Enzyme-loaded thermoplastic starch films incorporating papain and bromelain for active meat tenderization

• Functionalized edible films with papain improved meat tenderness within 30 min. • Interaction between starch and papain involved hydrogen bonding. • Film elasticity is controlled by enzyme concentrations and starch types. • Papain reduced water sensitivity and stabilized starch film structures in water. • Papain incorporated films improved cherry-red color of packaged beef. Incorporating proteolytic enzymes into thermoplastic starch (TPS)-based biopolymers presents a novel strategy for developing active packaging films capable of enhancing meat tenderness. However, preserving enzyme activity during high-temperature extrusion remains a major challenge. This study developed cassava TPS/poly (butylene adipate-co-terephthalate) (PBAT) films incorporating papain and bromelain at 10 %, 20 %, and 30 % (w/w) using a continuous extrusion process optimized to minimize thermal degradation. The films retained significant enzymatic activity, improving meat tenderness by up to sixfold, with hardness reduced from 4.20 ± 0.29 N (fresh meat) to 0.65 ± 0.07 N (10 % papain) and 1.10 ± 0.26 N (10 % bromelain) after 1 h of aging. FTIR spectra showed band shifts from 3323 cm⁻¹ to 3338 cm⁻¹ (papain) and 3329 cm⁻¹ (bromelain), confirming enzyme–polymer hydrogen-bond interactions. Dynamic mechanical thermal analysis (DMTA) revealed that papain decreased rigidity through plasticization, whereas bromelain increased the glass transition temperature due to network reinforcement. The water vapor permeability decreased from 5.49 ± 0.30 g·mm/m²·day·KPa (control) to 3.06 ± 0.16 g·mm/m²·day·KPa (30 % papain), while oxygen permeability increased from 4.71 ± 1.43 to 13.95 ± 0.57 cm³·mm/m²·day·KPa, indicating modified gas transport behavior. Films with 10 % enzyme loading exhibited the best balance between enzyme activity, barrier properties, and structural integrity. These findings demonstrate that enzyme-active TPS/PBAT films are promising for scalable, bio-based packaging applications that improve meat tenderness and quality during storage.