Enzymatic extraction and response surface method optimization for efficient extraction of bioactive peptides and functional activities from Leucopaxillus giganteus.

The Leucopaxillus giganteus is a typical high-protein wild edible fungus and is a high-quality natural plant protein source. In this study, the fruiting bodies of Leucopaxillus giganteus were adopted as experimental raw materials, and their intrinsic proteins were degraded via compound protease hydrolysis to produce fungus-derived bioactive peptides with antioxidant, antibacterial and anti-tumor properties. Single-factor tests were performed to determine the appropriate range of each hydrolysis factor, and Plackett-Burman design was adopted to screen three key variables significantly affecting peptide extraction efficiency, including hydrolysis temperature, time and pH. Box-Behnken response surface methodology was further applied to optimize the enzymatic hydrolysis process. The results indicated that the optimal compound protease ratio of alkaline protease to papain was 4:3. Under the optimal hydrolysis conditions (temperature 49 °C, enzyme dosage 5,870 U g-1, pH 7.1, solid-liquid ratio 1:25, hydrolysis time 5 h), the peptide extraction rate reached 28.93%. Subsequently, the activity of the peptide was determined. This peptide exhibited excellent DPPH scavenging ability and formed inhibition zones of 9.02 mm, 6.15 mm and 8.67 mm for Escherichia coli, Bacillus subtilis and Staphylococcus aureus, demonstrating good antibacterial activity. Moreover, the peptide could significantly inhibit the proliferation of Caco-2 colorectal cancer cells and HepG2 human liver cancer cells. In conclusion, the enzymatic hydrolysis process optimized by response surface methodology is simple and efficient, which lays a solid foundation for the development and comprehensive utilization of bioactive peptides derived from Leucopaxillus giganteus.