Frequency-dependent effects of multi-frequency ultrasound on the extraction, physicochemical properties, and antihypertensive activity of mushroom protein-peptide mixtures.

Mushroom protein-peptide mixtures (PPM) represent a high-quality resource with the potential to reduce reliance on animal protein sources. Multi-frequency ultrasound has been proven to be an efficient technique for extracting PPM; however, the influence of different frequency combinations on the extraction mechanism and structure-activity relationship of mushroom PPM remains unclear. This study investigated the use of multi-frequency ultrasound for extracting PPM from mushrooms. Different frequency combinations (23/25/40, 23/28/40, and 23/25/28/40 kHz) were compared. The quadruple-frequency mode yielded the highest amount of PPM. Multi-frequency synergy improved the thermal stability, reduced the particle size, altered the secondary structure, increased the hydrophobic amino acid content, and promoted the formation of C-terminal proline peptides of the PPM. The PPM exhibited potent angiotensin-converting enzyme inhibitory and antihypertensive activities, comparable to the drug captopril. Furthermore, quadruple-frequency ultrasound specifically enhanced the abundance of vascular protection-related proteins. This research elucidates the frequency synergy mechanism in ultrasound-assisted extraction.