Application of ultrasound-assisted physical mixing treatment improves in vitro protein digestibility of rapeseed napin

•IVPD was improved by ultrasonic treatment.•Ultrasound-assisted physical mixing is more effective than physical mixing in improving IVPD.•The secondary structure and hydrophobicity of napin were variational obviously by ultrasound-assisted physical mixing.•Napin digestibility was correlated with conformational changes of disulfide bonds. The in vitro protein digestibility (IVPD) of napin was studied using different pretreatment methods, including ultrasound, mixing napin with lactalbumin, and ultrasound-assisted protein mixing. The relationships between IVPD, molecular structure, and disulfide bonds were explored, showing that the IVPD of napin was the highest compared with the control when treated with 40% ultrasound power. When the proportion of napin to lactalbumin was 5:5, a synergistic influence between the two proteins was observed. Further investigation showed that the IVPD of napin was clearly improved by treatment with ultrasound-assisted protein mixing. Compared with the single protein in the control, the β-sheet content in the secondary structure of the mixed protein after sonication was reduced from 45.02% to 37.16%. The ordered protein structure was also disrupted by ultrasound, as supported by fluorescence intensity and surface hydrophobicity analyses. The decreased number of disulfide bonds and conformational changes indicated that the IVPD of rapeseed napin was closely related to the disulfide bond content. This study provides a theoretical basis for improving protein digestibility by combining ultrasound with physical mixing.