The morphology and life history of Mazoyerella gen. nov. (M. arachnoidea (Harvey) comb. nov.)—Rhodophyta,Ceramiaceae—from Southern Australia

Mazoyerella gen. nov., based on Corynospora arachnoidea Harvey from Tasmania, is placed provisionally in the Compsothamnieae tribe of the Ceramiaceae. All plants can produce elongate-ovoid monosporangia which reproduce the same phase. Sporophytes also bear polysporangia and sexual plants are dioecious, bearing either spermatangial heads or subterminal procarps followed by carposporophytes with nearly all gonimoblast cells becoming carposporangia and an involucrum of sterile filaments developing from sterile cells associated with the procarp. The life-history has been followed through successive generations in culture.     

Observations on Chrysophyceae from a Norwegian mountain lake

Chrysosphaerella coronasircumspina Wujek & Kristiansen and Spiniferomonas bilacunosa Takahashi have been shown by electron microscopy to be regular members of the phytoplankton society in an oligotrophic Norwegian mountain lake. This is the first published record of these species in Norway.     

Continuous Production of l-Alanine with NADH Regeneration by a Nanofiltration Membrane Reactor

A conjugated enzyme system, alanine dehydrogenase (AIDH) for stereospecific reduction of pyruvate to l-alanine and glucose dehydrogenase (GDH) for regeneration of NADH, were coimmobilized in a nanofiltration membrane bioreactor (NFMBR) for the continuous production of l-alanine from pyruvate with NADH regeneration. Since pyruvate was proved to be unstable at neutral pH, it was kept under acidic conditions and supplied to NFMBR separately from the other substrates. As 0.2 m pyruvate in HCl solution (pH 4), 10 mm NAD, 0.2 m glucose, and 0.2 m NH₄Cl in 0.5 m Tris buffer (pH 8) were continuously supplied to NFMBR with immobilized AIDH (100 U/ml) and GDH (140 U/ml) at the retention time of 80 min, the maximum conversion, reactor productivity, and NAD regeneration number were 100%, 320 g/liter/d, and 20,000, respectively. To avoid the effect of pyruvate instability, a consecutive reaction system, lactate dehydrogenase (l-LDH) and AIDH, was also used. In this system, the l-LDH provides pyruvate, the substrate for the AIDH reaction, from l-lactate regenerating NADH simultaneously, so the pyruvate could be consumed as soon as it was produced. As 0.2 m l-lactate, 10 mm NAD, 0.2 m NH₄Cl in 0.5 m Tris buffer (pH 8) were continuously supplied to NFMBR with immobilized l-LDH (100 U/ml) and AIDH (100 U/ml) at the retention time of 160 min, the maximum conversion, reactor productivity, and the NAD regeneration number were 100%, 160 g/Iiter/d, and 20,000, respectively.     

Characterization and immobilization on nickel-chelated Sepharose of a glutamate decarboxylase A from Lactobacillus brevis BH2 and its application for production of GABA

A gene encoding glutamate decarboxylase A (GadA) from Lactobacillus brevis BH2 was expressed in a His-tagged form in Escherichia coli cells, and recombinant protein exists as a homodimer consisting of identical subunits of 53?kDa. GadA was absolutely dependent on the ammonium sulfate concentration for catalytic activity and secondary structure formation. GadA was immobilized on the metal affinity resin with an immobilization yield of 95.8%. The pH optima of the immobilized enzyme were identical with those of the free enzyme. However, the optimum temperature for immobilized enzyme was 5?°C higher than that for the free enzyme. The immobilized GadA retained its relative activity of 41% after 30 reuses of reaction within 30?days and exhibited a half-life of 19 cycles within 19?days. A packed-bed bioreactor with immobilized GadA showed a maximum yield of 97.8% GABA from 50?mM l-glutamate in a flow-through system under conditions of pH 4.0 and 55?°C.     

Very high density of CHO cells in perfusion by ATF or TFF in WAVE bioreactor™. Part I. Effect of the cell density on the process

High cell density perfusion process of antibody producing CHO cells was developed in disposable WAVE Bioreactor? using external hollow fiber filter as cell separation device. Both “classical” tangential flow filtration (TFF) and alternating tangential flow system (ATF) equipment were used and compared. Consistency of both TFF‐ and ATF‐based cultures was shown at 20–35 × 10⁶ cells/mL density stabilized by cell bleeds. To minimize the nutrients deprivation and by‐product accumulation, a perfusion rate correlated to the cell density was applied. The cells were maintained by cell bleeds at density 0.9–1.3 × 10⁸ cells/mL in growing state and at high viability for more than 2 weeks. Finally, with the present settings, maximal cell densities of 2.14 × 10⁸ cells/mL, achieved for the first time in a wave‐induced bioreactor, and 1.32 × 10⁸ cells/mL were reached using TFF and ATF systems, respectively. Using TFF, the cell density was limited by the membrane capacity for the encountered high viscosity and by the pCO₂ level. Using ATF, the cell density was limited by the vacuum capacity failing to pull the highly viscous fluid. Thus, the TFF system allowed reaching higher cell densities. The TFF inlet pressure was highly correlated to the viscosity leading to the development of a model of this pressure, which is a useful tool for hollow fiber design of TFF and ATF. At very high cell density, the viscosity introduced physical limitations. This led us to recommend cell densities under 1.46 × 10⁸ cell/mL based on the analysis of the theoretical distance between the cells for the present cell line. © 2013 American Institute of Chemical Engineers Biotechnol. Prog., 29:754–767, 2013     

Three‐dimensional neural differentiation of embryonic stem cells with ACM induction in microfibrous matrices in bioreactors

The clinical use of pluripotent stem cell (PSC)‐derived neural cells requires an efficient differentiation process for mass production in a bioreactor. Toward this goal, neural differentiation of murine embryonic stem cells (ESCs) in three‐dimensional (3D) polyethylene terephthalate microfibrous matrices was investigated in this study. To streamline the process and provide a platform for process integration, the neural differentiation of ESCs was induced with astrocyte‐conditioned medium without the formation of embryoid bodies, starting from undifferentiated ESC aggregates expanded in a suspension bioreactor. The 3D neural differentiation was able to generate a complex neural network in the matrices. When compared to 2D differentiation, 3D differentiation in microfibrous matrices resulted in a higher percentage of nestin‐positive cells (68% vs. 54%) and upregulated gene expressions of nestin, Nurr1, and tyrosine hydroxylase. High purity of neural differentiation in 3D microfibrous matrix was also demonstrated in a spinner bioreactor with 74% nestin + cells. This study demonstrated the feasibility of a scalable process based on 3D differentiation in microfibrous matrices for the production of ESC‐derived neural cells. © 2013 American Institute of Chemical Engineers Biotechnol. Prog., 29:1013–1022, 2013     

Multispectral image analysis for algal biomass quantification

This article reports a novel multispectral image processing technique for rapid, noninvasive quantification of biomass concentration in attached and suspended algae cultures. Monitoring the biomass concentration is critical for efficient production of biofuel feedstocks, food supplements, and bioactive chemicals. Particularly, noninvasive and rapid detection techniques can significantly aid in providing delay‐free process control feedback in large‐scale cultivation platforms. In this technique, three‐band spectral images of Anabaena variabilis cultures were acquired and separated into their red, green, and blue components. A correlation between the magnitude of the green component and the areal biomass concentration was generated. The correlation predicted the biomass concentrations of independently prepared attached and suspended cultures with errors of 7 and 15%, respectively, and the effect of varying lighting conditions and background color were investigated. This method can provide necessary feedback for dilution and harvesting strategies to maximize photosynthetic conversion efficiency in large‐scale operation. © 2013 American Institute of Chemical Engineers Biotechnol. Prog., 29:808–816, 2013     

核桃壳提取液用于单针藻Monoraphidium sp.QLZ-3培养产油脂

[目的] 研究核桃壳提取液（walnut shell extracts,WSE）对单针藻Monoraphidium sp.QLZ-3生长和油脂积累的影响。[方法] 向BG-11培养基中添加不同量的WSE（培养基中保留有BG-11中全部营养成分）。[结果] 结果显示,当BG-11培养基中的WSE含量为40%时,单针藻的生物量产率及油脂产率达到（534.70±4.07）mg/（L·d）和（296.35±15.36）mg/（L·d）,相比对照组分别提高了的14.82%和33.50%,蛋白质和碳水化合物含量分别有不同程度的上调和下调。与对照组相比,微藻中谷胱甘肽（glutathione,GSH）和超氧化物歧化酶（superoxide dismutase,SOD）含量与活性均上调。此外,WSE作用下,微藻对多酚的移除达到84.37%,同时上调了核酮糖1,5-二磷酸羧化酶基因（ribulose 1,5-bisphosphate carboxylase/oxygenase,rbcL）和乙酰辅酶A羧化酶（acetyl coenzyme A carboxylase,accD）基因的表达量。[结论] 研究表明,WSE联合BG-11可以提高微藻的生物量产率和油脂产率,降低微藻培养的原料成本,为核桃壳的资源化利用及微藻的工业化生产提供了一定的技术支撑。     

GAP promoter‐based fed‐batch production of highly bioactive core streptavidin by Pichia pastoris

Streptavidin is a homotetrameric protein binding the vitamin biotin and peptide analogues with an extremely high affinity, which leads to a large variety of applications. The biotin‐auxotrophic yeast Pichia pastoris has recently been identified as a suitable host for the expression of the streptavidin gene, allowing both high product concentrations and productivities. However, so far only methanol‐based expression systems have been applied, bringing about increased oxygen demand, strong heat evolution and high requirements for process safety, causing increased cost. Moreover, common methanol‐based processes lead to large proportions of biotin‐blocked binding sites of streptavidin due to biotin‐supplemented media. Targeting these problems, this paper provides strategies for the methanol‐free production of highly bioactive core streptavidin by P. pastoris under control of the constitutive GAP promoter. Complex were superior to synthetic production media regarding the proportion of biotin‐blocked streptavidin. The optimized, easily scalable fed‐batch process led to a tetrameric product concentration of up to 4.16 ± 0.11 µM of biotin‐free streptavidin and a productivity of 57.8 nM h^?1 based on constant glucose feeding and a successive shift of temperature and pH throughout the cultivation, surpassing the concentration in un‐optimized conditions by a factor of 3.4. Parameter estimation indicates that the optimized conditions caused a strongly increased accumulation of product at diminishing specific growth rates (μ ≈ D < 0.01 h^?1), supporting the strategy of feeding. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:855–864, 2016     

Disposable orbitally shaken TubeSpin bioreactor 600 for Sf9 cell cultivation in suspension

Disposable orbitally shaken TubeSpin bioreactor 600 tubes (TS600s) were recently developed for the bench-scale cultivation of animal cells in suspension. Here we compared batch cultures of Sf9 insect cells in TS600s, spinner flasks, and shake flasks. Superior cell growth was observed in TS600s and shake flasks as compared with spinner flasks, and more favorable oxygen-enriched cell culture conditions were observed in TS600s as compared with either spinner or shake flasks. The results demonstrated the suitability of TS600s as a disposable vessel for the cultivation of Sf9 cells in suspension.