Carboxylated Cellulose Nanocrystals Developed by Cu-Assisted H2O2 Oxidation as Green Nanocarriers for Efficient Lysozyme Immobilization

Cellulose nanocrystals (CNCs), having a high specific surface area and versatile surface chemistry, provide considerable potential to interact by various mechanisms with enzymes for nano-immobilization purposes. However, engineering chemically safe CNCs, suitable for edible administrations, presents a significant challenge. A reliable carboxylate form of H-CNCs was formed using H2O2 oxidation of softwood pulp under mild thermal conditions. Negatively charged carboxyl groups (similar to 0.9 mmol g(-1)) played a key role in lysozyme immobilization via electrostatic interactions and covalent linkages, as evidenced by Fourier transform infrared and C-13 cross-polarization magic angle spinning nuclear magnetic resonance spectroscopies. Adsorption isotherms showed a high loading capacity of H-CNCs (similar to 240 mg g(-1)), and fitting the data to the Langmuir model confirmed monolayer coverage of lysozyme on their surface. Using a non-toxic coupling agent, 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium chloride, lysozyme-conjugated H-CNCs were developed with an immobilization yield of similar to 65% and relative catalytic activity of similar to 60%, similar to lysozyme adsorption on H-CNCs. These H-CNC-lysozyme nanohybrids, rationally processed via safe and green strategies, are specifically exploitable as catalytically active emulsifiers in food and pharmaceutical sectors.