Determination of agitation and aeration conditions for scale-up of cellulolytic enzymes production by <Emphasis Type="Italic">Trichoderma inhamatum</Emphasis> KSJ1

10 to 35 L jar fermentation scale-up cultures were performed to determine the optimum agitation and aeration rates in the cellulolytic enzymes production culture by Trichoderma inhamatum KSJ1. The optimum agitation rate in the 35 L jar fermenter was provisionally determined to be 150 rpm by using a geometrically resembled scale up method from the 10 L jar fermenter. The optimum aeration rate was determined to be 0.5 vvm by applying the mean values of superficial velocity and vvm. The DO (Dissolved Oxygen) concentration of the culture liquid was maintained below the critical DO concentration (2.336 mg/L) at 150 rpm in the 35 L jar fermenter. To increase the DO above the critical DO concentration, the agitation rate was increased from 150 to 200 rpm, with the aeration rate maintained at 0.5 vvm. As a result, the DO was maintained above critical DO concentration. The OUR (Oxygen Uptake Rate) and k _L a values were 0.91 mg-DO/L·min and 11.1 hr⁻¹, respectively. The amylase and FPase (filter paper activity) activities were 4.48 and 0.74 U/mL, respectively, in the 35 L jar fermenter, which was comparable to that in the 10 L fermenter (4.2 and 0.5 U/mL, respectively). Therefore, the scale-up conditions, 0.5 vvm and 200 rpm, were concluded to be the optimum aeration and agitation rates in the 35 L jar fermenter.     

氮源对酵母工程菌株生产α-淀粉酶的影响

在摇瓶培养条件下 ,研究了氮源对含重组质粒 p NA3的酿酒酵母 2 0 B- 12生产α-淀粉酶的影响。结果表明 :在葡萄糖去阻遏条件下必须有天然氮源存在 ,基因表达才能被诱导 ;几种天然氮源中酵母膏诱导效果最好 ,当酵母膏的添加量为 2 0 g/ L时 ,发酵液中α-淀粉酶的最大活力达 1.97U/ m L ,最大菌体密度为 0 .97g/L。各种天然氮源的添加量必须适中 ,否则会抑制工程菌的生长和基因表     

氮源对里氏木酶合成木聚糖酶的影响

分别以（ＮＨ４）２ＳＯ４、酵母浸膏及蛋白胨为产酶氮源制备木聚糖酶。试验结果表明,酵母浸膏的产酶效果最好,其次是（ＮＨ４）２ＳＯ４。当以酵母浸膏为氮源产酶时,酶活力最高达到２５．４７ＩＵ／ｍＬ,酶得率和酶产率分别为３６３８．６ＩＵ／ｇ木聚糖和８４９０．０ＩＵ／Ｌ·ｄ,酶活力分别是以（ＮＨ４）２ＳＯ４及蛋白胨为氮源时的１．５倍和２．０倍     

重组真菌腈水解酶的发酵工艺条件及生物催化特性初探【需加急】

对产真菌腈水解酶的重组大肠杆菌的培养基种类、培养基成分、诱导剂种类和浓度、诱导条件、p H和温度进行了系统考察。摇瓶发酵优化结果显示:以甘油作为主要碳源,蛋白胨和酵母膏作为主要氮源,并添加微量元素的SOC培养基作为发酵培养基,最适接种量为0.5%;较优的诱导剂诱导条件为:采用0.5 mmol/L的IPTG诱导12 h,发酵p H=7.5,诱导温度25℃时产酶效果最佳。经过优化后,重组酶的酶活得到了显著提高,总酶活最高达到了3.84 U/m L,相比初始水平(0.84 U/m L)提高约4倍。5 L发酵罐的放大实验表明,产酶效果良好,总酶活和比酶活均与摇瓶水平基本持平。全细胞催化性质考察研究结果表明,该菌株所产腈水解酶催化反应的最适催化反应温度是45℃,最适反应p H约为7.2。     

直接降解木质素的漆酶/木聚糖酶体系的合成

对一株高产漆酶及伴有木聚糖酶和少量纤维素酶的菌株,用不同的碳源和氮源对其调控培养,合成漆酶/木聚糖酶体系。实验结果表明,最佳的碳源是可溶性淀粉,用它作碳源,漆酶活性高达730IU/mL,木聚糖酶活性是4.49IU/mL,纤维素酶活性只有0.23IU/mL;最佳的氮源是蛋白胨,用其作氮源,合成漆酶活性可达812IU/mL,木聚糖酶活性是4.68IU/mL,纤维素酶活只有0.09IU/mL。     

Optimization of growth medium for the production of α‐amylase from Bacillus amyloliquefaciens using response surface methodology

The optimization of nutrient levels for the production of α‐amylase by Bacillus amyloliquefaciens was carried out using response surface methodology (RSM) based on the 2³ factorial central composite design (CCD). This procedure limited the number of actual experiments performed while allowing for possible interactions between three components. RSM was adopted to derive a statistical model for the effect of starch, peptone and yeast extract (YE) on α‐amylase production. The P‐value of the coefficient for linear effects of starch and YE concentration was <0.0001, suggesting that this was the principal experimental variable, having the greatest effect on the production of α‐amylase. The optimal combinations of media constituents for maximum α‐amylase production were determined as 12.61 g L^?1 starch, 2.83 g L^?1 peptone and 1.25 g L^?1 YE. The optimization of the medium resulted not only in a 34% higher enzyme activity than unoptimized medium but also in a reduced amount of the required medium constituents. Copyright © 2006 Society of Chemical Industry     

纳豆激酶液体发酵条件的优化

以纳豆枯草芽孢杆菌(Bacillus subtilis natto)为出发菌,研究了其液体发酵产生纳豆激酶(Nattokinase,NK)的培养基组成(碳源、氮源、碳氮比和金属离子组成)和培养条件(温度、初始pH值、发酵时间、接种量和装液量)对产酶量的影响.结果表明,液体发酵培养基的最佳碳源为麦芽糖,浓度为1.0%;最...     

Effect of mycelial morphology on bioreactor performance and avermectin production of Streptomyces avermitilis in submerged cultivations

Factors affecting the morphology of Streptomyces avermitilis and avermectin production in submerged cultivation, including nitrogen sources, inoculum level and DO (dissolved oxygen) tension in the broth were investigated in a 50-L bioreactor. It was found that a combination of soybean meal and yeast meal as nitrogen sources and 4.3% inoculum led to pellet formation, and the pellet morphology facilitated to maintain DO > 20% in the early stage of fermentation. With the aid of image analysis tools, area and density of pellets in different batches were calculated. Results show that higher dissolved oxygen tension was favorable for pellet formation and avermectin production.     

硫磺菌菌丝液体发酵适宜培养基配方研究

实验以硫磺菌为材料,在适宜的外部培养条件（培养温度为25℃、装液量75 mL/250 mL、摇瓶转速为150 r/min培养120 h）下,通过选取不同氮源（包括黄豆浆、尿素、牛肉膏、蛋白胨、（NH4）2SO4）、不同碳源（葡萄糖、蔗糖、淀粉、乳糖）、酸碱度几个单因素的控制以及正交实验,采用液体发酵技术培养菌丝体,以菌丝体湿重为指标,确定了硫磺菌的液体培养的适宜培养基配方。结果表明：以17.5 g/L的黄豆浆为氮源,20 g/L的葡萄糖为碳源,初始pH为5.5的配方既能获得高产又较为方便经济。     

夫西地酸生产菌复合诱变育种及发酵培养基的优化研究

利用常压室温等离子体射流诱变和紫外照射对夫西地酸生产菌株进行复合诱变,得到3株产量明显提高的突变菌株,3株菌的平均发酵效价较出发菌株提高14%;然后,采用均匀设计实验对其中发酵效价提高最多的AU-37菌株的发酵培养基碳、氮源进行了优化,得到适合该菌株的发酵培养基优化配方为:糊精1.86%、花生饼粉2.5%、蛋白胨0.3%、氯化铵0.5%、七水合硫酸镁0.3%、硫酸钾0.6%、碳酸钙1%、pH值7.2,在优化的发酵培养基中,AU-37菌株摇瓶发酵效价较出发菌株提高33.8%,为工业化生产奠定了基础。     

土壤中产壳聚糖酶霉菌的筛选及产酶条件优化

为筛选壳聚糖酶的高产菌株,利用壳聚糖水解圈作为筛选模型,从沈阳化工学院附近农田土壤中筛选得到了产壳聚糖酶能力较高的霉菌M19。对其产酶条件进行优化,最终得到培养基中最佳碳源为1．0％壳聚糖胶体,最佳氮源为1．0％（NH4）2SO4加1．0％蛋白胨,最适初始pH为6．0,最适溶氧量为50mL摇瓶装20mL培养基,最适接种量为7．5％,且发酵72h时该菌产酶能力最强。该方法简便、直观,很适合大量、快速筛选。     

Effects of oilseed storage proteins on aflatoxin production by aspergillus flavus

Potential involvement of seed storage proteins in susceptibility to aflatoxin contamination was assessed with in vitro tests. Initially, two oilseed storage proteins [cottonseed storage protein (CSP) and zein] were compared with bovine serum albumin (BSA) and collagen. Supplementation of a complete defined medium with either oilseed storage protein resulted in significantly more aflatoxin production by Aspergillus flavus than supplementation with either BSA or collagen. Little or no aflatoxin was produced when either BSA, CSP, or zein was employed (at 0.5%) as both the sole carbon and the sole nitrogen source. Media with collagen (0.5%) as the sole nitrogen and carbon source supported aflatoxin production similar to the complete defined medium. Although lower than levels observed with defined medium, aflatoxin production increased with both increasing CSP concentration (0 to 2.0%) and increasing zein concentration (0 to 6.0%) when these proteins served as both the sole carbon and sole nitrogen source. Denaturing polyacrylamide gel electrophoresis and protease activity assays indicated that fungal acquisition of protein carbon was probably via hydrolysis mediated by the 35 kD metalloprotease of A. flavus. Media lacking nitrogen but containing sucrose (5.0%) and supplemented with either zein (1.7%) or CSP (2.0%) supported three- to eightfold more aflatoxin production than the complete defined medium. The results suggest seed storage proteins, when present with an accessible carbon source, may predispose oilseed crops to support production of high levels of aflatoxins by A. flavus during seed infection.     

Effects of oilseed storage proteins on aflatoxin production by Aspergillus flavus

Potential involvement of seed storage proteins in susceptibility to aflatoxin contamination was assessed with in vitro tests. Initially, two oilseed storage proteins [cottonseed storage protein (CSP) and zein] were compared with bovine serum albumin (BSA) and collagen. Supplementation of a complete defined medium with either oilseed storage protein resulted in significantly more aflatoxin production by Aspergillus flavus than supplementation with either BSA or collagen. Little or no aflatoxin was produced when either BSA, CSP, or zein was employed (at 0.5%) as both the sole carbon and the sole nitrogen source. Media with collagen (0.5%) as the sole nitrogen and carbon source supported aflatoxin production similar to the complete defined medium. Although lower than levels observed with defined medium, aflatoxin production increased with both increasing CSP concentration (0 to 2.0%) and increasing zein concentration (0 to 6.0%) when these proteins served as both the sole carbon and sole nitrogen source. Denaturing polyacrylamide gel electrophoresis and protease activity assays indicated that fungal acquisition of protein carbon was probably via hydrolysis mediated by the 35 kD metalloprotease of A. flavus. Media lacking nitrogen but containing sucrose (5.0%) and supplemented with either zein (1.7%) or CSP (2.0%) supported three- to eightfold more aflatoxin production than the complete defined medium. The results suggest seed storage proteins, when present with an accessible carbon source, may predispose oilseed crops to support production of high levels of aflatoxins by A. flavus during seed infection.     

辅酶Q10高产菌Rhizobium radiobacter的选育及发酵条件优化

以放射型根瘤菌(Rhizobium radiobacter)WSH2601为出发菌株,经紫外线和亚硝基胍复合诱变,获得遗传稳定性好的抗放线菌素D突变株WSH-F06. 在摇瓶中考察了碳、氮源等营养条件以及接种量、装液量和初始pH等环境条件对突变株WSH-F06细胞生长和积累辅酶Q10的影响. 通过诱变和优化发酵条件,突变株WSH-F06的辅酶Q10产量和胞内含量分别达到34 mg/L和2.4 mg/g,比出发菌株在同样条件下提高了16%.     

Production of arachidonic acid by filamentous fungus, Mortierella alliacea strain YN-15

A filamentous fungus producing significant levels of arachidonic acid (AA, C20∶4n−6) was isolated from a freshwater pond sample and assigned to the species Mortierella alliacea. This strain, YN-15, accumulated AA mainly in the form of triglyceride in its mycelia. An optimized culture in 25 L of medium containing 12% glucose and 3% yeast extract yielded 46.1 g/L dry cell weight, 19.5 g/L total fatty acid, and 7.1 g/L AA by 7-d cultivation in a 50-L jar fermenter. Assimilation of soluble starch by YN-15 was notably enhanced by the addition of oleic acid, soybean oil, ammonium sulfate, or potassium phosphate to a starch-based medium. Using starch as a main carbon source in the pre-pilot scale cultivation improved the production of AA by up to 5.0 g/L. Mortierella alliacea strain YN-15 is therefore a promising fungal isolate for industrial production of AA and other polyunsaturated fatty acids.     

Thraustochytrid as a potential source of carotenoids

Thraustochytrids, marine protists whose dominant genera are Thraustochytrium and Schizochytrium, belong to the kingdom Chromista and are known as an industrial source of DHA. We describe here that thraustochytrid strain KH105, isolated as a DHA producer, also accumulates significant levels of β-carotene and xanthophylls including canthaxanthin and astaxanthin. A4-d cultivation using a medium composed of 10% glucose and less than 0.3% of nitrogen sources in a half-concentration of seawater gave an astaxanthin production up to 6.1 mg/L, and canthaxanthin content reached more than 10 mg/L under conditions where a higher concentration of nitrogen sources (6%) was employed. It might be advantageous in mass production systems for these carotenoids to be extracted readily by simply suspending the cells with organic solvents such as acetone and chloroform. Analyses on the morphological and life history features of the KH105 strain revealed that it belongs to the genus Schizochytrium. This particular species of thraustochytrids is thus considered to be a promising source of xanthophylls as well as DHA for use in the food industry.     

海洋微生物溶菌酶的发酵优化与中试生产

以海洋细菌S-12-86为试验菌株,采用摇瓶发酵优化的方式,研究培养基组分(碳源、氮源、碳源与氮源的比例、金属离子)与发酵条件(培养温度、接种体积分数、装液体积分数、起始pH值、产酶周期)对海洋微生物溶菌酶产量的影响,并进行中试放大试验。结果表明:该菌产酶最佳培养基组分为:葡萄糖10 g/L,蛋白胨5 g/L,MgSO45 g/L,CaCl22 g/L;最适发酵培养温度为30℃,接种体积分数为4.0%,装液体积分数为10.0%,起始pH值为8.0,发酵周期24 h。海洋细菌S-12-86发酵优化后的产酶量(25636.8 U/mL)较优化前的产酶量(14454.4 U/mL)提高了75.4%。海洋微生物溶菌酶中试发酵的产酶量达26697.87 U/mL。说明摇瓶发酵优化条件可以应用于海洋微生物溶菌酶中试生产上。     

虎奶菇菌丝体胞外多糖深层发酵培养基的优化

研究了营养性因子对虎奶菇菌丝体深层发酵的影响,结果表明蔗糖、马铃薯、蛋白胨和酵母膏有利于胞外多糖的形成。进一步的正交优化实验确定了虎奶菇多糖深层发酵的最佳培养基组成(g/L)为∶蔗糖2 g/100 mL,马铃薯25 g/100 mL,蛋白胨0.2 g/100 mL,酵母膏0.2 g/100 mL。     

运用试验设计方法提高胞外杂合b-葡聚糖酶在重组大肠杆菌中的表达

A genetically engineered Escherichia coli JM109 harboring pLF3 was used to produce a hybrid extracellular β-glucanase. Starting with enzyme production medium, glycerol and yeast extract combined with NaNO3 were screened to be the most suitable carbon and nitrogen source, respectively. Analysis of six components of the enzyme production medium by employing statistical optimization methods such as Plackett-Burman design and steepest ascent showed that yeast extract was the only significant variable and its best concentration for enzyme production was 12g·L^-1. After optimization of the medium, 297.71U·ml^-1 of β-glucanase activity in the medium and 352350U·g^-1 of β-glucanase selectivity could be obtained, which were 14 and 72 folds higher than those obtained from original medium, respectively. Even higher enzyme activities were achieved by batch cultivations in a conventional stirred bioreactor on the optimized medium.     

Production of α‐amylase under solid‐state fermentation utilizing coffee waste

Coffee industry substrates such as coffee pulp, coffee cherry husk, silver skin, spent coffee and mixtures of these coffee wastes (MC) were evaluated for their efficacy as sole carbon source for the synthesis of α‐amylase in solid‐state fermentation (SSF) using a fungal strain of Neurospora crassa CFR 308. For SSF with coffee pulp and with MC, α‐amylase activity of 3908 U g^?1 ds (units per gram of dry substrate) and 3870 U g^?1 ds, respectively, was observed. Parameters such as moisture (60%), pH (4.6), temperature (28 °C), particle size (1.0 mm), inoculum size (10⁷ spores g^?1 ds), and fermentation time (5 days) were optimized for enzyme synthesis, wherein 4981 and 4324 U g^?1 U g^?1 ds of α‐amylase activity was obtained in SSF with coffee pulp and MC, respectively. The enzyme production was further improved when the substrates were subjected to pre‐treatment by steaming. Accordingly, maximum α‐amylase activity of 7084 U g^?1 ds and 6342 U g^?1 ds was obtained with steam‐pretreated coffee pulp and MC, respectively, demonstrating them to be excellent sole carbon sources for synthesis of α‐amylase production. Copyright © 2009 Society of Chemical Industry