Cell/tissue culture bioreactors

For large-scale culture, inoculum size is one of the most significant factors affecting plant cell/tissue culture systems. Figure 8 shows the time course of the change in growth rate as inoculum size in bioreactor cultures. 

Light is an important regulatory factor in the production of alkaloids in plant cell cultures. The light is known not only to affect the alkaloid production but also the accumulation site in cell culture of C. roseus. Through various studies, it is known that in C. roseus cell/tissue cultures, light influences the ajmalicine/serpentine accumulation ratio and also vindoline and catharanthine production [38]. However, in large-scale production in stirred-tank bioreactors, such conditions are difficult and costly to realize. Therefore, production conditions on small scale should preferably be optimized with dark-grown cultures. 

In this review, we focus on the use of plant tissue culture to produce foreign proteins that have direct commercial or medical applications. The development of large-scale plant tissue culture systems for the production of biopharmaceutical proteins requires efficient, high-level expression of stable, biologically active products. To minimize the cost of protein recovery and purification, it is preferable that the expression system releases the product in a form that can be harvested from the culture medium. In addition, the relevant bioprocessing issues associated with bioreactor culture of plant cells and tissues must be addressed. 

The development of novel bioreactors and bioreactor operating strategies has the potential to improve the performance of cell culture systems. Some progress has been made in this area to enhance foreign protein production in plant tissue culture. 

Mammalian cell suspension cultures are the preferred choice for large-scale recombinant protein production in stirred-tank bioreactors. The most widely used systems are Chinese hamster ovary (CHO) cells and the murine myeloma fines NSO and SP2/0. In half of the biological license approvals from 1996-2000, CHO cells were used for the production of monoclonal antibodies and other recombinant glycosylated proteins, including tPA (tissue plasminogen activator) and an IgGl fusion with the tumor necrosis factor (TNF) receptor, the latter marketed as Enbrel [7]. 

Flower [25], Panda et al. [26], Doran [27]. and Payne et al. [28]. Several kinds of bioreactors, such as the stirred tank bioreactor with hollow paddle and flat blade impellers, the bubble column, the airlift bioreactor with internal and external loops, the rotating drum bioreactor, the stirred-tank with a draft tube, and the mist bioreactor have been attempted for plant cell, tissue and organ cultures (Fig. 1). 

In fermentation reactors, cell growth is promoted or maintained to produce metabolite, biomass, transformed substrate, or purified solvent. Systems based on macro-organism cultures are usually referred as tissue cultures. Those based on dispersed non-tissue forming cultures of micro-organisms are loosely referred as microbial reactors. In enzyme reactors, substrate transformation is promoted without the life-support system of whole cells. Frequently, these reactors employ immobilized enzymes, where an enzyme is supported on inert solids so that it can be reused in the process. Virtually all bioreactors of technological importance deal with a heterogeneous system involving more than two phases. 

BIOREACTORS FOR PLANT CELL TISSUE AND ORGAN CULTURES (by Shinsaku Takayama)... 

Air Driven Bioreactors. The simplest design is the air-driven bioreactor equipped with sparger at the bottom of the vessel. It is widely used for plant cell, tissue, and organ cultures. In cases where the cells grow rapidly and the cell mass occupies 40-60% of the reactor volume, the flow characteristics become non-Newtonian and the culture medium can no longer be agitated by simple aeration. 

Figure 20. Ninety-five liter automated bioreactor for plant cell, tissue and organ cultures. (Photo courtesy of K. F. Engineering Co., Ltd., Tokyo).

Gaseous Phase Bioreactor. As shown in Fig. 24, this type of bioreactor is equipped with filters on which the culture is supported and with a shower nozzle for spraying on the medium (Ushiyama et al., 1984 ti l Ushiyama, 1988).1 1 Seed cultures are inoculated on the filters and the medium is supplied to the culture by spraying from a shower nozzle. The drained medium is collected on the bottom of the bioreactor. This type of bioreactor is excellent for plant cell, tissue, and organ cultures because there is no mechanical agitation (e.g., driven impeller, aerator) and, therefore, the growth rate and the secondary metabolite production are enhanced. 

Yesil-Celiktas et al. (69) reviewed the literature concerning large-scale cultivation of plant cell and tissue culture in bioreactors, and Ziv (70) reviewed bioreactor technology for the micropropagation of plants. 

Yesil-Celiktas, O., Gurel, A., and Yardar-Sukan, F., Large Scale Cultivation of Plant Cell and Tissue Culture in Bioreactors, p. 3, Transworld Research Network, Kerala, India, 2010. 

Su, W. W, Bioreactor Engineering for Recombinant Protein Production Using Plant Cell Suspension Culture, in Dutta Gupta, S., and Ibaraki, Y. (Eds.), Plant Tissue Culture Engineering, pp. 135-159, Springer-Verlag, Beilin, 2006. 

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