Use of a Range of Analytical Techniques in Support of the Freeze Drying of Biologicals

Use of a Range of Analytical Techniques in Support of the Freeze Drying of Biologicals
May/June 2004
Paul Matejtschuk1, Julie Fleming1,2 & Roland Fleck2
Standards1 and Cell Biology & Imaging2 Divisions, National Institute for Biological Standards & Control, UK
APR
1. The Importance of Freeze Drying in Biological Product Development
Lyophilization is a key processing stage within the manufacture of a wide range of pharmaceuticals and as such its optimization is critical [1]. Lyophilization is desirable when the biological material is labile in the liquid state, and so unacceptable losses in potency or generation of undesirable and potentially harmful degradation products might otherwise occur over time in storage. A freeze dried presentation is also more convenient in terms of delivery and storage requirements. It may also allow for reconstitution at a higher concentration, which might not be stable for use with a liquid presentation.
Successful lyophilization delivers a functionally stable and visually acceptable presentation, and a batch of lyophilized biological may be worth tens or hundreds of thousand of dollars. However, the cost of purchasing and operating production scale lyophilization equipment is substantial and the added running costs resultant from a sub-optimal process cycle can be significant when considering the economic viability of a product in development. Even more detrimental is the effect of loss of product batches when a sub-optimized process results in occasional or consistently sub-standard freeze dried product either in part or all of a batch. Even in the diagnostic or reagents markets, where the value of an individual batch is considerably less, it is important to have a lyophilization process which consistently delivers highly reproducible and acceptable product.
One of the major factors in developing a successful and robust freeze drying cycle is a good understanding of the thermal properties of the material to be lyophilized. The individual components of a formulation may have been selected for a variety of reasons:
to deliver isotonicity on reconstitution,
stabilization of biological activity during the freezing or drying process,
prevention of product deposition on the container or denaturation at liquid surface or liquid air interfaces,
maintenance of optimal pH environment.
However, the overall formulation must be compatible with the lyophilization process if a successful product is to be delivered.
Dependent upon the formulation selected and the biological component to be lyophilized, the freeze dried material may have different physical properties. Inorganic buffer salts and some excipients such as mannitol may exhibit crystalline properties, others such as proteins are usually present in an amorphous state [2]. The propensity to crystallization may be a problem during lyophilization or even on later storage [3], whereas selective crystallization may be used to remove components from the amorphous phase so allowing use of higher primary drying temperatures [4].
For some biologicals, such as lyophilized intravenous immunoglobulin, the protein will be a large component of the final composition and will significantly contribute to the thermal properties of the whole. For others, such as recombinant cytokines or hormones, it may be a small percentage of the total bulk and the thermal properties of the material to be lyophilized will be governed by the excipients and bulking agents present.
In this paper the approach taken at NIBSC over recent years in the determination of thermal properties appertaining to a wide range of biological reference materials undergoing lyophilization will be presented. Compared to a therapeutics manufacturing situation, a wide variety of different formulations are presented for lyophilization. The biological materials themselves represent a wide range of biomolecules, proteins, glycans, nucleic acids, and may be processed only once or twice over the period of a number of years. Therefore a versatile approach is required which will result in a robust solution from a relatively short period of process optimization.
2. Thermal Analysis Methods
The thermal analysis methods used in lyophilization cycle development are well-established and are applications of analytical techniques which are well-used in a broad range of small molecule pharmaceutical formulation and stability studies.
Conductivity ? The electrical resistance or conductivity of a solution, during its freezing and subsequent warming to ambient conditions, was the first method to be applied to freeze drying optimization, several decades ago [5]. As such the apparatus is simple, consisting essentially of two electrodes of opposite charge and a thermocouple juxta-positioned to reference the temperature corresponding to the resistance of the frozen material being measured.
Equipment can be home-made or commercially obtained. Commercial
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