A design of experiment approach for optimized production of encapsulated trypsin using nano spray drying: Comparative physicochemical and kinetic characterization

Enzymes are challenging to formulate due to their inherent instability, particularly in solution. This study aims to use the design of experiment (DOE) approach to develop spray-dried encapsulated trypsin nano-powder (TrySP) with maximum activity and stability using a Büchi B-90 nano spray dryer. A 53 full factorial design with 26 randomly ordered experiments was created to study the impact of process variables on the quality attributes of TrySP. The additive concentration (%), mesh cap size (μm), inlet temperature (Tinlet) (°C), trypsin concentration (%), and additive type (Mannitol or trehalose) were selected as the independent variables. The dependent parameters were yield value, particle size, residual enzyme activity, and moisture content. Further, comparative physicochemical and kinetic characterization of TrySP was performed. TrySP had diverse qualities (yield value 60.2–96.8%, residual activity >90–50%, particle size 314–1030 nm, and moisture content 0.7–2.2%). Optimized TrySP exhibited 89% yield value and 95% residual activity in the presence of 8% (w/w) mannitol. Kcat of TypSP increased from 22.5/s to 25.6/s with improved operational and storage stabilities. The half-life (t1/2) of TrySP showed 6 folds increase. Furthermore, TrySP activation energy increased from 45,918 J/mol to 94,491 J/mol. The sustainable model presented in this study enables the development of a thermostable, encapsulated trypsin nano-powder with physical and chemical properties that are optimized for various biotechnological industrial applications.