Phase Transitions During Freezing and Freeze-Drying

When prelyophilization solutions are frozen, selective crystallization of a buffer component is known to cause a pH shift, which can sometimes be pronounced. Such pH shifts have been reported with both phosphate and succinate buffers. The presence of a noncrystallizing solute, sucrose or a monoclonal antibody, can prevent salt crystallization and mitigate the pH shift. In order to obtain an elegant lyophile, crystallization of the bulking agent is desired. Isothermal hold during cooling (annealing) maximized the mannitol crystallization and resulted in a homogenous freeze-concentrate of a constant composition characterized by a single glass transition temperature. Thus, in addition to a sugar as a stabilizer, the use of a crystallizing excipient coupled with an annealing step can provide an avenue for frozen storage of proteins. Mannitol can also crystallize as a hemihydrate which can then dehydrate on storage of the final lyophile. While the dehydration of the hemihydrate facilitated sucrose crystallization, there was no protein aggregation. On the other hand, while arginine hydrochloride, when retained amorphous in frozen solutions effectively inhibited protein aggregation, its stabilizing function was lost in the presence of mannitol wherein both solutes crystallized. Use of alternate arginine salts, allowed selective mannitol crystallization while arginine remained amorphous and exerted its stabilizing function. The ability of poloxamer 188 to decrease the interfacial stress and stabilize protein was enhanced in the presence of a sugar, possibly due to inhibition of surfactant crystallization. Presented by Dr. Raj Suryanarayanan, PhD. College of Pharmacy, University of Minnesota