In vitro and in vivo antioxidant activity of exopolysaccharide fractions from Bifidobacterium animalis RH

The aim of the study was to purify the exopolysaccharides (EPS) produced from Bifidobacterium animalis RH, which was isolated from the feces of Bama centenarians in Guangxi of China, and evaluate their antioxidant activities in vitro and in vivo. 2 fractions, a neutral EPS fraction (EPSa) and an acidic EPS fraction (EPSb), were obtained and compared for antioxidative activity. In vitro, they both showed remarkable inhibition effect on lipid peroxidation and strong DPPH radical scavenging activity, hydroxyl radical scavenging activity, superoxide radical scavenging activity, in which the last two were measured by the electron spin resonance (ESR). In vivo, EPSa and EPSb were orally administrated for 30 days in a d-galactose induced aged mice model. As results, they both could significantly increase the activities of SOD, CAT and total antioxidant capacity (TAOC) in serums and glutathione GST in livers. They also could inhibit significantly the formation of MDA in serums and livers, and reduce the activity of MAO and lipofuscin accumulation in mice brain. Moreover, EPSb exhibited much higher antioxidant activities than EPSa in vitro and in vivo. The results suggested that EPS fractions of Bifidobacterium animalis RH had direct and potent antioxidant activities.     

α-Galactosidase of Bifidobacterium adolescentis DSM 20083

Bifidobacterium adolescentis was grown anaerobically in medium enriched with α-D-galactosides. α-Galactosidase (EC 3.2.1.22) was released from the cells by ultrasonic treatment and purified 36-fold by ultrafiltration, ammonium-sulphate precipitation, anion-exchange chromatography, and size-exclusion chromatography. Two protein bands were consistantly observed after sodium-dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE). Electrophoretically homogeneous α-galactosidase was only obtained by electroelution. The enzyme had an apparent molecular mass of 344 kDa and 79 kDa as judged by size-exclusion chromatography and SDS-PAGE, respectively. Activity-staining after nondenaturing SDS-PAGE indicated an apparent molecular mass of 145 kDa. Thus, a tetrameric structure of the protein is suggested. The α-galactosidase showed optimal activity at pH 5.5 and 55°C. Lower pH values and higher temperatures rapidly inactivated α-galactosidase. The enzyme hydrolyzed specifically α-galactosidic linkages, and α-(1-3)-linkages were hydrolyzed at a higher rate compared to α-(1-6)-linkages. Hydrolysis of galactosides followed normal saturation kinetics; K_M-values for p-nitrophenyl-α-galactopyranoside (p-NPG) and raffinose were calculated with 0.957 mM and 4.12 mM, respectively. Received: 7 August 1998 / Accepted: 9 September 1998     

Serological identification and molecular characterization of B (A) 02 subtype in patients and blood donors from Eastern China

This study was designed to identify the rare?type?ABO?blood?groups, B(A) 02, from Eastern China. Three samples with discordant serological results during routine blood type identification and four samples from one sample’ family were selected. All of them were detected by serological method. The exon 6 and 7 of the ABO genes were amplified by PCR and sequenced. They were typed as AsubB by serology and as BO by genotype. In AsubB samples, nt 700C>G mutation was detected in B gene, which was previously defined as B(A)02 alleles. In these seven samples, six showed B(A)02/O01 and one showed B(A)02/O02.B(A)02 allele was found to be more common in this study than B(A)04 which is considered to be more frequent than B(A)02. The careful identification of rare blood types is important for the safety of clinical blood transfusion.     

The Production of β-Fructofuranosidase from Bifidobacterium spp.

The productions of β-fructofuranosidase from Bifidohacterium longum A1, B. adolescentis G1, and four other strains of Bifidobacteria were investigated. All strains used in this study were grown in modified BL broth containing a mixture of fructooligosaccharides [1^F (1-β-D-fructofuranosyl)_n-1sucrose, GF_n (n = 2 – 5)] as the only carbon source. Hydrolyses of 1-kestose, sucrose, and inulin were detected in the extract of the cell. The highest activity on 1-kestose was detected in the extract of B. longum A1 followed by B. adolescentis G1. The other extracts weakly attacked 1-kestose. The relative activities of the extract of B. adolescentis G1 for 1-kestose, nystose, 1^F-fructosylnystose, sucrose, and inulin were 100, 82.5, 50.8, 28.3, and 15.0, respectively. The relative activities for various substrates differed from invertases (yeast β-fructofuranosidases) and exo-inulinase from Penicillium trzehinskii.     

Transglycosidase activity of Bifidobacterium adolescentis DSM 20083 α-galactosidase

Bifidobacterium adolescentis, a gram-positive saccharolytic bacterium found in the human colon, can, alongside other bacteria, utilise stachyose in vitro thanks to the production of an α-galactosidase. The enzyme was purified from the cell-free extract of Bi. adolescentis DSM 20083^T. It was found to act with retention of configuration (α→α), releasing α-galactose from p-nitrophenyl galactoside. This hydrolysis probably operates with a double-displacement mechanism, and is consistent with the observed glycosyltransferase activity. As α-galactosides are interesting substrates for bifidobacteria, we focused on the production of new types of α-galactosides using the transgalactosylation activity of Bi. adolescentisα-galactosides. Starting from melibiose, raffinose and stachyose oligosaccharides could be formed. The transferase activity was highest at pH 7 and 40 °C. Starting from 300 mM melibiose a maximum yield of 33% oligosaccharides was obtained. The oligosaccharides formed from melibiose were purified by size-exclusion chromatography and their structure was elucidated by NMR spectroscopy in combination with enzymatic degradation and sugar linkage analysis. The trisaccharide α-d-Galp-(1 → 6)-α-d-Galp-(1 → 6)-d-Glcp and tetrasaccharide α-d-Galp-(1 → 6)-α-d-Galp-(1 → 6)-α-d-Galp-(1 → 6)-d-Glcp were identified, and this indicates that the transgalactosylation to melibiose occurred selectively at the C-6 hydroxyl group of the galactosyl residue. The trisaccaride α-d-Galp-(1 → 6)-α-d-Galp-(1 → 6)-d-Glcp formed could be utilised by various intestinal bacteria, including various bifidobacteria, and might be an interesting pre- and synbiotic substrate. Received: 15 March 1999 / Received revision: 8 June 1999 / Accepted: 11 June 1999     

Gigantonoclea guizhouensis Gu et Zhi 的叶部解剖研究

一、材料来源和研究方法解剖研究所用的D-1标本是笔者于1984年5月在贵州水城大河边煤矿二采区新开辟的运输巷侧壁砂岩中采到的,其层位经对比与田宝霖和张连武(1980)描述的水城汪家寨矿区综合地层柱中龙潭组下部第8分层的中部砂岩相当。该叶片沉积时受水流冲击,以中脉为对称轴向远轴侧对折成约20°角状(图版Ⅰ,图2)。     

Isolation and Characterization of Two β-d-Glucosidases from Bifidobacterium breve 203

Two β-d-glucosidases were purified to homogeneity from Bifidobacterium breve 203: one ( β-d-glucosidase I; molecular weight, 96,000) showed reactivity toward p-nitrophenyl (p-NP) β-d-fucoside, 74% of that to p-NP β-d-glucoside, and the other ( β-dglucosidase II; molecular weight, 450,000) did not. They also differed in their thermal and pH stabilities. Laminaribiose, cellobiose and gentiobiose were hydrolyzed by β-d-glucosidase I, with 53%, 34% and 3% of the reactivity in the case of p-NP β-d-glucoside, and by β-dglucosidase II, with 53%, 6% and 107% of the reactivity. The reaction of β-dglucosidase I with p-NP β-dfucoside was enhanced by the addition of glucose and other monosaccharides to the reaction mixture, whereas that with p-NP β-dglucoside was not affected. The activity of β-dglucosidase II with p-NP β-dglucoside was inhibited by glucose.     

Characterization of Two Novel α-Glucosidases from Bifidobacterium breve UCC2003

Two α-glucosidase-encoding genes (agl1 and agl2) from Bifidobacterium breve UCC2003 were identified and characterized. Based on their similarity to characterized carbohydrate hydrolases, the Agl1 and Agl2 enzymes are both assigned to a subgroup of the glycosyl hydrolase family 13, the α-1,6-glucosidases (EC 3.2.1.10). Recombinant Agl1 and Agl2 into which a His₁₂ sequence was incorporated (Agl1_His and Agl2_His, respectively) exhibited hydrolytic activity towards panose, isomaltose, isomaltotriose, and four sucrose isomers—palatinose, trehalulose, turanose, and maltulose—while also degrading trehalose and, to a lesser extent, nigerose. The preferred substrates for both enzymes were panose, isomaltose, and trehalulose. Furthermore, the pH and temperature optima for both enzymes were determined, showing that Agl1_His exhibits higher thermo and pH optima than Agl2_His. The two purified α-1,6-glucosidases were also shown to have transglycosylation activity, synthesizing oligosaccharides from palatinose, trehalulose, trehalose, panose, and isomaltotriose.The gastrointestinal tract is inhabited by a complex community of microorganisms, also referred to as the microbiota, which are believed to play an important role in human health and disease (39). This concept has been driving extensive attempts to positively influence the composition and/or activity of the intestinal microbiota through the use of so-called probiotics and/or prebiotics. A probiotic has been defined as “a preparation or a product containing viable, defined microorganisms in sufficient numbers, which alter the microflora (by implantation or colonization) in a compartment of the host and by that exert beneficial health effect in this host” (48). A prebiotic has recently been (re)defined as “a selectively fermented ingredient that allows specific changes, both in the composition and/or activity in the gastrointestinal microbiota that confers benefits upon host well-being and health” (42). Finally, a synbiotic is the combination of a probiotic and a prebiotic (16).One of the dominant bacteria of the intestinal microbiota of humans and animals (51) is bifidobacteria. These are gram-positive, pleomorphic, and anaerobic bacteria that have received increasing scientific attention in recent years due to their perceived probiotic activity (15, 27). The growth of gut-derived bifidobacteria has been shown to be selectively stimulated by various dietary carbohydrates that can thus be considered as prebiotics (30). In this context, it is interesting to note that more than 8% of the identified genes on bifidobacterial genomes are predicted to be involved in sugar metabolism, thus indicative of extensive carbon source-degrading abilities (55, 56).Carbohydrate degradation has been extensively studied in a variety of different Bifidobacterium species (reviewed in reference 53). For example, various α- and β-galactosidases have been characterized in Bifidobacterium breve 203 (60), Bifidobacterium adolescentis DSM20083 (20), Bifidobacterium bifidum NCIMB41171 (18), and Bifidobacterium longum MB219 (43). A number of studies have also shown that Bifidobacterium spp. produce various α- and β-glucosidase activities (reviewed in reference 53), while Bifidobacterium infantis ATCC 15697 (58), Bifidobacterium lactis DSM10140(T) (13), and B. breve UCC2003 (45) have been reported to produce β-fructofuranosidases during growth on fructooligosaccharides. Additionally, starch-, amylopectin-, and pullulan-degrading activities in bifidobacteria have been investigated (36, 44). Several β-glucosidases have been biochemically characterized from a number of strains of bifidobacteria, e.g., B. adolescentis Int-57 (8), B. breve clb (35,) and Bifidobacterium sp. strain SEN (59). To date, only two α-glucosidases (AglA and AglB) have been described from B. adolescentis DSM20083 (54). AglA was shown to preferentially hydrolyze isomaltotriose, while AglB exhibits a high preference to maltose. Both AglA and AglB were also demonstrated to have transglycosylation activity. Aside from this report, little is known about the biochemical characteristics of α-glucosidase enzymes from bifidobacteria, although it is a common activity observed among these bacteria (41).Carbohydrates other than the commercially exploited prebiotics, e.g., fructooligosaccharides (such as inulin) and trans-galactooligosaccharides (42), have received relatively little attention with regard to their possible prebiotic properties. Such potential prebiotics are, for example, honey oligosaccharides, some of which are also interesting because of their noncariogenic properties (14). One of the predominant fractions of noncariogenic sugars in honey is isomaltulose (5, 46), also called palatinose or 6-O-α-d-glucopyranosyl-d-fructose, which is a reducing disaccharide and a functional isomer of sucrose. Palatinose possesses approximately one-third of the sweetness of sucrose and is very resistant to acid and invertase hydrolysis (29, 32). The hydrolysis and adsorption of palatinose in the small intestine thus occurs at a much slower rate than does those of sucrose (17), which results in a reduction of the postprandial plasma glucose and insulin levels (3), which means that most palatinose passes through the small intestine to present a growth substrate for elements of the colonic microbiota.In this study, we describe the identification of two genes, agl1 and agl2, present in the genome of B. breve UCC2003 and responsible for the hydrolysis of α-glycosidic linkages, such as those present in palatinose.     

Modeling Proximal Tubule Cell Homeostasis: Tracking Changes in Luminal Flow

During normal kidney function, there are routinely wide swings in proximal tubule fluid flow and proportional changes in Na⁺ reabsorption across tubule epithelial cells. This “glomerulotubular balance” occurs in the absence of any substantial change in cell volume, and is thus a challenge to coordinate luminal membrane solute entry with peritubular membrane solute exit. In this work, linear optimal control theory is applied to generate a configuration of regulated transporters that could achieve this result. A previously developed model of rat proximal tubule epithelium is linearized about a physiologic reference condition; the approximate linear system is recast as a dynamical system; and a Riccati equation is solved to yield the optimal linear feedback that stabilizes Na⁺ flux, cell volume, and cell pH. The first observation is that optimal feedback control is largely consigned to three physiologic variables, cell volume, cell electrical potential, and lateral intercellular hydrostatic pressure. Parameter modulation by cell volume stabilizes cell volume; parameter modulation by electrical potential or interspace pressure act to stabilize Na⁺ flux and cell pH. This feedback control is utilized in a tracking problem, in which reabsorptive Na⁺ flux varies over a factor of two, in order to represent a substantial excursion of glomerulotubular balance. The resulting control parameters consist of two terms, an autonomous term and a feedback term, and both terms include transporters on both luminal and peritubular cell membranes. Overall, the increase in Na⁺ flux is achieved with upregulation of luminal Na⁺/H⁺ exchange and Na⁺–glucose cotransport, with increased peritubular Na⁺–3HCO₃⁻ and K⁺–Cl⁻ cotransport, and with increased Na⁺, K⁺–ATPase activity. The configuration of activated transporters emerges as a testable hypothesis of the molecular basis for glomerulotubular balance. It is suggested that the autonomous control component at each cell membrane could represent the cytoskeletal effects of luminal flow.     

N-glycan Utilization by Bifidobacterium Gut Symbionts Involves a Specialist β-Mannosidase

Bifidobacteria represent one of the first colonizers of human gut microbiota, providing to this ecosystem better health and nutrition. To maintain a mutualistic relationship, they have enzymes to degrade and use complex carbohydrates non-digestible by their hosts. To succeed in the densely populated gut environment, they evolved molecular strategies that remain poorly understood. Herein, we report a novel mechanism found in probiotic Bifidobacteria for the depolymerization of the ubiquitous 2-acetamido-2-deoxy-4-O-(β-d-mannopyranosyl)-d-glucopyranose (Man-β-1,4-GlcNAc), a disaccharide that composes the universal core of eukaryotic N-glycans. In contrast to Bacteroidetes, these Bifidobacteria have a specialist and strain-specific β-mannosidase that contains three distinctive structural elements conferring high selectivity for Man-β-1,4-GlcNAc: a lid that undergoes conformational changes upon substrate binding, a tryptophan residue swapped between the two dimeric subunits to accommodate the GlcNAc moiety, and a Rossmann fold subdomain strategically located near to the active site pocket. These key structural elements for Man-β-1,4-GlcNAc specificity are highly conserved in Bifidobacterium species adapted to the gut of a wide range of social animals, including bee, pig, rabbit, and human. Together, our findings uncover an unprecedented molecular strategy employed by Bifidobacteria to selectively uptake carbohydrates from N-glycans in social hosts.     

论四射珊瑚的内生长线——以新疆早二叠世 Kepingophyllum aksuense Wu et Zhou 为例

新疆早二叠世 Kepingophyllum aksuense Wu et Zhou的内生长线发育良好,它们是由年季节温度的变化而形成的.据内生长线,可推算出k. aksuense Wu et Zhou的平均生长率为5mm/年.运用珊瑚的生长率及年龄可计算出沉积事件发生的频率.     

In Crystallo Selection to Establish New RNA Crystal Contacts

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Emplectopteridium alatum Kawasaki 的脉序*

Emplectopteridium alatum Kaw. 的脉序历来被描述为叶脉结网并具邻脉,主要基于具有邻脉这一特征,该种被归于美羊齿类.经笔者研究,这种植物没有邻脉而具伴网眼.这样,不但 E. alatum Kaw. 的分类位置需重新考虑,而且所谓 Emplectopteridium 演化系的基础也就完全瓦解.     

High-efficiency synthesis of oligosaccharides with a truncated β-galactosidase from Bifidobacterium bifidum

An exceptionally large beta-galactosidase, BIF3, with a subunit molecular mass of 188 kDa (1,752 amino acid residues) was recently isolated from Bifidobacterium bifidum DSM20215 [M?ller et al. (2001) Appl Environ Microbiol 67:2276-2283]. The BIF3 polypeptide comprises a signal peptide followed by an N-terminal beta-galactosidase region and a C-terminal galactose-binding motif. We have investigated the functional importance of the C-terminal part of the BIF3 sequence by deletion mutagenesis and expression of truncated enzyme variants in Escherichia coli. Deletion of approximately 580 amino acid residues from the C-terminal end converted the enzyme from a normal, hydrolytic beta-galactosidase into a highly efficient, transgalactosylating enzyme. Quantitative analysis showed that the truncated beta-galactosidase utilised approximately 90% of the reacted lactose for the production of galacto-oligosaccharides, while hydrolysis constituted a 10% side reaction. This 9:1 ratio of transgalactosylation to hydrolysis was maintained at lactose concentrations ranging from 10% to 40%, implying that the truncated beta-galactosidase behaved as a "true" transgalactosylase even at low lactose concentrations.     

不同气体环境对益生菌Bifidobacterium lactis V9生长的影响

Bifidobacterium lactis V9(B.lactis V9)是一株具有良好益生特性且遗传稳定的益生茵,工业化生产环境中气体组成关系着益生菌活菌数量,进而影响其益生功效.[目的]研究不同气体环境对B.lactis V9生长及代谢的影响.[方法]在固体MRS培养基、液体MRS培养基及巴氏杀菌脱脂乳接种B.lactis V9,于不同的气体环境中培养.[结果]在固体MRS培养基上,B.lactis V9在混合气体(N2∶H2∶CO2=80∶10∶10)环境中菌落形成较氮气环境(N2∶99.99％)多,在空气环境(N2∶O2≈79∶21)中菌落形成极少.B.lactis V9在MRS液体中培养24 h,混合气体环境下其活菌数(9.11±0.11 log CFU/mL)显著高于空气环境下的活菌数(8.04±0.10 log CFU/mL) (P＜0.01),在混合气体环境下B.lactis V9代谢生成的乙酸和乳酸量分别为12.79±0.86 mmol/L和11.99±0.73 mmol/L,显著高于在空气环境中生成量0.65±0.07 mmol/L和2.75±0.57 mmol/L (P＜0.01),乙酸/乳酸比值分别为1.06∶1和0.24∶1.B.lactis V9在巴氏杀菌脱脂乳中发酵18h,混合气体环境下pH值(4.48±0.07)显著低于空气环境下的pH值(5.03±0.12) (P＜0.01),混合气体环境下其活菌数(9.02±0.15 log CFU/mL)显著高于空气环境下的活菌数(8.53±0.08 log CFU/mL) (P＜0.01).混合气体和空气环境下发酵脱脂乳产生的乙酸和乳酸量分别为60.52±2.30 mmol/L、5.17±1.02 mmol/L和16.86±0.34 mmol/L、5.92±0.81 mmol/L,乙酸/乳酸的值分别为11.71∶1和2.85∶1.[结论]在N2∶H2∶CO2=80∶10∶10混合气体环境下有利于B.lactis V9在液体MRS和脱脂乳中生长,其活菌数可以增加0.5-1个数量级.这一研究结果也可为B.lactis V9益生菌发酵乳的生产和产品中B.lactis V9活菌培养计数提供指导.     

Synthesis of α-galacto-oligosaccharides by a cloned α-galactosidase from Bifidobacterium adolescentis

A genomic library of Bifidobacterium adolescentis was constructed in Escherichia coli and a gene encoding an -galactosidase was isolated. The identified open reading frame showed high similarity and identity with bacterial -galactosidases, which belong to Family 36 of the glycosyl hydrolases. For the purification of the enzyme from the medium a single chromatography step was sufficient. The yield of the recombinant enzyme was 100 times higher than from B. adolescentis itself. In addition to hydrolytic activity the -galactosidase showed transglycosylation activity and can be used for the production of -galacto-oligosaccharides.     

Brachyspira asperella Yu et Zhang的个体发育

对Brachyspira asperella Yu et Zhang的40个个体的测量统计表明,此种在生长发育过程中可以区分出3个生长阶段:幼年期壳体生长速度缓慢,壳形卵圆形,胎壳乳突状,壳口圆形;成年期生长迅速,壳高显著大于壳宽的增长率,壳形卵圆形-长卵圆形,壳口卵圆形;老年期生长速度缓慢,壳形长卵圆形,壳饰强烈。这3个生长期代表了此种的个体发育过程。研究化石腹足类种的个体发育过程不仅能了解这一类群的系统发生史、生物演化过程及生物间的亲缘关系,而且能提高物种研究程度,避免分类上的错误。     

论双壳类的 Pteroperna 及其模式种 Pteroperna costatula (Deslongchamps)

翼股蛤(Pteroperna)是 J.Morris 和 J.Lycett 于1853年建立,但该属的分类位置当时并不明确,仅仅在讨论中提到可能为 Pterinea的亚属。Cox(1940)讨论 Pteroperna 时则将其作为翼蛤科(Pteriidae)的属而沿用至今。笔者于1984年在唐古拉山雁石坪地区测制侏罗纪地层剖面时,在同一层位中采集到许多隶属于 Pteroperna costatula 的标本,它们的壳饰特征、壳体形状和英国侏罗纪大鲕状灰岩(Great O(?)lite)中产出的同种标本几无差异。标本大多保存良好,特别是它们的壳层和其围岩在弱酸中可产生差异溶蚀,利用酸蚀法可得到许多暴露出铰系构造和软体印痕的标本,其中有一些反映出个体发育各个阶段的生长特     

The Final Size of an Epidemic and Its Relation to the Basic Reproduction Number

We study the final size equation for an epidemic in a subdivided population with general mixing patterns among subgroups. The equation is determined by a matrix with the same spectrum as the next generation matrix and it exhibits a threshold controlled by the common dominant eigenvalue, the basic reproduction number R₀{\mathcal{R}_{0}}: There is a unique positive solution giving the size of the epidemic if and only if R₀{\mathcal{R}_{0}} exceeds unity. When mixing heterogeneities arise only from variation in contact rates and proportionate mixing, the final size of the epidemic in a heterogeneously mixing population is always smaller than that in a homogeneously mixing population with the same basic reproduction number R₀{\mathcal{R}_{0}}. For other mixing patterns, the relation may be reversed.