recorded under high shear conditions, suggesting increased cellular maintenance
requirements in cells under hydrodynamic stress.
Shear SensitivityReductions in plant system productivity on scale-up from shake flasks have been
commonly attributed to the hydrodynamic stresses associated with aeration and agitation
in bioreactors. Many of the shear sensitivity studies performed on plant cell suspensions
have been highly specific, both in terms of cell line and operating conditions. However,
they clearly reveal that susceptibility to shearing is not only line-dependent (e.g. Meijer,
1989) but is also strongly influenced by culture age, history and maintenance conditions.
There is evidence that cultures may become adapted to high shear conditions (e.g. Allan
et al., 1988), possibly through regulation of cell wall synthesis (Tanaka et al., 1988) or by
preferential disruption of aggregates in a size-based sub-population which is more
susceptible to damage. Reviews of shear sensitivity in plant cell suspensions are
presented by Meijer et al. (1993) and Kieran et al. (2000). Early work focused on the
identification of conditions leading to loss of viability. More recent studies have
concentrated on sub-lethal metabolic responses (e.g. Takeda et al., 1994, 1998), which
may ultimately govern system productivity (Prokop and Bajpai, 1992; Namdev and
Dunlop, 1995). Steward et al. (1999a,b) describe a method, based on esterase activity, for
monitoring growth and lysis kinetics, as well as viability, in suspensions of Medicago
saliva L., which might be usefully extended to other cell lines.
Shear sensitivity studies of plant cell suspension cultures can be broadly categorised in
terms of the prevailing hydrodynamic environment, namely either well-or comparatively
poorly-defined. In the first case, cells are exposed to well-characterised and reproducible
flow conditions, in Couette-type, capillary or submerged-jet devices. Exposure may be
continuous (Couette-type) or intermittent (capillary, submerged jet). Shear stresses, shear
rates and, in turn, energy dissipation rates, may be many orders of magnitude higher than
those prevailing under normal cultivation conditions (e.g. Kieran et al., 1995;
MacLoughlin et al., 1998) As these devices are generally not designed for sterile
operation, exposure times are usually short and thus do not allow for system analysis
under growth conditions. Exceptions include the cultivation of Pirus malus suspensions
in a 2 L sparged, concentric cylinder bioreactor (Soule et al., 1987) and Taxas cuspidata
suspensions grown in a 110 mL (wv) horizontal, rotating wall vessel with bubble-free
aeration (Sun and Linden, 1999). In the short-term studies, suspension viability is most
commonly used as a response indicator, which, in the context of bioreactor design, masks
more subtle, non-lytic effects.
Shear studies under less well-defined conditions, in bioreactors designed for extended
sterile operation, arguably offer more immediately applicable data on system scale-up
prospects. Manipulation of the hydrodynamic environment is achieved via the rate or
method of agitation and/or aeration. A strategy developed by Smith et al. (1990) for
controlling dissolved oxygen and carbon dioxide concentrations, independently of total
gas flow rate and agitation speed, allows for de-coupling of mechanical agitation and
gassing effects. However, characterisation of the flow field in a multi-phase, bioreactor is
a complex problem (Nienow, 1998). In the context of the shear sensitivity of plant cells, a
number of approaches have been adopted. For STRs, the simplest analyses are based on
Multiphase bioreactor design 428