灵芝液态发酵胞内外多糖结构特征及其活性研究进展

灵芝是一味传统的中药材,具有很高的药用价值,多糖是其主要的活性成分之一,可从其子实体、孢子粉、发酵菌丝体和胞外液中获得。近年来,从灵芝菌丝体和胞外液中发现的多糖越来越得到研究者们的关注。本文从发酵培养基成分及发酵条件对灵芝胞内外多糖的影响、灵芝胞内外多糖的结构、灵芝胞内外多糖生物活性3个方面进行综述,为发酵来源灵芝多糖的开发利用提供科学理论依据。     

代谢工程改造大肠杆菌一碳模块高效合成L-甲硫氨酸

L-甲硫氨酸又名L-蛋氨酸,是人体必需8种氨基酸之一,在饲料、医药、食品领域具有重要应用。以实验室前期构建的M2(Escherichia coli W3110?IJAHFEBC/PAM)为出发菌株,以模块化代谢工程策略构建了一株L-甲硫氨酸高产菌株。首先通过过表达亚甲基四氢叶酸还原酶(methylenetetrahydrofolate reductase,MetF)和筛选不同来源的丝氨酸羟甲基转移酶(hydroxymethyltransferase,GlyA),增强了一碳模块甲基供体的生成,优化了一碳模块。随后针对一碳模块的前体供应,过表达了胱醚裂解酶(cysteamine lyase,MalY)和半胱氨酸内运基因(fliY),有效地提高了L-高半胱氨酸和L-半胱氨酸的供应。最终摇瓶发酵L-甲硫氨酸的产量由2.8 g/L提高至4.05 g/L,5 L发酵罐中达到18.26 g/L。研究结果表明,一碳模块对L-甲硫氨酸的生物合成具有十分重要的影响,在细胞内通过优化一碳模块,可以实现L-甲硫氨酸的高效生物合成。本研究为进一步提高微生物发酵生产L-甲硫氨酸的水平奠定了基础。     

Enrichment of homoacetogens converting H2/CO2 into acids and ethanol and simultaneous methane production

An anaerobic granular sludge was enriched to utilize H₂/CO₂ in a continuous gas-fed up-flow anaerobic sludge reactor by applying operating conditions expected to produce acetic acid, butyric acid, and ethanol. Three stages of fermentation were found: Stage I with acetic acid accumulation with the highest concentration of 35 mM along with a pH decrease from initial 6 to 4.5. In Stage II, H₂/CO₂ was replaced by 100% H₂ to induce solventogenesis, whereas butyric acid was produced with the highest concentration of 2.5 mM. At stage III with 10 µM tungsten (W) addition, iso-valeric acid, valeric acid, and caproic acid were produced at pH 4.5–5.0. In the batch tests inoculated with the enriched sludge taken from the bioreactor (day 70), however, methane production occurred at pH 6. Exogenous 15 mM acetate addition enhanced both the H₂ and CO₂ consumption rate compared to exogenous 10, 30, and 45 mM acetate by the enriched sludge. Exogenous acetate was failed to be converted to ethanol using H₂ as electron donor by the enriched acetogens.     

pH对灵芝液态发酵代谢物及抗氧化活性的影响

已有研究报道灵芝栽培生长的最适pH在中性偏酸环境,在碱性范围的生长及代谢情况鲜见报道。本研究主要探究广泛pH对灵芝液态发酵代谢物及其抗氧化活性的影响。采用摇瓶液态培养后分析代谢物中灵芝三萜、胞内外多糖、菌丝体蛋白及抗氧化活性等指标,系统比较灵芝菌丝体在pH值2-11的生长和代谢情况。研究结果表明,灵芝菌丝体生长、合成灵芝三萜、胞内多糖、30E胞外多糖、菌丝体蛋白和菌丝体水解氨基酸的最适pH值分别为10、3、2、7、2和2。对应结果分别为17.13 g/L、33.86 mg/g、72.73 mg/g、7.86 g/L、71.42 mg/g和107.10 mg/g。比对照分别提高28.5%、77.3%、22.4%、96.5%、97.1%和70.8%。胞内多糖组分1和组分2最高分子量均在初始pH 4,分别为1.016×10⁸ g/mol和9.280×10⁴ g/mol,胞外多糖组分1最高分子量在初始pH 10,为4.946×10⁶ g/mol;对菌丝体的总抗氧化能力、1,1-二苯基-2-三硝基苯肼(1,1-diphenyl-2- picrylhydrazyl,DPPH)自由基清除能力、羟自由基清除能力分析结果表明最佳的初始pH分别为3、7、9。本研究为液态发酵方式下灵芝生长及其代谢物定向调控发酵的工艺优化提供参考依据,同时发现灵芝菌丝体中优质蛋白及抗氧化活性可在功能性食品和化妆品领域推广应用。     

Soy-based medium for ethanol production byEscherichia coli KO11

An optimized soy-based medium was developed for ethanol production byEscherichia coli KO11. The medium consists of mineral salts, vitamins, crude enzymatic hydrolysate of soy and fermentable sugar. Ethanol produced after 24 h was used as an endpoint in bioassays to optimize hydrolysate preparation. Although longer fermentation times were required with soy medium than with LB medium, similar final ethanol concentrations were achieved (44–45 g ethanol L^–1 from 100 g glucose L^–1). The cost of materials for soy medium (excluding sugar) was estimated to be $0.003 L^–1 broth, $0.006 L^–1 ethanol.     

Use of agricultural surpluses for production of biomass by marine microalgae

The marine microalga Phaeodactylum tricornutum was cultivated in semi-continuous culture under mixotrophic conditions with the soluble fractions of potato, rye and wheat flours that had been naturally fermented, at 2% or 4% (w/v). The rye flour produced the highest microalgal cellular density of 90×10⁶ cells.ml^-1 when supplemented with NaNO₃ and NaH₂PO₄. The autotrophic control only gave 57×10⁶ cells.ml^-1. The value of agricultural surpluses, such as rye flour, can therefore be increased by its use in the production of valuable, microalgal biomass which is rich in protein, pigments and fatty acids.     

Oxytetracycline production by Streptomyces rimosus in solid-state fermentation of corncob

Corn-cob was used as a substrate in the production of oxytetracycline by Streptomyces rimosus TM-55 in a solidstate fermentation. Oxytetracycline was detected on day 4, and reached its maximum on day 8. Optimal conditions for oxytetracycline production were an initial pH of 5.2 to 6.3, an initial moisture content of 64% to 67%, supplementation with 20% (w/w) rice bran or 1.5% to 2.5% (w/w) (NH₄)₂SO₄ as sole N source, 1.0% (w/w) CaCO₃, 2% (w/w) MgSO₄.7H₂O, and 0.5% (w/w) KH₂PO₄, with incubation for 8 days at 25 to 30°C. Each g substrate produced 7 to 8 mg oxytetracycline.     

Production of pectinases by Aspergillus niger in solid state fermentation at high initial glucose concentrations

Exopectinase (exo-p) and endopectinase (endo-p) production by Aspergillus niger CH4 in solid state culture was studied at initial glucose concentrations of 100, 250, 350 and 450 g/l. The highest activity of exo-p (35 U/g) was produced at 72 and 120 h in the medium containing 100 and 250 g glucose/l, respectively. The maximum endo-p activity (9 U/g) was produced at 72 h in the medium with 250 g glucose/l. The reduction in pectinase production at 350 and 450 g/l initial glucose concentration was due neither to repression of the synthesis of the enzyme nor to the glucose consumption rate of the strain but due to a drastic drop in pH of the medium.S. Solis-Pereyra, E. Favela-Torres, M. Gutiérrez-Rojas, G. Saucedo-Castañeda and G. Viniegra-González are with the Departamento de Biotecnologia, Universidad Autónoma Metropolitana, A.P. 55-535, C.P. 09340, México D.F., México; S. Roussos is with the Laboratoire de Biotechnologie, ORSTOM, B.P. 5045, 34032, Montpellier Cedex, France, and P. Gunasekaran is with the Department of Microbial Technology, School of Biological Sciences, Madurai Kamaraj University, Madurai, 625-021, India.     

Microbiological and technological aspects of cassava-starch fermentation

The major genera found in the microflora of fermented, sour, cassava-starch were Streptococcus, Bacillus, Lactobacillus and Saccharomyces with amylase activity. Lactic acid bacteria predominated whereas the presence of moulds was not significant. No coliforms were detected. Electron microscopy showed bacteria and yeasts in contact with the starch granules and signs of erosion on the granule surface. Lactic acid was the main metabolite; no oligosaccharides, maltose or glucose were detected, indicating their rapid utilization. The degree of acidification, which correlated with the decrease in viscosity and the final quality of the product, was influenced by the variable microbial ecology.     

Anaerobic bioprocessing of organic wastes

Anaerobic digestion of dissolved, suspended and solid organics has rapidly evolved in the last decades but nevertheless still faces several scientific unknowns. In this review, some fundamentals of bacterial conversions and adhesion are addressed initially. It is argued in the light of G-values of reactions, and in view of the minimum energy quantum per mol, that anaerobic syntrophs must have special survival strategies in order to support their existence: redistributing the available energy between the partners, reduced end-product fermentation reactions and special cell-to-cell physiological interactions. In terms of kinetics, it appears that both reaction rates and residual substrate thresholds are strongly related to minimum G-values. These new fundamental insights open perspectives for efficient design and operation of anaerobic bioprocesses. Subsequently, an overview is given of the current anaerobic biotechnology. For treating wastewaters, a novel and high performance new system has been introduced during the last decade; the upflow anaerobic sludge blanket system (UASB). This reactor concept requires anaerobic consortia to grow in a dense and eco-physiologically well-organized way. The microbial principles of such granular sludge growth are presented. Using a thermodynamic approach, the formation of different types of aggregates is explained. The application of this bioprocess in worldwide wastewater treatment is indicated. Due to the long retention times of the active biomass, the UASB is also suitable for the development of bacterial consortia capable of degrading xenobiotics. Operating granular sludge reactors at high upflow velocities (5–6 m/h) in expanded granular sludge bed (EGSB) systems enlarges the application field to very low strength wastewaters (chemical oxygen demand < 1 g/l) and psychrophilic temperatures (10°C). For the treatment of organic suspensions, there is currently a tendency to evolve from the conventional mesophilic continuously stirred tank system to the thermophilic configuration, as the latter permits higher conversion rates and easier sanitation. Integration of ultrafiltration in anaerobic slurry digestion facilitates operation at higher volumetric loading rates and at shorter residence times. With respect to organic solids, the recent trend in society towards source separated collection of biowaste has opened a broad range of new application areas for solid state anaerobic fermentation.W. Verstraete and D. de Beer are with the Center for Environmental Sanitation, University of Gent, Coupure L 653, B-9000 Gent, Belgium; D. de Beer is also with the Max Plank Institut für Marine Mikrobiologie-Microzensor Group, Fahrenstrasse 1, 28359 Bremen, Germany. M. Pena is with the Groupo de Biotechnologia Ambiental, Departamento de Ingenieria Quimica, Universidad de Valladolid, Prado de la Magdalena, 47005 Valladolid, Spain. G. Lettinga is with the Department of Environmental Technology, Wageningen Agricultural University, Bomenweg 2, 6703 HD Wageningen, The Netherlands. P. Lens is with the Environmental Research Unit. Department of Microbiology, University College Galway, Galway, Ireland.