This study aimed to develop a microencapsulation formulation for efficient encapsulation of β-glucosidase to improve its stability in a rumen-like environment and sustain activity post-rumen in the ruminant gut. Various alginate-based formulations were evaluated to achieve high encapsulation efficiency (EE) and stability. These included control alginate beads (AB), microcapsules with chitosan (MCS), alginate–sucrose beads (AOS), alginate–sucrose–maltodextrin beads (AOMS), and alginate pectin beads (APB). The microcapsules were made using Buchi encapsulator B-390 with calcium chloride as the gelling solution. Alginate proved to be a suitable polymer for β-glucosidase encapsulation and <1 mm diameter microbeads were obtained across all formulations. Alginate alone (AB: 1% alginate, 0.2 U/mL β-glucosidase) showed low EE (3% ± 1.0) due to leakage and syneresis. Modifying the gelling solution with 0.1% chitosan (MCS) increased EE to 49 ± 2.64% by reducing alginate porosity. Further improvements were achieved by adding stabilizers to the alginate solution (AB), in addition to using the modified gelling solution (MCS): Adding sucrose (AOS) at 4% increased EE to 95.5 ± 2.08%, while adding sucrose (4%) and maltodextrin (2%) (AOMS) achieved 100 ± 2.16%. On the other hand, adding pectin (4%) (APB) to the alginate solution resulted in a lower EE of 40.5% ± 2.55, likely due to interference with alginate crosslinking. In vitro rumen fermentation showed a dry matter degradation of 42–54%, underscoring the need for more robust microcapsules. Encapsulation strategies, such as incorporation of additional protective layers, are essential to enhance bead stability, minimize degradation, and improve enzyme retention, to ensure efficient delivery and sustained enzymatic activity in the hindgut.

The need for effective enzyme delivery systems in ruminant nutrition is underscored by the challenge of protecting enzymes in the harsh rumen environment. This study focused on enhancing the encapsulation efficiency (EE) of β-glucosidase in alginate beads to improve its stability and protection within rumen-like conditions. The objective was to develop a microencapsulation system capable of sustaining enzyme activity post-ingestion, using β-glucosidase as a model enzyme. Various formulations were tested to optimize EE and stability. Data were collected by systematically incorporating different stabilizers and crosslinking agents into the alginate matrix. The addition of 0.1% chitosan into the gelling solution significantly improved EE to 49% by reducing the matrix’s porosity. Further improvements were achieved with stabilizers: 4% sucrose (AOS) increased EE to 95.5%, while the combination of 4% sucrose and 2% maltodextrin (AOMS) reached the highest EE at 100 ± 2.16%. In contrast, the addition of 4% pectin (APB) reduced EE to 40.5%, likely due to interference with alginate crosslinking and higher water content, weakening bead structure.In vitro rumen fermentation tests showed degradation rates of 42–54%, highlighting the need for further development of more robust microcapsules. These findings suggest that additional coating strategies are essential to improve bead stability in rumen environments. Future research should focus on creating bead formulations with enhanced protective layers to increase enzyme retention and recovery during delivery.