Relation between dissolved oxygen concentration and ajmalicine production rate in high-density cultures of Catharanthus roseus

The relation between dissolved oxygen (DO) and the ajmalicine production rate of Catharanthus roseus was investigated in 15-L tank reactors at constant stirrer speed and gas flow rate. Below a DO concentration of 29% of air saturation the ajmalicine production rate was less than 0.06 mumol/g/d. Above a DO of 43% the ajmalicine production rate was constant at 0.21 mumol/g/d. Between a DO of 29% and 43% there was a strong relation between the ajmalicine production rate and the DO concentration. After a period of at least 12 days at DO /=57%. A kinetic equation is proposed for the relation between DO and the specific ajmalicine production rate. (c) 1995 John Wiley & Sons, Inc.     

Photobioreactors for microalgal cultures: A Lagrangian model coupling hydrodynamics and kinetics

Closed photobioreactors have to be optimized in terms of light utilization and overall photosynthesis rate. A simple model coupling the hydrodynamics and the photosynthesis kinetics has been proposed to analyze the photosynthesis dynamics due to the continuous shuttle of microalgae between dark and lighted zones of the photobioreactor. Microalgal motion has been described according to a stochastic Lagrangian approach adopting the turbulence model suitable for the photobioreactor configuration (single vs. two‐phase flows). Effects of light path, biomass concentration, turbulence level and irradiance have been reported in terms of overall photosynthesis rate. Different irradiation strategies (internal, lateral and rounding) and several photobioreactor configurations (flat, tubular, bubble column, airlift) have been investigated. Photobioreactor configurations and the operating conditions to maximize the photosynthesis rate have been pointed out. Results confirmed and explained the common experimental observation that high concentrated cultures are not photoinhibited at high irradiance level. © 2015 American Institute of Chemical Engineers Biotechnol. Prog., 31:1259–1272, 2015     

High-density algal photobioreactors using light-emitting diodes

Lack of high-density algal photobioreactors (PBR) has been a limitation in exploiting the biotechnological potential of algae. Recent developments of highly efficient light-emitting diodes (LED using gallium aluminum arsenide chips) have made the development of a small LED-based PBR possible. We have calculated theoretical values of gas mass transfer requirements and light-intensity requirement to support high-density algal cultures for the 680 nm monochromatic red light from LED as a light source. A prototype PBR has been designed based on these calculations. A cell concentration of more than 2 x 10(9) cells/mL (more than 6.6% v%sol;v), cell doubling times as low as 12 h, and an oxygen production rate as high as 10 mmol oxygen/L culture/h were achieved using on-line ultrafiltration to periodically provide fresh medium. (c) 1994 John Wiley & Sons, Inc.     

微藻培养过程的光特性研究进展

微藻培养过程中光的吸收、衰减以及光暗循环等特性是影响微藻的生长速度及其产量的重要因素。本文分析了微藻的光吸收过程、光在微藻培养液中的衰减特性以及微藻培养过程中的光暗循环特性,重点综述了国内外各类光生物反应器中光特性的研究进展,并对其发展方向进行了展望,为微藻培养光生物反应器的设计提供参考依据。     

The oxygen evolution methodology affects photosynthetic rate measurements of microalgae in well‐defined light regimes

Designing photobioreactors correctly is a must for the success of microalgal mass production. Optimal photobioreactor design requires a precise knowledge of photosynthesis dynamics in fluctuating light conditions and hence a method for the measurement of photosynthetic rates in specific light regimes. However, it is not uncommon in literature that experimental protocols used to obtain oxygen generation rates are described ambiguously and the reported rates of photosynthesis vary widely depending on the methodology. Additionally, quite a number of methods overlook certain aspects that can affect the estimated rates significantly, and can therefore affect photobioreactor design. We have developed a method based on oxygen evolution measurements that accurately determines photosynthetic rates under well‐defined light regimes. Our experimental protocol takes into account most of the issues that can affect the rates of oxygen generation, such as depletion of nutrients during the measurements and precision of the measurements. We have focused on the basic applications in photobioreactor design and used a dynamic model of photosynthesis to analyze our results and compare them with available published data. The results suggest that our oxygen evolution method is consistent. Biotechnol. Bioeng. 2010;106: 228–237. © 2010 Wiley Periodicals, Inc.     

A novel in situ probe for oxygen uptake rate measurement in mammalian cell cultures

The newly developed in situ oxygen uptake rate (in situ OUR) probe presented in this article is based on the in situ microscope technology platform. It is designed to measure the oxygen uptake rate (OUR) of mammalian cells, an important parameter for metabolic flux analysis, inside a reactor (in situ) and in real-time. The system isolates a known volume of cell culture from the bulk inside the bioreactor, monitors the oxygen consumption over time, and releases the sample again. The sample is mixed during the measurement with a new agitation system to keep the cells in suspension and prevent oxygen concentration gradients. The OUR measurement system also doubles as a standard dissolved oxygen (DO) probe for process monitoring when it is not performing OUR measurements. It can be equipped with two different types of optical sensors (i.e., DO, pH) simultaneously or a conventional polarographic DO-probe (Clark type). This new probe was successfully tested in baby hamster kidney perfusion cell cultures.     

A new photobioreactor for continuous microalgal production in hatcheries based on external-loop airlift and swirling flow

This study deals with the scale of a new photobioreactor for continuous microalgal production in hatcheries. The combination of the state-of-art with the constraints inherent to hatcheries has turned the design into a closed, artificially illuminated and external-loop airlift configuration based on a succession of elementary modules, each one being composed of two transparent vertical interconnected columns. The liquid circulation is ensured pneumatically (air injections) with respect to a swirling motion (tangential inlets). A single module of the whole photobioreactor was built-up to scale its geometry (diameter and length) and to optimize its design (air sparger, tangential inlets). The volumetric productivities were predicted by modeling radiative transfer and growth of Isochrysis affinis galbana (clone Tahiti). The hydrodynamics of the liquid phase was modeled in terms of global flow behavior (circulation and mixing times, Péclet number) and of swirling motion decay along the column (Particle Image Velocimetry). The aeration performances were determined by overall volumetric mass transfer measurements. Continuous cultures of Isochrysis affinis galbana (clone Tahiti) were run in two geometrical configurations, generating either an axial or a swirling flow. Lastly, the definitive options of design are presented as well as a 120-L prototype, currently implemented in a French mollusk hatchery and commercialized.     

The effect of dissolved oxygen and aeration rate on antibiotic production of Streptomyces fradiae

Different dissolved oxygen concentrations and aeration rates were imposed on a stable mutant of Streptomyces fradiae during the antibiotic-producing phase. At high aeration rate (1 vvm), the tylosin yield in the fermentor broth with dissolved oxygen (DO) concentrations controlled close to 100% saturation (6-8 ppm) increased 10% as against uncontrolled. The rates of cellular growth, oil consumption, and tylosin production were severely reduced when DO concentration fell below 25% saturation, but all resumed to their initial rates when DO was raised to saturation level again. The DO concentration in combination with air flow rate affected the pattern of the antibiotics produced. At high DO levels, an additional macrolide antibiotic, macrocin, was synthesized to more than one-third the amount of tylosin at high aeration rate (1 vvm). On the other hand, tylosin production rate remained constant and no significant amount of macrocin was produced at low aeration rate (0.2 vvm).     

多鳞四须鲃耗氧率、排氨率和窒息点的初步研究

控制水温在(20±0.5)℃条件下,采用Winkler法和奈氏试剂法分别测定水中的溶氧含量和氨氮含量,通过比较流水呼吸室进、出口水中的溶解氧和氨氮含量之差以确定多鳞四须鲃(Barbodes schwanenfeldi)的耗氧率、排氨率及窒息点.结果表明,多鳞四须鲃耗氧率随体重增加而减小,关系式为Y=0.24X^-0.09(R²=0.9028);随放养密度的增大而减小,关系式为Y=-0.029X+0.3301 (R²=0.9291).排氨率随体重的增加而减小,关系式为Y=-0.0008X+0.2433 (R²=0.9817);随放养密度的增大而增加,关系式为Y=-0.050X+0.4979 (R²=0.9889).多鳞四须隹巴晚间(18:00～4:00)的耗氧率明显高于白天(6:00-1:00),排氨率则相反,白天的排氨率相对较高,表明多鳞四须鲃巴属于"昼伏夜出"型鱼类.多鳞四须啬巴的窒息点为1.2572mg·L^-1,耐氧性较差.     

Effect of the applied organic load rate on biodegradable polymer production by mixed microbial cultures in a sequencing batch reactor

This article studies the operation of a new process for the production of biopolymers (polyhydroxyalkanoates, PHAs) at different applied organic load rates (OLRs). The process is based on the aerobic enrichment of activated sludge to obtain mixed cultures able to store PHAs at high rates and yields. A mixture of acetic, lactic, and propionic acids at different concentrations (in the range 8.5-31.25 gCOD/L) was fed every 2 h in a sequencing batch reactor (SBR). The resulting applied OLR was in the range 8.5-31.25 gCOD/L/day. Even though, as expected, the increase in the OLR caused an increase in biomass concentration (up to about 8.7 g COD/L), it also caused a relevant decrease of maximal polymer production rate. This decrease in polymer production rate was related to the different extent of "feast and famine" conditions, as function of the applied OLR and of the start-up conditions. As a consequence the best performance of the process was obtained at an intermediate OLR (20 gCOD/L/day) where both biomass productivity and PHA storage were high enough. However, at this high OLR the process was unstable and sudden decrease of performance was also observed. The sludge characterized by the highest PHA storage response was investigated by 16S rDNA clone library. The clone library contained sequences mostly from PHA producers (e.g., Alcaligenes and Comamonas genera); however many genera and among them, one of the dominant (Thauera), were never described before in relation to PHA storage response.     

光生物反应器脱除空气中CO2的模型研究

微藻光生物反应器具有脱除空气中CO_2能力。从光生物反应器构型、进气流速、混合传质,及微藻光合/呼吸速率等方面,探讨气升式光生物反应器脱除空气中CO_2效果,提出了时间离散化和集中参数法两种分析方法。运用集中参数法建立了气升式柱型光生物反应器脱除CO_2的数学模型,模拟了藻液中溶氧浓度(DO)、pH随时间的变化情况,及进气CO_2浓度影响,预测并验证了光照条件下出气CO_2、O_2浓度的变化趋势。模拟结果和实验数据基本吻合,所提出的模型对光生物反应器的优化设计、微藻的高密度培养,及CO_2去除能力预测具有参考意义。     

Effects of oxygen and nutrients limitation on ajmalicine production and related enzyme activities in high density cultures of Catharanthus roseus

Oxygen and nutrient limitation was investigated in order to identify the origin of a lower specific ajmalicine production in Catharanthus roseus cultures at high cell densities in an induction medium. The effect of oxygen limitation was explored by comparing two identically aerated and agitated high cell density bioreactor cultures with dissolved oxygen (DO) concentration of 15% and 85% of air saturation, with respect to alkaloid formation and related enzymes activities. Oxygen had an evident effect on ajmalicine production: in the high DO cultures production was more than 5 times higher than in the low DO cultures. The difference in ajmalicine production between high and low DO could not be explained by the enzyme activity profiles. Moreover, the productivity in the high density culture could not restored to the level of a low density culture (at a high DO) by increasing the DO alone. The effect of nutrient limitation was studied with response surface methodology in shake flask cultures. Nutrient limitation could not be demonstrated to be responsible for the productivity loss. Alkaloid and enzyme measurements in the shake flask cultures supported previous findings that the tryptamine pathway may regulate alkaloid production, provided that the terpenoid pathway is sufficiently active. (c) 1994 John Wiley & Sons, Inc.     

Aerobic phosphorus release linked to acetate uptake in bio-P sludge: process modeling using oxygen uptake rate

The main processes involved in enhanced biological phosphorus removal (EBPR) under anaerobic and subsequently aerobic conditions are widely described in the literature. Polyphosphate accumulating organisms (PAO) are the organisms responsible for this process. However, the mechanisms of PAO are not fully established yet under conditions that differ from the classical anaerobic/aerobic conditions. In this work, we made a comparison between the behavior of PAO under classical EBPR conditions and its behavior when consuming substrate under only aerobic conditions. In addition, oxygen uptake rate (OUR) was measured in the set of experiments under aerobic conditions to improve the characterization of the process. A kinetic and stoichiometric model based on Activated Sludge Model No.2 (ASM2) and including glycogen economy (AnOx model), calibrated for classical anaerobic/aerobic conditions, was not able to describe the experimental data since it underestimated the acetate consumption, the PHB storage, and the OUR. Two different hypotheses for describing the experimental measurements were proposed and modeled. Both hypotheses considered that PAO, under aerobic conditions, uptake acetate coupled to PHB storage, glycogen degradation, and phosphorus release as in anaerobic conditions. Moreover, the first hypothesis (PAO-hypothesis) considered that PAO were able to store acetate as PHB linked to oxygen consumption and the second one (OHO hypothesis) considered that this storage was due to ordinary heterotrophic organisms (OHO). Both hypotheses were evaluated by simulation extending the AnOx model with additional equations. The main differences observed were the predictions for PHB degradation during the famine phase and the OUR profile during both feast and famine phases. The OHO hypothesis described the experimental profiles more accurately than the PAO hypothesis.     

溶氧对Candida glycerinogenes产甘油发酵过程的影响

研究了溶氧浓度对产甘油假丝酵母分批发酵生产甘油过程的影响。实验结果表明：当溶氧浓度控制在30%时,C. glycerinogenes的甘油产量、得率和产率达到最高,分别为120.7 g/L、0.575 g/g和1.69 g/(L•h),而糖酵解代谢副产物形成最少。当溶氧浓度为10%时,发酵过程呈现出“巴斯德效应”的特征,生成的酵解代谢副产物维持在较高水平。在快速生长阶段,随着溶氧从10%增加到60%,细胞呼吸类型表现为从厌氧呼吸向好氧呼吸转变,酵解代谢副产物依次减少。在生长稳定期,控制的溶氧浓度越高,酵解代谢副产物乙醇、乙酸等的生成减少。分别选用Logistic方程、Luedeking-Piret方程和Luedeking-Piret-like方程,能较好地模拟细胞生长、甘油合成和葡萄糖消耗的动力学过程。     

氧对膜生物反应器短程硝化的影响

为了研究膜生物反应器的短程硝化性能以及氧对短程硝化的影响,通过对比耗氧率和供氧率,提出了膜生物反应器短程硝化的控制优化建议。在膜生物反应器硝化过程中,DO小于1 mg/L开始出现亚硝氮积累;DO降到0.5 mg/L,出水氨氮浓度与亚硝氮浓度之比接近1∶1;DO调控在0.5-1 mg/L范围内,有利于前置硝化反应器与后续厌氧氨氧化反应器衔接。膜生物反应器中污泥浓度可达20 g/L,耗氧能力可达19.86 mg O2/(L·s),但最大供氧能力仅为0.369 mg O2/(L·s),供氧成为反应器运行的制约瓶颈,"低DO高流量"曝气是继续提高短程硝化效能的控制策略。     

Control of recombinant human endostatin production in fed-batch cultures of Pichia pastoris using the methanol feeding rate

Endostatin is a 20 kDa carboxyl-terminal fragment of collagen XVIII that strongly inhibits angiogenesis and tumor growth. The methylotrophic yeast, Pichia pastoris, is a robust expression system that can be used to study methods to improve the yields of rhEndostatin. We expressed rhEndostatin in P. pastoris under the control of the alcohol oxidase 1 (aox 1) promoter (Mut⁺ phenotype) as a model, and used a cell biomass of about 50 g l^–1 dry cell wt as a starting point for the induction phase and varied the methanol feed rate at 8 ml l^–1 h^–1, 11 ml l^–1 h^–1 and 15 ml l^–1 h^–1. While the cell growth rate was proportional to the rate of methanol delivery, protein production rate was not. These findings could be used to guide parameters for large-scale production of recombinant proteins in the P. pastoris system.     

The effect of dissolved oxygen on PHB accumulation in activated sludge cultures

Nitrogen removal from wastewater is often limited by the availability of reducing power to perform denitrification, especially when treating wastewaters with a low carbon:nitrogen ratio. In the increasingly popular sequencing batch reactor (SBR), bacteria have the opportunity to preserve reducing power from incoming chemical oxygen demand (COD) as poly-beta-hydroxybutyrate (PHB). The current study uses laboratory experiments and mathematical modeling in an attempt to generate a better understanding of the effect of oxygen on microbial conversion of COD into PHB. Results from a laboratory SBR with acetate as the organic carbon source showed that the aerobic acetate uptake process was oxygen-dependent, producing higher uptake rates at higher dissolved oxygen (DO) supply rates. However, at the lower DO supply rates (k(L)a 6 to 16 h(-1), 0 mg L(-1) DO), a higher proportion of the substrate was preserved as PHB than at higher DO supply rates (k(L)a 30, 51 h(-1), DO >0.9 mg L(-1)). Up to 77% of the reducing equivalents available from acetate were converted to PHB under oxygen limitation (Y(PHB/Ac) 0.68 Cmol/Cmol), as opposed to only 54% under oxygen-excess conditions (Y(PHB/Ac) 0.48 Cmol/Cmol), where a higher fraction of acetate was used for biomass growth. It was calculated that, by oxygen management during the feast phase, the amount of PHB preserved (1.4 Cmmol L(-1) PHB) accounted for an additional denitrification potential of up to 18 mg L(-1) nitrate-nitrogen. The trends of the effect of oxygen (and hence ATP availability) on PHB accumulation could be reproduced by the simulation model, which was based on biochemical stoichiometry and maximum rates obtained from experiments. Simulated data showed that, at low DO concentrations, the limited availability of adenosine triphosphate (ATP) prevented significant biomass growth and most ATP was used for acetate transport into the cell. In contrast, high DO supply rates provided surplus ATP and hence higher growth rates, resulting in decreased PHB yields. The results suggest that oxygen management is crucial to conserving reducing power during the feast phase of SBR operation, as excessive aeration rates decrease the PHB yield and allow higher biomass growth.     

温度和个体大小对单齿螺耗氧率和排氨率的影响

采用静水法测定了不同温度、不同个体大小的单齿螺耗氧率和排氨率。结果表明:在16-33℃的实验温度范围内单齿螺的耗氧率(R_O)和排氨率(R_N)与软体部干重(W)都呈负相关,它们之间关系可以分别用幂函数R_O=aW^-b和R_N=a₁W^-b1表示。16～29℃温度范围内单齿螺的耗氧率和排氨率均随温度的升高而增加,29℃时耗氧率和排氨率达到最大值,当温度继续升高超过29℃后,耗氧率和排氨率则随温度的升高而下降,耗氧率、排氨率与温度之间呈显著的指数函数关系R_O=ce^dT和R_N=c₁e^d1T;不同个体大小单齿螺的O:N比在16～20℃时较大,Q₁₀取值范围0.56-3.74,平均值为1.64。方差分析表明,温度、软体部干重对单齿螺的耗氧率和排氨率均有极显著的影响(P<0.01)。     

Continuous, real-time monitoring of the oxygen uptake rate (OUR) in animal cell bioreactors

A new method for real-time monitoring of the oxygen uptake rate (OUR) in bioreactors, based on dissolved oxygen (DO) measurement at two points, has been developed and tested extensively. The method has several distinct advantages over known techniques.It enables the continuous and undisturbed monitoring of OUR, which is conventionally impossible without gas analyzers. The technique does not require knowledge of k(L)a. It provides smooth, robust, and reliable signal. The monitoring scheme is applicable to both microbial and mammalian cell bioprocesses of laboratory or industrial scale. The method was successfully used in the cultivation of NSO-derived murine myeloma cell line producing monoclonal antibody. It was found that while the OUR increased with the cell density, the specific OUR decreased to approximately one-half at cell concentrations of 16 x 10(6) cells/mL, indicating gradual reduction of cell respiration activity. Apart from the laboratory scale cultivation, the method was applied to industrial scale perfusion culture, as well as to processes using other cell lines. (c) 1994 John Wiley & Sons, Inc.     

The effect of dissolved oxygen tension and the utility of oxygen uptake rate in insect cell culture

Dissolved oxygen tension and oxygen uptake rate are critical parameters in animal cell culture. However, only scarce information of such variables is available for insect cell culture. In this work, the effect of dissolved oxygen tension (DOT) and the utility of on-line oxygen uptake rate (OUR) measurements in monitoring Spodoptera frugiperda (Sf9) cultures were determined. Sf9 cells were grown at constant dissolved oxygen tensions in the range of 0 to 30%. Sf9 metabolism was affected only at DOT below 10%, as no significant differences on specific growth rate, cell concentration, amino acid consumption/production nor carbohydrates consumption rates were found at DOT between 10 and 30%. The specific growth rate and specific oxygen uptake rate followed typical Monod kinetics with respect to DOT. The calculated _max and max were 0.033 h^-1 and 3.82×10^-10 mole cell^-1h^-1, respectively, and the corresponding saturation constants were 1.91 and 1.57%, respectively. In all aerated cultures, lactate was consumed only after glucose and fructose had been exhausted. The yield of lactate increased with decreasing DOT. It is proposed, that an apparent DOT in non-instrumented cultures can be inferred from the lactate yield of bioreactors as a function of DOT. Such a concept, can be a useful and important tool for determining the average dissolved oxygen tension in non-instrumented cultures. It was shown that the dynamic behavior of OUR can be correlated with monosaccharide (fructose and glucose) depletion and viable cell concentration. Accordingly, OUR can have two important applications in insect cell culture: for on-line estimation of viable cells, and as a possible feed-back control variable in automatic strategies of nutrient addition.Abbreviations DOT Dissolved oxygen tension - OUR Oxygen uptake rate - specific oxygen uptake rate - specific growth rate - X_v viable cell concentration - C_L, C^*, and oxygen concentrations in liquid phase, in equilibrium with gas phase, and medium molar concentration, respectively - H Henry's constant - K_La volumetric oxygen transfer coefficient - P_T total pressure - oxygen partial pressure - oxygen molar fraction - i discrete element