Chapter 37
Freeze-Drying of Proteins
Baolin Liu and Xinli Zhou
Abstract
Freeze-drying has become one of the most important processes for the preservation of biological products.
This chapter provides protocols for freeze-drying of proteins and discusses the importance of formulation,
cycle development, and validation. Specific formulations for stabilization of proteins are presented as well as
advice on common problems with freeze-drying of proteins.
Key words Freeze-drying, Proteins, Formulation, Stabilizer, Lyophilization
1 Introduction
Proteins are becoming an important class of drugs in human health
care. However, their marginal stability always presents challenges in
their development for pharmaceutical and other industrial use.
Since many proteins do not have adequate long-term storage stability in the aqueous state, a solid-state dosage form is preferred.
Freeze-drying is most commonly used to prepare dehydrated
proteins, in the areas of protein purification, protein reagent preparation, and manufacturing of protein biomolecules for therapeutic
and diagnostic applications. Freeze-dried proteins have long-term
stability at ambient temperatures, which offers advantages for storage as well as for shipping and distribution.
1.1 Theory of FreezeDrying
The freeze-drying process primarily consists of three stages. The
first stage is freezing, which involves freezing the product and
creating a solid matrix suitable for drying. The second stage is
primary drying, sometimes preceded by an additional step called
annealing or thermal cycling. Primary drying involves the removal
of ice through sublimation by reducing the pressure of the product
environment while maintaining the product temperature at a low
target level. The third stage in the process is called secondary
drying, in which bound water is removed until the residual moisture content reaches its targeted level. Although freeze-drying is
Willem F. Wolkers and Harrie ¨ tte Oldenhof (eds.), Cryopreservation and Freeze-Drying Protocols, Methods in Molecular Biology,
vol. 2180, https://doi.org/10.1007/978-1-0716-0783-1_37, © Springer Science+Business Media, LLC, part of Springer Nature 2021
683
Freeze-Drying of Proteins
Baolin Liu and Xinli Zhou
Abstract
Freeze-drying has become one of the most important processes for the preservation of biological products.
This chapter provides protocols for freeze-drying of proteins and discusses the importance of formulation,
cycle development, and validation. Specific formulations for stabilization of proteins are presented as well as
advice on common problems with freeze-drying of proteins.
Key words Freeze-drying, Proteins, Formulation, Stabilizer, Lyophilization
1 Introduction
Proteins are becoming an important class of drugs in human health
care. However, their marginal stability always presents challenges in
their development for pharmaceutical and other industrial use.
Since many proteins do not have adequate long-term storage stability in the aqueous state, a solid-state dosage form is preferred.
Freeze-drying is most commonly used to prepare dehydrated
proteins, in the areas of protein purification, protein reagent preparation, and manufacturing of protein biomolecules for therapeutic
and diagnostic applications. Freeze-dried proteins have long-term
stability at ambient temperatures, which offers advantages for storage as well as for shipping and distribution.
1.1 Theory of FreezeDrying
The freeze-drying process primarily consists of three stages. The
first stage is freezing, which involves freezing the product and
creating a solid matrix suitable for drying. The second stage is
primary drying, sometimes preceded by an additional step called
annealing or thermal cycling. Primary drying involves the removal
of ice through sublimation by reducing the pressure of the product
environment while maintaining the product temperature at a low
target level. The third stage in the process is called secondary
drying, in which bound water is removed until the residual moisture content reaches its targeted level. Although freeze-drying is
Willem F. Wolkers and Harrie ¨ tte Oldenhof (eds.), Cryopreservation and Freeze-Drying Protocols, Methods in Molecular Biology,
vol. 2180, https://doi.org/10.1007/978-1-0716-0783-1_37, © Springer Science+Business Media, LLC, part of Springer Nature 2021
683