In order to solve the mechanism about the stabilization by freeze drying the protein used for medical treatments, basic Fibroblast Growth Factor (bFGF: M. W. 17000) was used as a model protein, as well as mannitol for a crystalline freeze-drying base. bFGF was freeze dried with mannitol at concentrations ranging from 0% to 100% (w/w). Differential scanning calorimetry (DSC), Fourier transform infrared spectroscopy (FTIR) and high performance liquid chromatography (HPLC) were used to evaluate bFGF/mannitol interaction by the samples after freeze drying and storage samples through stored at 30°C, 40°C and 50°C for 10 days. The result of DSC showed that according to the ratio of bFGF and mannitol, each formulation had different crystallinity and melting points respectively. Moreover, FTIR results showed that in each formulation after freeze drying, the random structure peak of bFGF shifted to a higher wave number as the mannitol ratio increased. Also, in the storage samples, it was found that the random structure peak of bFGF shifted to a higher wave number in heat dependence. Furthermore, HPLC results showed that it was the same degradation pathway of bFGF in each formulation. This work suggests that although the interaction of bFGF and mannitol influenced the crystallinity of formulation, stabilization of bFGF with mannitol is difficult. At the same time, it is thought that an amorphous structure has contributed to the stabilization of bFGF since secondary structure change of bFGF is in inverse proportion to an amorphous quantity in formulation.
Vidarabine is an antiviral agent and marketed as a lyophilized form for injection (ARASENA-A, 300 mg/vial). Because the solubility of vidarabine is low in intravenous solutions such as isotonic sodium chloride solution and 5% glucose solution, ARASENA-A has poor solubility at room temperature, and it is therefore necessary to heat to 40°C when preparing the drug solution to achieve complete dissolution. This heating process has constituted a burden for health care professionals. Raising the buffer capacity by adjusting the pH to a slightly lower region, achieved by adding glycine and strong acids, we have produced an improved formulation that permits dissolution of 2 vials (600 mg of vidarabine) at a temperature of 20°C or higher in 500 mL of isotonic sodium chloride solution or 5% glucose solution for injection. It was previously known that the solubility of vidarabine is increased by lowering the pH of the solution, but concern was also expressed that this may cause hydrolysis of vidarabine. Contrary to this concern, however, is that the new formulation of ARASENA-A is stable. To evaluate the mechanism of this stability, we analyzed the lyophilized product using an X-ray powder diffractometer. The findings suggested that hydrogen chloride, which probably plays a role in hydrolysis, is captured by glycine; this appears to be the mechanism behind the stability. The new formulation of ARASENA-A promises to reduce the burden on health care professionals.