Evaluating the impact of using heat and polyethylene glycol in the yield of albumin purification from cryo poor plasma (CPP)

Abstract

Introduction: Human plasma is a suitable source for different proteins and enzymes with a variety of therapeutic effects. Examples include coagulation factors VIII and IX, immunoglobulin, and albumin. The aim of this study is to evaluation of the impact of using heat and polyethylene glycol in the yield of albumin purification from cryo poor plasma (CPP). Materials and Methods: CPP was prepared as frozen from the Blood Transfusion Organization and after defrosting at 4 ° C, it was elicited in 15 cc volumes for experiments. First, the effect of different percentages of polyethylene glycols of 4000, 6000, 8000 and 10000 kDa on 8% gel was investigated and the percentages with acceptable results were selected for use in combination with the desired conditions to investigate the effect of heat. At each step after adding PEG to CPP, the resulting mixture was incubated for 1 hour while stirring at room temperature and then centrifuged at 13000rpm for half an hour at room temperature and the precipitate was removed. Trial and error methods were used to prepare the best percentage of gel required for electrophoresis. Given that the stability of albumin to heat is about 60 ° C, and most proteins are denatured before reaching this temperature; therefore, the use of temperature increase by Benmari method, regardless of the effect of the polymer, on the amount of albumin separation was investigated. Samples (2 cc microtubules) were placed at 37, 40, 45, 50, 55, 60 and 65 ° C for half an hour and then incubated at room temperature for two hours. The temperature of 50 ° C was selected according to the results of electrophoresis and observation of albumin retention in the supernatant. In the final step, the selected temperature and the effect of polyethylene glycol 8000 kDa in combination were applied in certain percentages to determine the best effect in separating albumin from other proteins. One-dimensional electrophoresis method on polyacrylamide gel (SDS-PAGE) on 8% gel was used to investigate the deposition pattern of proteins, especially albumin, and sampling was performed from both the precipitate and the supernatant. Experimental design was not performed because the required data limits were known. Results: In this study, among the polyethylene glycols with molecular weights of 4000, 6000, 8000 and 10000 kDa, polyethylene glycol 8000 was the best molar mass for purification of albumin from CPP. Also, among the temperatures of 37° C to 65 ° C, 50 ° C was the best temperature for the deposition of CPP proteins. Finally, the best temperature and molar mass of polyethylene glycol in the purification of albumin from Cryo Poor Plasma (CPP) was the integration of 50 ° C and polyethylene glycol 8000 kDa at 15 to 22 percent. Conclusion: According to the results of this study, it seems that the best temperature and molar mass of polyethylene glycol in the purification of albumin from Cryo Poor Plasma (CPP) is the integration of 50 ° C and polyethylene glycol 8000 kDa at 15 to 22 percent. It is suggested that these findings be examined on a semi-industrial scale by estimating the amount of albumin extracted (albumin extraction efficiency by this method).