CLEAs of amidase from Bacillus smithii IIIMB2907: Development and application in hydroxamic acid synthesis

Cross-linked enzyme aggregates (CLEAs) offer a promising strategy for enzyme immobilization, enhancing reusability, high enzyme loading capacity, carrier-free nature, enhanced thermal or operational stability, and ease of preparation and suitability for industrial-scale applications. This study presents the development and optimization of CLEAs of crude amidase from Bacillus smithii IIIMB2907 for efficient biocatalytic synthesis of pharmaceutically important hydroxamic acids from amides. By optimizing key preparation parameters, a robust immobilized biocatalyst was established that retains high activity and stability during repeated use. The CLEAs demonstrated strong catalytic performance in a 1 L bioreactor, successfully converting benzamide to benzohydroxamic acids with sustained activity over multiple subsequent cycles retaining ~ 70 % of their initial activity. This work highlights the practical potential of using immobilized crude enzymes for cost-effective biocatalysis and provides a foundation for further scale-up and process development in sustainable hydroxamic acid production.