Biomedical Engineering Reference
In-Depth Information
5.2 APPROACH TOWARD PROCESS CHARACTERIZATION
The objective of process characterization studies is to evaluate the robustness of the
process and to define the “design space” within which the process can operate and still
perform in an acceptable fashion with respect to product quality and process consistency
[4, 5]. Data generated in these studies serves as a basis for the following:
1. Categorization of parameters into critical (those that impact product quality), key
(those that impact process consistency), and non-key (those that are neither
critical nor key).
2. Identification of acceptable ranges (ARs) that define the design space.
3. Establishing acceptance criteria for process validation.
Technical information from the characterization study has become a regulatory
expectation in recent years as an important precursor a pre-requisite for manufacturing
process validation as well as for the long-term manufacturing support [13]. Since
performing a characterization study at the manufacturing scale is not practically feasible
due to cost of operation and limited availability of facility and equipment, small scale
models that represent the performance of manufacturing scale process are usually
employed inprocesscharacterizationstudiesat the laboratoryscale.Theoverall procedure
of process characterization is shown in Fig. 5.1.
The general process flow diagram for the case study is shown in Fig. 5.2. In brief, the
case study involves a recombinant human therapeutic protein that is produced in
Escherichia coli grown in a complex medium under the control of a temperature--
sensitive promoter. This protein is expressed intracellularly as insoluble inclusion bodies
in high cell density fed-batch fermentations. This protein is expressed as intracellular,
insoluble form and this allows removal of several fermentation process contaminants by
centrifugation and washing of the inclusion bodies suspension during the downstream
operation. The manufacturing process begins with an inoculum scale-up from a shake
flask to a seed fermenter, which is then used to seed the production fermenter. Three
phases occur during the production fermentation. After inoculation of the production
fermenter, the culture grows in batch mode to a target optical density. By then, glucose in
the fermentation broth is nearly depleted, and the fed-batch phase begins with the
addition of Feed-1 solution. During the fed-batch phase, the agitation speed is fixed. The
Feed-1 solution is added at an exponential rate to provide sufficient nutrients for
exponential cell growth until the target OD is reached. Once the cell density reaches
this point, Feed-1 addition is discontinued, and the induction phase begins with changes
in temperature to activate the temperature-sensitive promoter and induce protein
expression. During the induction phase, the Feed-2 solution is added at a fixed rate
to provide additional amino acids for product synthesis. After a fixed period, the
induction phase ends. The fermentation broth is then chilled. Subsequently, microfiltra-
tion/diafiltration and disc-stack centrifugation steps are performed to generate cell paste
intermediate (CPI) that is stored frozen until the start of the purification process up to the
final formulated drug product. In the following sections, we use this case study to
