One‐Pot Encapsulation of Enzymes in a Calcium Carboxylate Metal‐Organic Framework for Improved Buffer Stability

Abstract Diverse metal‐organic frameworks (MOFs) have been actively studied for enzyme encapsulation to enhance encapsulation efficiency (EE), retained enzymatic activity (REA), and stability. This study develops a biocompatible synthesis protocol for a MOF constructed from Ca ² ⁺ and 4,5‐imidazoledicarboxylate ligand (termed CaIDC) in water at room temperature. This method enabled in situ, one‐pot encapsulation of various enzymes, including bovine serum albumin (BSA), glucose oxidase (GOx), catalase (CAT), and esterase (EST). Compared to widely studied zeolitic imidazolate frameworks (ZIFs), CaIDC‐based biocomposites demonstrated enhanced chemical robustness in phosphate buffer at pH 6.0 and 7.4. To optimize the synthesis process, a machine learning‐assisted workflow incorporating Latin hypercube sampling (LHS) is developed for an efficient exploration of the entire synthesis space. As a model system, the optimized EST@CaIDC sample (EC19) exhibited EE, REA, and protein loading (P _loading ) values of 28.7%, 20.1%, and 4.2 wt.%, respectively. In all, this study presents the development of a robust CaIDC‐based platform for enzyme encapsulation and the implementation of an efficient ML‐assisted optimization strategy, offering a pathway to advance enzyme encapsulation technologies with enhanced catalytic performance and sustainability.