植物生物反应器研究进展

植物生物反应器是近年来生物技术领域新的研究方向,利用农作物进行疫苗、药用蛋白的生产,具有广阔的市场前景和商业价值。研究证明,用各种农作物为载体的植物生物反应器产品可通过种子、果实或块茎表达,便于贮藏、运输和利用。它拓宽了传统农业概念,成为现代生物农业重要的研究方向之一,推动了生物经济快速健康的前进,促进农业可持续发展。综述植物生物反应器的研究与应用现状,并对转基因作物作为植物生物反应器的发展前景作分析和展望。     

动物细胞培养用生物反应器设计原理

动物细胞培养用生物反应器设计和放大的关键问题是细胞破损与供氧和混合的矛盾,在分析细胞破损机理基础上,提出了动物细胞培养生物反应器的设计原理——设计模型和有关设计条件,从而清楚地确立了细胞死亡速度与培养基组成、反应器设计和操作参数间的定量关系,以及反应器设计应遵循的保证细胞生长和满足传质要求的条件。还对强化传质和抑制细胞破损这一矛盾作了简要分析和讨论。     

封闭式光生物反应器研究进展

国际上80～90年代,封闭式光生物反应器是微藻生物技术的重要研究热点,也是微藻生物技术产业化的关键技术之一。本文较全面地介绍了用于微藻大规模培养的封闭式光生物反应器研究现状。将封闭式光生物反应器分为柱式、管式、板式和光导纤维反应器等类型。工业放大前景的管式和板式光生物反应器采取了典型个案分析的方法,列表比较了典型反应器的主要技术参数,并对它们的技术发展趋势进行了归纳总结。     

新型生物反应器结构研究进展

生物反应器是生物工程的核心设备,其结构的合理性直接决定反应器生物加工的效率。生物反应器的研究一直是生物工程的核心问题之一。随着青霉素的工业化生产,机械搅拌式生物反应器应运而生,此后,随着动植物细胞培养,高等真菌培养,藻类培养等生物过程的发展,人们相应开发了大量的生物反应器,其中以机械搅拌式生物反应器和气升式样生物反应器尤为突出,本文总结了近年来文献报道的新型生物反应器,主要阐述了机械搅拌式和气升式两类生物反应器结构的研究进展,对目前国内外报道的11种新型反应器典型结构进行了总结与分析。     

植物生物反应器研究现状、瓶颈及策略

近10年,植物作为重组蛋白生产系统是生命科学中研究最活跃领域之一。植物系统具有低成本、安全和易规模化优势,其表达生物活性药用蛋白能力已被许多研究所证实;同时,植物药用蛋白产品还表现出潜在的市场和广阔应用前景。鉴于此,回顾了植物生物反应器兴起,介绍了植物表达系统和重组蛋白研究现状,综述了植物生物反应器面临瓶颈问题、解决对策和未来一段时间内研究热点;在展望植物生物反应器前景同时,对我国研究现状、与国外差距和未来发展应采取策略进行了讨论。     

植物细胞培养生物反应器的种类特点及展望

生物反应器技术应用于植物细胞培养既可以打破环境条件的限制,又有助于生产过程的人为调控,为植物细胞大规模培养或工厂化直接生产植物细胞有用代谢产物创造了条件,是当前植物细胞培养工作的研究热点。在介绍植物细胞培养特点的基础上,对适用于植物细胞培养的各类生物反应器(搅拌式生物反应器、非搅拌式生物反应器、用于植物细胞固定化培养的生物反应器、光生物反应器以及一次性培养生物反应器)的原理、优缺点等进行比较分析,最后提出了植物细胞培养生物反应器研究的发展方向,以期为植物细胞培养生物反应器的选择及改良提供参考。     

中国生物工程学会将举办全国生物反应器学术研讨会

生物反应器是多学科交叉的生物技术领域,是21世纪生物工程发展的重要前沿之一。为总结交流近年来生物反应器研究领域在基础、应用基础和应用研究方面的进展与成果,促进农业、医学和工业生物技术的交叉与渗透,推动产、学、研协作,加快东北老工业基地传统产业的技术升级和高科技产业的发展,中国生物工程学会农业生物技术专业委员会、医学生物技术专业委员会、产业促进会联合吉林农业大学共同发起召开“全国生物反应器学术研讨会”。     

我国生物信息产业的现状及问题分析

针对我国生物信息产业的现状及存在的问题进行分析,介绍了生物信息学以及生物芯片研究的现状和新技术、生物信息产业的发展,并对生物信息产业的知识产权保护问题进行了分析和讨论。对于今后如何发展我国生物信息产业以及如何采取策略和措施提供参考。     

利用动物乳腺生物反应器生产药用蛋白

动物乳腺生物反应器是利用动物乳腺特异性启动子调控元件指导外源基因在乳腺中特异性表达,并从转基因动物奶液中获取重组蛋白。应用动物乳腺生物反应器生产药用蛋白具有生产方式简单,产量大,蛋白能进行翻译后修饰等优点,是具有广阔前景的生物医药产业。本文仅就动物乳腺生物反应器的建立、检测、目的蛋白的分离纯化以及存在的问题等作一综述。     

高通量植物生物反应器及其在遗传资源挖掘中的应用

生物反应器(bioreactor)是一种以表达目标产物或获得繁殖体为目的的设备系统,包括微生物、动物、植物生物反应器以及相关设备。植物生物反应器(phytobioreactor)是借鉴植物组织培养和微生物发酵原理制作的设备系统。其中,应用较广泛的是间歇浸没式植物生物反应器。与传统植物组织培养相比,该方法具备可换气、无需转接和大容量培养等特点。国内制作的BIOF系列新型植物生物反应器还可以利用串/并联方法,实现更高通量培养能力,其应用于植物种苗繁育、代谢产物的表达、耐盐等变异的定向筛选、植物生长发育的动态分析等方面均具备显著优势。现代植物生物技术在基础研究和产业方面的应用对植物生物反应器提出了新要求,新型生物反应器应用方法的持续改进和设备系统的不断完善,使其成为植物学领域的高效研究平台,并将促进植物育种和植物源化合物的发掘等方面研究效率的提高。     

Packed Bed Bioreactor for the Isolation and Expansion of Placental-Derived Mesenchymal Stromal Cells

Large numbers of Mesenchymal stem/stromal cells (MSCs) are required for clinical relevant doses to treat a number of diseases. To economically manufacture these MSCs, an automated bioreactor system will be required. Herein we describe the development of a scalable closed-system, packed bed bioreactor suitable for large-scale MSCs expansion. The packed bed was formed from fused polystyrene pellets that were air plasma treated to endow them with a surface chemistry similar to traditional tissue culture plastic. The packed bed was encased within a gas permeable shell to decouple the medium nutrient supply and gas exchange. This enabled a significant reduction in medium flow rates, thus reducing shear and even facilitating single pass medium exchange. The system was optimised in a small-scale bioreactor format (160 cm²) with murine-derived green fluorescent protein-expressing MSCs, and then scaled-up to a 2800 cm² format. We demonstrated that placental derived MSCs could be isolated directly within the bioreactor and subsequently expanded. Our results demonstrate that the closed system large-scale packed bed bioreactor is an effective and scalable tool for large-scale isolation and expansion of MSCs.     

用于重组蛋白生产的哺乳动物细胞表达新技术

近年来,用于重组蛋白生产的哺乳动物细胞表达领域涌现出一系列革命性的新技术。优化的工程细胞为表达重组蛋白提供了优良的宿主;基于荧光的筛选方法可以快捷地得到高表达细胞株;高通量的培养工艺能够预测适合外源蛋白表达的细胞培养条件;可抛弃式生物反应器为大规模细胞培养提供了更多的选择;大规模瞬时表达技术节省了重组蛋白的生产时间。这些新技术提高了重组蛋白的研发和生产效率,加快了蛋白药物的工业化进程。     

Manufacture of l-amino acids with bioreactors

Several l-amino acids are currently manufactured by using enzymes or whole-cells contained within bioreactors. Production of l-aspartic acid in Japan is one of the earliest examples of this technology. A bioreactor method recently scaled-up for manufacture of l-phenylalanine from cinnamic acid and ammonia achieves product titers of over 60 g/l and a raw material conversion of about 90%. A bioreactor process in development for making l-serine from glycine and formaldehyde exploits rDNA technology and reaches an l-serine titer of over 400 g/l. The l-serine can be enzymatically converted to L-tryptophan to achieve a bioreactor titer of about 200 g/l.     

Bioreactors for 3-dimensional high-density culture of human cells

A bioreactor was developed as an instrument to culture human or animal cells that require attachment in a large quantity or at a high density. The purpose for developing such a bioreactor is two-fold: to produce a large quantity of animal or human cells that have been modified by gene recombination technology to accommodate manufacture of physiologically-active substances or human proteins on an industrial scale; and for research to culture animal cells to form a high-density 3-dimensional structure as a morphological or functional tissue or organ entity. In the current report, the circulatory flow bioreactor and radial flow bioreactor (RFB) are introduced, in which the former can be scaled up. As a small bioreactor produced for the latter purpose, a rotary cell culture system and novel multicoaxial hollow-fiber bioreactor are introduced. Finally, a small RFB culture system that was scaled down by the present author and his collaborators for the study of a 3-dimensional high density culture system is described. The RFB can be readily scaled up for manufacturing or scaled down for research purposes. This is a cell culturing system that can induce the functions of human tissues by preparing a high density 3-dimensional organization of cells of human origin.     

Design and development of a new ambr250® bioreactor vessel for improved cell and gene therapy applications

The emergence of cell and gene therapies has generated significant interest in their clinical and commercial potential. However, these therapies are prohibitively expensive to manufacture and can require extensive time for development due to our limited process knowledge and understanding. The automated ambr250® stirred-tank bioreactor platform provides an effective platform for high-throughput process development. However, the original dual pitched-blade 20 mm impeller and baffles proved sub-optimal for cell therapy candidates that require suspension of microcarriers (e.g. for the culture of adherent human mesenchymal stem cells) or other particles such as activating Dynabeads® (e.g. for the culture of human T-cells). We demonstrate the development of a new ambr250® stirred-tank bioreactor vessel which has been designed specifically to improve the suspension of microcarriers/beads and thereby improve the culture of such cellular systems. The new design is unbaffled and has a single, larger elephant ear impeller. We undertook a range of engineering and physical characterizations to determine which vessel and impeller configuration would be most suitable for suspension based on the minimum agitation speed (N_JS) and associated specific power input (P/V)_JS. A vessel (diameter, T, = 60 mm) without baffles and incorporating a single elephant ear impeller (diameter 30 mm and 45° pitch-blade angle) was selected as it had the lowest (P/V)_JS and therefore potentially, based on Kolmogorov concepts, was the most flexible system. These experimentally-based conclusions were further validated firstly with computational fluid dynamic (CFD) simulations and secondly experimental studies involving the culture of both T-cells with Dynabeads® and hMSCs on microcarriers. The new ambr250® stirred-tank bioreactor successfully supported the culture of both cell types, with the T-cell culture demonstrating significant improvements compared to the original ambr250® and the hMSC-microcarrier culture gave significantly higher yields compared with spinner flask cultures. The new ambr250® bioreactor vessel design is an effective process development tool for cell and gene therapy candidates and potentially for autologous manufacture too.

     

Successful development of small diameter tissue-engineering vascular vessels by our novel integrally designed pulsatile perfusion-based bioreactor

Small-diameter (<4 mm) vascular constructs are urgently needed for patients requiring replacement of their peripheral vessels. However, successful development of constructs remains a significant challenge. In this study, we successfully developed small-diameter vascular constructs with high patency using our integrally designed computer-controlled bioreactor system. This computer-controlled bioreactor system can confer physiological mechanical stimuli and fluid flow similar to physiological stimuli to the cultured grafts. The medium circulating system optimizes the culture conditions by maintaining fixed concentration of O(2) and CO(2) in the medium flow and constant delivery of nutrients and waste metabolites, as well as eliminates the complicated replacement of culture medium in traditional vascular tissue engineering. Biochemical and mechanical assay of newly developed grafts confirm the feasibility of the bioreactor system for small-diameter vascular engineering. Furthermore, the computer-controlled bioreactor is superior for cultured cell proliferation compared with the traditional non-computer-controlled bioreactor. Specifically, our novel bioreactor system may be a potential alternative for tissue engineering of large-scale small-diameter vascular vessels for clinical use.     

植物组织培养生物反应器技术研究进展

从植物大规模组织培养的特点、反应器类型和反应器中微环境等方面介绍了生物反应器技术在药用植物微繁殖和天然产物细胞生产中的研究进展。     

An assessment of the impact of Raman based glucose feedback control on CHO cell bioreactor process development

Process analytical technology (PAT) tools such as Raman Spectroscopy have become established tools for real time measurement of CHO cell bioreactor process variables and are aligned with the QbD approach to manufacturing. These tools can have a significant impact on process development if adopted early, creating an end-to-end PAT/QbD focused process. This study assessed the impact of Raman based feedback control on early and late phase development bioreactors by using a Raman based PLS model and PAT management system to control glucose in two CHO cell line bioreactor processes. The impact was then compared to bioreactor processes which used manual bolus fed methods for glucose feed delivery. Process improvements were observed in terms of overall bioreactor health, product output and product quality. Raman controlled batches for Cell Line 1 showed a reduction in glycation of 43.4% and 57.9%, respectively. Cell Line 2 batches with Raman based feedback control showed an improved growth profile with higher VCD and viability and a resulting 25% increase in overall product titer with an improved glycation profile. The results presented here demonstrate that Raman spectroscopy can be used in both early and late-stage process development and design for consistent and controlled glucose feed delivery.     

New developments in solid-state fermentation: II. Rational approaches to the design, operation and scale-up of bioreactors

Over the last decade there has been a significant improvement in understanding how to design, operate and scale-up solid-state fermentation bioreactors. The key to these advances has been the application of mathematical modeling techniques to describe the biological and transport phenomena within the system. This review focuses on the advances in understanding that have come from this modeling work, and the insights it has given us into bioreactor design, operation and scale-up. It also highlights two promising bioreactor designs that have emerged over the last decade or so. For processes in which the substrate bed must remain static throughout the fermentation, the most promising design is the Zymotis design of ORSTOM at Montpellier, France, which involves closely spaced internal heat transfer plates within a packed-bed bioreactor. For those processes in which mixing can be tolerated, the stirred bioreactor developed at INRA, in Dijon, France, has been successfully demonstrated at scales of 1–25 t of substrate. Theoretical work suggests that mathematical models will be useful tools in the scale-up process, however, there are no reports that they have been used in the development of any current large-scale process. Rather, the models have been validated against data obtained from laboratory-scale bioreactors. There is an urgent need to test the accuracy and robustness of the models by applying them within real process development.     

Design of a large-scale surface-aerated bioreactor for biomass production using a VOC substrate

The design of a large-scale bioreactor for the production of bacterial biomass adapted to the biodegradation of volatile organic compounds was carried out. The bioreactor model used integrated the microbial kinetics and fluid dynamics described by the compartment model approach. The process conditions and kinetic parameters were adopted from the laboratory experimental study of (León, E., Seignez, C., Adler, N., Péringer, P., 1999. Growth inhibition of biomass adapted to the degradation of toluene and xylenes in mixture in a batch reactor with substrates supplied by pulses. Biodegradation 10, 245-250). The performance of the pulsed-batch stirred bioreactor under surface aeration conditions was simulated for different mixing configurations and conditions such as the impeller diameter, number of impellers, stirring speed, and oxygen pressure. The simulations were used for the cost analysis which resulted in the optimal design of the bioreactor.