Optimization of Pheretima Protein Extraction by Response Surface Methodology, Structural Characterization of the Resulting Enzymatic Peptides, and Evaluation of Their Skin-Whitening and Anti-UVB Photodamage Effects in Mice

The growing demand for naturally derived multifunctional skincare agents has directed attention toward underutilized protein-rich bioresources. Pheretima, a traditional medicinal resource, is a promising but underexplored source of bioactive peptides. Here, response surface methodology (RSM) was used to optimize the extraction of Pheretima proteins (PPs). The optimal conditions (material-to-liquid ratio 1:7.1 (g/mL), pH 7.5, homogenization time 131.0 s) gave an extraction rate of 19.67%, in good agreement with the predicted 19.80%. The derived enzymatic peptides (PPEPs) were fractionated by ultrafiltration. The PPEPs > 3 kDa fraction showed the highest proline content, greater colloidal stability and a β-sheet-rich secondary structure. Crude PPEPs and all fractions showed strong antioxidant activity, and the PPEPs >3 kDa fraction most strongly inhibited tyrosinase and hyaluronidase. In female ICR mice with acute UVB-induced photodamage, topical PPEPs (repair and protection regimens) attenuated erythema, erosion, epidermal thickening and collagen fragmentation, reduced ROS and returned the UVB-induced compensatory elevation of Nrf2 toward control levels, suppressed IL-1β, IL-6 and TNF-α, and downregulated MMP9 and TRP1, with modulation of TGF-β1/Smad, p38 and AKT signaling. These results indicate that PPEPs are sustainable multifunctional ingredients with anti-photodamage and skin-whitening potential.