Influence of growth conditions on production of capsular and extracellular polysaccharides byRhizobium leguminosarum

The influence of growth rate and medium composition on exopolymer production byRhizobium leguminosarum was studied. When grown in medium containing 10g/l mannitol and 1g/l glutamic acid,Rhizobium leguminosarum biovartrifolii TA-1 synthesized up to 2.0g/l of extracellular polysaccharide (EPS), and up to 1.6g/l of capsular polysaccharide (CPS). Under non-growing cell conditions in medium without glutamic acid, CPS synthesis by strain TA-1 could proceed to 2.1g/l, while EPS-production remained relatively low (0.8g/l). Maximal CPS-yield was 2.9g CPS/l medium in a medium containing 20g/l mannitol and 2g/l glutamic acid. TheEPS-deficient strain R. leguminosarum RBL5515,exo4::Tn5 was able to produce CPS to similar levels as strain TA-1, but CPS-recovery was easier because of the low viscosity of the medium and growth of the cells in pellets. With strain TA-1 in nitrogen-limited continuous cultures with a constant biomass of 500mg cell protein/l, EPS was the most abundant polysaccharide present at every dilution rate D (between 0.12 and 0.02 h^–1). The production rates were 50–100mg/g protein/h for EPS and 15–20mg/g protein/h for CPS. Only low amounts of cyclic -(1,2)-glucans were excreted (10–30 mg/l) over the entire range of growth rates.Abbreviations bv biovar - CPS capsular polysaccharide - EPS extracellular polysaccharide - HM_r high molecular mass - LM_r low molecular mass - YEMCR Yeast Extract-Mannitol-Congo Red agar     

Production of a hypoglycemic, extracellular polysaccharide from the submerged culture of the mushroom, Phellinus linteus

Mycelial growth and extracellular polysaccharide production of Phellinus linteus were optimal at pH 5 and 25 °C. Maximum biomass production (14.2 g l^–1) was after 15 d of cultivation, whereas, extracellular polysaccharide was maximal (3.5 g l^–1) after 21 d. The hypoglycemic effect of the polysaccharide, investigated in streptozotocin-induced diabetic rats, decreased plasma glucose, total cholesterol and triacylglycerol concentrations by 49%, 32%, and 28%, respectively, and aspartate aminotransferase activity by 20%. The results indicate the potential of this polysaccharide to prevent hyperglycemia in diabetic patients.     

A thermostable lipase produced by a newly isolated <Emphasis Type="Italic">Geotrichum</Emphasis>-like strain,R59

Growth and production of lipase by a new Geotrichum-like strain, R59, were studied. Production of extracellular lipase was substantially enhanced when the initial pH of the culture medium, types of carbon and nitrogen sources, substances probably stimulating the lipase biosynthesis, the temperature, and time of growth were optimized. Sucrose and triolein were the most effective carbon sources for lipase production. Maximum lipase activity (146 U/ml^–1) was obtained with urea as the nitrogen source. Growth at 30°C, an initial pH of 6.0 and incubation time of 48 h were found as optimum conditions for cell growth and production of lipase by Geotrichum-like strain R59. The enzyme was thermostable and exhibited very high activity after 1 h incubation at 60°C.     

Effects of photon flux density,CO2, aeration rate,and inoculum density on growth and extracellular polysaccharide production byPorphyridium cruentum

Growth and extracellular polysaccharide production byPorphyridium cruentum were measured as a function of several culture parameters. Photon flux density of 75 μmol m⁻² s⁻¹ and CO₂ concentration of 2.5% were found to be optimum for both growth and extracellular polysaccharide production. Interactive studies on these two parameters further confirmed that at these levels of photon flux density and CO₂, when applied together, both growth (5.9·10⁷ cells per mL) and extracellular polysaccharide production (1.9 g/L) were at the maximum. Maximum growth and extracellular polysaccharide production were observed at inoculum density of 10⁶ cells per mL and aeration rate of 500 mL air per min per liter.     

Photoproduction of Hydrogen by the Marine Heterocystous Cyanobacterium Anabaena Species TU37-1 Under a Nitrogen Atmosphere

Hydrogen production rates by Anabaena sp. strain TU37-1 obtained after an initial 1-day incubation period were approximately 70 to 80 and 3 to 9 µmol (mg chl)^–1 h^–1 under argon and nitrogen atmospheres, respectively. Hydrogen production under argon was not enhanced by addition of carbon dioxide, but was enhanced to some extent under nitrogen by increasing the initial carbon dioxide concentration. Rates of hydrogen and oxygen production during the initial 7-hour period were 15 and 220 µmol (mg chl)^–1 h^–1, respectively, in vessels with 18.5% initial carbon dioxide. Hydrogen production under nitrogen was enhanced by addition of carbon monoxide (1%). The rate obtained from the initial 1-day incubation period was about 40 µmol (mg chl)^–1 h^–1, which corresponded to about 60% of that under argon. On the basis of these observations, a possible strategy for hydrogen production by nitrogen-fixing cyanobacteria under nitrogen in the presence of carbon monoxide is indicated.     

大兴安岭北部天然针叶林土壤氮矿化特征

采用顶盖埋管法对大兴安岭地区天然针叶林(樟子松林、樟子松-兴安落叶松混交林和兴安落叶松林)土壤铵态氮(NH~+_4-N)、硝态氮(NO~-_3-N)、净氮矿化速率进行研究,并探索土壤理化性质与氮矿化之间的相关性,为大兴安岭地区森林生态系统土壤养分管理及森林经营提供帮助。结果表明:观测期内(5—10月)3种林型土壤无机氮变化范围为31.51—70.42 mg/kg,以NH~+_4-N形式存在为主,占比达90%以上,且与纯林相比混交林土壤无机氮含量较高。3种林型土壤净氮矿化、净氨化、净硝化速率月变化趋势呈V型,7、8月表现为负值,其他月份为正值。净氮矿化速率变化范围樟子松林为-0.54—1.28 mg kg~(-1) d~(-1)、樟子松-兴安落叶松混交林为-0.13—0.55 mg kg~(-1) d~(-1)、兴安落叶松林为-0.80—1.05 mg kg~(-1) d~(-1)。土壤净氨化过程在土壤氮矿化中占主要地位,占比达60%以上。3种林型土壤净氮矿化、净氨化及净硝化速率垂直差异显著,0—10 cm土层矿化作用明显高于10—20 cm土层(P0.05)。土壤氮矿化速率与土壤含水量、土壤有机碳含量、土壤C/N、枯落物全氮含量和枯落物C/N均存在显著相关性。不同类型的森林土壤及枯落物的质量也存在差异,进而影响土壤氮矿化特征。     

Temporal separation of nitrogen fixation and photosynthesis in the filamentous,non-heterocystous cyanobacterium Oscillatoria sp.

When growing in laternating light-dark cycles, nitrogenase activity (acetylene reduction) in the filamentous, non-heterocystous cyanobacterium Oscillatoria sp. strain 23 (Oldenburg) is predominantly present during the dark period. Dark respiration followed the same pattern as nitrogenase. Maximum activities of nitrogenase and respiration appeared at the same time and were 3.6 mol C₂H₄ and 1.4 mg O₂ mg Chl a ^-1·h^-1, respectively. Cultures, adapted to light-dark cycles, but transferred to continuous light, retained their reciprocal rhythm of oxygenic photosynthesis and nitrogen fixation. Moreover, even in the light, oxygen uptake was observed at the same rate as in the dark. Oxygen uptake and nitrogenase activity coincided. However, nitrogenase activity in the light was 6 times as high (22 mol C₂H₄ mg Chl a ^-1·h^-1) as compared to the dark activity. Although some overlap was observed in which both oxygen evolution and nitrogenase activity occurred simultaneously, it was concluded that in Oscillatoria nitrogen fixation and photosynthesis are separated temporary. If present, light covered the energy demand of nitrogenase and respiration very probably fulfilled a protective function.     

Exopolysaccharide production by a unicellular cyanobacterium isolated from a hypersaline habitat

The unicellular cyanobacterial strain 16Som2, isolated from a Somaliland saltpan and identified asCyanothece sp., is characterized by cells surrounded by a thick polysaccharidic capsule, the external part of which dissolves into the medium during growth, causing a progressive increase in culture viscosity. In spite of this, the thickness of the capsule remained almost constant under all the culture conditions tested, demonstrating that the processes of its synthesis and solubilization occurred at a similar rate. The synthesis of carbohydrates was neither enhanced by increasing salinity (sea-water enriched with NaCl in the range 0 to 2.0 M) nor by Mg²⁺, K⁺ or Ca²⁺ deficiencies. In contrast, N-limitation and, to a lesser extent, P-limitation induced a significant enhancement of carbohydrate synthesis; in particular, N-deficiency stimulated the synthesis of all the carbohydrate fractions (intracellular, capsular and soluble). The soluble polysaccharide, separated from the culture medium and hydrolyzed with 2N trifluoroacetic acid, showed a sugar composition consisting of glucuronic acid: galacturonic acid: galactose: glucose: mannose: xylose: fucose in a molar ratio of 1: 2: 2.4: 6.8: 4.8: 2.9: 1.6.Cyanothece sp. culture subjected to nitrogen starvation synthesized polysaccharide with a mean productivity of 115 mg (EPS) l^–1d^–1, for the polymer solubilized into the medium, and of 15 mg (CPS) l^–1d^–1 for the capsular polysaccharide.     

Production of Penicillin G acylase from Bacillus sp.: Effect of medium components

A Bacillus sp. producing a high level of intracellular penicillin G acylase (PAC) was isolated. The PAC production in this strain was induced by phenylacetic acid. Various carbon and nitrogen sources were evaluated for their effect on growth and PAC production at 28 °C and pH 7.0. Cells grown in medium supplemented with sucrose as carbon source and tryptone as nitrogen source produced maximum activity of 6.45 and 8.92 U mg^–1, respectively. Maximum concentration of PAC (10.1 Umg^–1) was produced by the cells grown in the medium containing sucrose and tryptone, which was twofold higher than the production in basal medium.     

Ulva rigida (Ulvales,Chlorophyta) tank culture as biofilters for dissolved inorganic nitrogen from fishpond effluents

Ulva rigida was cultivated in 7501 tanks at different densities with direct and continuous inflow (at 2, 4, 8 and 12 volumes d^–1) of the effluents from a commercial marine fishpond (40 metric tonnes, Tm, of Sparus aurata, water exchange rate of 16 m³ Tm^–1) in order to assess the maximum and optimum dissolved inorganic nitrogen (DIN) uptake rate and the annual stability of the Ulva tank biofiltering system. Maximum yields (40 g DW m^–2 d^–1) were obtained at a density of 2.5 g FW 1^–1 and at a DIN inflow rate of 1.7 g DIN m^–2 d^–1. Maximum DIN uptake rates were obtained during summer (2.2 g DIN M^–2 d^–1), and minimum in winter (1.1 g DIN m^–2 d^–1) with a yearly average DIN uptake rate of 1.77 g DIN m^–2 d^–1 At yearly average DIN removal efficiency (2.0 g DIN m^–2 d^–1, if winter period is excluded), 153 m² of Ulva tank surface would be needed to recover 100% of the DIN produced by 1 Tm of fish.Abbreviations DIN= dissolved inorganic nitrogen (NH _inf4 ^sup+ + NO _inf3 ^sup– + NO _inf2 ^sup– ); - FW= fresh weight; - DW= dry weight; - PFD= photon flux density; - V= DIN uptake rate     

精胺对霍山石斛类原球茎悬浮培养细胞生长和多糖合成的影响

研究了在添加外源精胺时,霍山石斛类原球茎细胞生长、多糖积累、主要营养物质消耗以及细胞内多胺含量的变化。结果表明,0.6mmol/L的精胺明显促进霍山石斛类原球茎细胞的生长和多糖的合成。细胞的比生长速率从0.046d^-1提高到0.054d^-1。培养30d时,类原球茎干重达32.4g DW/L,多糖总产量为2.46g/L ,分别是对照的1.32和1.31倍。添加外源精胺能够提高内源多胺的含量,同时,蔗糖酶和硝酸还原酶等相关代谢酶的活性增强,促进了碳、氮的吸收和利用。     

Change in colony morphology and kinetics of tylosin production after UV and gamma irradiation mutagenesis of Streptomyces fradiae NRRL-2702

Tylosin is a macrolide antibiotic used as veterinary drug and growth promoter. Attempts were made for hyper production of tylosin by a strain of Streptomyces fradiae NRRL-2702 through irradiation mutagenesis. Ultraviolet (UV) irradiation of wild-type strain caused development of six morphologically altered colony types on agar plates. After screening using Bacillus subtilis bioassay only morphological mutants indicated the production of tylosin. An increase of 2.7±0.22-fold in tylosin production (1500 mg/l) in case of mutant UV-2 in complex medium was achieved as compared to wild-type strain (550 mg/l). Gamma irradiation of mutant UV-2 using ⁶⁰Co gave one morphologically altered colony type γ-1, which gave 2500 mg/l tylosin yield in complex medium. Chemically defined media promoted tylosin production upto 3800 mg/l. Maximum value of q_p (3.34 mg/gh) was observed by mutant γ-1 as compared to wild strain (0.81 mg/gh). Moreover, UV irradiation associated changes were unstable with loss of tylosin activity whereas mutant γ-1 displayed high stability on subsequent culturing.     

Exopolysaccharide, pigment and protein production by the marine microalga Chroomonas sp. in semicontinuous cultures

The marine microalga Chroomonas sp. isolated from Venezuela was grown in semicontinuous culture in order to study the effect of renewal rate and nutrient concentration on alloxanthin, chlorophyll a, carotenoid, carbohydrate, exopolysaccharide, protein and cell productivity. Maximal cell productivity of 8.43 ± 1.8 and 8.81 ± 2.3 × 10⁹ cell l^–1 day^–1 were achieved with renewal rates of 30 and 40%. Maximal protein and chlorophyll productivity of 64.64 ± 2.3 and 2.72 ± 0.3 mg l^–1 day^–1 were obtained with renewal rate of 20 and 30%. Biochemical composition of Chroomonas sp. was influenced by renewal rate. Nutrient concentration seems not to affect cell, protein, chlorophyll and carotenoid productivity. However, carbohydrate and exopolysaccharide productivity of 7.56 ± 0.4 and 9.57 ± 1.2 mg l^–1 day^–1 were increased at 12 mM NaNO₃(P < 0.05). Also, alloxanthin and chlorophyll a production analysed by HPLC, were higher between 8 and 12 mM NaNO₃ at a renewal rate of 30%. Results demonstrated that a renewal rate of 30% and nutrient concentration at 8 mM NaNO₃ optimize the cell, protein, carbohydrate, chlorophyll a, and exopolysaccharide productivity in semicontinuous cultures of Chroomonas. This microalga, as biological source of commercially valuable compounds, shows high capacity for changing its productivity and biochemical composition in semicontinuous system on the basis of nutrient concentration and the renewal rate.     

Biotransformation of phenylpyruvic acid to l-phenylalanine using a strain of Pseudomonas fluorescens ATCC 11250 with high transaminase activity

The rate of l-phenylalanine production from phenylpyruvic acid by whole cells of Pseudomonas fluorescens strain ATCC 11250 was greater than 3 g·l^-1 h^-1. Synthesis of transaminase was constitutive but activity was greatest in medium containing d- or l- phenylalanine as sole nitrogen source. Maximum conversion was observed at 34–40° C and at alkaline pH, with over six times initial rate of conversion at pH 12 than at pH 5. The optimum catalyst (cell) concentration was between 10–20 mg ml^-1 dry weight. The initial rate of conversion was directly proportional to phenylpyruvate concentration, up to 4%, but the conversion yield steadily decreased between 2% and 4% substrate concentration. The rate of conversion, as expected, increased as the concentration of glutamate increased. Whole cells were still capable of over 63% conversion after 40 days providing reactions were supplemented with pyridoxal phosphate. Immobilisation of cells in calcium alginate and operation of a packed bed bioreactor enabled the continuous production of l-phenylalanine in concentrations greater than 15 g·l^-1 after 60 days operation.     

碳氮源对花榈木胚性愈伤组织诱导、发育及有机物积累的影响

为探讨花榈木体胚发生过程中不同碳氮源处理对胚性愈伤组织诱导、发育和有机物积累的影响,并筛选出有利于花榈木体胚发生的碳氮源,优化体胚发生体系,该研究以成熟胚为外植体,通过单因素试验分析3种碳源、4种蔗糖浓度和6种氮源处理下胚性愈伤组织诱导、发育和有机物积累的差异。结果表明：(1)蔗糖中胚性愈伤组织诱导率显著高于葡萄糖和麦芽糖,但其体胚诱导率、体胚分化率、胚性愈伤组织可溶性糖、淀粉和可溶性蛋白含量差异不显著。(2)随着蔗糖浓度的升高,胚性愈伤组织、体细胞胚(体胚)诱导率、体胚分化率、胚性愈伤组织重量和可溶性蛋白含量呈先升高后降低的趋势,均以添加30 g·L^-1蔗糖最高,而胚性愈伤组织可溶性糖和淀粉含量呈增加的趋势。(3)在6种氮源处理中,胚性愈伤组织诱导率以添加500 mg·L^-1谷氨酰胺的处理最高,体胚诱导率则以添加谷氨酰胺和水解酪蛋白的处理较高,但不同氮源处理间体胚分化率无差异;添加有机氮源的处理其胚性愈伤组织可溶性蛋白含量显著高于无氮源处理。总之,不同的碳氮源通过影响花榈木胚性愈伤组织的诱导、发育和有机物的积累,从而影响其体胚诱导率,但对体...     

Role of Light Intensity and Temperature in the Regulation of Hydrogen Photoproduction by the Marine Cyanobacterium Oscillatoria sp. Strain Miami BG7

The effects of several key environmental factors on the development and control of hydrogen production in the marine blue-green alga (cyanobacterium) Oscillatoria sp. strain Miami BG7 were studied in relation to the potential application of this strain to a bio-solar energy technology. The production of cellular biomass capable of evolving hydrogen gas was strongly affected by light intensity, temperature, and the input of ammonia as a nutrient. Depletion of combined nitrogen from the growth media was a prerequisite for the initiation of hydrogen production. Maximum hydrogen-producing capability coincided with the end of the linear phase of growth. Hydrogen production exhibited considerable flexibility to environmental extremes. The rate of production saturated at low light intensities (i.e., 15 to 30 μEinsteins/m² per s), and no photoinhibition was observed at high light intensity (i.e., 1,000 μEinsteins/m² per s). The upper temperature limit for production was 46°C. Above the light compensation point for O₂ evolution H₂ production was inhibited. However, this problem was alleviated by two related phenomena. (i) The capacity of cells to evolve oxygen deteriorated with increasing culture age and nitrogen depletion, and (ii) the ability of these cells to produce oxygen in closed anaerobic hydrogen production systems was temporally limited.     

四种添加物对铁皮石斛原球茎生长及多糖含量的影响

为探讨铁皮石斛(Dendrobium officinale)培养基中添加物的作用,在1/2MS培养基中加入椰肉、甘蔗渣、香蕉皮和麦麸等4种添加物,研究不同浓度添加物和培养时间对原球茎生长和多糖含量的影响。结果表明,4种添加物对铁皮石斛原球茎的增殖、分化和多糖含量均有一定影响,其中添加15.0 g L–1甘蔗渣,培养60 d能明显促进铁皮石斛原球茎的增殖与分化(146.1%);而添加20.0 g L–1甘蔗渣,培养40 d能显著提高铁皮石斛原球茎多糖含量(50.4%)。这说明甘蔗渣是培养铁皮石斛原球茎的适宜添加物,既能促进铁皮石斛原球茎的生长发育,还能降低生产成本。     

3种水稻土中7株固氮蓝细菌的分离与特征

【背景】蓝细菌是水生和陆地生态系统中生物固氮的主要贡献者。【目的】增加对稻田土壤固氮蓝细菌的了解,获得用于进一步研究的可培养固氮蓝细菌菌株。【方法】选择3种具有不同固氮能力的水稻土,采用BG11-N培养基分离培养固氮蓝细菌菌株,对新分离菌株进行形态特征观察,通过基因组DNA的nifH基因扩增明确其固氮潜力,进一步采用乙炔还原法和~(15)N_2示踪法定量测定其固氮能力,通过基因组DNA的16SrRNA基因序列比对进行鉴定。【结果】在光照培养条件下,采用BG11-N培养基共分离纯化得到自养菌株7株,细胞呈圆形或椭圆形、单列、无分枝、丝状和念珠状,在固体培养基上形成团垫状菌落。新分离菌株在BG11-N培养基中生长状况良好,以基因组DNA为模板可扩增出nifH基因,乙炔还原法和~(15)N_2示踪法测定结果显示具有较高固氮能力,同时具有铁载体生成能力。结合16S rRNA基因序列比对和形态特征,7株菌被初步鉴定隶属于念珠藻科(Nostocaceae)。【结论】从水稻土中分离到在稻田生物固氮中发挥重要作用的蓝细菌(念珠藻科)菌株,可培养固氮蓝细菌菌株固氮能力较高,兼具铁载体生成能力,可作为进一步深入研究的微生物资源,具有潜在的研究应用价值。     

Preparation and use of media for protease-producing bacterial strains based on by-products from Cuttlefish (Sepia officinalis) and wastewaters from marine-products processing factories

Cuttlefish powder (CFP) from Sepia officinalis by-products was prepared and tested as a fermentation substrate for microbial growth and protease production by several species of bacteria: Bacillus licheniformis, Bacillus subtilis, Pseudomonas aeruginosa, Bacillus cereus BG1, and Vibrio parahaemolyticus. All microorganisms studied grew well and produced protease activity when cultivated in medium containing only CFP indicating that the strains can obtain their carbon and nitrogen source requirements directly from whole by-product proteins. Moreover, it was found that the addition to the cuttlefish medium of diluted fishery wastewaters (FWW), generated by marine-products processing factories, enhanced the production of protease. Maximum activity was obtained when cells were grown in cuttlefish media containing 5-times or 10-times diluted FWW. Five-times diluted FWW enhanced protease production by B. cereus BG1 and B. subtilis by 467% and 75% more than control media, respectively. The enhancement could have been due to the high organic content or high salts in FWW.As a result, cuttlefish by-products powder enriched with diluted FWW was found to be a suitable growth media for protease-producing strains. This new process, which converts underutilized wastes (liquid and solid) into more marketable and acceptable forms, coupled with protease production, can be an alternative way to the biological treatment of solid and liquid wastes generated by the cuttlefish processing industry.     

Exopolysaccharide production by a new Halomonas strain CRSS isolated from saline lake Cape Russell in Antarctica growing on complex and defined media

A haloalkalophilic Halomonas strain CRSS, isolated from salt sediments in Antarctica, produced exocellular polysaccharides (EPS) up to 2.9gg^-1 dry cells. Acetate was the most efficient carbon source for EPS production. The composition of media strongly affected the nature of the polymers; a mannan and a xylo-mannan, were obtained when cells were grown on complex media. Acetate was the most efficient carbon source for EPS production and in presence of this substrate, a new polysaccharide, a fructo-glucan, was produced. The EPS fraction was composed by glucose, fructose, glucosamine and galactosamine in relative proportions of 1:0.7:0.3:trace.Revisions requested; Revisions received 6 September 2004