Mechanical enhancement and fish freshness monitoring performance of starch biofilms reinforced with α-cellulose and cellulose nanocrystals

Starch-based biofilms immobilized with butterfly pea ( Clitoria ternatea L.) anthocyanins have attracted considerable attention as colorimetric indicators for intelligent packaging. However, their relatively poor mechanical properties remain a major limitation for practical application. This study aimed to compare the reinforcing effects of cellulose nanocrystals (CNCs) and α-cellulose on starch biofilms and evaluate their performance as colorimetric indicators for monitoring aquatic product freshness. Biofilms were prepared by solution casting using CNCs or α-cellulose at 0.05%, 0.10%, and 0.25%, followed by immobilization with butterfly pea anthocyanins. Tensile strength, elongation at break, Young's modulus, and yield strength were evaluated, followed by ammonia vapor testing and application to catfish, shrimp, and oysters stored at room temperature for 24 h. Indicator performance was validated using total volatile basic nitrogen (TVB-N) and pH. Filler type and concentration significantly affected the mechanical properties of the biofilms (p < 0.05). CNCs provided greater reinforcement than α-cellulose. Increasing CNC concentration significantly decreased tensile strength and elongation at break while increasing Young's modulus (p < 0.05), indicating reduced flexibility and increased stiffness. In contrast, increasing α-cellulose concentration significantly improved tensile strength and elongation at break (p < 0.05), although the resulting films exhibited relatively high Young's modulus and lower mechanical performance than CNC-reinforced films. Among the tested formulations, F1C3 (0.05% CNCs) exhibited the most suitable characteristics for a colorimetric biofilm, with the highest tensile strength (6.18 MPa) and elongation at break (33.49%), representing relative increases of 142.35% and 150.90%, respectively, compared with the control, while maintaining a relatively low Young's modulus. F1C3 was therefore selected as the most suitable formulation within the tested concentration range. The anthocyanin-immobilized F1C3 responded to NH₃ vapor with a distinct color change from purple to greenish-blue. Similar color transitions occurred in catfish, shrimp, and oysters, corresponding to TVB-N increases to 68.01 ± 1.89, 135.16 ± 1.76, and 181.42 ± 2.80 mg/100 g, respectively, with pH increasing from approximately 6.1–6.5–7.5–8.5. These findings demonstrate that CNCs are more effective than α-cellulose for reinforcing starch biofilms, while F1C3 provides a favorable balance of strength, flexibility, and sensitivity to volatile basic compounds, supporting its potential as an environmentally friendly colorimetric indicator for real-time, non-destructive seafood freshness monitoring.