RESUMO
The effects of different extraction methods (traditional extraction, ultrasound extraction, cellulase extraction, and ultrasound-assisted cellulase extraction) on the yield of mulberry leaf protein (MLP) were investigated, and the results revealed that multi-frequency ultrasound-assisted cellulase extraction was the most efficient extraction method. The mechanism of the synergistic extraction method used to efficiently extract protein from mulberry leave was investigated, focusing on the kinetics and thermodynamics of the enzymatic process. The results revealed that kinetic parameters KM decreased by 14.07% and kA increased by 5.02%, and the thermodynamic parameters Ea, ΔH, and ΔS decreased by 44.81%, 48.41%, and 21.12 %, respectively, following the process of multi-frequency ultrasound (MFU) pretreatment. The spectral analysis with fluorescence spectra manifested that ultrasound exposed hydrophobic groups and induced molecular unfolding of MLP. Atomic force microscope showed that ultrasound decreased particle size while increasing flexibility of MLP. The effect of ultrasound increases the binding frequency of cellulase and substrates, resulting in greater affinity between the two and promoting the solubilization of MLP. This study provides a theoretical basis to improve the application prospects of MLP.