the role of various stabilizers in the acquisition of thermo-toler ance in factor viii activity

we prepared a highly purified and relatively heat stable form of factor viii which contained 25 units per ml (u/ml) activity using peg-4000 and developed an effective and new manufacturing process. heat treatment was performed at 80°c for 72 hrs in the presence of different stabilizers. in our studies, we used different organic solvents as preservatives to maintain factor viii activity, since few enzymes have been observed to function perfectly well in such conditions. our results show that the activity of factor viii in the presence of organic solvent at 80°c for 72 hrs as viral inactivation is significantly decreased. it seems that irreversible aggregation of protein is the major mechanism in cessation of factor viii activity. on the other hand, protein stability may be increased by certain low molecular weight substances. we therefore used different sugars and amino-acids to protect factor viii activity in the process of heat treatment. our results showed that glycine, glucose, saccharose, mannitol and bsa were not effective stabilizers of factor viii. but the loss of factor viii activity at 80°c for 72 hrs, in the presence of 10, 15 and 20% concentrations of trehalose, was about 15-20%, which is acceptable for large-scale processes. thus, the condition obtained in the present study seems to be a more suitable condition than that reported previously.