Review

Advances in product release strategies

and impact on bioprocess design
Bangaru Balasundaram1, Sue Harrison2 and Daniel G. Bracewell1
1

The Advanced Centre for Biochemical Engineering, Department of Biochemical Engineering, University College London,
Torrington Place, London WC1E 7JE, UK
2
Centre for Bioprocess Engineering Research, Department of Chemical Engineering, University of Cape Town 7701, South Africa

Intracellular products such as recombinant insulin,

which are typically produced in microbial host cells,
demand a product release step to remove them from
the cell. How this is performed determines the quantity
of released contaminants, the particle size distribution of
cell debris and the physical properties of the resultant
process stream, which all impact on the performance of
the downstream operations. Thus, achieving selective
release of the desired product is crucial for improving the
process economics. Advances in upstream processing
(the bioreactor phase) have been successful in achieving
high product titres, and downstream costs now typically
dominate the overall manufacturing costs. Here, we
review and discuss the selective release of products as
a possible means of improving the efficiency of downstream processing.
Cell disruption in the context of an entire bioprocess
Biologics-based medicines, such as hormones (e.g. human
growth hormone), vaccines (e.g. against hepatitis B) and
monoclonal antibodies (e.g. for the treatment of rheumatoid arthritis), are projected to form an increasing proportion of new products and hence are becoming
increasingly important to large and small pharmaceutical
companies. This is exemplified by the large number of
biopharmaceutical products (44% of all pharmaceutics)
that are currently in various phases of investigation
[1,2]. These biopharmaceuticals are usually manufactured
using mammalian, fungal or bacterial production systems
but are associated with inherently significant manufacturing costs owing to the regulatory requirements aimed at
ensuring their safety and efficacy. Downstream processing
(in which the products are purified) accounts for a large
proportion of the production costs. For proprietary reasons,
literature on the optimization of the biopharmaceutical
production process is scarce; however, reports using surrogate systems (typically yeast and Escherichia coli) and
endogenous enzymes as model proteins that mimic the key
features of the process can be found.
Cell engineering has been successfully applied to
increase product titres during the cell culture phase [3].
However, the economic gain through increases in the
product titres in the bioreactor cannot be directly passed
on to downstream processing, when operational steps,
such as chromatography and associated factors including
buffer volumes, sizes of ancillary vessels, sanitization and
Corresponding author: Bracewell, D.G. (d.bracewell@ucl.ac.uk)

infrastructure, need to be scaled up in proportion to the

increased product titres. Thus, improvements in bioreactor
productivity have caused a bottleneck in downstream processing and means of improving the overall process
economics are being sought [4], as discussed in several
recent reviews on this subject [59]. Most papers focus
largely on the chromatography step and the search for
potential cost-saving alternatives, whereas Roush and Lu
highlighted the potential of membrane separations [9].
Apart from increasing product titres, process economics
could also be improved by selectively recovering the products during the initial recovery steps; via this method,
some of the key contaminants that are frequently presented
to the subsequent purification operations could be eliminated. In the case of an intracellular product, this could be
achieved by releasing the product selectively during the first
recovery operation, cell disruption. In this article, selective
product release (SPR) is defined as the ratio between
released product and contaminants (mg of product/gm of
contaminants). In yeast and Gram-negative bacteria, such
as E. coli, there is a region of space in between the cytoplasmic membrane and the outer cell wall that is termed the
periplasm (Figure 1). The products located in the periplasm
can be released selectively by interrupting the integrity of
the outer cell wall while keeping the cytoplasm intact.
This selective release of products has several benefits: (i)
it reduces the release of contaminant proteins, thus
increasing the adsorption capacity during chromatography; (ii) it avoids the micronization of cell debris with
concomitant benefits for clarification and filtration steps
(micronization is a phenomenon encountered typically
with mechanical methods of cell disruption, where the cell
debris generated in the initial stages of disruption is
broken down into finer particles during the prolonged
disruption times required to maximize product recovery);
and (iii) it avoids the release of DNA, thus preventing an
increase in viscosity of the process stream (low viscosity
benefits most of the operational steps involved in purification). Taken together, these advantages make selective
product recovery during cell disruption one of the most
promising means of improving the process economics for
intracellular products.
Current cell disruption strategies
The need for cell disruption
S. cerevisiae and E. coli are widely used production
hosts for a variety of recombinant proteins [10], although

0167-7799/$ see front matter 2009 Elsevier Ltd. All rights reserved. doi:10.1016/j.tibtech.2009.04.004 Available online 1 July 2009

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Figure 1. Compartmentalization of products in yeast and Gram-negative bacteria as hosts. Examples of products and the corresponding cell disruption techniques used for
their release are shown. Abbreviations: ER, endoplasmic reticulum; IB, inclusion bodies; S, secretory vesicles.

frequently the products of interest are not secreted into the

culture medium, which would be ideal, but are instead
retained at various locations within the cell, as shown in
Figure 1. Genetic engineering has been employed in an
attempt to force the secretion of the recombinant proteins
into the extracellular medium by various means, such as
co-expression of the kil gene (which induces cell lysis upon
induction) and the generation of leaky mutants, among
others [11]. This strategy was found to be more useful for
transporting the product actively across the cytoplasmic
membrane than for passive transfer across the outer cell
wall [11,12]. The disadvantages of such protein secretion
systems include a reduced stability of the secreted protein
in the bioreactor environment, possible entrapment of the
protein in the periplasmic space, economically unviable
levels of secretion and lack of glycosylation (when using a
prokaryotic host such as E. coli). Consequently, attempts to
secrete the product to the extracellular medium when
using yeast or E. coli as hosts have not attracted significant
commercial interest [13]. Hence either partial or complete
cell disruption remains inevitable for the release of recombinant products.
Criteria for an optimal cell disruption process
The physical characteristics of the suspension that results
from cell disruption have a significant impact on the downstream processing operations involved (Figure 2). The
particle size of the cell debris, dp, strongly influences
the particle settling velocity, y, in the centrifuges
(y / d2p ), and the viscosity of the suspension is inversely
related to the particle settling velocity. Attempts to maximize product recovery through increases in the intensity of
478

cell disruption usually result in micronization of cell debris

and increased viscosity of the product stream caused by the
release of DNA and the co-release of contaminants. Micronization of cell debris reduces the efficiency of particulate
removal during the subsequent centrifugation step [14,15].
The increase in the release of DNA with increasing cell
disruption [16] translates into difficulties in the following
solidliquid separation steps, such as a reduction in flux
during membrane filtration [17]. In addition, the contaminant load, which is formed as a result of intense disruption
conditions breaking down cellular structure, impacts on
the performance of chromatography, adversely affecting
not only the separation performance but also other key
factors, such as the ease of sanitization and the lifetime of
the column. Thus, based on the above discussion (summarized in Figure 2), cell disruption should be optimized for the
competing requirements of all unit operations in a process.
Cell disruption would thus include the selective and efficient release of the product to achieve high recovery,
reduced contaminants and minimal micronization of cell
debris. This frequently means that a trade-off between
relevant parameters, such as product and contaminant
release, must be accepted.
Current cell disruption techniques
Cell disruption can be achieved by a variety of means
[10,18], and available methods can be classified broadly into
mechanical and non-mechanical methods. Furthermore,
mechanical methods can also be combined with non-mechanical methods to increase the overall efficiency of cell
disruption [19,20]. Mechanical methods include bead mills,
high-pressure homogenizers and cavitation, which can be

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Trends in Biotechnology

Vol.27 No.8

Figure 2. Generic flow sheet for purification of intracellular recombinant therapeutic proteins from microbial host cells, indicating the impact of product release on whole
bioprocess. Abbreviations: AEX, anion exchange; CEX, cation exchange; HIC, hydrophobic interaction chromatography. In the case of an insoluble product, such as
inclusion bodies, an additional resolubilization step is included after cell disruption.

generated through either sonication or fluid flow (termed as

hydrodynamic cavitation) [18]. Non-mechanical methods
include chemical treatments, cell lysis using enzymes, heat
or alkaline conditions, osmotic shock, freeze/thaw cycles or
cell engineering using tightly regulated genes, such as the
kil gene [11,12].
Techniques used at process scale
Although a variety of techniques are available for the
release of intracellular products, mechanical methods have
found greater commercial application compared with nonmechanical methods, because the latter infer operational
and economic limitations at process scale. Bead mills and
high-pressure homogenizers tend to be used in large-scale
microbial cell disruption, whereas other mechanical and
non-mechanical methods are predominantly used at the

laboratory scale, notable exceptions being heat lysis for the

release of periplasmic antibody fragments [21] and
alkaline lysis for the release of cytoplasmic plasmid
DNA [22]. The disadvantages of the use of bead mills
and homogenizers include a non-selective release of the
product (i.e. the co-release of contaminants) and micronization of the cell debris.
Ultrasonication is currently not used for large-scale cell
disruption because of the difficulty in transmitting sufficient
power to large volumes of cell suspension [23]. In addition,
most of the energy that is absorbed by the cell suspension
during ultrasonication is transformed into heat, thereby
challenging the ability to control the temperature. Hydrodynamic cavitation, on the other hand, is a relatively new
technology that so far has been mainly studied at pilot scale
[24], and its applications are currently in the research stage.
479

Review
Chemical and enzymatic methods have been limited to
laboratory scale mainly because of the costs of the chemicals or enzymes required for disruption at large scale but
also because of difficulties in removing exogenous chemicals from the product of interest or the potential denaturation of the product on exposure to specific chemicals [25].
For instance, in a large-scale process for the production of
polyhydroxybutyrate from Cupriavidus necator (previously known as Alcaligenes eutrophus), mechanical
methods were found to be superior to chemical lysis
because the latter required large amount of chemicals
and also longer processing time [18,26,27].
The relative selectivity and recovery that can be
achieved with the above-mentioned methods of cell disruption is summarized in Figure 3. Although the industrially
preferred mechanical methods, such as bead mills and
high-pressure homogenizers, are superior in terms of product recovery, their poor selectivity is a major drawback.
Taken together, it is apparent that an industrially
applicable method for product release with enhanced selectivity and reduced micronization will be vital to address
the future needs of the bioprocess industry.
Optimization of upstream processing for efficient cell
disruption
Microbial cells in the rapid-growing phase (i.e. those that
are cultivated at higher growth rate) have been reported to
disrupt more easily than cells in the stationary or slowgrowing phase [25,27,28]. By contrast, cells cultivated at
low growth rate were resilient to disruption [29,30], most
likely due to an increased production of peptidoglycans,

Trends in Biotechnology Vol.27 No.8

which results in stronger cell walls [28,3133]. Although

lower growth rates typically promote product formation
and also help to reduce a leakage of periplasmic products
into the culture media, the resilience of these cells to
product liberation, which implies the need for enhanced
lysis conditions, and the potential accumulation of endotoxins need to be considered when attempting to optimize
the production process [34].
Impact of cell disruption on subsequent purification
steps
As mentioned above, cell disruption has a significant
impact on other subsequent unit operations (see
Figure 2), and its detailed effects on centrifugation, filtration and chromatography are further discussed below.
Agerkvist and Enfors [15] reported that increasing the
intensity of disruption of E. coli cells by increasing the
number of homogenization cycles [15] from one to three
increased the product recovery from 58% to 78%, while at
the same time the mean particle size of the cell debris was
reduced from 0.5 mm to 0.2 mm. This increased micronization of cell debris resulted in a 20% reduction in the
sedimentable solids during centrifugation [35].
Studies have also shown that increasing the number of
homogenization cycles will lead to an increase in the
viscosity of the homogenates due to the increased release
of glycans from yeast cell walls [36] and of DNA from the
nuclei of bacterial and yeast cells [15,31]. However, if the
number of homogenization cycles is increased beyond what
is required for maximum product release, the viscosity of
the suspension will decrease due to shear damage, albeit

Figure 3. Relationship between the selectivity of product release and the ease of product recovery for various disruption techniques based on literature reports.

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Trends in Biotechnology

with a simultaneous micronization of cell debris [15],

which as discussed above has undesirable effects on the
process. For example, the recovery of inclusion bodies from
E. coli, which were pretreated with ammonia before homogenization, was found to generate cell debris that was at
the limit of separation by centrifugation [37]. In another
study, increasing the detergent concentration for the
recovery of virus-like particles (VLPs) from the endoplasmic reticulum of yeast was found to result in a co-release of
contaminant lipids that fouled the membrane and
increased its resistance during the subsequent ultrafiltration step [38]. Finally, the influence of the particle size of
the cell debris during expanded bed adsorption chromatography (EBA) has been reported in several studies [39
41]. Because intact cellular structures tend to adsorb to
the chromatography media to a greater extent than finer
cell debris, they will interfere with product adsorption
[39,40]. The benefits of a periplasmic location of the product for achieving selective product release, which minimizes the above-mentioned adverse effects, are discussed
below.
Advantages of a periplasmic product location
In the production of heterologous proteins, the gene encoding the product of interest can be cloned along with a signal
sequence to transport the product from the cytoplasm to
the periplasm in Gram-negative bacteria. The periplasmic
location of proteins in bacteria has several advantages. The
periplasm contains only seven out of 25 known cellular
proteases and comprises only 4% to 8% of total cell
protein (potential contaminants) [42]. Furthermore, the
oxidative environment of the periplasm facilitates correct

Vol.27 No.8

disulfide bonding and protein folding and, moreover,

intense disruption conditions or a complete disruption of
the cell are not required because this outer compartment
can be accessed easily by only damaging the outer cell wall.
During cell disruption, the contents of the periplasm are
released faster than those of the cytoplasm [43]. The ratio
between the release rate of periplasmic products and the
release rate of the total soluble protein (as a marker of
cytoplasmic contents) has been termed the location factor
and was used by Balasundaram and Pandit [44] to systematically compare the efficiency of different cell disruption
methods for selective product release. For example, the
location factor values for the release of periplasmic penicillin acylase from E. coli were found to be 1.736 for
sonication and 1.481 for high-pressure homogenization
at 5000 psi. The higher value for sonication was caused
by a faster release of periplasmic enzymes, thus indicating
a more selective release by sonication than by homogenization [44].
Therefore, an optimal two-phase strategy for the production of recombinant proteins would be to: (i) incorporate
a translocation signal into the DNA sequence of the host
(e.g. E. coli or S. cerevisiae) to direct the desired product to
the periplasm; and (ii) selectively disrupt the host cells to
recover the product without releasing significant amounts
of contaminant cytoplasmic proteins [33]. Recent results
and important advances of this approach are further discussed below.
Selective product release
Traditionally, unit operations involved in the purification
of intracellular products are optimized individually

Table 1. Selective release of cellular components by mechanical disruption techniques

Microorganism Enzyme (location)
Bead mill
Invertase (cell wall bound), a-glucosidase (periplasmic),
S. cerevisiae
ADH (cytoplasmic), fumarase (mitochondria)

S. cerevisiae

Invertase (cell wall bound), acid phosphatase (periplasmic),

ADH (cytoplasmic), alkaline phosphatase (cytoplasmic
membrane)

S. cerevisiae

Invertase (cell wall bound), G6PDH (cytoplasmic membrane)

S. cerevisiae

G6PDH (cytoplasmic), fumarase (mitochondria)

S. cerevisiae

a-Glucosidase (periplasmic), G6PDH (cytoplasmic)

S. cerevisiae

G6PDH

High-pressure homogenization
Invertase (cell wall bound), acid phosphatase (periplasmic),
S. cerevisiae
G6PDH and 6PGDH (cytoplasmic), alkaline phosphatase,
(cytoplasmic membrane bound), fumarase (mitochondrial)
Ultrasonication
Acid phosphatase (periplasmic), b-galactosidase (periplasmic),
E. coli
G6PDH (cytoplasmic)
Hydrodynamic cavitation
Invertase (cell wall bound), a-glucosidase (periplasmic),
S. cerevisiae
ADH (cytoplasmic)
Acid phosphatase (periplasmic), soluble protein (cytoplasmic)
E. coli

Results and comments

Refs

Periplasmic and outer-cell-wall-bound products are

released faster, but this difference was not sufficient for
selective release. kproteins (0.029 s1) > kinvertase
(0.026 s1) > ka-glucosidase (0.023 s1) > kADH (0.023 s1) >
kfumarase (0.011 s1).
Periplasmic and outer-cell-wall-bound products are
released faster, but this difference was not sufficient for
selective release. kinvertase > kacid phosphatase > kADH >
kalkaline phosphatase
Maximum release of the soluble protein and G6PDH
in four passes, maximum invertase release in one pass
G6PDH release profile mimicked soluble protein, whereas
fumarase release was retarded owing to its mitochondrial
location
0.550.85 mm diameter beads required for cytoplasmic
product, 1 mm beads suffice for periplasmic product
Higher specific activity of G6PDH was obtained at an
agitation speed of 2300 rpm than at 3100 rpm

[45]

Release rates were in the order of acid phosphatase =

invertase > G6PDH = 6PGDH > fumarase and alkaline
phosphatase

[51]

Release selectivity of acid phosphatase > b-galactosidase

> G6PDH

[53]

39% of invertase, 27% of a-glucosidase and 10% of ADH

release was observed
90% of acid phosphatase was released, and only 45% of
soluble protein was co-released

[24]

[43]

[46]
[47]

[48]
[49]

[29]

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Trends in Biotechnology Vol.27 No.8

Table 2. Selective release of cellular components by non-mechanical methods

Microorganism Enzyme (location)
Chemicals/enzymes
Chemical methods
a-Amylase (periplasmic)
1% glycine
E. coli
Long-R3-IGF-I (cytoplasmic) Two-stage extraction with
E. coli
urea as permeabilizing agent
Penicillin acylase
Reverse micellar extraction
E. coli
(periplasmic)
with anionic surfactant
Asparaginase (periplasmic) K2HPO4 and Triton X-100
E. coli
Invertase (cell wall bound) Dithiothreitol (DTT)
S. cerevisiae
Enzymatic methods
Alkaline phosphatase
Lysozyme/EDTA
E. coli
(periplasmic)
Osmotic shock
Penicillin acylase
Osmotic shock
E. coli

and separately. However, the relationship between cell

disruption and other unit operations implies that it is
crucial to optimize this step in the context of an entire
bioprocess. This has become more and more important as
product titres have increased dramatically along with the
number and amounts of contaminants. The focus of early
downstream processing operations has thus shifted
from product isolation to bulk contaminant removal, and
this has led to recent key results for selective product
release that are discussed below and summarized in
Tables 1 and 2.

Results and comments

Refs

7080% of a-amylase from the periplasm released selectively

46% purity and 81% recovery by two-stage extraction,
41% purity and 88% recovery with conventional process
Enzyme purity increased eightfold compared with sonication,
but yield decreased to 60%
70% release of asparaginase compared with sonication
Reduction of disulfide bonds of the cell wall creates pores

[57]
[70]

93% of alkaline phosphatase

[61]

94% recovery with a specific activity of 3.9 U/mg compared

with 0.31 U/mg of protein by sonication

[60]

[71]
[58]
[55]

be insufficient for an effective fractionation, that is,

selective release was not possible with the conditions
studied. However, recently Sehdev and Spitali [52]
showed that a combination of homogenization and heat
lysis techniques has potential for the selective release of
periplasmic products from E. coli. Mild homogenization
discharge pressures were used under which no significant
cell lysis was observed (i.e. cell integrity was maintained),
which was followed by heat lysis during which the
periplasmic Fab0 antibody fragments were released.

Bead mill Several studies have investigated the effect

of the use of bead mills on different components of the
cell and have provided evidence for variation of product
release rates in dependence of their location within the
host cell; for example, cell-wall-bound and periplasmic
products are released faster than cytoplasmic products
[4549]. However, these studies were not able to suggest
particular parameters for cell disruption that might favour
the selective release of cell-wall-bound or periplasmic
products. In the case of a cytoplasmic product, hepatitis
B surface antigen (HBcAg), a bead mill operated on a
continuous mode was found to be less selective,
releasing only 0.26 mg per mg of total protein compared
with 0.3 mg per mg of total protein by high-pressure
homogenization [50]. It seems that the use of bead mill
disruption approaches has declined, which might be
connected to issues with cleaning and scale-up.

Cavitation Sonication is more selective than homogenization in terms of releasing periplasmic enzymes
[53]. The related hydrodynamic cavitation approach also
has higher selectivity than high-pressure homogenization
when used to release periplasmic products such as acid
phosphatase from E. coli [29] and a-glucosidase from
brewers yeast [24]. For instance, during the disruption
of E. coli by hydrodynamic cavitation, 90% of the acid
phosphatase was released from the periplasm and only
45% of the soluble proteins (i.e. the contaminants) were coreleased. This effect might be attributed to the cell
disruption mechanism, which constitutes pressure
fluctuations of oscillating or collapsing vapour cavities
interacting with the cell wall and acting on its exterior
surface, thus damaging the cell wall but leaving the
cytoplasm largely intact. This method has also been
applied to yeast cell invertase, a cell-wall-bound yeast
enzyme, which could be released selectively [24,54].
However, the efficiency of the release of cytoplasmic
constituents
from
yeast
requires
considerable
improvement to expand the applicability of this
technique to the large number of yeast products that are
retained in the cytoplasm.

High-pressure homogenisation High-pressure homogenization is a widely used technique for cell disruption
because complete cell disruption and maximum product
recovery can be achieved, albeit with a poor selectivity
(Figure 3). During high-pressure homogenization of S.
cerevisiae over the discharge pressure 1046 MPa,
periplasmic enzyme was found to be released faster,
followed by cell-wall-bound, cytoplasmic and membranebound enzymes in this order [51]. The differences in
the release rates of the enzymes were acknowledged to

Non-mechanical methods for selective product release

Proteins that are associated with the outer cell wall can be
released selectively by weakening the cell wall structure by
either mechanical [24] or chemical means. Experiments
have shown that the disulfide bonds in the yeast cell wall
can be reduced using common chemicals, such as dithiothreitol (DTT), and this leads to the release of proteins, for
example invertase, that are entrapped in the cell wall [55].
To selectively release periplasmic enzymes, pores could be
created in the outer cell wall. This approach has been

Mechanical methods for selective product release

Mechanical methods of product release are preferred over
non-mechanical methods in process-scale operations and
hence are discussed separately.

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demonstrated using different chemicals, such as anionic
surfactants for the release of several proteins, including
penicillin acylase [56], glycine for the release of a-amylase
[57], Triton X-100 and K2HPO4 for the release of asparaginase [58] and glycol ether for the release of a proprietary
protein product [59]. Furthermore, in E. coli, penicillin
acylase released via osmotic shock conditions had a higher
specific activity (3.9 U/mg) than penicillin acylase released
via sonication (0.3 U/mg) [60]. Furthermore, spheroplasts,
which are obtained after a partial cell wall loss, were also
able to selectively release periplasmic enzymes [61,62];
however, this method was found to be cost prohibitive at
pilot scale [62].
Future trends
Based on the above discussion, we have identified three key
areas that could provide improvement in intracellular
product release strategies in the immediate future.
Innovations in cell disruption technology
The mechanical forces involved in the traditional disruption techniques, bead mills and high-pressure homogenizers, are often very intense, resulting in a complete
rupture of the cell, which presents a greater challenge
for the selective release of a periplasmic product. Instead,
a combination of disruption techniques, such as homogenization and heat lysis, as reported by Sehdev and Spitali
[52], constitutes a feasible alternative approach for achieving selective product release. Hydrodynamic cavitation,
which has similar underlying principles to homogenization, has shown promising results for selective release
of periplasmic products [29]. Its mode of action targets the
cell envelope from the outside, thereby facilitating incremental breakage. In our view, technologies that enable cell
disruption conditions to be varied over a range at process
scale to selectively liberate products depending on their
cellular location will be required to meet the future needs
of the bioprocess industry.
Novel process strategies
Unit operations can also be integrated, so that product
release and concentration can be achieved in the same
step. This is possible through the use of aqueous two-phase
systems [63], polymers such as ethylene butyl glycol [59] or
heat lysis for a thermostable product [64]. Further work is
required to inform process design where multiple objectives or a sequence of operations is crucial.
Microscale bioprocess development
To assess the benefits of alternative process strategies, a
large number of process variables need to be investigated,
which implies a large number of experiments involving
large amounts of resources and time. Thus microscale
technologies that are informative of large-scale processes
might have a key role in the development of advanced
industrial bioprocesses [65,66]. In particular, pertaining to
the cell disruption unit operation, an innovative ultrasonication device for increased throughput has shown promising results in investigating the impact of cell disruption
on other unit operations at greater speed than previous
approaches [67].

Trends in Biotechnology

Vol.27 No.8

Looking further ahead, the field of cell engineering

holds the promise of dramatically improving our ability
to target bioproducts to certain cellular structures, and
specific organelles could even be engineered for this purpose. But as this review illustrates, those considering such
approaches would be well advised to consider the current
limitations of large-scale product release, such as corelease of contaminants and micronization of cell debris.
A successful approach could radically improve process
economics by simplification of the downstream process.
This demands a holistic approach to process design that
considers the key factors for manufacturability: for
example the reduction of viscosity [68], the removal of
product-related contaminants [69] and the elimination of
enzymes, such as proteases, that might reduce product
quality. Therefore, selective release strategies of the future
will consider the operations ability to manipulate these
attributes in addition to those of the product itself.
Acknowledgements
B.B. and D.G.B. would like to acknowledge the support for the Innovative
Manufacturing Research Centre (IMRC) for Bioprocessing housed in The
Advanced Centre for Biochemical Engineering by the Engineering and
Physical Sciences Research Council (EPSRC) under the Innovative
Manufacturing Research initiative. S.T.L.H. would like to acknowledge
support from the Department of Science and Technology (DST) and
National Research Funding (NRF), South Africa for the Bioprocess
Engineering Research Centre at the Chemical Engineering Department
of the University of Cape Town.

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