Effects of ultrasound pretreatment with different frequencies and working modes on the enzymolysis and the structure characterization of rice protein

The effects of ultrasound pretreatment with different frequencies and working modes, including mono-frequency ultrasound (MFU), dual-frequency ultrasound (DFU) and tri-frequency ultrasound (TFU), on the degree of hydrolysis (DH) of rice protein (RP) and angiotensin-I-converting enzyme (ACE) inhibitory activity of RP hydrolysate were investigated. Ultraviolet-visible (UV) spectroscopy, fourier transform infrared (FTIR) spectroscopy, surface hydrophobicity and scanning electron microscopy (SEM) of RP pretreated with ultrasound were measured. The results showed that ultrasound pretreatment did not increase DH of RP significantly (p>0.05). However, all the ultrasound pretreatment increased the ACE inhibitory activity of RP hydrolysate significantly (p<0.05). The MFU of 20kHz showed higher ACE inhibitory activity compared to that of other MFU. The ACE inhibitory activity of sequential DFU was higher than that of simultaneous with the same frequency combination. Sequential TFU of 20/35/50kHz produced the highest increase in ACE inhibitory activity in contrast with other ultrasound frequencies and working modes. All the results under ultrasound pretreatment showed that ultrasound frequencies and working modes were of great effect on the ACE inhibitory activity of RP. The changes in UV-Vis spectra and surface hydrophobicity indicated the unfolding of protein and exposure of hydrophobic groups by ultrasound. The FTIR analysis showed that all the ultrasound pretreatment with different frequencies and working modes decreased α-helix, β-turn content and increased β-sheet, random coil content of RP. The SEM results indicated that ultrasound pretreatment resulted in the deformation of RP. In conclusion, the frequency selection of ultrasound pretreatment of RP is essential for the preparation of ACE inhibitory peptide.