补料分批培养提高转氨酶发酵产酶密度的研究

转氨酶是苯丙酮酸酶法制备L-苯丙氨酸的关键酶源,为提高转氨酶的发酵产酶密度,文章采用补料分批培养方式对大肠杆菌A5发酵产酶进行了研究。优化的补料培养工艺为:初始葡萄糖质量浓度5 g/L,初始氮源体积分数为玉米浆5 mL/L、蛋白胨质量浓度1.5 g/L,控制发酵过程pH值7.5,当葡萄糖质量浓度下降为2 g/L,开始每隔2 h补加质量浓度为120 g/L的葡萄糖溶液,从8 h起每隔2 h补加20 mL/L玉米浆+6 g/L蛋白胨及0.6 g/L的4种氨基酸溶液(L-甲硫氨酸、L-缬氨酸、L-异亮氨酸和L-谷氨酸)。在此条件下发酵培养24 h,菌体干质量浓度达10.5 g/L,比优化前产酶量提高了126%。     

粘质沙雷氏菌G1利用玉米浆干粉和磷酸氢二铵生产2,3-丁二醇的研究

研究了几种工业氮源对粘质沙雷氏菌G1发酵生产2,3-丁二醇的影响,在确定玉米浆干粉为氮源的基础上,利用Plackett-Burman(PB)实验和响应面法(RSM)实验对玉米浆干粉和磷酸氢二铵[(NH4)2HPO4]的浓度进行了优化,确定优化培养基(g·L-1)为:蔗糖90,玉米浆干粉20.32,(NH4)2HPO47.21,NaAc 4,柠檬酸钠14,MgSO40.5,Fe-SO40.02,MnSO40.01。并以此优化培养基进行了摇瓶和分批补料发酵,结果表明,摇瓶发酵中,90g·L-1的蔗糖最终被转化成43.06g·L-1的2,3-丁二醇;分批补料发酵中,2,3-丁二醇浓度为128.28g·L-1,产率为2.67g·L-1·h-1,转化率为0.48g·g-1蔗糖。以玉米浆干粉和(NH4)2HPO4为氮源,2,3-丁二醇浓度较高,培养基的成本大幅降低,为工业化生产奠定了基础。     

玉米浆对VB_(12)发酵的影响

玉米浆质量和添加浓度对VB12发酵有很大的影响,为了选取最优玉米浆作为培养基氮源,本实验比较了3个生产厂家和5个不同浓度的玉米浆。结果表明:厂家Ⅲ的玉米浆为最优玉米浆,最适浓度为3.7%。     

玉米浆对大肠杆菌JH-B22发酵产L-丙氨酸的影响

为了进一步降低L-丙氨酸的生产成本,以大肠杆菌JH-B22为发酵菌株,以玉米浆为氮源、葡萄糖结晶废糖液为碳源进行L-丙氨酸的发酵。结果表明,发酵培养基中玉米浆添加量为20g·L~(-1)较为适宜。在玉米浆发酵培养基中进行L-丙氨酸的发酵,L-丙氨酸产量和糖酸转化率分别为54.30g·L~(-1)和90.5%;其L-丙氨酸产量略低于无机盐发酵培养基的56.12g·L~(-1)和LB发酵培养基的56.48g·L~(-1),但玉米浆发酵培养基具有配制更简单、无需额外添加无机盐或者成本较高的酵母粉等优点,可作为L-丙氨酸发酵生产的不错选择。     

枯草芽孢杆菌产芽孢发酵培养基的优化研究

在三角瓶培养条件下,采用单因素实验对枯草芽孢杆菌B53发酵培养基碳源、氮源进行了优化。在此基础上进行了正交实验,确定了菌株B53最佳的产孢发酵培养基为葡萄糖25g/L、大豆蛋白胨14g/L、玉米浆17g/L、NaCl 3g/L,MgSO_4 0.2g/L。在最优条件下发酵培养,芽孢浓度为9.6×10~9个/mL。     

产蛋白酶菌Planomicrobium sp.L-2发酵条件的优化

通过优化从长蛸胃肠道筛选得到的产蛋白酶菌Planomicrobiumsp.L-2菌株的发酵条件,提高其产蛋白酶的活力。采用单因素实验研究发酵培养基的最佳碳源、氮源以及最佳发酵条件,采用Box-Behnken设计方法进行响应面实验设计,进一步优化培养基组成。结果表明,最佳培养基组成及发酵条件为:可溶性淀粉0.73%、蛋白胨0.68%、酵母膏0.15%、磷酸高铁0.01%、海水,培养基初始pH值8.0,接种量8%,发酵温度25℃,发酵时间3.5d。优化后,所产蛋白酶的最高酶活力为1 457U·mL-1,比优化前提高约2倍。     

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。     

玉米浆为有机氮源的L-乳酸发酵的研究

利用细菌厌氧发酵生产L-乳酸,以降低L-乳酸生产成本为主要目的,实验以玉米浆为有机氮源,以硫酸铵为主要无机氮源,研究了不同接种量对发酵过程的影响;并就以玉米浆替代酵母粉、豆粕水解液、生物素为有机氮源的L-乳酸发酵进行了对比研究实验,实验证明了玉米浆做为有机氮源用于L-乳酸发酵的可行性。在保证生产能力的同时,使用玉米浆为有机氮源对进一步降低L-乳酸生产成本,减少玉米浆对环境的污染进行了探索。     

绿色木霉菌产黄色素液态发酵条件的优化

以绿色木霉菌(Trichoderm aviride)为试验菌株,在培养基碳源、氮源组成,装液量,摇床转速,接种量,培养时间及培养温度等单因素试验的基础上,采用Box-Behnken设计及响应面分析法优化T.viride产天然黄色素的培养基组成和发酵工艺条件。结果表明,在试验范围内,T.viride产黄色素的优化培养条件为:马铃薯液态培养基(250g.L-1马铃薯浸汁,2g.L-1Na2HPO4),外加碳源选择21.7g.L-1木糖,摇瓶装液量为50ml.(250ml)-1,培养温度35℃,接种量6%(体积),摇床转速120r.min-1,培养时间152h。在优化培养条件下发酵液黄色素的色价为(52.8±0.8)U.ml-1。马铃薯培养基中添加尿素、硫酸铵、蛋白胨、牛肉浸粉等氮源会抑制黄色素的生成。     

桦剥管孔菌固态发酵纤维素酶及发酵基质酶解

建立纤维素酶固态发酵与生物预处理相耦合工艺。实验优化固态发酵条件,测定发酵基质结晶度及酶解糖化得率。结果表明3 g稻草粉为基质,0.5%淀粉为碳源,1%蛋白胨为氮源,0.5%芦丁,初始pH值为5,发酵14d,褐腐真菌Piptoporus betulinus产CMC酶活力达到76.46 U/g,滤纸酶活力达到7.75 U/g;酶解糖化阶段减少外源纤维素酶量36.05%;P.betulinus降解固态基质中的无定型纤维素,暴露结晶纤维素,提高发酵基质的酶解糖化得率184%。     

抗生素AGPM摇瓶发酵条件的正交实验

采用正交实验设计考察了培养基组成对新型抗生素AGPM发酵的影响,改进后在培养基组成为(g/L)：葡萄糖5, 玉米淀粉40, 黄豆饼粉16, 玉米浆2 ml, K2HPO4 1.0, MgSO4×7H2O 0.5, NaCl 0.5, 淀粉酶0.05及pH 5.5的条件下,单位发酵液的活性提高了18.9倍;同时表明较高的碳氮比对抗生素AGPM的合成有利.     

Fermentative production of L(+)‐lactic acid from starch hydrolyzate and corn steep liquor as inexpensive nutrients by batch culture of Enterococcus faecalis RKY1

BACKGROUND: Attempts were made to determine the lactic acid production efficiency of novel isolate, Enterococcus faecalis RKY1 using four different starches (corn, tapioca, potato, and wheat starch) with different concentrations (50, 75, 100, and 125 g L^?1) and corn steep liquor as an inexpensive nitrogen source. RESULTS: The yield of lactic acid from each starch was higher than 95% based on initial starch concentrations. High lactic acid concentration (129.9 g L^?1) and yield (1.04 g‐lactic acid g^?1‐starch) were achieved faster (84 h) from 125 g L^?1 of corn starch. Among the starches used, tapioca starch fermentation usually completed in a shorter incubation period. The final dry cell weight was highest (7.0 g L^?1) for the medium containing 75 g L^?1 of corn starch, which resulted in maximum volumetric productivity of lactic acid (3.6 g L^?1 h^?1). The addition of 30 g L^?1 corn steep liquor supplemented with a minimal amount of yeast extract supported both cell growth and lactic acid fermentation. CONCLUSION: Enterococcus faecalis RKY1 was found to be capable of growing well on inexpensive nutrients and producing maximum lactic acid from starches and corn steep liquor as lower‐cost raw materials than conventionally‐used refined sugars such as glucose, and yeast extract as an organic nitrogen source in laboratory‐scale studies. These fermentation characteristics are prerequisites for the industrial scale production of lactic acid. Copyright © 2008 Society of Chemical Industry     

Production of bacterial nanocellulose from wine industry residues: Importance of fermentation time on pellicle characteristics

Bacterial nanocellulose (BNC) was produced by Gluconacetobacter xylinus under static conditions using grape pomace extract (the most abundant residue of the wine industry) as a carbon source and corn steep liquor (a byproduct of corn wet‐milling) as the main nitrogen source. Carbon and nitrogen source concentrations, as well as inocula size, fermentation time, and temperature, were all considered in order to maximize BNC production by the use of statistically designed experiments and the response surface methodology. At optimum production conditions, the effect of fermentation time on morphology, solids content, chemical structure, crystallinity, thermal decomposition pattern, and storage modulus of dried BNC pellicles was analyzed. The results evidenced that dried BNC pellicles that were incubated for longer times showed higher thermal stability, higher crystallinity, and higher storage modulus, resulting from a denser nanoribbons network. All of these characteristics will certainly play a role in the performance of BNC in practical applications. © 2015 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2016 , 133, 43109.     

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 Lipase from Geotrichum candidum Using Corn Steep Liquor in Different Bioreactors

The production of an extracellular lipase using corn steep liquor (CSL) as the nitrogen source in the cultivation of Geotrichum candidum NRRLY‐552 was evaluated. The optimized conditions in shake flasks were CSL, 8.0 % w/v, soybean oil, 0.6 % w/v, pH 7.0, 30 °C, 250 rpm, and 48 h, resulting in a maximum lipase productivity of 0.438 U mL^?1 h^?1(U = the amount of enzyme required to liberate 1 μmol of fatty acid per minute). Scale‐up was evaluated with airlift and stirred tank reactors; the best conditions, respectively, were 1 vvm(volume of gas per volume of medium per minute) of aeration which resulted in 0.535 U mL^?1 h^?1 (32 h) and 1 vvm and 300 rpm resulting in 0.563 U mL^?1 h^?1 (16 h). To facilitate downstream processes, lipase production was also evaluated using CSL previously clarified with activated charcoal resulting in 0.275 U mL^?1 h^?1 (24 h) using 12 % (w/v) of clarified CSL in shake flasks. The obtained results showed that CSL leads to similar productivity compared to peptone using the same microorganism under similar conditions. In addition the cost of fermentation medium using CSL is much lower because it is a very inexpensive by‐product from corn processing.     

Fermentative production of mitomycins by Streptomyces caespitosus

Production of mitomycin antibiotics by Streptomyces caespitosus was increased by the utilisation of molasses (1.0%) treated with 0.01 % potassium ferrocyanide and corn steep liquor (0.9 %). Three different fermentation media were used for the fermentation production of mitomycins. Appropriate amounts of ammonium sulphate, soluble starch, ferrous sulphate, magnesium sulphate, potassium dihydrogen phosphate, sodium chloride and calcium carbonate increased the antibiotic yield when added to the fermentation medium containing molasses and corn steep liquor. The addition of some individual amino acids, vitamins and organic acids to a base fermentation medium did not improve or increase its performance.     

The fermentative production of magnamycin by Streptomyces halstedii

Production of magnamycin in different fermentation media by Streptomyces halstedii was studied to select the most suitable for the formation of the antibiotic. Two distinct phases were observed during the fermentative production of magnamycin. The first phase was characterised by vegetative growth, while the second phase was distinguished by formation of the antibiotic. The optimum initial pH value of the medium was 6.12. Glucose was used as carbon source. The best concentrations of glucose and starch supporting biosynthesis of magnamycin were 8.0 and 15.0 g/litre respectively. Soyabean meal was the most suitable organic nitrogen source. The most suitable concentrations of soyabean meal, yeast extract and corn steep liquor were 8.0, 2.0 and 16.0 g/litre respectively.     

Enhanced production of cellobiose dehydrogenase and β-glucosidase by Phanerochaete chrysosporium

The production of cellobiose dehydrogenase (CDH) and β-glucosidase by Phanerochaete chrysosporium ATCC 32629 was assessed during submerged fermentation. The maximum concentrations of CDH and β-glucosidase were obtained using rice straw as the carbon source. Organic nitrogen sources were more effective in enzyme production than inorganic nitrogen sources. Corn steep liquor (CSL) for CDH production and soy bean meal (SBM) for β-glucosidase production were the most appropriate organic nitrogen sources. Using optimum medium obtained by response surface methodology (RSM), the maximum concentrations of CDH and β-glucosidase achieved in the stirred-tank reactor (STR) were 204 U/L and 140 U/L, respectively. CDH productivity (22.7 U/L·day) was the highest at 9 days.     

重组大肠杆菌共表达D-海因酶和N-氨甲酰基-D-氨基酸酰胺水解酶

通过PCR分别扩增得到N-carbamoylase基因和上游带有RBS区的D-hydantoinase基因,并将其依次克隆入pBAD/His A质粒中,得到呈多顺反子结构的双酶表达质粒pBAD-CRH,转化E. coli Top10F￠得到重组菌TaraCRH. 实验证明,E. coli TaraCRH对外源蛋白的表达控制严紧,而加入阿拉伯糖诱导后可以成功表达两酶基因. 诱导条件优化结果表明,诱导温度采用29℃,培养基中阿拉伯糖浓度0.05 g/L可达到最高酶活. 进一步尝试采用玉米浆培养基代替LB培养基,在经过优化的玉米浆培养基中培养TaraCRH菌株,经阿拉伯糖诱导,海因酶和水解酶酶活分别达到3.52和4.21 U/mL.     

固态发酵植物药提取残渣生产饲料蛋白工艺条件

以某制药厂双黄连制剂生产过程中产生的废弃药渣作为研究对象,在对药渣成份进行了全分析的基础上,利用正交实验法研究了混菌固态发酵植物药提取残渣生产饲料蛋白工艺参数的不同,对发酵产物蛋白含量的影响。结果表明:以40g药渣,35g玉米浆,8g玉米面,15g麸皮,2g尿素组成固态发酵培养基,产朊假丝酵母与扣囊拟内孢霉的菌种配比为2∶1时,最佳发酵条件为:发酵温度30℃,接种量15%,含水量65%,培养时间60h。在最佳发酵条件下,发酵产物的粗蛋白含量可达29.98%。以废弃药渣为主要原料发酵生产蛋白饲料是切实可行的。