An efficient and industrially feasible process for purification and immobilization of orange-peel-based lipase for biocatalytic applications

Abstract

Lipase is an important enzyme with wide industrial applications in various sectors. The present study focuses on the development of an efficient and industrially feasible process for the purification and immobilization of orange-peel-based lipase. The integration of ultrafiltration (UF), adsorptive chromatography, and immobilization on synthetic resins offers an efficient method for producing purified, immobilized lipases. Ultrafiltration membranes with varying molecular weight cutoffs were evaluated for concentrating crude lipase. Among the membranes tested, the 5 kDa UF membrane demonstrated over 94% enzyme activity retention. Various microporous adsorbents (hydrophilic, hydrophobic and ion exchange) were tested for removing color impurities from concentrated crude lipase. The hydrophobic adsorbent achieved over 90% removal of color components. Overall, the integration of UF membranes and adsorptive chromatography showed a 2.29-fold increase in production efficiency. Different microporous adsorbents were tested for the immobilization of purified lipase to produce an immobilized lipase biocatalyst. The use of hydrophobic adsorbents with lower porosity and a larger surface provides maximum binding efficiency (95%). The characterization study of purified lipase was conducted by using Fourier transform infrared (FTIR) spectroscopy and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The purification process yielded a lipase enzyme concentration of 0.28 mg mL⁻¹ with a specific activity of 250.54 U mL⁻¹, achieving an approximately 2.29-fold increase in purity. This study presents an efficient and industrially viable method for the purification and immobilization of lipases, enabling their use in biocatalytic applications.