“pH Swing” in Frozen Solutions—Consequence of Sequential Crystallization of Buffer Components

Succinate buffer solutions of different initial pH values and concentrations were cooled. The solution pH and the phases crystallizing from solution were monitored as a function of temperature. In a solution buffered to pH 4.0 (200 mM), the freeze-concentrate pH initially increased to 8.0 and then decreased to 2.2. On the basis of X-ray diffractometry (synchrotron source), the “pH swing” was attributed to the sequential crystallization of succinic acid, monosodium succinate, and disodium succinate. A similar swing, but in the opposite direction, was seen when a solution with an initial pH of 6.0 was cooled. In this case, crystallization of the basic buffer component occurred first. The direction and magnitude of the pH shift depended on both the initial pH and the buffer concentration. In light of the pH-sensitive nature of a significant fraction of pharmaceuticals (especially proteins), extreme care is needed, both in the buffer selection and in its concentration.