Abstract
The purpose of the present study was to elucidate the effects of sodium tetraborate (borax) and boric acid on the crystallization of mannitol in frozen aqueous solutions and freeze-dried solids. Thermal analysis of frozen solutions showed that sodium tetraborate inhibits mannitol crystallization at sodium tetraborate/mannitol molar concentration ratios of approximately 0.05, which is much lower than the other co-solutes studied (boric acid, sucrose, sodium phosphate buffer). Inhibition of the mannitol crystallization in frozen solutions resulted in highly amorphous mannitol in the freeze-dried solids. Mannitol remained in an amorphous state in some of the combination freeze-dried solids, even at elevated temperatures. Changes in the thermal transition temperatures (glass transition temperature of maximally freeze-concentrated solute ( T g′) and glass transition temperature of freeze-dried solid ( T g)) suggested reduced mannitol molecular mobility with increases in the sodium tetraborate ratio. Fourier-transform infrared spectroscopy (FT-IR) analysis of the bovine serum albumin secondary structure showed apparent protein structure-stabilizing effects of the amorphous mannitol and sodium tetraborate combination during the freeze-drying process. The mannitol and sodium tetraborate combination also protected lactate dehydrogenase (LDH) from inactivation during freeze-drying. We conclude that the complex formation and the accompanying reduction in molecular mobility make sodium tetraborate an effective mannitol crystallization inhibitor in frozen solutions and freeze-dried solids.
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