链霉菌CPCC 203577产生的napyradiomycins的分离与鉴定

目的对一株具有抗菌活性的链霉菌CPCC 203577产生的次级代谢产物进行系统研究。方法采用ISP2琼脂平板发酵培养链霉菌CPCC 203577,乙酸乙酯提取发酵培养物,获得粗提物;粗提物经反相色谱柱、凝胶色谱柱、制备型TLC和半制备HPLC等分离纯化获得目标化合物纯品;MS和NMR等确定化合物结构;琼脂稀释法测定抗菌活性。结果从链霉菌CPCC 203577中分离鉴定了含萘醌并吡喃母核的3个已知化合物,3-chloro-6,8-dihydroxy-8-α-lapachone(1)、16-dechloro-16-hydroxynapyradiomycin C2(2)和napyradiomycin A2(3)。化合物1具有较强的抗革兰氏阳性细菌活性,最低抑菌浓度(MIC)值为4~8μg/ml;化合物2和3没有抗菌活性。结论链霉菌CPCC 203577具有丰富的次级代谢产物合成能力,产生napyradiomycins类化合物。     

Purification,characterization, and biological cytotoxic activity of the extracellular cholesterol oxidase produced by Castellaniella sp. COX

A new bacterial strain producing extracellular cholesterol oxidase (ChOx) was isolated and identified as Castellaniella sp. COX. The ChOx was purified by salting-out and ion-exchange chromatography up to 10.4-fold, with a specific activity of 15 U/mg with a molecular mass of 59 kDa. The purified ChOx exhibited pH 8.0 and temperature 40°C for its optimum activity. The enzyme showed stability over a wide pH range and was most stable at pH value 7.0, and at pH 8.0, it retained almost 86% of its initial activity after 3 h of incubation at 37°C. The enzyme possessed a half-life of 8 h at 37°C, 7 h at 40°C, and 3 h at 50°C. A Lineweaver–Burk plot was calibrated to determine its K_m (0.16 mM) and V_max (18.7 μmol·mg⁻¹·min⁻¹). The ChOx activity was enhanced with Ca²⁺, Mg²⁺, and Mn²⁺ while it was inhibited by Hg²⁺, Ba²⁺, Fe²⁺, Cu²⁺, and Zn²⁺ ions. Organic solvents like acetone, n-butanol, toluene, dimethyl sulfoxide, chloroform, benzene, and methanol were well tolerated by the enzyme while iso-propanol and ethanol were found to enhance the activity of purified ChOx. ChOx induced cytotoxicity with an IC₅₀ value of 1.78 and 1.88 U/ml against human RD and U87MG established cell lines, respectively, while broadly sparing the normal human cells.     

Anti-inflammatory activity of lansais from endophytic Streptomyces sp. SUC1 in LPS-induced RAW 264.7 cells

This research was undertaken to find the in vitro inflammatory action of lansai A–D produced by Streptomyces sp. SUC1. We investigated the effects of lansai C not only on the formation of nitric oxide (NO), prostaglandin E₂ (PGE₂), tumour necrosis factor (TNF-α), interleukin (IL)-1α, IL-6 and IL-10, but also on inducible NO synthase and cyclooxygenase-2 (COX-2) in lipopolysaccharide (LPS)-induced murine macrophage RAW 264.7 cells. The data obtained were consistent with the modulation of inducible nitric oxide synthase enzyme expression. A similar fashion was also observed when LPS-induced PGE₂ release and COX-2 expression were tested. The significant inhibitory effects were shown in concentration-dependent manners. In addition, lansai C also mildly but significantly reduced the formation of TNF-α. These findings support the application of lansai C as anti-inflammatory agent.     

Antibacterial,antioxidant and anticancer activities of biphenyls from Streptomyces sp. BO-07: an endophyte in Boesenbergia rotunda (L.) Mansf A.

Strain BO-07 was isolated from the root tissue of Boesenbergia rotunda (L.) Mansf A. and identified as Streptomyces sp. on the basis of morphology, chemotaxonomy and 16S rDNA sequencing. The fractionation of the crude extract from strain BO-07 cultures led to the isolation of two biphenyls: 3′-hydroxy-5-methoxy-3,4-methylenedioxybiphenyl (1) and 3′-hydroxy-5,5′-dimethoxy-3,4-methylenedioxybiphenyl (2); these compounds and the crude extract had potent antibacterial activity against Gram-positive bacteria, and antioxidant and anticancer activities. These compounds showed the highest activity against Staphylococcus aureus ATCC25932, Bacillus cereus ATCC7064 and Bacillus subtilis ATCC6633 with a minimum inhibitory concentration value of 0.5?µg/ml and minimum bactericidal concentration of 2–8?µg/ml. Compounds 1 and 2 showed the highest (1, 1diphenyl-2-picryl hydrazyl) DPPH antioxidant activity with a scavenging concentration (SC₅₀) value of 85.84 and 88.26?µg/ml, respectively, and also showed strong cytotoxicity against all the three cancer cell lines (HeLa, HepG2 and Huh7) at an IC₅₀ value of 3.04–20.30?μg/ml. Both the compounds were less toxic on normal cells (L929) than on the investigated cancer cell lines.     

Purification, characterization and thermodynamics of antifungal protease from Streptomyces sp. A6

A 20 kDa antifungal serine protease from Streptomyces sp. A6 was purified to 34.56 folds by gel permeation chromatography. The enzyme exhibited highest activity at neutral to near alka- line pH 7-9 and 55 °C. Neutral surfactant triton X-100 enhanced the activity by 4.12 fold. The protease activity also increased (109.9-119%) with increasing concentration of urea (2-8 mole/l). The enzyme was identified as serine protease with 67% similarity to SFase 2 of Streptomyces fradiae by MALDI-LC-MS/MS analysis. Determination of kinetic constants k(m) , V(max) , k(cat) and k(cat) /k(m) suggested higher affinity of enzyme for N-Suc-Ala-Ala-Val-Ala-p NA (synthetic substrate for chymotrypsin activity). The enzyme was highly stable at temperature prevailing under field conditions (40 °C) as apparent from K(d) and t(1/2) values, 0.0065 and 106.75 min, respectively and high ΔG* and negative ΔS * values, 87.17 KJ/mole and -126.95 J/mole, respectively. Thermal stability and increased activity of protease in presence of commonly used chemical fertilizer, urea, suggested its feasibility for agricultural applications. The present study is the first report on thermodynamic and kinetic properties of an antifungal protease from Streptomyces sp. A6. The study reflects potential of this enzyme for biocontrol of fungal plant pathogens.     

Scanning electron microscopy of red blood cells from eleven species of marsupial

Samples of red of blood cells (RBC), washed free of plasma, from eleven marsupial species were examined in a Jeol JSM-6300 F scanning electron microscope. The diameters of the RBC, lying completely flat or exactly on edge, were measured on photographs using a binocular enlarging optical system with a calibrated eye piece. RBC from the following species were studied: bandicoot (Isoodon macrourus), bilby (Macrotis lagotis sagitta), Bennett's wallaby (Macropus rufogriseus), Goodfellow's tree kangaroo (Dendrolagus goodfellowi), koala (Phascolarctos cinereus), parma wallaby (Macropus parma), red kangaroo (Macropus rufus), swamp wallaby (Wallabia bicolor), Tammar wallaby (Macropus eugenii), Tasmanian devil (Sarcophilus harrish) and whiptail wallaby (Macropus parryi). In all species the RBC were biconcave discs. The major morphological difference was in the size of the cells. Taking the human RBC as a reference (with a diameter of 8.0 m), the RBC of the Tasmanian devil, bilby, bandicoot and Goodfellow's tree kangaroo were 7 m in diameter, whereas those of the other marsupials ranged from 7.8 to 8.6 m.     

Comparative studies of the same adenocarcinoma cells,macrophages, and mesothelial cells by light microscopy,scanning electron microscopy,and transmission electron microscopy

The identification of cells in body cavities of cancer patients is sometimes difficult to make. In order to make a definite cytological diagnosis, we observed the same cells by using light microscopy (LM)-scanning electron microscopy (SEM)-transmission electron microscopy (TEM). In this study we first stained cells by the Papanicolaou method after fixation in 1% glutaraldehyde for LM, and then attempted to observe them successively by SEM-TEM after fixation in 1% paraformaldehyde and 1.25% O^s0⁴. Our method and procedures in examining successively one and the same cells in body cavity fluids by using LM, SEM, and TEM ensured accurate discrimination among adenocarcinoma cells, mesothelial cells, and macrophages. The results of this study suggest that LM-SEM-TEM may be of diagnostic value in distinguishing among mesothelial cells, macrophages, and adenocarcinoma cells. This method also succeeded in disclosing differences between the ultrastructure of the cell surfaces, and those of the cytoplasm, and of the nuclei It is desirable that LM-SEM-TEM observation can be introduced into various aspects in order to obtain an improvement in the diagnosis by cytologic examination, the judgment of therapeutical effects, drug selection, and prognostic presumption. Diagn Cytopathol 1994; 11:333–342. © 1994 Wiley-Liss, Inc.     

Effect of carbon source on polysaccharide production by alginate-entrapped Aureobasidium pullulans ATCC 42023 cells

The production of the exopolysaccharide pullulan using entrapped cells of the fungus Aureobasidium pullulans ATCC 42023 was investigated relative to carbon source. Fungal cells grown on glucose or sucrose as a carbon source were entrapped in calcium alginate beads and found to be capable of synthesizing the polysaccharide for two production cycles. Using 2.5% glucose or sucrose as a carbon source, productivity was 18.3 or 21.9 mg polysaccharide/g cells × h, respectively after the initial production cycle and decreased to 9.6 or 8.5 mg polysaccharide/g cells × h, respectively, after the second production cycle. Independent of carbon source, the entrapped fungal cells exhibited a higher yield during the initial cycle than the second production cycle while the entrapped ATCC 42023 cells elaborated a polysaccharide with a higher pullulan content during the second production cycle compared to the initial production cycle.     

Optimization of indigo production by a newly isolated Pseudomonas sp. QM

Optimization of indigo production process from indole using a newly isolated phenol‐degrading bacterial strain was performed by Plackett‐Burman design and response surface methodology. The strain designated as QM was identified as Pseudomonas sp. according to 16S rDNA analysis. Spectrum analysis of indole biotransformation products revealed the presence of indigo and a by‐product indirubin. To improve indigo yield, Plackett‐Burman design was used to select significant factors from 8 viriables. Then response surface methodology based on a 2³ central composite design was used to further optimize the transformation process. Under the optimal conditons, strain QM can produce 27.20 mg/l indigo after 24 h cultivation at 30 °C, which was 151.3% higher than that from the initial conversion condition. The results indicated that Pseudomonas sp. QM should be a potential candidate for indigo industial production. (© 2012 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)     

Bioremediation of chromium(VI) contaminated soil by Streptomyces sp. MC1

This work provides quantitative information on Cr(VI) reduction in soil samples by an indigenous actinomycete. Streptomyces sp. MC1, previously isolated from sugarcane, has shown ability to reduce Cr(VI) in liquid minimal medium. A reduction of 100 and 75% was obtained at initial Cr(VI) concentrations of 5 and 50 mg l^–1, respectively, after 48 h of incubation. Bioremediation ability of Streptomyces sp. MC1 was assayed in soil extracts and soil samples. Relative growth of Streptomyces sp. MC1 was 77 and 38% when grown in soil extract with 10 and 50 mg l^–1 of Cr(VI), respectively. MC1 was able to reduce 30% of Cr(VI) after 96 h of incubation with 10 mg l^–1 of Cr(VI), and reduction coincided with the exponential growth phase at pH 7 and 30 °C. In soil samples, Streptomyces sp. MC1 was able to reduce up to 94% of the Cr(VI) bioavailability (50 mg kg^–1) after 7 d. These results were compared with non‐inoculated soil samples with Cr(VI). Bioremediation activity of Streptomyces sp. MC1 was not inhibited by natural soil microbial flora. Besides, Streptomyces sp. MC1 growth was not inhibited by 50 mg kg^–1 of Cr(VI). In contrast to findings obtained by other authors, our results showed almost complete Cr(VI) removal from soil without any previous treatment, and without addition of any substrate and with a normal soil humidity level. These results confirm the Cr(VI)‐contaminated soil bioremediation potential of Streptomyces sp. MC1. (© 2009 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)     

The effect of cytochalasin B on chick mesoderm cells as studied by scanning electron microscopy

Summary The effects of cytochalasin B (CCB) on chick mesoderm cells in vivo was examined by scanning electron microscopy (SEM). The embryos were mounted for New Culture and the mesoderm exposed by dissecting off the endoderm. Cytochalasin B was suspended in saline and the embryos flooded with the suspension. Control embryos were treated with saline alone. The embryos were reincubated for varying times at 37°C.In the treated embryos the mesoderm cells were rounded and separated from each other. Many had long branched processes and rough surfaces. These changes became more pronounced as treatment time was increased. They were also reversible on reincubating treated embryos in the absence of cytochalasin B. The morphological changes produced by CCB are thought to be due to an effect on the cytoskeleton, either a direct disruptive effect or detachment of skeletal microfilaments from the cell membrane. There may also be a direct removal of cell surface materials leading to the observed surface roughening of treated cells.     

Scanning electron microscopic studies of ileal epithelial cells in suckling rats

Summary The ileal epithelial cells containing the tubulo-vacuolar systems and supranuclear vacuoles in suckling rats were investigated with scanning electron microscopy, using specimens treated with osmium-thiocarbohydrazide-osmium staining methods, and critical point drying and cracking.The cracked surface of the apical cytoplasm is seen as irregular and small hollows and pores of the anastomosing and branching tubulo-vacuolar system. The cracked surface of the supranuclear vacuoles shows the ellipsoidal structures. Numerous pores of various size and irregular shape are present on the apical inner surface of the supranuclear vacuole. These pores are clearly the openings from the tubulo-vacuolar system to the supranuclear vacuole. Some small pores are visible on the inner lateral surface of the supranuclear vacuole, especially near the nucleus. They are probably the pathways of the absorbed materials from the supranuclear vacuole into the lateral cytoplasm. Usually, the inclusions of the supranuclear vacuole reveal the globes or coarse and sponge-like networks.     

The extremophile Anoxybacillus sp. PC2 isolated from Brazilian semiarid region (Caatinga) produces a thermostable keratinase

The aim of this study was to select and identify thermophilic bacteria from Caatinga biome (Brazil) able to produce thermoactive keratinases and characterize the keratinase produced by the selected isolate. After enrichment in keratin culture media, an Anoxybacillus caldiproteolyticus PC2 was isolated. This thermotolerant isolate presents a remarkable feature producing a thermostable keratinase at 60°C. The partially purified keratinase, identified as a thermolysin-like peptidase, was active at a pH range of 5.0–10.0 with maximal activity at a temperature range of 50–80°C. The optimal activity was observed at pH 7.0 and 50–60°C. These characteristics are potentially useful for biotechnological purposes such as processing and bioconversion of keratin.     

Endothelial cell heterogeneity in experimentally-induced rabbit atherosclerosis. Demonstration of multinucleated giant endothelial cells by scanning electron microscopy and cell culture

We investigated the aortic endothelial cells of cholesterol-fed rabbits, using scanning electron microscopy and a cell culture technique. Rabbits were given a 1% cholesterol diet intermittently for up to 40 weeks. In these animals, the area of endothelial cells was increased and the cells showed polymorphism in relation to the progression of atherosclerosis. In animals fed the cholesterol diet for 12, 28 and 40 weeks, the average area of the endothelial cells was 436±15, 762±153, and 836±165 m², respectively. In the cholesterol-fed 40-week group, in particular, giant endothelial cells, measuring more than 1200 m², accounted for 14% of the population. In animals fed a standard diet there was no significant difference in endothelial cell morphology between control 0-week and control 40-week groups; in both, the luminal surface of the thoracic aorta formed a homogeneous sheet covered by small rhomboidal endothelial cells, the area of most being less than 400 m². Primary cultured endothelial cells harvested from those control groups were mononuclear typical small cells with a centrally located nucleus; the proportion of binucleated cells was less than 2% and no multinucleated giant cells with three or more nuclei were detected. Endothelial cells from the cholesterol-fed groups, however, contained larger numbers of binucleated cells, with the number increasing in proportion to the duration of cholesterol feeding. The major distinguishing feature of the endothelial cells in the cholesterol-fed groups was the presence of multinucleated giant cells with three or more nuclei; these accounted for 2.3% and 3.3% of the total cell population in the cholesterol-fed 28- and 40-week groups, respectively. No bromodeoxyuridine uptake was found in the nuclei of the cultured multinucleated giant cells. Heterogeneity of endothelial cells, with the concomitant appearance of multinucleated giant cells, emerges with the progression of diet-induced atherosclerosis. The morphological alterations of endothelial cells observed in the present study intimately reflect changes in their function associated with the progression of atherosclerotic lesions.     

体外诱导成骨细胞在降解性镁合金表面的附着与增殖

目的 研究能够与骨形成建立良好动态重建平衡关系的降解性镁合金,探讨WE43镁合金作为骨科固定材料的可行性。 方法 在18只SD大鼠股骨内分别植入WE43镁合金棒(9只)及Mg-Mn合金棒(9只),术后不同时间处死动物,取肝、肾行病理分析;体视镜及扫描电镜下观察移植物/新骨界面区,并计算降解率;电子探针全扫描分析合金元素分布状态。 结果 两种镁合金均发生不同程度降解,WE43比Mg-Mn合金具有更好的新骨诱导性和耐腐蚀性能,两种材料降解均未对大鼠肝、肾造成不良影响。实验组WE43中的稀土元素可在大鼠股骨体周围形成少量存留,并对骨生成起促进作用,两种合金中的镁元素及Mg-Mn合金中的锰元素在大鼠股骨体周围均不形成存留。 结论 WE43镁合金比Mg-Mn合金的降解速率慢,更适合与新骨重建形成良好平衡关系,是一种可降解性内固定材料。