Lyophilization, or freeze drying, is an important manufacturing step widely used to increase the stability of pharmaceutical
and biological products. Developed during the 1940s, lyophilization produces a dry product that can be readily reconstituted
to its original form by adding water when required (1). It prolongs product shelf life by inhibiting chemical, microbiological,
and physical degradation pathways that occur in the presence of moisture, particularly where long periods of storage and transit
prior to use are involved (1, 2).
PHOTO 24/MARIA TOUTOUDAKI/GETTY IMAGES; DAN WARD
The lyophilization process consists of three main stages—freezing (solidification), primary drying (ice sublimation), and
secondary drying (moisture desorption)—and usually takes several days to complete. Multiple vials containing a liquid drug
formulation are loaded on temperature-controlled shelves within a sterile chamber and cooled to low temperatures until completely
solidified (2). After that, chamber pressure is reduced and shelf temperature is raised to remove the frozen solvent through
sublimation. The remaining unfrozen solvent that is chemically bound to the solid product is removed by a desorption process
(3). The drying process is concluded by stoppering the vials in the chamber, generally under a subambient pressure of inert
gas. The final dry product, called a cake, usually occupies approx-imately the same volume as the initial liquid fill because
of its high porosity (2). "To ensure that high quality products are consistently produced, it is crucial to be able to control
and provide repeatability of the lyophilization cycles," says Joseph Brower, technology manager at IMA Life North America.
The freezing step
The freezing step is one of the most important steps in lyophilization because it determines the texture of the frozen material
and consequently, the final morphological characteristics of the freeze-dried material and its biological activity stability
(4). "Proper freezing creates the foundation for efficient and consistent freeze-drying cycles," says T.N. Thompson, president
of Millrock Technology, a company that develops freeze drying/lyophilization systems for laboratory applications and cGMP
The three steps in the freezing process are nucleation, crystallization of the freeze concentrate, and for the maximal freeze
concentrate, either freeze separation in eutectic products or concentration in amorphous products. The parameters of the freezing
protocol directly affect pore size distribution and pore connectivity of the porous network of the freeze-dried matrix. The
ice-crystal morphology determines both mass and heat transfer rates through the dry layer and as a result, freezing parameters
have a strong influence on the total duration of the primary and secondary drying steps (4).
The nucleation process
During the freezing phase of a typical freeze-drying cycle, the nucleation process of which the first solid domains are formed
occurs randomly in the vials. "In an uncontrolled environment, due to the lack of nucleation sites in pure systems, the formulation
solution must be cooled down to temperatures that are significantly lower than the equilibrium freezing point (i.e., supercooled)
to initialize formation of ice crystals," Brower explains.
The contents of individual vials often nucleate or begin freezing over a broad range of temperatures, "usually spanning 10–15
°C below the formulation's thermo-dynamic freezing point in a laboratory freeze dryer and 20 °C or greater in a cGMP Class
100 production dryer," says Mark Shon, vice-president of sales and marketing at SP Scientific. This supercooling phenomenon
creates significant vial-to-vial heterogeneity in the solid microstructure, which significantly affects the subsequent drying
processes. "To accommodate this heterogeneity, today's best practice is to design lyophilization processes for the worst-case
scenarios; however, this strategy can result in excessively long drying cycles, broad product specifications, longer process
development times, and nonoptimal product preservation," comments Cheryl Thierfelder, Praxair's business development manager.
It has long been recognized that one of the most important goals during the freezing step of lyophilization is to produce
a uniform batch, which is a challenge due to the stochastic nature of nucleation. The random nature of nucleation, however,
makes it difficult to control the nucleation temperature and maintain it within the desirable supercooling range.