was removed in the bay, especially in summer. Correlation analyses indicated the functional microbial communities and environmental variables, such as temperature, dissolved oxygen, salinity, played important roles for connecting the seasonal variation of N*. Among them, temperature was the trigger factor. High temperature in the summer induced the growth of functional microbes, which subsequently consumed a large portion of DIN inputted from the tributaries and reduced the N*. The current study provided the relative importance of microbial communities and environmental variables in driving the DIN loss in the bay.