The freezing step

The extent of solid dehydration achieved with freeze-drying depends on the particular type of material and on the temperature reached by the sample during the freezing step. In any case, some water (containing dissolved solids) will always remain unfrozen in the sample. 

Effectively preserving the initial structure of the frozen sample relies on a combination of stiffness in the frozen and unfrozen regions. Ice is quite resistant to deformation, so it acts as a support for the unfrozen region — the stiffness of which is more widely variable. The amount of ice formed and that of material which remains unfrozen depend on the sample temperature and the initial solid concentration in the sample, which are thus the two variables most markedly influencing the efficiency of ice as a support. 

---

Lyophilization is a very common method to prepare enzymes for use in organic media, but the procedure often results in preparations having low catalytic activity. The method can cause inactivation of the enzyme both in the freezing step and in the drying step [7]. FT-IR spectroscopy has been used to study secondary structure in enzyme preparations, and lyophilization has been shown to decrease the a-helix content and increase the (i-sheet content compared to native enzyme and... 

Allison, S.D., Molina, M.C., Anchordoquy, TJ, (2000). Stabilization of lipid/DNA complexes during the freezing step of the lyophilization process the particle isolation hypothesis. Biochim. Biophys. Acta, 1468, 127-138. 

Even in an imaginary idealized form, the process would involve heat transfer through one wall and two films, while a process using expansion of a light hydrocarbon in direct contact with saline liquor for the freezing step, and also direct contact of the hydrocarbon and ice for the melting step, would involve only two films. Detailed calcu-... 

In any event, the freezing step is considered to be most critical in the multi-step processes, and it should be carried out according to well-defined methods specifically designed for each individual species. 

In some cases, freezing or freeze-drying a highly concentrated solution may prove difficult and require diluting the sample prior to freezing so that an adequate amount of ice is formed in the freezing step. 

The sublimation step removes the water molecules that were separated from the solids as ice crystals in the freezing step. Sublimation is the process by which solid water is directly transformed into vapour water. When sublimation occurs during the freeze-drying step, it is commonly referred to as the primary drying process . The sublimation step does not affect unfrozen water associated to the solids (sorbed water), which is removed during the desorption step. 

The sublimation step separates the ice crystals formed during the freezing step. When an ice crystal forms, that which remains is the concentrated solute phase called the dry layer . This will become the freeze-dried material at the end of the process. Immediately following passage through the interface, however, the solids in the dry layer still contain a substantial amount of water (about 25-30 g per 100 g of solids), which continues to be strongly bound to the solids. Most sample materials will not be structurally or chemically stable unless most of this water (called sorbed water ) is removed. The process... 

The freezing step is conducted at atmospheric pressure. It is crucial to ensure that the product will be thoroughly frozen by the end. This entails maintaining the temperature below the eutectic or collapse point of the product. 

This step is intended to reduce residual moisture to levels allowing no microbial growth or chemical reactions of the end product. The amount of residual moisture present in a product depends on its desorption isotherms. Such isotherms in turn depend on various factors including the product temperature, pressure chamber, partial vapour pressure in the container and nature of the interaction of the water vapour with the interstitial material formed in the freezing step. The computer should be fed with information on the target sample component. For example, if the component of interest is a protein, then overdrying may alter its configuration and decrease the potency of the end product. Consequently, the computer should control not only the final product temperature but also the partial water vapour pressure and the duration of the desorption step. 

The solubility of salts in water affects the freezing and drying steps. If the salt is sparingly soluble, then large quantities of water must be sublimed to dry the salt, an expensive process that may lead to excessively dusty powder. On the other hand, if the salt is highly soluble, ice crystals will not precipitate during the freezing step rather, the... 

More recently, in an attempt to make this cell culture system more user friendly, this model was adapted in a ready-to-use, frozen format. Because of the freezing step, the final user does not need specific knowledge of brain endothelial cell culture and subculture. The time necessary for the whole experiment is substantially reduced, and the new model is ready for use 4 days after thawing (Fig. 2c) [21]. 

---