Journal of Biotechnology 59 (1997) 39 – 52


Plant cell suspension cultures: some engineering considerations
P.M. Kieran a,*, P.F. MacLoughlin b, D.M. Malone b

Biochemical Engineering Research Group, School of Biological Sciences, Dublin City Uni6ersity, Dublin 9, Ireland b Department of Chemical Engineering, Uni6ersity College Dublin, Belfield, Dublin 4, Ireland Received 1 October 1996; received in revised form 3 February 1997; accepted 25 July 1997

Abstract Higher plants are the source of a vast array of biochemicals which are used as drugs, pesticides, flavourings and fragrances. For some of these compounds, plant cell culture can provide a potential production alternative to traditional cultivation methods or chemical synthesis routes. Many systems have been patented and the last 20 years have seen considerable industrial and academic interest in the development of large scale cultures to produce pharmaceutically active, high value substances. However, the industrial application of plant cell suspension cultures has, to date, been limited. Commercialisation has essentially been impeded by economic feasibility, arising from both biological and engineering considerations. This paper reviews the commercial development of the technology to date and focuses on the impact of specific engineering-related factors, in particular, the shear sensitivity of plant cell suspension cultures. Evidence of sensitivity to hydrodynamic shear in bioreactors has generally been attributed to the physical characteristics of the suspended cells. Recent studies indicate that shear sensitivity may not be as important, in some cases, as initially anticipated. © 1997 Elsevier Science B.V. Keywords: Plant cells; Shear sensitivity; Bioreactor; Scale-up

1. Introduction Higher plants are recognised as important sources of a wide range of biochemicals, used as drugs, pesticides, flavourings and fragrances. Tra* Corresponding author. Tel.: + 353 1 7045584; fax: + 353 1 7045412; e-mail:

ditionally, these substances have been extracted from naturally grown whole plants. On a commercial basis, this approach involves large-scale crop cultivation (e.g. alkaloids from Catharanthus roseus). Many plant products can now be produced by chemical synthesis, which can be a more reliable, consistent and cost-effective method. Plant cell culture provides an alternative ap-

0168-1656/97/$17.00 © 1997 Elsevier Science B.V. All rights reserved. PII S 0 1 6 8 - 1 6 5 6 ( 9 7 ) 0 0 1 6 3 - 6

1992.. Scragg.40 P. workers at Shiseido have alternatively employed C. 1994). 1995. In Japan. Dornenburg and ¨ Knorr. industrial processes involving plant cell cultures have been limited to a handful of applications. few have proven to be economically viable. Plant cell culture may yet provide another production route (Fett-Neto and DiCosmo. 1996. 1991. in some instances. Verpoorte et al. And large-scale processes for the use of cell cultures as a source of biomass (Hashimoto and Azechi. If the biosynthetic pathway for Taxol® can be fully elucidated. To obtain 1 kg of Taxol® requires the bark of more than 1000 trees. using this technology. As a source of enzymes for the genetically engineered synthesis of natural products (Scott. 1987) and ginsenosides from Panax ginseng (Ushiyama. The anti-cancer drug Taxol® (a registered trademark of Bristol-Myers Squibb) is a very important example of a potential candidate for production by cell culture methods. which may be attractive under certain circumstances: if. Nicolaou et al. 1993. 1992) offer genetic stability as well as. 1993.. cell suspension culture has more immediate potential for industrial application than plant tissue or organ cultures. Total synthesis of Taxol® has recently been reported (Holton et al. they offer great promise. Metabolite yield by the cell culture may significantly exceed that observed in the parent plant. Zhong et al. Thus. Despite its enormous potential. 1995. each up to 100 years old. 1995). ¨ 1992). Plant cell cultures have predominantly been valued as a source of naturally occurring secondary metabolites. DiCosmo and Misawa. Seki et al. the metabolite can be produced under controlled and reproducible conditions. due to the extensive body of expertise which has been amassed for the treatment of submerged microbial cultures. to date. 1997).. Many other processes have been investigated and patented but. Kreis. has a long cultivation period or has a low metabolite yield. which have hampered its exploitation on an industrial scale. the development of appropriate reactors and processing techniques for these systems will involve enormous de novo investment. Su. berberine from Coptis japonica (Fujita and Tabata. 1994) but is not economically sustainable on a large scale and Bristol-Myers Squibb currently employs a semi-synthetic process for the production of Taxol®. From an engineering perspective. The purpose of this paper is to provide an overview of the commercial development of plant cell suspension culture technology to date and to focus on a limited number of engineering issues. is evidenced in many recent reviews concerning the technology and strategies for its optimisation (Zenk.. 1991). Taxus bre6ifolia. but as yet largely unattained prospect of long-term commercial success. if chemical synthesis has not been achieved or if it is technically problematic. 1995. 1991).. Inomata et al. Taxol® was originally isolated from the bark of the Pacific Yew tree. . Kieran et al. for example. superior metabolic performance over suspension cultures of the same lines. most research effort has been directed towards the commercialisation of plant cell suspension culture. However. roseus for the production of arbutin by a similar process (Yokoyama and Yanagi. This slow growing tree is principally found in the Pacific North-West. / Journal of Biotechnology 59 (1997) 39–52 proach. 1995. 1991. including the production of shikonin from Lithospermum erythrorhizon (Fujita. they can also be used for biotransformations such as the glucosylation of hydroquinone to arbutin by Rau6olfia serpentina (Lutterbach and Stockigt. Accordingly. 1996). 1988) have been investigated. Shuler. genetically engineered synthesis may be possible. specifically hydrodynamic shear sensitivity. There is also preliminary evidence to suggest that fungi growing on the Pacific Yew can produce both Taxol® and other taxanes (Stierle et al.. involving taxane precursors extracted from various yew species. 1995. independent of geographical and climatic factors. An alternative approach to production is essential if Taxol® supplies are to be assured and if the Pacific Yew population is not to be destroyed.. While tissue and root cultures (Flores and Curtis. Buitelaar and Tramper.. 2. 1994. the source plant is difficult to cultivate.M. 1991. 1988). Schlatmann et al. Commercial development That plant cell culture offers the tantalising. 1992.

g. in addition to the commercial processes mentioned earlier. How- ever. Verpoorte et al. 1991. 1995). its use can only be justified if it offers an economic advantage over chemical synthesis or traditional extraction processes. in an ideal system the cells would actively secrete the product into the suspending process fluid (Buitelaar and Tramper. 1995). woody plants (e. including Taxol® and the indole alkaloids. It has been reported (Anonymous. / Journal of Biotechnology 59 (1997) 39–52 41 Reviews on the potential of plant systems for the production of valuable products by these routes are presented by Parr (1989). the calculated product prices vary substantially with the choice of production strategy (e. 1991). etc. 1994) that ESCAgenetics. which filed a patent for the production of vanillin using tissue culture (Knuth and Sahai. or if no other alternative production route exists. is an inherently capital intensive process. cell–cell interaction and product synthesis. albeit non-commercial systems. there have been fewer commercial applications and most research effort has been concentrated on a limited number of high value products. Nicotiana tabacum cultures have been successfully cultivated in a 20 m3 stirred tank reactor (Noguchi et al. it is clear that there is a role for both biologists and engineers in improving system performance. However. research in Japan has received strong support from both government and industry (Misawa.. With regard to secondary metabolites. For example. batch culture with product retained intracellularly. Plant cell suspensions can now be almost routinely cultivated in small-scale configurations. Their analysis confirmed that while product price would be high and process optimisation essential. (1987) estimated that a 40-fold increase in the ajmalicine productivity of C. ¨ (1995). Engineering considerations From an engineering perspective.g. system morphology. 1995) reveal the predominance of Japanese companies in the search for commercial applications of plant culture technology. In the USA and Europe. the primary challenges lie in the area of process scale-up. The question of economic feasibility could be largely resolved by the development of stable. 3. Product concentrations and productivities are typically low. 1993) compared the results of a number of economic evaluation studies for the production of alkaloids by cell culture. 1996) and has produced almost all of the commercialised processes. Westphal (1990) reported on long-term cul- . Kieran et al. Alfermann and Petersen (1995) and Stockigt et al. the productivity of plant cell suspensions is of the order of 10 − 2 –10 − 1 g l − 1 per day (Scragg. Spearheaded by the company Plant Cell Culture Technology. Drapeau et al. there are a number of examples of largescale. (1991. Phyton Catalytic recently filed a patent for the production of Taxol® from highyielding cell cultures of Taxus chinensis (Bringi et al. in turn. Shuler (1993) predicted that pharmaceuticals produced naturally by slow-growing. roseus would be required to justify the production of this compound by cell culture methods. 1991. thereby simplifying downstream processing..). in addition to a clearer picture of the interactions between growth kinetics. In comprehensive reviews on the production of alkaloids. a fuller understanding of the biosynthetic pathways is essential.P. however. On this basis. Due to the fact that productivity depends on product yield. Taxol®) are the most likely candidates for development. high-yielding strains (Berlin.. cell culture processes could be feasible for speciality chemicals.M. Su. was also involved in Taxol® production scale-up trials. to achieve these objectives. 1977). Pras (1992). However. Moreover. economic feasibility studies are unanimous in acknowledging productivity as a limiting factor. Hara. 1992). The recent literature and patent libraries (Verpoorte et al. on an industrial scale. 1988) and the identification of optimal cultivation conditions. Plant cell culture. the organism growth rate and prevailing biomass levels. spontaneous or induced product release. derive largely from a combination of biological and engineering factors. Limited commercialisation of the technology to date can essentially be attributed to matters of economic feasibility which. While the high biomass levels required for economic viability necessarily limit the volume of free medium available.

Aggregation patterns. Hooker et al. whereas. For example. / Journal of Biotechnology 59 (1997) 39–52 Table 1 Summary of biocatalyst characteristics Biocatalyst Plant cells Animal cells Bacteria Shape Spherical Cylindrical Spherical Spherical Cylindrical Spiral Spherical Mycelial Size (vm) l: 100–500 d: 20–50 d: 10–20 d: B1 l: B5 d: 5–10 d: 5–10 l: B100 Cell wall Yes No Yes Aggregates Yes No Yes/No Doubling time (h) 20 – 100 20 0. 1987. Ikeda. employing a variety of impeller designs. although the secretion of extracellular polysaccharides (ECP). N.. 1987) techniques. under certain conditions as unbranched chains of up to 50 cells (DeJong et al. due to differences between the characteristics of both individual cells and suspensions of the respective systems. Doran. Hegglin et al. System characteristics and bioreactor design A number of reviews have dealt with the choice of bioreactors for plant cell suspension culture (Kargi and Rosenberg.. By way of illustration. Panda et al.. variously attributed to mixing and mass transfer problems in the characteristically nonNewtonian broths. (1993) the corresponding figure lay between approximately 10% (lag and stationary . 3. have high biomass concentrations. 3. Product recovery is complicated. 1995) and sieving (Mavituna and Park. 1989. ideally.1. 1991. equipment normally employed for microbial cultivation may not be immediately applicable to plant cells. Individual plant cells have a typically characteristic length of the order of 101 –102 vm and may be spherical to cylindrical in shape. The aggregation phenomenon has been used in the development of self-immobilisation methods (Prenosil et al. 1990) exists. may contribute to increased aggregation (Taticek et al. a summary of some of the physical properties of biological systems is presented in Table 1. by the fact that most systems retain the product of interest intracellularly. (1975) reported that 60% of a Morinda citrifolia culture existed as single cells or cells of two chains. largely due to the failure of cells to separate after division. variously studied using image analysis (Kieran et al. 1991).. Payne et al. vary significantly between cell lines and also as a consequence of culture age and cultivation conditions. 1967). Zenk et al. Aggregation Plant cells are significantly larger and slower growing than most microbial organisms.42 P. However. 1988. in many cases.. tivation of a number of lines in 50 m3 vessels. 1993) and most emphasis has been placed on modifications of the conventional stirred tank reactor (STR). both technically and economically. in the vacuole.1. particularly in the later stages of batch growth. which exhibit varying degrees of aggregation and which. 1987. scale-up has been accompanied by a reduction in system productivity (Schiel and Berlin. with bubble aeration.. 1989. 1991). comprising up to 102 cells. 1990).. thereby necessitating either cell lysis or non-destructive permeabilisation. However. may be many millimetres in diameter (Tanaka. Aggregation is common. in a study by Kieran et al. Aggregates.1.M. 1987. which is frequently described as highly aggregated (Hashimoto and Azechi.5 – 10 Qa (mmol l−1 h−1) o 10−1 10−1 103 Yeasts Moulds Yes Yes Yes/No Yes/No 10 10 – 20 102 102 a Characteristic oxygen consumption rate for biocatalyst in liquid culture. Kieran et al. 1987. 1982) and typically exhibit a tendency to settle.. Taticek et al.. tabacum.

(Bond et al. 1995) point to the importance of dissolved oxygen concentration for metabolite productivity. are of the order of 10 − 6 g g − 1 per second. Curtis and Emery. roseus from pelleted to single cells on scale-up from shake flasks to an air-lift reactor. Using the concept of the apparent viscosity of a fluid (Metzner and Otto. 1991)) and. 1957) rotational devices fitted with purpose-built impellers have also been used for the rheological characterisation of plant cell suspensions: for example.. Kieran et al. Ho et al. 1977). in some cases. There have been no comprehensive on-line rheological studies. on a dry weight basis. Specific oxygen consumption rates. Data have been collected for a variety of plant cell systems using conventional viscometers.. Kato et al.P. 1987). aggregate interactions.. Although critical dissolved oxygen concentrations of approximately 15–20% air saturation are commonly quoted for plant cell suspension cultures (Payne et al.g. Tanaka (1981). be affected by variations in aggregation patterns arising on scale-up. in which the combined effects of aeration and agitation are inextricably linked. Oxygen and aeration effects The oxygen requirements of plant cells are comparatively modest. In common with many microbial suspensions. as with other microbial suspensions. 1982. the system mass transfer coefficient. designed to satisfy the need for good suspension mixing at the widest possible range of laminar flow conditions (Jolicouer et al.M. although.. Kieran. although the data reported in the literature do not. it was generally concluded that for each system. 1992). high cell densities and high fluid viscosities can reduce oxygen mass transfer efficiencies in bioreactor systems. Thixotropic behaviour has been observed.. therefore. The influence of the morphology of the suspended cells and/or aggregates on the apparent viscosity of the suspension requires further investigation (Zhong et al. below which the . Smart and Fowler (1981) and Leckie et al. although only in isolated samples (Wagner and 3. 1993).. (1991) all investigated the effect of initial mass transfer coefficients on system performance in a variety of bioreactor configurations. in general.1.1. 1993). helical ribbon impellers. result in high whole-broth viscosities for plant cell suspensions. the critical value for cell growth may be significantly lower than that for metabolite synthesis and recent studies (Schlatmann et al.. Many systems also show evidence of a yield stress. Although the results are system specific.3. Deviations from expected aggregate distributions may be indicative of culture variations in response to environmental factors. These examples emphasise the importance of identifying morphological trends under actual growth conditions and with due reference to culture age. up to 70 g l − 1 on a dry weight basis (Matsubara and Fujita. 1986) and/or aggregate disruption (Rosenberg. 1992a. high biomass concentrations (e. Studies of the effects of aeration on plant suspension cultures have focused largely on the influence of kLa. Shuler (1993) discusses evidence to suggest that metabolite productivity may be significantly influenced by the degree of cellular association and may. the apparent viscosity is found to be strongly dependent on biomass concentration (Tanaka. A summary of relevant studies is presented in Table 2 and it is apparent that the majority of suspension cultures investigated exhibit non-Newtonian. 1988. 1992a. shear-thinning characteristics. it should also be noted that the use of biomass concentration (on a dry weight basis) as a correlating factor for viscosity can mask effects attributable to variations in individual cell size and water content over the course of the growth cycle. 3. However.2. ECP secretions. Zhong et al. 1995.. Dubuis et al. 1993). Rheology Aggregation. Wagner and Vogelmann (1977) observed a change in the morphology of suspensions of C. 1992. a lower limiting value of kLa exists. Vogelmann. Although the role of cell – cell interactions in undifferentiated systems has yet to be conclusively established. (1975). care must be taken to avoid sedimentation effects (Scragg et al.. / Journal of Biotechnology 59 (1997) 39–52 43 phases) and 50% (exponential phase). 1995). Kieran. refer to the high density suspensions required for economically viable largescale processing.

1992b. n. 1981. in some cases (Smart and Fowler. modified Stormer concentric cylinder Curtis and Emery (1993) Curtis and Emery (1993) = 9.9 Pseudoplastic n: 0. However.8 Pseudoplastic.9 513 B20 5450 (fresh weight) Pseudoplastic. .. (1978) Zhong et al. 1986) or shear-related effects (Ballica and Ryu. Wongasmuth and Doran. flow behaviour index.0 =7. Aeration of bioreactors can lead to foaming and in plant cell suspension cultures this prob- lem can be particularly severe (Zhong et al. (1992) Ballica and Ryu (1993) Kieran (1993) Jolicouer et al.. n :0. 1993)..5 Characterisationb Viscometric device Reference Papa6er somniferum Nicotiana tabacum Batch cultivation Semi-continuous Nicotiana tabacum Perilla frutescens Datura stramonium Newtonian Brookfield. 1994).M. Wongasmuth and Doran. Zhong et al. Schlatmann et al.44 P.6 Newtonian Pseudoplastic.. 1992b).1BnB0.53 Double-helical ribbon impeller Helical ribbon impeller Dry weight basis unless otherwise indicated. Ducos and Pareilleux. the contribution of extracellular polysaccharides and other medium components to foaming potential or foam stability has yet to be established. 1994. Using a gas recirculation bioreactor for scale-up studies involving C. reduced productivity observed at higher gassing rates is variously attributed to the stripping of CO2 and essential volatiles from the system (Kato et al. 1994) that loss of an unidentified essential volatile factor was responsible for the reduced ajmalicine synthesis observed on scale-up from a shake flask to an aerated bioreactor. n= 0. 1975. Brookfield Kato et al. 1994) in reduced system productivities. roseus. (1993) confirmed the importance of dissolved gaseous components for system performance and concluded (Schlatmann et al. A number of antifoams have been used to control foaming in plant cell suspensions (Smart and Fowler. n: 0. / Journal of Biotechnology 59 (1997) 39–52 Table 2 Rheological characterisation of plant cell suspension cultures System Biomass concentrationa (g l−1) B14. modified Stormer concentric cylinder Brookfield. Although there have been few comprehensive studies of this phenomenon and all have been limited to laboratory or pilot scale systems. Li et al.. 0. foaming has typically been correlated with aeration rates and extracellular protein concentrations (Wongasmuth and Doran. culture is inhibited. 1981.7 Bingham plastic Casson plastic Brookfield-type Brookfield-type Modified Weissenberg rheogonimeter Contraves rheomat.515 data Catharantus roseus Nicotiana tabacum a b Pseudoplastic. (1992) Tanaka (1982) Morinda citrifolia 5450 (fresh weight) Catharanthus 527 roseus Cudriana tricuspi. 1995) resulting. (1992a) Ballica et al. Kieran et al.

classified in terms of the prevailing shear environ- . 1994. However. ¨ 1991....5. Kieran et al.. Matsushita et al. slow-moving impellers often provide good mixing at relatively low rotational speeds.. tabacum (Shibasaki et al. 1991).. Investigations of shear sensitive systems. There have been many reports on the development of novel bioreactors for plant cell suspension culture. N. Here again..M. 1993. These systems encompass a range of particle sizes. in an 11 l surface-baffled vessel used for the cultivation of C. Shear studies on biological systems. including prokaryotic and eukaryotic suspensions and enzyme solutions. The response of a biological system to any hydrodynamic environment depends on the duration and intensity of the applied conditions as well as on the physiological characteristics of the system itself. can be broadly divided into two categories. 1993. 1994). Merchuk. the intensity of the aeration and agitation conditions. 1988) and they have been used by a number of workers for plant cell suspension systems (Smart and Fowler. Bioreactor design As with microbial systems. low-shear environment and reduced wall growth. rotating drum reactors (RDR) have been used for the cultivation of C. roseus (Tanaka et al. in particular of sub-lytic effects.. Khlebnikov et al. large. but its integration into the operation of large-scale bioreactors is problematic. erythrorhizon (Takahashi and Fujita. However. 1981. it is emphasised that the quantitative evaluation of shear-related effects. 3.. Markl et al. mass transfer and mixing-related problems typically include improved reactor design and. While few submerged systems are adversely affected by the hydrodynamic environment associated with laboratory maintenance conditions. 1986. Hua et al.g. Shear sensiti6ity in plant cell suspension cultures The hydrodynamic shear sensitivity of biological systems. 1993). Techniques employed for plant cell systems are summarised in Table 3. In STRs. has attracted considerable research attention in recent years and has been comprehensively reviewed by a number of authors (Thomas. 1990. Bubble columns and air-lift loop reactors offer the promise of a low-shear environment (Moo-Young and Chisti. as well as varying degrees of structural and metabolic complexity. which frequently precludes the direct comparison of results collected from different studies. 1994). Joshi et al. with conventional processing equipment.1. in general.. In a number of studies (Dunlop et al. a low-shear environment and increased cell – cell interaction..1. all of which impact on their sensitivity to a given shear environment and on the extent of their responses. due to poor bubble dispersion. The response may be positive (e. 1993) which is reduced at the broth viscosities associated with the high biomass levels necessary for an economically viable process. however. 1995). solutions must be achieved without concomitant negative shearrelated effects. the choice of an RDR in preference to either an air-lift reactor or a paddle-agitated STR was based on its superior performance in terms of suspension homogeneity. 1995. With a view to system scale-up.. 1983). / Journal of Biotechnology 59 (1997) 39–52 45 3.P. 1996). 1991. more commonly. Jolicouer et al. In the latter case. System response to shear may be measured and assessed in a number of ways.4. (1992) reported on the successful use of a double helical ribbon impeller. For example. it is interesting to note how few of the measurement techniques listed lend themselves to on-line applications. 1994. roseus cultures. Furusaki et al. Oxygen transfer may be limiting. an increase in biomass yield or secondary metabolite synthesis). generally focus on the negative or damaging effects associated with the hydrodynamic conditions encountered in process equipment. Fluidised-bed reactors have been proposed for industrial scale use (Dubuis et al. performance is limited by mixing efficiency (Doran. Takeda et al. offering the possibility of perfusion operation with a facility for cell harvesting. Fulzele and Heble.. increasing agitation and aeration intensity. Hegarty et al. approaches to overcoming suspension. Light irradiation has been shown to have a stimulating effect on the secondary metabolism of some systems (Zhong et al. 1992) and L. process scale-up and the cultivation of high-density suspensions increase the mass transfer requirements of the system and accordingly. depends crucially on the choice of damage indicator. 1992.

Zhong et al. for the duration of cultivation or a significant portion thereof (Scragg et al. moreover. the intensity of the environment has been generally related to impeller speed or power input. Kieran et al. (1993) Rosenberg (1987). due to the variety of bioreactors employed. (1995) Rosenberg (1987).. Studies involving plant cell suspension cultures are summarised in Table 4.. (1989) Ho et al. (1988) Takeda et al. (1984). (1993) Hooker et al. (1994). Due to the difficulties involved in quantifying the levels of shear to which an organism is exposed in the turbulent field of an agitated bioreactor. (1994). generally under non-growth conditions. (1994) Hooker et al. Kieran et al. for short periods of time. Kieran (1993) Markx et al. Meijer et al. Kieran et al.. may be less appropriate for systems yielding non-growth associated metabolites.. in general.. 1988. 1995).g. (1989). 1994. 1995). (1994).. it is arguable that analysis under actual growth conditions. (1995) Zhong et al. (1995) Parr et al. Use of chemostat culture (Meijer et al. An exponential decay model is commonly employed to describe viability loss with increasing exposure time to the imposed shearing conditions. As the apparatus is normally designed for sterile opera- tion. reflect actual production conditions. Takeda et al. laminar or turbulent flow conditions (e. or in a scaled down version of a production bioreactor. / Journal of Biotechnology 59 (1997) 39–52 Table 3 Methods used for the assessment of shear-related effects in plant cell suspension cultures System response Reduction in viability Parameter measured Growth rate Regrowth potential Membrane integrity Dye exclusion Dual isotope labelling Dielectric permitivity pH variation Protein release Total organic carbon Secondary metabolite release Oxygen uptake rate Respiration activity (TTC reduction) ATP concentration Metabolite productivity Cell wall composition Aggregate size/shape Expansion index Reference Rosenberg (1987). Scragg et al. In the second type of study. (1994) Tanaka et al..M. 1994. on the basis of system response to lami- . (1991) Wagner and Vogelmann (1977) Meijer et al.46 P. the concept of a critical shear stress is employed. where batch processing is most commonly employed and. The hydrodynamic environment is generally regulated by changing the rate or method of agitation and/or aeration. 1995). 1993) facilitates the investigation of long-term effects. Frequently.. (1994) Takeda et al. offers the greatest potential for successful scale-up of results (Meijer et al.. Although these studies tend to be highly system-specific. the resultant death or decay rate is used to quantify system response to a given hydrodynamic environment (Kieran et al. (1993) Meijer et al. (1988) Takeda et al. capillary and submerged jet devices). 1994. but does not. For example. 1994. the biological suspension is exposed to shear forces under growth conditions. The benefits of examining the shear sensitivity of a system under actual or proposed operating conditions are not to be underestimated. (1994) Release of intracellular components Change in metabolism Changes in morphology and/or aggregation patterns ment and the duration of cell exposure. Takeda et al. cells are exposed to well-defined. in couette. Meijer et al. Zhong et al. In the first category. Zhong et al. Kieran et al. Ho et al. the experimental exposure time is limited only by the kinetics of the system itself and by the ability of the organism to survive under the prevailing conditions.

as conducted to date and proposed a model for a more systematic and holistic evaluation of the response of plant cell systems to hydrodynamic environments. stress protein expression. 1987). (1989) Cell lysis Metabolite production Respiration activity Zhong et al. Kieran et al. 1990) and biomass productivity (Ho et al. In subsequent studies by Meijer et al. and may also depend on culture age and cultivation history. a critical shear stress of 50 N m − 2 was identified for suspensions of Daucus carota (Rosenberg. rather than low-shear bioreactors. (1995) MacLoughlin et al.. however. roseus.. Namdev and Dunlop (1995) recently highlighted limitations of shear-related studies. system response is most frequently monitored in terms of loss of viability and accordingly. 1995.. it appears that plant cell suspensions are more robust than initially anticipated and that significant losses of viability are not. However. System response may vary significantly between cell lines. more subtle.. C. 1997) have pointed to the use of energy dissipation as a correlating factor for shear-related damage. in general. no single mechanism for cell damage has been conclusively identified. given the variety of aggregate morphologies exhibited by plant cell systems. Wagner and Vogelmann (1977) reported evidence of shear sensitivity in cultures of C. 1993. non-lytic effects may be overlooked (Namdev and Dunlop. (1993). Because these devices are generally operated under non-sterile conditions. Vogelmann et al. (1978) suggested a critical shear stress of between 80 and 200 N m − 2 for M. it appears as if research effort might be more profitably devoted to the development of shear-resistant cell lines. 4. (1995) Perilla frutescens Couette-type nar flow conditions in a viscometric device. Interestingly. / Journal of Biotechnology 59 (1997) 39–52 Table 4 Defined flow-field shear experiments using plant cell suspension cultures Cell suspension Morinda citrifolia Apparatus Recirculating flow capillary Submerged jet Daucus carota Couette viscometer Flow regime Laminar/turbulent Turbulent Laminar Damage indicator Dye exclusion Morphology Dye exclusion Morphology Morphology Regrowth Cell lysis Reference Kieran et al. However. There are a number of important conclusions to be drawn from these studies which have significant consequences for the success of commercial systems in conventional bioreactors. to be expected under normal operating conditions in a conventional STR. using regrowth potential as an indicator of system response. (1997) Rosenberg (1987) 47 Nicotiana tabacum Couette-type Transitional/turbulent Viability Hooker et al. citrifolia.P. Recent studies (Dunlop and Namdev. osmo-regulation and aggregation. focusing on calcium transport. Conclusions This paper has considered only a limited number of engineering-related issues which impinge . 1995). roseus was found to be considerably more robust than cultures of Cinchona robusta and Tabernaemontana di6aricata. Kieran et al. other less dramatic shear-related effects may be observed. Metabolic responses may be related to changes in cell–cell interactions effected by shear-induced aggregate disruption. MacLoughlin et al. 1995). including reductions in metabolite yield (Hooker et al.M. For example. Overall.

M. The product can be concentrated and by removing feed-back inhibition effects. 674 – 684. Dubuis. DiCosmo.. Kieran et al. Microb. Ryu. Drapeau. 43. Jansen.W.. 1995. Despite the obvious attractions of these integrated processing approaches.M. pp. / Journal of Biotechnology 59 (1997) 39–52 upon the commercialisation of plant cell suspension technology.. problems of both economic and technical feasibility must be resolved before their application on an industrial scale can be considered. Growth of Catharanthus roseus cell suspensions in bioreactors: on-line analysis of oxygen and carbon dioxide levels in inlet and outlet gas streams.. Khlebnikov et al. In the area of process control.M. including plant cell suspension cultures (Hooker and Lee. B.W. Strategies to improve the production of secondary metabolites with plant cell cultures: a literature review. a facility for product removal. J. continuous.F. Kut..). Cell Res. BHR Group Conference Series. Buitelaar. London. 1988. wide-scale commercialisation of plant cell suspension technology may soon be feasible. 1992. Ballica.E. 1988.C.D.Y. Rheological properties of plant cell suspensions. High yield production of taxol and taxane production from Taxus species cell cultures: by cultivation in a nutrient medium cells derived from callus and/or suspension cultures of Taxus species useful for cancer treatment. 1995. R. W.. Enzym. A. Technol.. enhanced cell – cell contact. Biotechnol. 47. Kadkade. ¨ J. References Alfermann. 48. 7. D. 1993. May 24. Biotechnology in Agriculture and Forestry: Medicinal and Aromatic Plants I.R. DeJong.A. Formation of secondary metabolite in cultured plant cells and its impact on pharmacy. 42. (Ed. 37 – 59. 1989) offers the possibility of a low-shear environment. Design of reactors for plant cells and organs.G. Bond. Y. but there is scope for development of models which take account of the fact that suspensions may not be homogeneous with respect to metabolite synthesis... Tramper. Powell. 1992... In: Nienow. Scragg. D. Bourne.R. Prenosil. 17.. 1993). B. Exp. if product secretion can be achieved. Different operating strategies (e. a large-scale operation of plant systems is hampered by difficulties associated with the online analysis of parameters such as cell or aggregate size distributions which may play an important role in metabolite productivity.. Bioeng. Blanch. A.. M. Romette. P.. B. A. Fowler. (Ed.. Economic assessment of plant cell culture for the production of ajmalicine. 1181 – 1189. 10. The overall focus of related research is the achievement of large-scale cultivation of high-yielding cell lines and efficient product recovery. batch. J.. 1992) can facilitate continuous processing without loss of biomass. Berlin. Adv. E. D.. biomass re-use and. Immobilisation (Hulst and Tramper. 89 – 100.W. 4. Kane. Adv. 43. D. Dornenburg. 1993.F.. van Gulik et al.. 1995. Biotechnol. Natural product formation by plant cell biotechnology.D. Petersen. Olson. 111 – 141. etc. R. Biotechnol. Misawa. growth associated and non-growth associated) and which either store the product of interest intracellularly or. D. Emery. Plant Cell Tissue Organ Cult. J. 1987. Plant cell and tissue culture: alternatives for metabolite production.S.g. Curtis.. Effects of rheological properties and mass transfer on plant cell bioreactor performance: production of tropane alkaloids.W. . V. In: Bajaj. 199 – 205. Bioeng.Y. Roach. vol. Springer-Verlag. Process modelling has been successfully used as a tool for the design.. Pilot-scale cultures of Coffea arabica in a novel loop fluidised-bed reactor.H.. Dubuis. 1992. preferably. 1993. A. Prog. C.. J. Wilke.. 117 – 168. E. vol. 30. Prenosil...M. Bioreactor and Bioprocess Fluid Dynamics.L. Biotechnol.. Anonymous.g. Non-lethal methods for inducing product release and the use of two-phase systems (Buitelaar and Tramper.). H. Eng.) may be required for cell lines exhibiting different metabolite synthesis kinetics (e.L.M. R. J.. M. 1995). 171 – 183... 425 – 453. MEP.R. 1993. A. Lett. 520 – 526. Physical factors affecting the design and scale-up of fluidized-bed bioreactors for plant cell culture. Biotechnol. A. 139 – 156. Plant cell suspension culture rheology. 13.. Ballica. Chemical Marketing Reporter. Schubmehl. pp. p. Bringi. Knorr. analysis and optimisation of many biological systems. Doran. R. through the integration of biologicallyand engineering-based approaches. Through concerted research efforts in the areas discussed in this paper and in those highlighted above... B. two-stage. Biotechnol. P. F. R.H. secrete it into the suspending fluid. 42.E.. H. 1967. Prince.. Biotechnol. 946 – 953.P.. P... Bioeng. 23. Plant Cell Tissue Organ Cult. Some of these advantages have already been demonstrated using self-immobilised plant cell systems in fluidised-bed reactors (Dubuis et al. 713 – 718. 1993. 1995. ESCA scales up taxol.J.W.. Strategies for the improve¨ ment of secondary metabolite production in plant cell cultures. D... Berlin. Biotechnol. US Patent 5 407 816. Owen. 413 – 420..48 P. O.. Pluss. 8. 1995. productivity may be improved. C.. Oxidoreductive hydrolytic enzyme patterns in plant suspension culture cells. 5. 1994. J. Ryu. Biochem.M. Kut.

224 – 228. 1975... Shimizu.M. D. (Ed. Yiin. 49 Holton. Effect of aeration rate and influence of pCO2 in large-scale cultures of Catharanthus roseus cells. 1994. Berlin.A.. Yokoyama.. 45 – 57.E. O’Donnell. New York Acad. Lee. B. In: Komamine. Y. Prenosil. Growth and productivity of Beta 6ulgaris cell culture in fluidized-bed reactors. Malone. 271 – 282. pp. Lee. A. M. Large-scale cultivation of Catharanthus roseus cells: production of ajmalicine in a 20-l airlift bioreactor. Liu..S. Ireland.. Kawazoe. M. Don suspension cultures in airlift bioreactors: the inhibitory effect at high aeration rates on culture growth.). P. Effect of fluid shear forces on plant cell suspensions. J.H. Curtis. Biotechnol.R. (Eds. Yanagi.. 26–32. 116.. Jolicouer.J. Biotechnol. A.. J. 315 – 319. 1992. Furusaki... H. Plant Tissue and Cell Culture. R. S. Bioeng. 225 – 236. 1–7.N. plant and microbial cells....H... Biotechnol. In: DiCosmo.). Kieran.S. 1988.M.F. Soc. 511 – 521. C..K. 1995. Plant Cell Culture Secondary Metabolism – Toward Industrial Application. Liang.).J.. pp.-B. P.P. 1991. Y. Cultivation of plant cells in a stirred vessel: effect of impeller design. P. DiCosmo.. Y.. Crit. Rorrer. P. Bioreactor and Bioprocess Fluid Dynamics. H. Kieran. T. Bioeng.. Alan R.. Chem. Eng. Suzuki. Reaktorsystem zur Massenkultivation von pflanzlichen Zellkulturen bei niedrigem hydrodynamischen Stress. Scragg. J. Food and bioproducts processing.). Ho.. F. Nadizadeh. F. MacLoughlin. Ferment.-P. H.. K.P.M.. pp. Zhang. F.. 2263 – 2276. 13.. Biotechnol. Research on the production of useful compounds by plant cell cultures in Japan. Biesboer. C. 546 – 558... Dunlop. C. M. M. Eng.. Biotechnol. J.. Flores. Gentile.M...K. A... Enzym... Approaches to understanding and manipulating the biosynthetic potential of plant roots.P. Dunlop. F. P. Kim. Sci. pp. Shindo. Y. Technol. 140 – 145. Ikeda. Biotechnology in Agriculture and Forestry: Medicinal and Aromatic Plants I. 11. pp.. Henderson. Berlin. J.. Exp.A. K. cells in suspension culture. J. D.A. Y. Glasgow.. 1978. Immobilized plant cells: a literature survey. T. Smith. CRC Press. A. Role of hydrodynamic shear in the cultivation of animal.D. The effect of light irradiation on secondary metabolite production by Coffea arabica cells. J. vol. D. Hooker.. 56. 1987.. C. P. Somoza.-Y. Bioeng. 1597 – 1598. Am. 2. 744 – 751.. Rosenberg. G. D... Technol. Technol. Biotechnol.. Biotechnol. P. Fowler.. Application of a new structured model to tobacco cell cultures. 3. pp.E. Fulzele. Y. C. Hegarty. N.M. Fujita. Somers. Boca Raton. vol. J.. Cell damage and oxygen mass transfer during cultivation of Nicotiana tabacum in a stirred-tank bioreactor. Kieran. 39. J...M. 1992. Misawa. M. Y. Hooker. Ann.. Bioproducts and Bioprocesses. B.. P. 1989. Y.P. J.. Shikonin: production by plant (Lithospermum erythrorhizon) cell cultures.H. J.. Eng. M. Hashimoto. R.. Lee. 1987. M.. pp. Biotechnology in Agriculture and Forestry: Medicinal and Aromatic Plants I. Murthi.S. A. S. 51 – 56. Boca Raton. Kargi. Berlin. Bourne.. Trans. Misawa. Prenosil. Pareilleux.B. Tao.. MacLoughlin.J. 169–185. Microb. 35. S.. In: Nienow. Kim.S. An. An.W. E. DiCosmo. Appl.D.. Thesis. (C) 71.E. Microbiol. G. 296 – 304. Effect of fluid forces on plant cell suspensions. Response of plant tissue culture to a high-shear environment. Variation of aggregate size in plant cell suspension batch and semi-continuous cultures. Microb. P. M. Functionalization of the B ring. Plant Cell Culture in Japan. Development of a helical-ribbon impeller bioreactor for high-density plant cell suspension culture. Namdev.. Bot. B.-H. H. Hegglin.R. J. Kato. 14. (Ed.. Patole.G. The aeration of Catharanthus roseus L. 39. Kato. Secondary metabolites from plant cells: pharmaceutical applications and progress in commercial production. Malone... Tramper.E. Soh. Erickson. Heble. Chem.F.R.). Chavarie. Viscosity of the broth of tobacco cells in suspension culture.C.L. Biotechnol. M.S... Prog.).S. R. 45. Nagai. 305 – 328. In: Bajaj. 1993. Elias. Production of anti-plant-viral protein by Mirabilis jalapa L. vol. 1995. Appl. Hooker. J. Seki. Joshi. F. 765 – 774. 1 – 8. FL. Hackett.G. Prog. E. G.. Tanner.. F.. Azechi.. Ph. 37. Production of Paclitaxel and related Taxoids in cell cultures of Taxus cuspidata: perspectives for industrial applications. 1993.. M. 1995. M. 11..B.P. 36.. Kut. 121 – 141.E. / Journal of Biotechnology 59 (1997) 39–52 Ducos. 40 – 46. S.M.. A. Bioeng.. P. 1992.. Biediger. First total synthesis of taxol 1. Fett-Neto. High-level production of arbutin from hydroquinone in suspension cultures of Catharanthus roseus plant cells.. In: Yoshida.. Springer-Verlag. S. D. (Eds.). T. 139–166. S. Liss. A. Tabata.. . L. 1986. C. Technol.. 4.Z. S. Hulst. In: Green. CMC.A.. 53. J. Seto. Smart.. I. 1990. M. H. E.. 1993.W. O. 1991. Effect of initial kLa on the growth of tobacco cells in batch culture. Tokyo. 25. Chimia 44. J.. B. Springer-Verlag. (Eds. 484 – 490. Khlebnikov. 1994. 1993.. 1994. Bioreactors for large-scale culture of plant cells. W... 49. Biotechnol. (Ed.. CRC Press. A. 101–105. 665. pp. P.. 1986. J. 11. (Eds.. T.C. Fujita. 4.M. 1996. Rev. W. M.. Rosenberg. An investigation of the hydrodynamic shear susceptibility of suspension cultures of Morinda citrofolia. Bioproc.. In: Bajaj. Furuya. Hara.). Fluid shear effects on suspension cultures of Morinda citrifolia. Sci. 104–122. 1989. Plant cell bioreactors: present status and future trends. MEP.. 447– 455. M. Carreau. 62. L. 1988. Springer-Verlag. 415 – 425. Ferment. 1911 – 1920. J.D. Review of the effects of shear and interfacial phenomena on cell viability.. 187–202. Dubuis. London. 1993. Kurata. Biotechnol... Boatman. Namdev.Z. FL. J. Misawa. S. A. 1996. (Eds.. 1990.. P. Archambault. Enzym. Plant Cell Culture Secondary Metabolism – Toward Industrial Application... Tang. Chem.K. 35. T.H.J.K. In: DiCosmo. C.M. Kieran et al. University College Dublin. Inomata. M. 188–209.. Chem. New York.D.M. Vu. 1996. Microbiol.. L... Hua..

N. E. Bisaria.. Couladouros.. 109– 131. (Eds. Schlatmann. A. Thesis.E. Kieran. A. A. J. R.. J. Int. A. J.. Ueno. 386 – 397.. Libbenga.). Apoth. 54.. W. (in press). DiCosmo. A. 1992..F.. pp.. 1995.. H. Metzner. K. 435 – 439. Biotechnol. Prenosil..E. H.. Meijer. 1987.. J... B9–B14.J. 1994. J.F. Y. A.E.F.. 16. Plant Tissue Culture and its Biotechnological Application. Louis.L. 467–477.E. / Journal of Biotechnology 59 (1997) 39–52 Nicolaou. Furusaki. R. 501. Eng. Biotechnol.R. Biotechnol. Agitation of non-Newtonian fluids.L..M. 3. Hirata...). Noguchi. Microb. In: DiCosmo. Nuutila. M. 1997. ten Hoopen.50 P. Meijer.. 1991. 85 – 94. van Gulik.J...H. Shuler. Ph. J. Yang. Azechi. Misawa. Moo-Young.J. K..M.. R. Amsterdam. Verpoorte. Murtagh.J.G.. J. Berlin... 1987. B. A.. R. R.. H. E. Matsubara. O.. H.J. (Eds. Enzym. Misawa. Considerations for designing bioreactors for shear-sensitive culture.. Bringi.. pp. ten Hoopen..R.E.. Luyben. Effects of hydrodynamic stress on cultured plant cells: a literature survey.. 253 – 262.C. 1988. 1993. F.. M. In: Ryu. 1991. Technol.R.. Bhojwani. Kato.. J.. (Eds. Otto. M. Rhodes. Yamamoto. J. 1996. D. Lee. 1995. Renaud. J. CMC. (Eds.M. P. (Ed. Tokyo. 6... M.. Plant Cell Culture in Japan.K. J. M. 9. Panda. A. Nat. Scaleup of ajmalicine production by plant cell cultures of Catharanthus roseus. Cult. P. Scragg. Heijnen. K. Total synthesis of taxol.. Technol. 1989. Enzym. R.. ten Hoopen. Markx.. Payne. Org.. pp. Heijnen.. Sorensen. K. Appl. Large scale fermentation and alkaloid production of cell suspension cultures of Catharanthus roseus. M. 1994.E. Microb.H.Q.. C. 234 – 238. MacLoughlin. Namdev. Y. Bioeng. 1993.). 3–10. Acta. Li. Berlin. Heijnen. Production of berberine. Improvement of growth rate of plant cell cultures. V. Mineral oil addition as a means of foam control for plant cell cultures in stirred tank fermenters.M.S. pp. MO. Parr. Munich. St.P. (Eds. Mavituna. CMC. Technol. Shear sensitivity of plant cells in suspension.. 339 – 353. DiCosmo.. 630 – 634. F..A. The hydrodynamic shear sensitivity of suspension cultured plant cells. US Patent 5 057 424.). Hamilton.... 185 – 197.F. 1994.. ¨ Ztg.H..J. Nantermet. Libbenga. 193 – 229. T. Fujino. 1992. Biotechnol.. Genetically engineered synthesis of natural products. F. Eng. 145–158. . J.G. O. Plant Cell Tiss. 1991. Robins. Bio/Technol. Kieran et al...M.J.C. K. G.M.. 1991. 39–44. Pras.I. Munich. V.. pp.M. 37.. Ogawa. Shear effects on suspended cells.Ch.. Sci. Z. 57. E.. W. In: Lydersen.. A..L.G. 10. An investigation into the role of initial kLa on the growth and alkaloid accumulation by cultures of Catharanthus roseus. Shin. Microb. In: Barz. 31.. W. Washington University. 45. Biotechnol. 41. pp.A. Fonck.. Plant Cell Culture Secondary Metabolism – Toward Industrial Application. 133. Chem. Dielectric spectroscopy as a novel and convenient tool for the study of the shear sensitivity of plant cells in suspension culture. Apparent free space and cell volume estimation: a non-destructive method for assessing the growth and membrane integrity/viability of immobilised plant cells.J. 2009–2011. M. 1987. Prince. Biotechnol. Leckie.J. Chim. M. Plant cell reactors: a perspective.E. Hegglin.. Biochem.. In: Komamine. van Gameren. Biochem.. Plant Cell Rep. Research activities in Japan.. Schlatmann.G. M. Scott. Purine alkaloid production by free and immobilized Coffea arabica cells. J. C.S. Biotechnol. Payne. H. Claiborne..C. New York Acad.. Large scale plant cell culture. J.M. Schlatmann. Luyben. 44... Katsuyama..G.. 75. 713–718. J. Effects of hydrodynamic stress on the growth of plant cells in batch and continuous culture.). Markl. T. Chem. Bioeng. High density culture of plant cells using an air lift column for production of valuable metabolites. 1977.M. P.. The negligible role of carbon dioxide and ethylene in ajmalicine production by Catharanthus roseus cell suspensions.. Tokyo. 1994. Bioeng. K.I. A.. Wittek. 1991. S. 17 – 42.. J.. Brodelius.. 1993. G. Vinke. Adv. Plant Cell Rep. ten Hoopen. Zenk. Parr. Bourne.. K... Fujita. Elsevier. S. 35. Knuth. Plant Cell Culture in Japan. Matsushita. J. F. Funatsu. In: Komamine. Flavor composition and method. B.. Y.. Dtsch.J. H. K.M. Merchuk.. 1291 – 1296.G. Cliffe.L. J.J. Park. 557 – 573.J. Large-scale production of secondary metabolites by plant cell cultures. Schiel. Biotechnol. Eng... Misawa.J. 3. 8. Schlatmann. Paulvannan. Guy. J. 157 – 160.. 1984.. Reinhard. 364 –370. The production of secondary metabolites by plant cell cultures. Misawa.J. 29 – 62... SpringerVerlag.). Dunlop.G.Y.D. Ann. Sahai.K. J. H.K. Lutterbach.J. 14. Large Scale Cell Culture Technology.. Shuler. Malone. Rosenberg. Koga. Boca Raton. 3–7. J. Saroj. Helv. Meijer. 19. H. J. The resistance ¨ of micro-organisms to hydrodynamic stress. Y. FL.H. Heijnen. ten Hoopen. S. 11. The effects of turbulent jet flows on plant cell suspension cultures. Biotechnol. N.D. M.. P. Studies in Plant Science 4: Advances in Plant Biotechnology.G. ten Hoopen. M. P.K. Smith. Matsumoto. 390 – 394.J. Hanser. Enzym. M.. Stockigt. J. J. K. E. Bioeng.und Gewebekulturen: Moglichkeiten und Grenzen.A. J. K. K.E.Z. Tech. Biotechnol. 153 – 161. F. Chisti. Arzneistoffe aus Pflanzlichen Zell. Nature 367. M. 1992. Relation between dissolved oxygen concentration and ajmalicine production rate in high-density cultures of Catharanthus roseus. R. Kreis. Plant Cell and Tissue Culture in Liquid Systems. K.J.. H. J....J. J. 161 – 164. ten Hoopen. J.. Liu.J. 1987.C. Size distribution of plant cell aggregates in batch culture. Bronnenmeier. A. K.. 15. J..B. E.. M. High yield formation of ¨ arbutin from hydroquinone by cell-suspension cultures of Rauwolfia serpentina. 65–122. 1957.Ch.Ch. 26.. 11 – 52. 1987.. CRC Press. Vinke. A. 1991.H. D.J. K. Bioconversion of naturally occurring precursors and related synthetic compounds using plant cell cultures. G. 1992. J...L.. 1995. Kato. 1 – 26. G... Hanser. 1989. Prod...T. 1994..

M. M. Westphal. 27. E. Lett. In: Komamine.. Munich. S. 425–442. 29. 1991.A. F. 41..J. 58. 1315–1324. Scragg. Eng.M.. van Gulik. P. D. 2 – 187. (Eds. 10. H.. Misawa. F. Nat.M. pp. Ohzora. Bioeng. F. Appl. van der Heijden. T.. T.. Strobel. Anthraquinone production by cell suspension cultures of Morinda citrifolia. Scragg.. Biotechnol. Pape. Fowler.. C. R. H. K.H. F.... Springer-Verlag. J. F. 130 – 146. 1988. Misawa. Bioeng.. Doran.W. R. 10. ten Hoopen. Enzym. M. 1988. van der Heijden. Biotechnol. Progress in plant cellular and molecular biology... Bond. High-level production of arbutin by biotransformation. 97–109. A..). H. Strategies for improving productivity in plant cell. Verpoorte. ten Hoopen.P. Taxus bre6ifolia.M.. Technol. Tokyo. 139 – 159. Large scale production of new biologically active compounds in plant cell cultures. (Ed. Takeda.. 3. pp. Furusaki. Production of Shikonin.. Yokoyama. CMC. 1981. Bignami. 50.. Grothaus. M. 245 – 252. Reinhard. Bioprocessing technology for plant cell suspension cultures. Verpoorte. R.. Hoge.. 1991. Kluwer.H. Phytochemistry 30. A.. van Aartrijk. Leckie. pp. 43. 51 Taticek. Bioeng. Seki. CMC. W. (Eds.W.J. M.H.J. Prod. Jpn. Taxol (Paclitaxel) production using free and immobilized cells of Taxus cuspidata. (Eds..H. U. R. Berlin....G. The Alkaloids. M. The effect of physical stress on plant cells in suspension cultures.). 1995. tissue and organ culture in bioreactors. Tokyo... S.A. Plant Tissue Culture and Its Biotechnological Application.. Prod. In: Nijkamp. M. 1992. K. 1995. R. Production of Natural Compounds by Cell Culture Methods.. Iwamoto. Allan. In: Yoshida.H. 3. T. El-Shagi. Some aspects on mass cultivation. DiCosmo. . 79 – 101. (Eds. H. Suspension culture of Nicotiana tabacum cells in a rotarydrum bioreactor. Springer-Verlag. Critical Reports on Applied Chemistry: Chemical Engineering Problems in Biotechnology.).J..H. applications and products. Fowler. Biotechnol. Chem. M.. van Gulik. Foaming and cell flotation in suspended plant cell cultures and the effect of chemical antifoams. Lett. 43... Bioeng. Elsevier Applied Science. Curr. 485–490.. Stafford.. 653 – 658. Wagner. T. 24. 361 – 367. 189– 230. Biotechnol. DiCosmo. Scragg. In: Brossi.H. Technological problems in cultivation of plant cells at high density..J. Academic Press.. J. 105 – 109.L.. (Ed. 214–219. pp. In: Barz. 24. A. 1203– 1218. pp. M. / Journal of Biotechnology 59 (1997) 39–52 Scragg.. ten Hoopen. Large scale plant cell culture: methods. J.. Zenk.. P. P. Technol. J. Suppl. Biochem. Bischof. Cultivation of plant tissue cultures in bioreactors and formation of secondary metabolites. 1995. Smart. A structured model describing carbon and phosphate limited growth of Catharanthus roseus plant cell suspensions in batch and chemostat cultures... 1992. Tokyo. Zenk. 79 – 91.. Biotechnol.).J. Tanaka. M.H. 38. Shibasaki.. Seki. H. Large scale culture of ginseng.. Cambridge. 53. Bioeng. Vogelmann.. H.. (Eds. Biotechnol. C. A. Berlin. G. (Eds. The problems associated with high biomass levels in plant cell suspension cultures. S.. 23 – 94. London. J. Rotating drum fermenter for plant cell suspension cultures.G.. 72–78. Opin. M. Planta Med. (Eds. vol. G. Plant Cell Tissue Organ Cult. D.. 1994. Cambridge University Press. P. vol. Yoshida. Secondary Metabolism in Plant Cell Cultures. H. Thomas.. Biotechnol.. pp. Lutterbach. Biotechnol. 23. Bioeng.. 1993.. Oxygen transfer in broths of plant cells at high density. Harada. Chasing the enzymes of secondary metabolism: plant cell cultures as a pot of gold... T. F. Biotechnol.. H. Schripsema. Heijnen. 92 – 98. In: Morris. Plant biotechnology for the production of alkaloids: present status and prospects. Zenk. 1983. Plant cell biotechnology for the production of alkaloids: present status and prospects. M. Plant Cell Culture in Japan.. Stockigt. L. vander Plan..). 1986... Hirose.. pp.. 171 – 176.. T..). 1977. Falkenhagen. Vogelmann. 1994.. 56.. Biotechnol. Obitz. 1991. 1993. Problems of shear in biotechnology.. A..J. 163–170. Ushiyama.. Y. Yonemoto. 1991. Smart. E. In: Winkler. F. Nishijima. M. London.. 1991. The search for a taxol-producing micro-organism among the endophytic fungi of the Pacific Yew. E. E. 186 – 207. Wagner. 2. A. Stierle. J. DiCosmo. Plant Cell Culture in Japan.. R.).W. J. Seki. Furusaki.. Rheological properties of plant cell suspension cultures.. S. Zhong. In: Komamine. 44. 601 – 608. A. Effect of light irradiation on anthocyanin production by suspended culture of Perilla frutescens..R.-J. Kinoshita. 481 – 488. 1990. Semba. A. N. The scale-up of plant cell culture: engineering considerations. Schulte.. Effect of aeration on large-scale cultures of plant cells.H. H. Reinhard. Suwa. (Eds. Kieran et al.J. A. 1991. Yanagi. M. M. CMC. H.. Tanner. Effect of shear on the viability of plant cell suspensions. T. Jitsufuchi. Tanaka. Biotechnol.W. S.. J. GSF. Takadi. Natural products and enzymes from plant cell cultures.. 1991. En¨ dress.J. 178–194.. Su.).. 1975. 2359–2370.. Shuler.. M. vol. 1982.. A. Allan.. W. A.W. Scragg... P. 3861 – 3863. In: Alfermann. 771 – 780. 359– 363. Takeda. M. 1995. In: Komamine. Tanaka. P.). 235–245. W. Biotechnol. H. Misawa. 1993. Plant Cell Tissue Organ Cult.. 1997. W. M. Bioeng. Chem. pp. Stierle. Legge. R. Plant Cell Culture in Japan.J. N.. N.L. J.R. H. Takahashi. Wongasmuth.D. Bioproducts and Bioprocesses. 1978. T.A. Plant Cell Tissue Organ Cult.G. pp. A. Fujita. Microb. Dordrecht. 25. J.H.). Hydrodynamic damage of cultured cells of Catharanthus tinctorius in a stirred tank reactor... H. H.. Moo-Young. pp. R. 1981.. Nat. 466–471. A..C. 40. 53. Tanaka.. 1990.

Zhong. Bioeng..-J..... 1994.52 P. J. 106–109. / Journal of Biotechnology 59 (1997) 39–52 Zhong.-J. J. Yoshida. Rheological characterisation of cell suspension and cell culture of Perilla frutescens. 1992a. 40. Effects of surfactants on cell growth and pigment production in suspension cultures of Perilla frutescens. Microbiol... Yoshida. T. J. T. S.. 1995. 1256– 1262. S. T.-J. Zhong. Fujiyama. T.-T. J. Bioeng.-J.. 11. World J. Yu. K. Yoshida. 649 – 654.. Zhong. Kieran et al. 44. A quantitative analysis of shear effects on cell suspension and cell cultures of Perilla frutescens in bioreactors.. Yoshida. Microbiol. Kinoshita.M. Recent advances in plant cell cultures in bioreactors. Seki. Seki. Biotechnol. . . 481 – 487. World J. T. Kinoshita.. T. 1992b. Biotechnol. T.. J. Seki. Biotechnol.. Biotechnol. 8.

Sign up to vote on this title
UsefulNot useful