Redistribution of Protein Biological Activity in a Freeze-Dried Cake

Abstract

Lactate dehydrogenase (LDH) solutions were freeze dried in order to investigate the redistribution of the remaining LDH activity in the dried cake. D-Mannitol and a polysaccharide (κ-carrageenan, ι-carrageenan, or dextran) were mixed and used as stabilizers in freeze drying. It was found that the remaining LDH activity was significantly redistributed in the freeze-dried cake, and the enzyme concentration was not uniform along the axis of the dried cake due to variations in the freeze-drying conditions and stabilizer concentration. In some cases, active enzymes were highly localized in a specific portion of the dried cake. This was mainly due to the redistribution of the protein content. Further freeze-drying tests employing bovine serum albumin (BSA) and the above-mentioned polysaccharides showed that proteins (BSA) tended to accumulate in the lower portions of the dried cake, forming concentration gradients. Therefore, the concentration gradient in the freeze-dried cake was mainly due to the freezing process. This redistribution of protein content contributed considerably to the apparent redistribution of enzyme activity. The results suggest that a process that can control the redistribution of protein content in a freeze-dried cake would further improve the biological activity of freeze-dried proteins.

Keywords:

• Biological activity
• Freeze drying
• Pharmaceuticals
• Proteins
• Redistribution

Notes

The sample ID is composed of the cooling rate (R: 2.0°C/min, S: 0.5°C/min), carrageenan content (0: 0%, 1: 0.1%, 2: 0.2%, 3: 0.3%), shelf temperature (T20: −20°C, T10: −10°C), and chamber pressure (P05: 5 Pa, P25: 25 Pa).