Different strategies to obtain laccase-based time-temperature indicator for monitoring of cold chain storage and management

Abstract

In this work, we have developed a battery of enzymatic time-temperature indicators (TTIs) based on the oxidation of 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) catalyzed by laccase from Trametes versicolor. For this, enzyme concentration, pH value, and addition of sodium azide (a non-competitive inhibitor of the enzyme) were utilized to modulate the apparent activation energy of the reaction catalyzed by the biocatalyst (Ea) and the reaction rate constant, tailoring them to match specific product requirements. The color change (∆E) was used as the response parameter. To check the feasibility of this biosensor, ∆E was modeled using a modified Arrhenius equation to extract kinetic parameters for each TTI formulation. TTI prototypes were evaluated under a wide range of isothermal (4–45 °C) and dynamic temperature conditions (4–30 °C). The results showed that enzyme concentration primarily influenced the (∆E) rate, while reaction Ea was significantly affected by pH and laccase inhibitor concentration. Under dynamic conditions simulating real cold chain scenarios, the model accurately predicted the TTI response, demonstrating high correlation between experimental and simulated data (R2 > 0.94). These findings confirm the feasibility of customizing TTIs for different perishable products. These enzymatic TTIs offer a scalable and cost-effective tool for integration into intelligent packaging systems, enhancing cold chain management and reducing food waste