CHAPTER 4

PACKED BED BIOREACTORS

J. N. WARNOCK, K. BRATCH AND M. AL-RUBEAI

Department of Chemical Engineering, University of Birmingham, Birmingham, U.K.

1. INTRODUCTION
Intensive cell culture systems, such as packed bed bioreactors, have attracted
considerable interest for the commercial production of biopharmaceutical products
from mammalian cells (Thelwell and Brindle 1999) and their potential for long term
cultivation of high cell densities makes them an attractive option for tissue
engineering applications. The large cell numbers obtained can subsequently be used
for transplant, or the bioreactor can serve as a surrogate organ such as a bioartificial
liver (BAL) (Jauregui et al. 1996). However, to fully utilise these systems a
comprehensive understanding of the influence of reactor design on cell growth and
physiology is required (Banik and Heath 1995; Knight 1989).
The principle behind a packed bed bioreactor (PBR) is that cells are immobilised
within a suitable stationary matrix that forms the bed. Several types of matrices have
been used, including macroporous microcarriers (Fassnacht et al. 1999; McTaggart
and Al Rubeai 2000), porous ceramic beads (Park and Stephanopoulos 1993),
porous glass beads (Racher and Griffiths 1993), glass fibres (Rodrigues et al. 1999),
and polyester discs (Hu et al. 2000; Kadouri and Zipori 1989; Kaufman et al. 2000;
Wang et al. 1992; Warnock 2003). These provide a large surface area to allow cell
attachment, resulting in cell concentrations as high as 5 x 108 cells ml-1 of matrix
(Park and Stephanopoulos 1993). The porous structures also offer a protective
environment for cells from local shear forces (Cong et al. 2001). Culture medium is
perfused through the bed to supply cells with nutrients and to remove toxic
metabolites. This allows the reactor to be run in batch, fed-batch or a continuous
mode of operation, enabling long term cultivation of cells, and production runs in
excess of 30 days are not uncommon (Cong et al. 2001; Ducommun et al. 2001;
Fassnacht et al. 1999; McTaggart 2000).
The main problem associated with PBR is the heterogeneity of the reactor caused
by concentration gradients of nutrients and waste products (Chresand et al. 1988;

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J.B. Chaudhuri & M. Al-Rubeai (eds.), Bioreactors for Tissue Engineering, 87 - 113.
© 2005 Springer. Printed in the Netherlands.
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RUBEAI

Drury et al. 1988; Piret and Cooney 1990; Piret and Cooney 1991). These will have
an adverse effect on the function of a bioartificial
r tissue, such as a BAL (bioartificial
liver), where the metabolic activity of the cells is intrinsically linked to their
function (Thelwell and Brindle 1999). Transport limitations are the main limiting
factor in the linear scale up of PBRs. Additionally, direct sampling of cells is often
not possible (Ducommun et al. 2001; Kaufman et al. 2000) which causes problems
in the calculation of culture parameters such as specific growth rate. Instead, cell
numbers are estimated indirectly from either the glucose or oxygen uptake rates or
the lactate production rate (Mancuso et al. 1990; Pörtner et al. 1994; Rodrigues et
al. 1999). Unfortunately, these parameters can vary greatly with changes in growth
conditions (Lin et al. 1993; Ljunggren and Häggström 1994; Neermann and Wagner
1996). PBRs are often used to culture anchorage dependent cells. Therefore, cells
for inoculation are prepared from roller bottles. Hence, the development of large-
scale systems is difficult due to the number of cells required for seeding (Cong et al.
2001).
Since the development of the packed bed bioreactor by Spier and Whiteside
(1976), many advances have been made. This review will present the general
characteristics of packed beds with respect to mass transport, fluid flow and changes
in pressure over the bed and how these affect
f cellular activity. It also describes the
various types of reactors that have been developed and their application to the
engineering of living tissues.

2. MASS TRANSPORT
Adequate mass transfer is necessary in packed bed bioreactors in order to achieve
maximal cell growth. It is also desirable that no detrimental effects, such as damage
from hydrodynamic forces, result from techniques used to achieve efficient transport
of nutrients. Proficient mass transfer is easily attainable at laboratory scales but the
nutrient and metabolite gradients that develop often limit the scale-up of
immobilised cell bioreactors (Piret and Cooney 1991). The environment surrounding
a cell will determine the level of cellular activity. Hence, mass transfer often has a
decisive role in the selection of a suitable bioreactor (Bratch 2000).
There are two criteria for mass transport in a bioreactor. Firstly, the system must
be capable of transferring nutrients to cells. Cells require glucose as their primary
carbon source for energy, as well as various other nutrients contained in culture
medium. In addition, oxygen concentration will affect cellular viability and function.
Secondly, glucose is metabolised into pyruvate, which in turn is converted into
lactate. Similarly, L-glutamine is metabolised into ammonia. At high concentrations
these may become toxic and it is therefore necessary that they be transported away
from cells. Transport to and from cells is especially important in the design of a
BAL assist device as it needs to ensure the successful detoxification of blood borne
endogenous toxins, the unhindered return of liver specific factors back into patient’s
blood and the long term maintenance of cell viability and function (Bratch 2000).
Oxygen and glucose limitations have both been correlated with necrotic regions
in tumours (Tannock 1972; Vaupel 1977), as well as spheroids of packed cells