AN IMPROVED AND INTEGRATED MEMBRANE-BASED CLARIFICATION AND CONTROLLED CRYSTALLIZATION STRATEGY FOR THE EFFICIENT PURIFICATION OF PHARMACEUTICAL-GRADE SERRATIOPEPTIDASE

Serratiopeptidase, a zinc-dependent metalloprotease produced by Serratia marcescens, is widely recognized for its therapeutic applications, notably in anti-inflammatory and fibrinolytic treatments. Despite its pharmaceutical significance, efficient large-scale purification remains a challenge due to complex downstream processing requirements. This study presents a systematic approach to optimize the downstream processing of serratiopeptidase, focusing on membrane-based clarification and controlled crystallization. The fermentation broth was first clarified using microfiltration to remove biomass and insoluble particulates, followed by ultrafiltration to concentrate the enzyme and reduce process volume while preserving enzymatic activity. Crystallization parameters—including pH, temperature, and solvent system—were individually evaluated to enhance product yield and purity. The optimal conditions were identified as pH 6.0, a temperature range of 0–5°C, and acetone as the anti-solvent, resulting in a maximum yield of 62.86% and activity of 2535 units/mg, meeting pharmacopeial standards. The integrated process significantly improved recovery, reproducibility, and scalability, offering a robust purification strategy suitable for pharmaceutical production. This work provides valuable insights for developing efficient and compliant purification platforms for industrial-scale enzyme manufacturing.