Evaluation of the performance of affinity matrix produced from improved variants of S3 peptide to remove endotoxin from Active pharmaceutical ingredient of streptokinase and comparison with commercial matrices

Introduction: Endotoxin or lipopolysaccharide (LPS) is one of the components of the wall of gram-negative bacteria, which, when in contact with blood, stimulates the immune system and causes fever and even serious adverse effects or death. Removing endotoxin is one of the most daunting challenges presented to the purification process in the production of recombinant biological drugs. Bacterial endotoxin (LPS) is resistant to heat and passes through sterilizing filters, and due to its dangerous side effects in living organisms, part of the target protein purification process is always related to removing this disturbing factor from the product. Materials & Methods: The affinity matrix S3E3-S-Sepharose, which was produced by immobilizing the improved variant of antimicrobial peptide S3 called S3E3 peptide on Sepharose chromatography resin, was used to remove endotoxin from active pharmaceutical ingredient (API) of streptokinase, and the performance of this matrix was compared with the performance of a disposable commercial matrix (containing S3 peptide as its ligand) and with ion exchange chromatography resin. Results: The statistical analysis of the results revealed that compared to commercial S3 matrices and ion exchange chromatography, the S3E3-S-Sepharose matrix had higher protein recovery (84.07% compared to 81.50 and 75.31%, respectively) and higher streptokinase biological activity recovery (81.95% vs. 27 76.76 and 61.54%, respectively). Conclusion: As evidenced by the obtained results, the S3E3-S-Sepharose matrix seems to be a suitable candidate for use in the purification processes of Streptokinase API and other recombinant biopharmaceuticals.