Statistical Optimization of Culture Components for Enhanced Bacteriocin Production by Lactobacillus plantarum LR/14

Statistical optimization of bacteriocin production by a natural isolate of Lactobacillus plantarum LR/14 was carried out in TGYE medium at 37°C and 200 rpm for 20h. In the first step of optimization using Plackett-Burman design, yeast extract, glucose and incubation period were identified as the most important factors for bacteriocin production. These factors were further optimized by response surface methodology (RSM) to understand their interaction and to determine their optimal levels. Results indicated that the maximum bacteriocin production was achieved in a medium containing glucose 2.0% and yeast extract 2.5%, and after an incubation period of 20h. An overall approximatley eightfold improvement in bacteriocin production was achieved as a result of optimization. These results indicated the importance of statistical tools in designing culture conditions for enhancing the production of bacteriocin from L. plantarum LR/14. Such an improved production will facilitate the application of bacteriocin, especially in food preservation.     

Optimization for the maximum bacteriocin production of Lactobacillus brevis DF01 using response surface methodology

To enhance the production of bacteriocin DF01, produced by Lactobacillus brevis DF01, cultivation conditions and medium composition were optimized by using response surface methodology (RSM). The selected 5 factors based on MRS medium were glucose, yeast extract, MgSO₄, temperature, and initial pH. Fractional factorial design (FFD) was effective in searching for the main factors. By a 2^5?1 FFD, glucose, yeast extract, and initial pH were found to be significant factors and had positive effects on bacteriocin production. The effects of the 3 main factors on bacteriocin production were further investigated by a central composite design (CCD). RSM revealed that the maximum bacteriocin production was achieved at yeast extract concentration of 14.56 g/L, glucose concentration of 28.95 g/L, and initial pH of 6.8. After RSM, the titer of bacteriocin was increased by 4-fold.     

Antibacterial activity of Lactobacillus plantarum UG1 isolated from dry sausage: characterization, production and bactericidal action of plantaricin UG1

Lactobacillus plantarum UG1 isolated from dry sausage produced an antimicrobial substance that inhibited other strains of the genera Lactobacillus and Lactococcus, and some foodborne pathogens including Listeria monocytogenes, Bacillus cereus, Clostridium perfringens and Clostridium sporogenes. This antibacterial substance was inactivated by proteolytic enzymes and showed a bactericidal mode of action. Consequently, it was characterized as a bacteriocin, and was designated plantaricin UG1. This bacteriocin was stable in the pH range 4.5 to 7.0, partially inactivated by amylolytic enzymes and relatively thermostable. It was not affected by organic or lipolytic enzymes. Production of plantaricin UG1 was pH-and temperature-dependant and maximum yields were obtained in MRS broth cultures maintained at initial pH 6.5, and incubated at 25 °C to 30 °C, in the exponential to the early stationary growth phase of the producer organism. Ultrafiltration studies indicated that plantaricin UG1 has a molecular weight between 3 and 10 KDa. Curing experiments with L. plantarum UG1 resulted in the appearance of variants that lost bacteriocin production ability but were still immune to the bacteriocin. Plantaricin UG1 production appeared to be chromosomal encoded. Sensitive and insensitive Gram-positive bacteria adsorbed plantaricin UG1 irrespective of their susceptibility to it. In contrast, Gram-negative bacteria did not adsorb plantaricin UG1. The bactericidal action of plantaricin UG1 did not depend on the physiological state of the indicator culture and did not cause cell lysis. The resistance of two indicator strains to plantaricin UG1 has been studied.     

植物乳杆菌P158产细菌素培养基及培养条件的优化

为提高乳酸菌细菌素产量,以藤黄微球菌、铜绿假单胞杆菌为指示菌,通过单因素和正交试验优化植物乳杆菌P158产细菌素的培养基和培养条件。结果表明,5种乳酸菌培养基中MRS培养基为该菌株产细菌素的适宜培养基;最佳培养条件为种子液接种量3%(V/V)、培养基初始pH 6.0、34℃静置培养42 h;最佳培养基配方为葡萄糖添加量2 g/100 mL、酵母浸膏添加量2 g/100 mL、大豆蛋白胨添加量1.5 g/100 mL、MgSO_4添加量0.058 g/100 mL、MnSO_4添加量0.025 g/100 mL、FeSO_4添加量0.02 g/100 mL、Tween 80添加量0.08 g/100 mL、乙酸钠添加量0.5 g/100 mL、K_2HPO_4添加量0.2 g/100 mL。在此条件下,细菌素效价为1 145 IU/mL,较优化前(362 IU/mL)提高了216%     

植物乳杆菌(Lactobacillus plantarum)8-6产细菌素发酵条件的优化

对植物乳杆菌(Lactobacillus plantarum)8-6产细菌素的发酵条件进行了优化,分别研究了培养时间、温度、接种量、培养基起始pH值、培养基碳源、氮源等因素对细菌素产生的影响,通过单因素水平试验和正交试验,确定产细菌素的最佳培养基组合和最佳发酵条件为葡萄糖3%,胰蛋白胨2%,蛋白胨1%,酵母膏1%,硫酸镁0.058%,吐温-80 0.2%,30℃培养24h,培养基起始pH值为6.5,接种量2%。乳杆菌8-6优化后效价为1825.56IU/mL,比优化前提高了373.15%。     

响应面法优化根霉菌产麦角固醇的液体发酵工艺

以米根霉(Rhizopus oryzae ZW017)发酵产麦角固醇的产量为响应值,对其液体发酵工艺进行优化。采用HPLC法检测菌株产麦角固醇含量,在单因素筛选试验基础上,以PDB液体发酵培养为基础条件,应用响应面分析法(RSM)对碳源、氮源及发酵时间进行优化。结果表明:以葡萄糖、酵母膏分别为最佳碳、氮源;最佳工艺条件为:PDB基础培养基中添加葡萄糖3g/L、酵母膏5g/L、发酵培养9.64d,麦角固醇平均产量达5761.83μg/100mL,较优化前提高了247.86%,与构建模型理论预测值(5818.39μg/100mL)相吻合,且100mL液体培养基中麦角固醇产量占菌体细胞干质量(0.36g)的1.60%。     

产细菌素屎肠球菌TRS5特性分析及其编码基因的扩增

屎肠球菌TRS5在37℃、p H 6.5的MRS培养基中经过24 h的培养后,其细菌素生成量达到最大。培养基中添加胰蛋白胨或葡萄糖有利于促进TRS5细菌素的生成,而添加麦芽糖、乳糖或甘露糖(20 g/L)后细菌素活性减少50%。外源添加5 g/L的甘油和吐温-80会抑制TRS5细菌素的产生,而添加K_2HPO_4或VB_1、VB_2、VB_6、VC则对细菌素的生成没有影响。药敏实验证实屎肠球菌TRS5对红霉素、氯霉素、万古霉素、替考拉宁、四环素、青霉素敏感。聚合酶链式反应及测序结果证实屎肠球菌TRS5含有肠球菌素enterocin P和类L50的结构基因。     

植物乳杆菌CGMCC.5297产细菌素条件的优化及应用

用响应面法对植物乳杆菌CGMCC.5297生产细菌素的培养基进行了优化。通过Plackett-Burman设计和中心组合试验设计,植物乳杆菌CGMCC.5297代谢产细菌素的最佳培养条件为酵母粉5.09g/L,牛肉膏10.85g/L,葡萄糖55.34g/L。此时的细菌素上清与指示菌单核细胞增多性李斯特菌CVCC1595共培养4h后,指示菌OD600nm为0.001 73,效价为4499 IU/mL,提高了1.4倍。在最优发酵条件下获得的试验结果与模型预测值吻合,说明所建立的模型是切实可行的。将优化后的植物乳杆菌上清加入到自制酸奶中,发现此细菌素对酸奶具有良好的保鲜效果。     

Lactobacillus plantarum 163产细菌素食品级培养基筛选及发酵条件优化

为获得Lactobacillus plantarum 163最佳食品级培养基,首先筛选出Lb. plantarum 163食品级培养基配方,即白菜汁200 mL/L、番茄汁50 mL/L、葡萄糖10 g/L、K2HPO4 2 g/L,蒸馏水补足至1 L。Plackett-Burman试验设计筛选出Lb. plantarum 163食品级培养基配方的关键因子,即接种量、K2HPO4添加量、pH值和大白菜汁添加量。通过Box-Behnken试验构建了Lb. plantarum 163食品级培养基二次多项式模型,得到理想发酵条件,即K2HPO4 1.89 g/L、大白菜汁341.5 mL/L、接种量3.56%、pH 6.95,其抑菌活性比优化前增加30%以上。     

抗单增李斯特菌枯草芽孢杆菌C3增菌培养条件优化

枯草芽孢杆菌对单增李斯特菌有强烈抑制作用,该文对其增菌培养基和培养条件进行优化,为今后将其应用于食品和饲料奠定基础。在对培养基成分和培养条件进行单因素试验的基础上,设计5因素4水平正交试验对培养基成分进行优化。结果表明,培养基成分优化为葡萄糖1.0%,酵母浸粉1.0%,NaCl 0.5%,KH2PO4 0.2%,MgSO4·7H2O 0.2%,培养条件优化为pH5.4,装液量50 mL/250 mL,接种量3%,温度37 ℃,150 r/min培养14 h,在此条件下,活菌数可达到7.1×1010 CFU/mL,比优化前提高了7.89倍。     

金属离子对低糖酵母发酵活性的影响

该研究考察了3种金属离子对低糖酵母发酵活力的影响。首先,采用响应面设计法对酵母发酵培养基进行了优化。通过Plackett-Burman设计试验筛选出3个主要因素：镁离子（Mg2+）、锌离子（Zn2+）、锰离子（Mn2+）。在这个基础上应用最陡爬坡路径法逼近最大响应值区域,然后利用响应面分析法确定最佳培养基配方为酵母抽提物（FM888） 10 g/L、蛋白胨（FM318） 20 g/L、葡萄糖20 g/L、六水合氯化镁 6.95 g/L、氯化锌1.78 mg/L、一水合硫酸锰0.069 mg/L。其次,将优化培养基配方应用于低糖酵母发酵,干酵母活力可达426.86 mL/g。经过3次平行试验的验证,实际的平均发酵活力与预测的发酵活力值相近,比优化前提高了24.8%。此研究对低糖酵母的工业化生产具有一定的指导意义。     

嗜酸乳杆菌NX2-6产细菌素的发酵条件优化

在Plackett-Burman试验结果基础上,采用响应曲面法(Box-Behnken设计)对嗜酸乳杆菌NX2-6发酵产细菌素的关键影响因素,即葡萄糖质量浓度、乙酸钠质量浓度、柠檬酸三钠水合物质量浓度及培养时间的最佳水平范围进行研究和探讨。通过对发酵液抑菌圈直径的二次多项回归方程求解得知,在葡萄糖质量浓度、乙酸钠质量浓度、柠檬酸三钠水合物质量浓度和培养时间分别为60.0、8.0、5.0g/L和36h时,菌株NX2-6的发酵液抑菌圈直径预测值为21.37mm,验证实验抑菌圈直径实测值与预测值的相关系数R2为0.9918。优化后培养基与基础培养基相比,发酵液抗菌活性增加约80.0%,由此可见,利用响应曲面法对嗜酸乳杆菌NX2-6发酵产细菌素条件进行优化是经济有效且科学合理的。     

液体培养的桑黄胞外多糖发酵培养基成分的优化

通过27-3IV部分因子设计(FFD),对葡萄糖、蛋白胨、酵母膏、磷酸二氢钾、硫酸镁、VB1以及草酸铵7个营养因子进行筛选。在此基础上,采用中心组合设计对部分因子设计,筛选出对桑黄胞外多糖产量的关键因子,确定培养基的组成和水平。在葡萄糖34.12g/L、酵母膏5.01g/L、蛋白胨4g/L、MgSO4 0.75g/L、KH2PO4 1g/L、VB1 0.0075g/L、草酸铵0.88g/L的条件下进行5次平行发酵实验。结果发现桑黄胞外多糖平均产量为(2.363±0.04)g/L,与预测值(2.342g/L)基本相符。可见,用该回归模型优化产桑黄胞外多糖的发酵培养基是可行的。     

均匀设计法优化廉价型牛凝乳酶工程菌发酵培养基

为了获得生产用廉价型牛凝乳酶工程菌发酵培养基,通过单因素试验考察发酵培养基中各组分对产酶的影响。结果显示：葡萄糖、玉米浆、酵母提取物、尿素质量浓度对产酶影响显著。以上述因素作为随机因子,进行均匀设计试验,采用逐步回归方法对试验结果进行分析。结果表明：在葡萄糖45g/L、玉米浆17g/L、酵母提取物6g/L、尿素12g/L的条件下,凝乳酶活性达342.86SU/mL,比优化前提高了1.22倍。所得培养基为重组牛凝乳酶的高效低成本生产提供了参考。     

Sporidiobolus pararoseus WZ012产类胡萝卜素发酵条件的优化

以响应面方法对Sporidiobolus pararoseus WZ012产类胡萝卜素的摇瓶发酵条件进行了优化。在Plackett-Burman实验的基础上,利用中心组合设计研究了4个主要因素(葡萄糖、酵母膏、KH2PO4、接种量)对产类胡萝卜素的影响。结果表明,较高浓度的酵母膏和较大的接种量有助于类胡萝卜素的生产,而较高浓度的葡萄糖和KH2PO4有助于生物量的提高,生物量与类胡萝卜素产量之间没有必然的联系。优化得到的发酵培养基组成(g/L)为:葡萄糖60.23、酵母膏12.21、KH2PO41.68、接种量为8.38%。在此条件下红酵母产类胡萝卜素的最大产量为29.3mg/L,较优化前提高了54.1%。类胡萝卜素成分分析表明主要含有β-胡萝卜素、圆酵母素和红酵母红素这3种类胡萝卜素物质,其中β-胡萝卜素占62.5%。     

响应曲面法优化乳杆菌产细菌素的条件研究

以分离自泡菜可产细菌素的乳杆菌作为实验菌,优化其产细菌素的最佳培养条件,以提高其产细菌素的能力。通过Plackett-Burman实验筛选出对乳杆菌产细菌素有显著影响的3个因素,分别为装液量、葡萄糖质量浓度以及蛋白胨质量浓度。以抑菌圈直径大小作为产细菌素能力大小的判断依据,通过最陡爬坡实验和Box-Behnken实验进一步优化,并对优化的结果进行验证,验证结果表明,预测值和实际值有良好的拟合性,此优化模型可靠。最后确定的乳杆菌产细菌素的优化培养基组成为:蛋白胨30g/L、葡萄糖15g/L、柠檬酸氢二铵2g/L、K2HPO42g/L、乙酸钠5g/L、MnSO4.4H2O0.25g/L、MgSO4.7H2O0.58g/L、吐温800.1%;最佳培养条件为:装液量25mL、温度30℃、培养时间24h、接种量1%、pH6.0。在此优化发酵条件下,细菌素的产量提高了30%。     

Comparison of the performances of different fermentation strategies on cell growth and bacteriocin production by Lactobacillus curvatus CWBI‐B28

The dynamics of cell growth and bacteriocin production by Lactobacillus curvatus CWBI‐B28 in modified De Man/Rogosa/Sharp (mMRS) broth with various concentrations of glucose and complex nitrogen source (CNS; peptone, yeast extract and meat extract) was investigated in flask fermentations and in a laboratory fermentor using batch and fed‐batch cultivations. In fed‐batch fermentation the rate of feeding of the reactor with the substrates was either maintained constant (0.12 L h^?1) or varied exponentially as a function of time. The results showed that both cell growth and bacteriocin activity were influenced by changes in the concentrations of glucose and CNS. Optimal growth and bacteriocin activity were obtained in mMRS broth containing 40 g L^?1 glucose and 40 g L^?1 CNS (mMRS_40/40). A bacteriocin titre of 4266 AU mL^?1 and a cell count of 8.7 log colony‐forming units (cfu) mL^?1 were recorded when this medium was used for cultivation. In batch fermentation using the same medium, a higher cell count (9.5 log cfu mL^?1) and twice as much bacteriocin as in flask fermentation were produced. The highest bacteriocin titre (8533 AU mL^?1) was obtained with fed‐batch fermentation at an exponentially varying rate of feeding. Bacteriocin activity and cell dry mass did not always correlate. Copyright © 2007 Society of Chemical Industry     

Effect of Culture Variables on Mycelial Arachidonic acid Production by Mortierella alpina

Effect of culture conditions on biomass, lipid, and arachidonic acid production was investigated in the oleaginous fungus Mortierella alpina CBS 528.72 under shake flask conditions. Several factors have been found to affect the biomass buildup and lipogenesis in this fungus, complicated by the fact that different strains demonstrate varying optimization conditions. Growth, lipid accumulation, and arachidonic acid production in the strain investigated were influenced by media, pH, temperature, carbon source, nitrogen source, etc. The results indicated that the most effective medium for growth and arachidonic acid production was glucose yeast extract medium. The optimum pH and temperature were found to be 6.5 and 28°C, respectively. On the same weight basis, glucose was the most efficient carbon source for biomass and lipid production in this fungal strain which yielded 6.8 g/L dry biomass and 40.2% (w/w) total lipid after 7 days of cultivation. Maximum arachidonic acid (ARA) production of 40.41% achieved in rhamnose-containing media was not concomitant with higher biomass and lipid yields. Efficacy of organic carbon sources, viz, yeast extract and peptone over inorganic sources like sodium nitrate, ammonium sulfate, ammonium chloride, etc, was established in the present study. M. alpina CBS 528.72 grown in peptone acquired the highest lipid content (42.0% (w/w)). However, the ARA content (28.74%) proved to be significantly less than that grown in yeast extract (35.28%). Furthermore, it was found that the biomass and ARA production declined drastically in a medium with vegetable oils as the sole carbon source but triggered the lipogenic pathway leading to higher accumulation of total lipids. Under the ideal conditions mentioned above, the maximum biomass, total lipid, and arachidonic acid production were 6.8 g/L, 41.6%, and 35.28% total fatty acid, respectively, in shake flask system.     

柳小皮伞深层发酵培养基的筛选

采用Plackett-Burman设计对影响柳小皮伞（Marasmius salicicola）AS5．166发酵胞外多糖培养基的组成成分进行了筛选，所选取的10个相关因素为：葡萄糖、蔗糖、酵母膏、蛋白胨、K2HPO4、KH2PO4、CaCl2、FeSO4、MgSO4、VB1。结果表明，影响柳小皮伞（Marasmius salicicola）AS5．166胞外多糖含量的主要因素为葡萄糖、蔗糖、酵母膏、KH2PO4、MgSO4和FeSO4。     

罗伊氏乳杆菌培养基优化及其生长代谢研究

罗伊氏乳杆菌的MRS 培养基是富培养基,简化培养基组分,优化培养基组成和培养条件是微生物工业化培养的重要基础。本实验在传统MRS 培养基基础上,首先对氮源进行单因素优化,然后采用Plackett-Burman 和中心组合试验设计对影响罗伊氏乳杆菌CG001 生长的MRS 培养基和培养条件进行筛选优化,并在最优条件下研究该菌的生长代谢情况。结果表明：酵母膏质量浓度、葡萄糖质量浓度、硫酸锰质量浓度和温度是影响菌体质量浓度的4 个关键因素;经响应面法分析确定该菌的最优培养条件为：酵母膏质量浓度20g/L,葡萄糖质量浓度20g/L,醋酸钠质量浓度7g/L,柠檬酸铵质量浓度1g/L,磷酸氢二钾质量浓度3g/L,硫酸镁质量浓度0.2g/L,硫酸锰质量浓度为0.23g/L,吐温-80 质量浓度1g/L,初始pH6.2,培养温度38.6℃,最终菌体质量浓度达0.984g/L,相对优化前提高1.102 倍。发酵罐中菌体生长曲线呈S 型,4～12h 菌体处于对数生长期,之后趋于平衡,葡萄糖的代谢与菌体的生长速率相对应。