正十六烷对节杆菌发酵产环磷酸腺苷的影响

溶氧在节杆菌发酵产环磷酸腺苷(c AMP)过程中起着重要作用。本研究中,笔者首先对4种氧载体(正癸烷、正十二烷、正十四烷和正十六烷)的最佳添加浓度和添加时间进行筛选。结果表明:在0 h添加2%(体积分数)正十六烷促进c AMP生产效果最佳。在5 L发酵罐中添加2%(体积分数)正十六烷后,细胞干质量(DCW)和c AMP产量分别达到10. 85 g/L和8. 87 g/L,比对照分别提高了19. 4%和23. 2%。氧载体的加入提高了发酵液中的氧传递系数(K_La),降低了胞内NADH/NAD~+比率以及胞内三磷酸腺苷(ATP)水平,而对关键酶的酶活影响不显著。     

赭曲霉转化甾体过程中色素的分析和控制

通过对甾体Ｃ_１１α羟基化菌株赭曲霉发酵液中色素的分离纯化,得到一种砖红色的色素。经紫外－可见分光光度法、红外光谱法和核磁共振谱图分析,证明该色素为苯醌类色素,与初级代谢过程中的芳香族氨基酸有相同的代谢途径,利用芳香族氨基酸的代谢模式,在加有２％沃氏氧化物的发酵体系中添加５ｍｍｏｌ／Ｌ的色氨酸,或５ｍｍｏｌ／Ｌ等体积的苯丙氨酸和酪氨酸混合液,均能明显抑制发酵过程中色素的产生,提高产物的收率。同时通过诱变选育的莽草酸缺陷型菌株发酵后,甾体转化力均有所下降,但发酵液不再产生砖红色色素,进一     

非常规介质中细胞生化反应研究进展

对完整细胞在非常规介质中的生物催化反应进行了回顾 ,分别总结了产物为醇 ,甾体 ,有机酸 ,生物大分子及其它各类反应的研究进展。并从溶剂和细胞两种角度对主要的研究方法进行了阐述。     

非常规介质中细胞生化反应研究进展

对完整细胞在非常规介质中的生物催化反应进行了回顾,分别总结了产物为醇,甾体,有机酸,生物大分子及其它各类反应的研究进展。并从溶剂和细胞两种角度对主要的研究方法进行了阐述 。     

枯草芽胞杆菌PnbA酯酶选择性拆分制备l-薄荷醇

利用重组大肠杆菌表达来源于枯草芽胞杆菌CICC 20034中的PnbA酯酶,不对称催化水解dl-薄荷醇丙酸酯制备l-薄荷醇。考察助溶剂种类、助溶剂添加浓度、温度、催化剂用量、底物浓度以及p H等对反应的影响。结果表明:添加25%(体积分数)助溶剂乙醇可显著提升该酯酶对l-薄荷醇的立体选择性,对映选择率(E)由2.4提高到99.43,为不加乙醇条件下的40倍。酶催化最佳条件:25%乙醇作为助溶剂,反应温度37℃,缓冲液为0.1 mol/L Tris-HCl(p H 8.0)并保持p H 8.0反应条件,底物量50 mmol/L,反应体系中酶的添加量750 U/m L,在此条件下,酶促反应30 min后,l-薄荷醇转化率可达34%,产物光学纯度对映体过量值(e.e._p)达95%。     

火菇属野生菌株发酵液对栽培菌株的化感效应

为探究火菇属Flammulina野生菌株发酵液对栽培菌株的影响,以火菇属野生柳生金针菇F. rossica F52、野生金针菇F. filiformis F39和栽培金针菇F. filiformis F47为供试菌株,采用固体平板培养和液体发酵的方法,对比了添加不同体积分数火菇属野生菌株发酵液对栽培菌株平板中菌丝长速、液体发酵中菌丝生物量、干质量及漆酶和淀粉酶活力的影响。结果表明：不同体积分数野生金针菇菌株F39发酵液对栽培菌株F47菌丝长速有抑制作用,对发酵液中菌丝生物量无显著影响,当添加发酵液体积分数为50%时可显著提高菌株F47发酵液中漆酶活力,当体积分数为40%时,可显著提高菌株F47发酵液中淀粉酶的活力（P<0.05）;添加不同体积分数的野生柳生金针菇菌株F52发酵液对菌株F47除淀粉酶外其他指标均影响显著,添加体积分数为30%的发酵液对栽培金针菇F47菌丝长速具有显著的促进作用,当添加发酵液体积分数为40%时,对菌株F47液体发酵中菌丝干质量和漆酶活力均有显著促进作用（P<0.05）。证明了化感效应不仅存在于属间,在同属下的种间也存在。     

基于微生物法甾体羟基化反应

甾体化合物又称类固醇,是重要的药物活性成分和药物合成中间体,因其具有环戊烷多氢菲的基本骨架,反应类型丰富,其中羟基化反应因产品具有广阔的市场应用前景而受到广泛关注。羟基化反应有化学法和生物法两种,生物法具有区域和立体专一性、对映体专一性等特点而成为目前主要的生产方法。首先从反应原理、类型及机制3个方面介绍了甾体微生物羟基化过程;其次,基于文献及自身研究工作,从甾体羟基化反应的发酵条件、底物溶解性、跨膜运输及反应器内流体力学特性4个角度对羟基化过程的影响进行了综述;最后,基于甾体羟基化反应特性及当前研究进展,对该反应过程后续研究提出展望,旨在为后期甾体羟基化反应的相关研究提供一定参考依据。     

非常规介质中细胞生化反应研究进展

对完整细胞在非常规介质中的生物催化反应进行了回顾，分别总结了产物为醇，甾体，有机酸，生物大分子及其它各类反应的研究进展。并从溶剂和细胞两种角度对主要的研究方法进行了阐述。     

Penicillium raistrickii 15α羟基化左旋乙基甾烯双酮工艺研究

通过生物转化技术对甾体化合物左旋乙基甾烯双酮进行15α位羟基化,合成了重要的药物中间体15α-羟基左旋乙基甾烯双酮,对生物转化工艺进行了优化。重点对底物的助溶剂进行了筛选,同时对培养基成分,接种量,初始pH,通气量,投料浓度,投料时间,转化时间等转化条件进行了优化。结果表明:在摇瓶发酵中,Penicilliumraistrickii对甾体化合物左旋乙基甾烯双酮生物转化,产物15α-羟基左旋乙基甾烯双酮转化率达到60%,在发酵罐放大试验中,转化率达到50%以上。具有工业生产前景。     

短梗霉多糖发酵过程特征的研究

本文针对短梗霉多糖发酵过程,经研究建立了基于逻辑方程和Luedeking—Piret方程的动力学模型： dx／dt=肛x(1一x／xm) dP／dt=m1x十m2(dx/dt) ds／dt=-b1x-b2(dx／dt)-b3(dP／dt) 其流变特性由初始时的牛顿流体转变为典型的假塑性非牛顿流体并遵从指数方程,即：τ=Kγn随发酵过程进行,发酵液的表观粘度增大,体积氧传质系数减小。搅拌转速的增加有利于提高体积氧传质系数。流变指数n、稠度系数K、气液传质系数Kla。与菌体浓度x、多糖浓度P、搅拌转速N及表观粘度ηa间分别有如下经验方程; k=1．2×10-2X2.43 n=0．461(P／Pm)0.07(x／xm)0.216 KLaDi2=1．48×104(D:N2)0.71(ηW)0.15 Dg ηa     

Prozeßtechnische Untersuchungen zur Ausschöpfung mikrobieller Leistungsgrenzen

The knowledge of the upper limits of growth rate and cell density is necessary to define the maximum possible productivity of a growth process. After a brief survey of the theoretical basis of microbial growth in dependence of mass transfer of oxygen and rheological properties of fermentation medium recent experimental results on the influence of higher pressure in the fermenter, of different substrate and dissolved oxygen concentrations on growth rate and yield coefficients of methanol utilizing bacteria are presented. Also some correlations between cell concentration and viscosity of fermentation broth are considered.     

A comparative evaluation of oxygen mass transfer and broth viscosity using Cephalosporin-C production as a case strategy

The production of Cephalosporin-C (CPC) a secondary metabolite, using a mold Acremonium chrysogenum was studied in a lab scale Internal loop air lift reactor. Cephalosporin-C production process is a highly aerobic fermentation process. Volumetric gas–liquid mass transfer coefficient (k_La) and viscosity (η) were evaluated, during the growth and production phases of the microbial physiology. An attempt has been made to correlate the broth viscosity, η and volumetric oxygen transfer coefficient, k_La during the Cephalosporin-C production in an air lift reactor. The impact of biomass concentration and mycelial morphology on broth viscosity has been also evaluated. The broth exhibits a typical non-Newtonian fermentation broth. Rheology parameters like consistency index and fluidity index are also studied.     

以摇瓶所得摄氧率为基准进行发酵放大

通过设计特殊摇瓶,用亚硫酸盐法测出摇瓶口纱布层氧通透率的基础上,在实际发酵情况下通过测定瓶内气、液相氧的变化得出其发酵过程中的摄氧率(OUR)及氧传递系数(K_La)。以特制摇瓶取得的菌体CUR为基准进行发酵过程和发酵罐的放大。通过质谱仪在线检测及采样分析,研究了3种不同供气流量及搅拌转速下的放大结果。摇瓶与发酵罐在菌体OUR、菌体产量方面吻合很好,而在整个放大过程中,发现摇瓶与发酵罐内的氧传递系数(K_La)、溶解氧(C_L)差异较大。     

Production of phytase (myo-inositolhexakisphosphate phosphohydrolase) by Aspergillus niger van Teighem in laboratory-scale fermenter

The growth and production pattern of phytase by a filamentous fungus, Aspergillus niger van Teighem, were studied in submerged culture at varying agitation rates and controlled and uncontrolled pH conditions. Allowing the initial culture to grow under neutral condition with subsequent decline in pH resulted in increased phytase productivity. A maximum of 141 nkat/mL phytase was obtained when the broth pH was maintained at pH 2.5 as compared to 17 nkat/mL units at controlled pH 5.5. The culture morphology and rheological properties of the fermentation broth significantly varied with the agitation rate. The volumetric oxygen transfer coefficient was determined at different phases of fungal growth during batch fermentation using static gassing out and dynamic gassing out methods. The oxygen transfer coefficient (k(L)a) of the fermenter was found to be 125 h(-)(1) at 500 rpm as compared to 38 h(-)(1) at 200 rpm. The oxygen transfer rates at different phases of growth were significantly affected by cell mass concentration and fungal morphology. During the course of fermentation there was a gradual decline of k(L)a from 97 h(-)(1) on day 2 to 63 h(-)(1) on day 6 of fermentation, after which no significant change was observed. The degree of agitation considerably influenced the culture morphology where shear thinning of filamentous fungus was observed with the increase in agitation.     

Effects of microorganisms on effective oxygen diffusion coefficients and solubilities in fermentation media

Effective oxygen diffusion coefficients and solubilities were measured for submerged cultures of Saccharomyces cerevisiae, Escherichia coli, and Penicillium chrysogenum. Both effective oxygen diffusion coefficients and solubilities were found to decrease with increasing cell concentrations in the fermentation media. Comparison of the experimental results of effective oxygen diffusion coefficients in fermentation media with values theoretically predicted on the assumption of unpenetrable microbial cells indicates that oxygen molecules diffuse through the cells during the diffusion process. Within the cell concentration range of typical submerged fermentations, the effective oxygen diffusion coefficient of the fermentation media can be described as D(e) = A(1)f + A(2)f(2). In this equation, fis the cell volume fraction and both A(1) and A(2) are functions of the shape of the cells and the ratio of effective oxygen diffusion coefficient in microbial cells to that in the medium.     

Rheological characterization of a fungal fermentation for the production of pneumocandins

The filamentous fungus Glarea lozoyensis produces a novel, pharmaceutically important pneumocandin (B₀) that is used to synthesize a polypeptide, which demonstrates fungicidal activity against clinically relevant pathogens. The scale-up of the pneumocandin fermentation requires an understanding of the rheological properties of the broth and the factors that influence flow behavior. A systematic approach for characterizing the rheological properties using a standard methodology is presented here. An appropriate rheometer was chosen and the effects of shear rate ramping, broth handling, creep and yield testing were examined. The fed-batch fermentation used a soluble production medium that allowed the relationship between biomass and rheological properties to be studied up to the 19-m³ scale. The morphologically heterogeneous broth demonstrated time-dependent shear thinning behavior with thixotropy and a yield stress. The flow curves were described by the power law model, with flow behavior of 0.35-0.4 and consistency index up to 10 Pa.sⁿ. The use of a cup and bob rheometer was preferable to alternative techniques, including turbine and spindle systems defined by Mitschka's technique. The consistency index and flow behavior were shown to have a non-linear relationship with biomass concentrations up to 140 g/L. The consistency index continually increased with biomass during the fermentation, while the flow behavior initially decreased rapidly and then remained at low values for the remainder of the batch cultivation. The consistency index and yield stress were influenced by temperature, osmotic pressure, and pH, while the flow behavior remained independent of these factors.     

Oxygen transfer in broths of plant cells at high density

The rheological properties of the culture broths of some plant cells (Cudrania tricuspidata, Vinca rosea, and Agrostemma githago) at high density (10-18 g dry wt/L) were measured, and oxygen transfer in the broths in various bioreactors was investigated. The rheological properties of the broths were dependent on the size, specific gravity, and concentration of the cell aggregates contained in the broths. The broths were non-Newtonian and pseudoplastic fluids. The flow behavior index n was fairly constant (0.53) and the consistency index K varied in proportion to the sixth-to-seventh power of the cell mass concentration M. The apparent viscosity mu(a) of the broths was in proportion to the 6.5th power of M. The oxygen transfer in the broths was discussed on the basis of the results obtained for suspensions of granulated agars (agar concentration, 5.8%) in water, which were similar to the broths in rheological properties. The volumetric oxygen transfer coefficient k(L)a in the broths was dependent on mu(a)(k(L)a proportional, variant mu(a) (-m)) and decreased greatly at a certain apparent viscosity, mu(ac). The values of m and mu(ac) were closely related to the aeration-agitation mechanisms of the bioreactors. The values of mu(ac) in aeration-agitation type bioreactors was larger than that in aeration-type bioreactors, whereas for m, the reverse was true.     

Dimensional analysis for modeling oxygen transfer in rotating biological contactor

In rotating biological contactors (RBC), the oxygen mass transfer coefficient (KLa) is often inadequately predicted by the available models. Hence, dimensional analysis based empirical models were developed for predicting KLa and the component of KLa due to turbulence (KLat) using data available in the literature. The overall oxygen transfer number (OTN) and its component due to turbulence (OTNt) were defined as dimensionless groups based on KLa and KLat, respectively. They were expressed as a function of dimensionless groups, formed using disc diameter (D), area of discs (Ad), rotational speed (omega) and cross-sectional area of the tank (At). OTN was also a function of thickness of the water film on the disc (delta) and working volume of the reactor (V). Decrease in number of discs and decrease in (Ad/At) resulted in decreasing OTN but increasing OTNt. Both OTNt and OTN increased with increase in omega. The proposed empirical models will facilitate scale-up of RBCs.     

Oxygen transfer kinetics of riboflavin fermentation by Ashbya gossypii in agitated fermentors

Effects of agitation and aeration rates on volumetric oxygen transfer coefficient and oxygen uptake rate of a riboflavin broth containing Ashbya gossypii were investigated in three batch, sparged, and agitated fermentors having the working volumes of 0.42, 0.85, and 2.5 l. The change of oxygen uptake rate with time at 250 rev min⁻¹ stirring and vvm aeration rates was shown. The volumetric oxygen transfer coefficients and maximum oxygen uptake rates obtained have been correlated to mechanical power inputs per unit volume of the fermentation broth and the superficial air velocities.     

Evaluation of oxygen transfer rates in stirred-tank bioreactors for clinical manufacturing

Several methods are available for determining the volumetric oxygen transfer coefficient in bioreactors, though their application in industrial bioprocess has been limited. To be practically useful, mass transfer measurements made in nonfermenting systems must be consistent with observed microbial respiration rates. This report details a procedure for quantifying the relationship between agitation frequency and oxygen transfer rate that was applied in stirred-tank bioreactors used for clinical biologics manufacturing. The intrinsic delay in dissolved oxygen (DO) measurement was evaluated by shifting the bioreactor pressure and fitting a first-order mathematical model to the DO response. The dynamic method was coupled with the DO lag results to determine the oxygen transfer rate in Water for Injection (WFI) and a complete culture medium. A range of agitation frequencies was investigated at a fixed air sparge flow rate, replicating operating conditions used in Pichia pastoris fermentation. Oxygen transfer rates determined by this method were in excellent agreement with off-gas calculations from cultivation of the organism (P = 0.1). Fermentation of Escherichia coli at different operating parameters also produced respiration rates that agreed with the corresponding dynamic method results in WFI (P = 0.02). The consistency of the dynamic method results with the off-gas data suggests that compensation for the delay in DO measurement can be combined with dynamic gassing to provide a practical, viable model of bioreactor oxygen transfer under conditions of microbial fermentation.