In-situ immobilized laccase on bacteria cellulose: varying the laccase concentration for isotherm and kinetic models determination / Thivvinesh A/L Raja

Faculty of Chemical and Process Engineering Technology

Final Year Report (Bachelor of Chemical Engineering ) -- Universiti Malaysia Pahang Al-Sultan Abdullah - 2025

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Enhanced enzyme activity and the potential of in-situ immobilization to reduce time and cost have motivated this study, alongside the need to deepen understanding of kinetic and isotherm models. This work aims to determine the interaction of in-situ immobilized laccase on bacterial cellulose using the best-fit isotherm model by varying enzyme concentration and to elucidate the immobilization mechanism of in-situ immobilized laccase on bacterial cellulose using the suitable-fit kinetic model by varying immobilization time. Bacteria cellulose was prepared through the fermentation of Gluconacetobacter xylinus in nutrient-rich media under static conditions. For in-situ immobilization, laccase (50-800 U/ml) was added to sterilized pineapple waste media during bacteria cellulose fermentation, and immobilization was performed over periods (0 -14 days). Experimental data obtained used to calculate for bound laccase. The bound laccase data was used to calculate 𝑞𝑒 and 𝐶𝑒 that were fitted to isotherm models, including Langmuir, Freundlich, Temkin, Dubinin-Radushkevich, Sips, Redlich-Peterson, and Hill, to evaluate the interactions governing immobilization. The Langmuir model emerged as the best fit, supported by the Sips model, which exhibited Langmuir-like behavior, indicating monolayer adsorption and the parameters values obtained for maximum adsorption capacity was 0.002 and bonding energy was ranged 0.728 and 1.042. The bound laccase obtained from experimental data was used to fit in to the kinetic models. In Kinetic studies using pseudo-first-order, pseudo-second-order, and intra-particle diffusion models revealed that the pseudo-first-order model described the physical adsorption of laccase onto bacterial cellulose, characterized by weak bonding. Additionally, the intra-particle diffusion model was validated with a high regression coefficient (0.96), identifying intra-particle diffusion as the rate-limiting step. The maximum adsorption capacity and equilibrium laccase adsorbed showed consistency between experimental and calculated values. These results confirm that the in-situ immobilization of laccase onto bacterial cellulose involves monolayer, physical adsorption with diffusion-limited kinetics.