Production of Ammonium Lactate by Fed-batch Fermentation of Rhizopus oryzaefrom Corncob Hydrolysate

L-(+)-Lactic acid production from corncob hydrolysate as a cheap carbohydrate source by fed-batch fermentation of Rhizopus oryzaeHZS6 was studied. After 96 h of fermentation in a 5 L fermentor, the final concentration of ammonium L-(+)-lactate, average productivity(based on initial xylose concentration) and maximum dry cell weight were 132.4 g/L, 1.38 g/(L·h), and 8.9 g/L respectively. The optical purity of L-(+)-lactate was 98.8%.

Fed-Batch Fermentation for Spinosad Production in an Improved Reactor

As a kind of aerobic bacteria, Saccharopolyspora spinosa exhibits a high demand for oxygen. In the fermentation process, the methods of increasing ventilation and improving agitation speed are usually adopted to achieve higher values of dissolved oxygen. These methods decrease the efficiency of spinosad biosynthesis. In this study, an improved reactor was designed to solve these problems. The exhaust gas reflux device, impellers, and baffles were improved. Furthermore, we established the kinetic models for the cell growth, substrate consumption and spinosad generation in batch fermentation process. The simulation results were in good agreement with the experimental data. Spinosad production reached 583.86 mg/L after employing the suitable feeding strategy by fed-batch fermentation in the improved reactor, whereas it was only 157.01 mg/L before optimization. The method described can provide insight to strengthen spinosad production and can be extended to the culturing process of filamentous aerobic bacteria. © 2017 Tianjin University and Springer-Verlag Berlin Heidelberg

Strategies for Optimizing Feed Rate of Fed-Batch Yeast Fermentation by Fuzzy-Neural Network

In this paper,a novel fuzzy neural network model,in which an adjustable fuzzy sub-space was designed by uniform design,has been established and used in fed-batch yeast fermentationas an example.A brand-new optimization sub-network with special structure has been built andgenetic algorithm,guaranteeing the optimization in overall space,is introduced for the feed rateoptimization.On the basis of the model network,the optimal substrate concentration and theoptimal amount of fed-batch at different periods have been studied,aided with the optimizationnetwork and the genetic algorithm separately.The above results can be used as a basis for theestablishment of a fuzzy neural network controller.

Principles and Case Studies of Fed Batch Fermentation and Continuous Fermentation

Modern fermentation processes include a variety of fermentation methods,such as fed batch fermentation and continuous fermentation.This paper will focus on the principles and case studies of these two methods.Fed batch fermentation originates from fractionation fermentation with closed culture and adjustment of the pH value of the carbon source,to which the process of feeding the carbon source to the cell culture in a controlled manner has been added.This type of fermentation is more commonly used compared to the other.Continuous fermentation is also a closed fermentation system,which can operate without restrictions by continuous or intermittent addition of fresh nutrient media to the fermenter;however,it is susceptible to contamination by stray bacteria and metabolic inconvenience.

Breeding of Coenzyme Q_(10) Produced Strain by Low-Energy Ion Implantation and Optimization of Coenzyme Q_(10) Fermentation

In order to increase the production efficiency of coenzyme Q10, the original strain Agrobacterium tumefaciens ATCC 4452 was mutated by means of Nitrogen ions implantation. A mutant strain, ATX 12, with high contents of coenzyme Q10 was selected. Subsequently, the conditions such as carbohydrate concentration, nitrogen source concentration, inoculum＇s size, seed age, aeration and temperature which might affect the production of CoQ10 were investigated in detail. Under optimal conditions, the maximum concentration of the intracellular CoQ10 reached 200.3 mg/L after 80 h fed-batch fermentation, about 245% increasing in CoQ10 production after ion implantation, compared to the original strain.

An Approach to Continuous Approximation of Pareto Front Using Geometric Support Vector Regression for Multi-objective Optimization of Fermentation Process

The approaches to discrete approximation of Pareto front using multi-objective evolutionary algorithms have the problems of heavy computation burden, long running time and missing Pareto optimal points. In order to overcome these problems, an approach to continuous approximation of Pareto front using geometric support vector regression is presented. The regression model of the small size approximate discrete Pareto front is constructed by geometric support vector regression modeling and is described as the approximate continuous Pareto front. In the process of geometric support vector regression modeling, considering the distribution characteristic of Pareto optimal points, the separable augmented training sample sets are constructed by shifting original training sample points along multiple coordinated axes. Besides, an interactive decision-making(DM)procedure, in which the continuous approximation of Pareto front and decision-making is performed interactively, is designed for improving the accuracy of the preferred Pareto optimal point. The correctness of the continuous approximation of Pareto front is demonstrated with a typical multi-objective optimization problem. In addition,combined with the interactive decision-making procedure, the continuous approximation of Pareto front is applied in the multi-objective optimization for an industrial fed-batch yeast fermentation process. The experimental results show that the generated approximate continuous Pareto front has good accuracy and completeness. Compared with the multi-objective evolutionary algorithm with large size population, a more accurate preferred Pareto optimal point can be obtained from the approximate continuous Pareto front with less computation and shorter running time. The operation strategy corresponding to the final preferred Pareto optimal point generated by the interactive DM procedure can improve the production indexes of the fermentation process effectively.

Batch, Fed Batch Production and Characterization of Glutaminase Free L-Asparaginase II of <i>Pectobacterium carotovorum</i>MTCC 1428 in <i>Escherichia coli</i>

The present study describes the production optimization of recombinant L-asparaginase II of Pectobacterium carotovorum MTCC 1428 in Escherichia coli BL21 (DE3) at batch and fed batch bioreactor level. Production of recombinant L-asparaginase II in batch and fed batch mode was found to be 1.34 and 5.38 folds higher, respectively as compared to shake flask culture. SDS-PAGE and native PAGE of the purified enzyme revealed that molecular mass of the subunits and native enzyme are ~37.5 kDa and ~150 kDa, respectively. Optimum range of pH and temperature for hydrolysis of L-asparagine were found to be 7.5 - 8.5 and 47°C - 52°C, respectively. The recombinant enzyme is very specific for its natural substrate, L-asparagine. The activity of recombinant L-asparaginase II is improved by mono cations and diverse effectors including Na+, K+, L-cystine, L-histidine, glutathione and 2-mercaptoethanol whereas, it is moderately inhibited by different divalent cations and thiol group blocking reagent. The kinetic parameters Km, Vmax, kcat and Km/Kcat of purified recombinant L-asparaginase II were determined. The purified L-asparaginase II possesses no partial glutaminase activity, which is prerequisite to reduce the possibility of side effects during the course of anti-cancer therapy.

油脂酵母Trichosporon fermentans HWZ004的分批补料法高密度培养

为提高发酵性丝孢酵母Trichosporon fermentans HWZ004发酵产油脂的能力,采用分批补料的培养方式对其进行高密度培养,考察了初始糖浓度对发酵性丝孢酵母生长及油脂积累的影响,确定初始糖浓度为100 g/L,在该浓度下Trichosporon fermentans HWZ004可获得较高的生物量及油脂产量,且不存在底物抑制现象;然后,在5 L发酵罐中进行分批补料培养,发酵132h后,Trichosporon fermentans HWZ004的生物量、油脂含量及油脂生产强度分别为102g/L、48%和0.37g/(L·h).气相色谱分析结果表明,所得油脂的脂肪酸组成主要为硬脂酸、棕榈酸、油酸、亚油酸,不饱和脂肪酸含量达65%以上,适用于生物柴油的生产.

南荻纤维素分批补料同步糖化发酵产乙醇工艺优化及其结构表征

文章以南荻纤维素为原料,采用分批补料同步糖化发酵工艺生产生物乙醇,分析了底物初始质量分数、底物最终质量分数、补料次数对乙醇产量的影响,并在此基础上采用正交试验进行工艺优化,获得最佳工艺参数:底物初始质量分数为17.5%,分批补料2次至底物最终质量分数为32.5%。在此条件下,乙醇含量最高可达70.15 g/L,较未补料同步糖化发酵过程获得的最大乙醇含量提高23.36%。在分批补料同步糖化发酵最佳工艺的基础上,做了物料平衡分析,并采用扫描电镜、傅里叶变换红外光谱、X-衍射对发酵前、后的南荻纤维素结构进行表征,结果显示,酶解糖化及发酵过程会改变南荻纤维素的内部结构,使氢键或共价键断裂,底物结晶度提高。

批式流加发酵中的鲁棒脉冲时滞最优控制

本文研究了批式流加发酵中的鲁棒脉冲时滞最优控制问题。首先,提出一个非线性状态依赖的脉冲时滞系统描述批式流加发酵甘油生产1,3-丙二醇(1,3-PD)过程。由于批式流加发酵过程中的动力学参数难以准确估计,本文建立了一个具有连续状态不等式约束的鲁棒脉冲时滞最优控制模型。这里,目标函数为终端时刻1,3-PD浓度及其关于动力学参数的灵敏性的加权和,控制向量为流加发生时甘油的临界浓度及每次流加时甘油的流加体积。然后,通过引入辅助脉冲系统,将该鲁棒最优控制问题转化为等价的标准最优控制问题。进一步,通过约束转换技术,将等价的最优控制问题转化为仅具有盒式约束的罚问题。最后,设计了一种并行差分进化算法求解转化后的罚问题。数值结果表明:当参数受到微小扰动时,尽管牺牲了少量的1,3-PD浓度,但是明显提高了系统的鲁棒性。

高温产2,3-丁二醇甲基芽孢杆菌的筛选及其发酵优化

2,3-丁二醇是一种典型的大宗化学品,广泛应用于化工、食品、医药、燃料等领域。生物法发酵制备2,3-丁二醇是当下的主要方法,然而,大多数报道的菌株为条件致病菌且发酵条件严格,限制了生物法制备2,3-丁二醇的发展。该研究以温泉底土壤为试验材料,经过分离、筛选与鉴定获得了1株高温条件下产2,3-丁二醇的甲基芽孢杆菌。通过摇瓶实验优化发酵工艺,确定最适发酵条件与最优培养基组成,利用优化后的发酵工艺甲基芽孢杆菌发酵24 h产物产量达到32.57 g/L。进一步进行上罐试验,基于不同发酵罐转速条件下甲基芽孢杆菌的产量积累情况,理性设计两阶段发酵策略,并以最优策略分批补料发酵制备2,3-丁二醇,2,3-丁二醇的产量在甲基芽孢杆菌发酵45 h时达到73.19 g/L,转化率达到90.25%。

热带假丝酵母β-紫罗兰酮工程菌的构建及其发酵优化

β-紫罗兰酮是一种来源于植物的不规则单萜化合物,具有良好的驱虫、抗菌、抗癌和抗氧化等生物活性,广泛应用于农业、食品、医药和香精香料等行业。热带假丝酵母是潜在的萜类高效合成底盘细胞。为了提高β-紫罗兰酮产量,该研究在课题组前期构建的β-紫罗兰酮工程菌的基础上,通过酶的亚细胞区室组合,构建了一株β-紫罗兰酮合成关键基因PhCCD1,同时在其胞质、过氧化物酶体和脂滴表达的β-紫罗兰酮工程菌PCL-01;然后,对其摇瓶发酵培养基装液量、培养基碳氮比、发酵温度和辅因子Fe^(2+)浓度等条件进行优化;最后进行5 L发酵罐小试。工程菌PCL-01的β-紫罗兰酮摇瓶产量为152.4 mg/L,较出发菌株增加50%;经过摇瓶发酵优化,β-紫罗兰酮产量为185.8 mg/L,提升22%;最后,在5 L发酵罐上可达403.9 mg/L。该研究对于其他萜类物质合成的菌株代谢改造具有借鉴意义,亦可为其他菌种发酵性能的研究提供参考价值。

补料分批发酵优化氧化葡糖杆菌UZ209发酵产酸工艺

醋酸菌作为氧化乙醇生成乙酸的主要菌种,其产酸能力备受关注。氧化葡糖杆菌具有不完全氧化糖和糖醇的特性,可以产生多种类的有机酸,提升食醋风味,但相较于醋酸杆菌属,其产酸能力和乙醇耐受能力较低,限制了其在食醋工业中的应用。本研究通过优化氧化葡糖杆菌UZ209的培养基成分和补料分批发酵方式提高其发酵液总酸含量。结果表明,最佳发酵培养基为:2%葡萄糖、1%酵母浸粉、8%乙醇。补料分批发酵可以显著提高氧化葡糖杆菌UZ209的生长和发酵特性,在乙醇含量为3%的液体发酵培养基中补料分批发酵的最佳方式为按照吸光值补料,最佳补料吸光值为OD6000.8±0.025。在此条件下,5%和6%乙醇补料分批发酵8 d的总酸含量分别为(7.42±0.16)和(6.95±0.45)g/100mL,分别比对照组提高了18.91%和15.04%。

HIP1过表达对酿酒酵母工程菌麦角硫因产量的影响

麦角硫因是一种普遍存在于高等动植物体内的天然抗氧化剂,具有减少细胞氧化损伤、抗炎和治疗相关疾病的能力,被广泛应用于食品、化妆品和医药行业。与传统的植物提取法及化学合成法相比,通过微生物合成法制备麦角硫因具有周期短、成本低和产量高等优势。该研究以酿酒酵母Saccharomyces cerevisiae CEN.PK113-5D为出发菌株,利用CRISPR/Cas9系统向其基因组中引入异源酶基因,构建具有完整麦角硫因生物合成途径的酿酒酵母工程菌。为进一步提高其产量,过表达组氨酸特异性转运蛋白Hip1p后,在仅有0.5 mmol/L组氨酸供给条件下,菌株胞内游离组氨酸水平显著提高,且麦角硫因产量提高10%。最终经摇瓶发酵72 h产量达(6.88±0.11)mg/L,而在5 L罐补料分批发酵策略下,培养72 h后麦角硫因产量达237.34 mg/L。该研究为后续代谢工程改造产麦角硫因酿酒酵母工程菌株提供靶基因参考。

补料发酵工艺的应用及其研究进展

综述了补料工艺在发酵工业中应用和研究。介绍了补料发酵工艺及其优点,着重讨论了补料发酵动力学和控制理论研究,以期为补料发酵的应用提供充分的参考依据。

癸酸抗性突变株流加发酵法生产达托霉素

采用He-Ne激光对达托霉素产生菌玫瑰孢链霉菌LC-54进行诱变,筛选前体癸酸抗性突变株,在此基础上进一步考察了一次补料、间歇补料和恒速流加补料等不同补料策略对达托霉素发酵生产的影响．结果表明,选育前体癸酸抗性突变株可以提高达托霉素的生产能力,并最终获得了达托霉素发酵效价较出发菌株提高了37．2%的高产突变株LC-KS-54．采用恒速流加策略,能够有效地提高达托霉素产量,经过130 h的恒速流加培养,细胞干重达到15．4g/L,达托霉素产量达到258 mg/L,明显优于其他前体补料策略．

分批补料合成纤维素酶扩大试验

研究了里氏木霉以纸浆为碳源分批补料制备纤维素酶。里氏木霉在10L发酵罐中以纸浆为碳源间歇式发酵合成纤维素酶,培养基中碳源浓度为15g/L时,滤纸酶活力、纤维二糖酶活力、酶产率和酶得率分别为2.15FPIU/mL、0.20IU/mL、16.3FPIU/(L·h)和143.3FPIU/g;碳源浓度提高到27.5g/L,采用分批补加碳源的方法,滤纸酶活力、纤维二糖酶活力、酶产率和每克纸浆酶得率分别为3.90FPIU/mL、0.35IU/mL、23.2FPIU/(L·h)和141.8FPIU。研究表明,提高培养基中碳源浓度,采用补料分批发酵技术,产酶时酶活力和酶产率随着培养基中碳源浓度的提高而提高,且保持酶得率不变,达到了降低产酶成本的目的。

不同甲醇流加策略对重组毕赤酵母高密度发酵生产水蛭素的影响

考察了不同甲醇流加策略对毕赤酵母高密度发酵生产水蛭素的影响。溶氧控制法不能有效地防止甲醇的过量流加。气相色谱离线检测法虽然防止了甲醇流加过量 ,但甲醇浓度的波动较大。利用甲醇传感器在线检测控制甲醇的流加可维持较恒定的甲醇浓度。在流加甲醇的同时 ,以限制性速率流加甘油可以增加表达期间的能量供应 ,提高产物的表达量。经优化后 ,采用甲醇甘油混合流加时细胞干重达到 16 2g L ,水蛭素活性达到 2 4×10 4ATU mL ,即 1 7g L。

木质纤维素稀酸水解液乙醇发酵的新方法

为了降低木质纤维素水解液发酵抑制剂对乙醇发酵的负影响,采用混合菌种对木质纤维素稀酸水解液乙醇发酵方式进行了研究。对批式发酵、补料批式发酵和间隔补料批式发酵3种发酵方式进行了比较。实验结果表明,间隔补料批式发酵可以有效地减弱水解液中抑制因子对菌种的影响,乙醇产量明显高于其他两种发酵方式,利用酿酒酵母(Saccaromyces cerevisiae 2.535)和嗜鞣管囊酵母(Pachysolen tannophilis ATCC 32728)混合发酵,乙醇产量最终达到14.4g/L,乙醇产率(Yp/s)为0.47g/g,相当于最大理论产率的92.2%。利用酿酒酵母和重组大肠杆菌混合菌种发酵,乙醇产量达到了14.5g/L。对木质纤维素稀酸水解液采用间隔补料批式乙醇发酵方法,可进一步减少抑制剂对乙醇发酵的影响,使发酵顺利进行。

Alcaligenes sp．NX-3产威兰胶的补料分批发酵工艺研究

在7.5L发酵罐上考察了Alcaligenes sp.NX-3产威兰胶的发酵工艺。选用葡萄糖为碳源,通过分析比较不同初糖浓度下的细胞比生长速率和产物比合成速率,进一步研究了不同补料方式对产胶的影响。结果表明,采用分批补糖发酵工艺,威兰胶产量较分批发酵提高了13.6%,而且有效地缩短了发酵周期。在50L发酵罐上进行补料分批发酵放大实验,威兰胶产量高达27.0g/L,葡萄糖转化率由初始的44%提高到54%。