Optimization and Characterization of TSG-Enriched Polygonum multiflorum Extract and Its Dual Mechanism Against...

This study aimed to optimize the extraction and purification of TSG-enriched Polygonum multiflorum extract and evaluate its therapeutic potential against androgenetic alopecia. P. multiflorum Thunb. has long been used in traditional Chinese medicine to promote hair growth and preserve hair pigmentation. Microwave-assisted extraction (MAE) was optimized to obtain a 2,3,5,4'-tetrahydroxystilbene-2-O-β-D-glucoside (TSG)-enriched P. multiflorum extract (PME). The crude extract was further purified by medium-pressure liquid chromatography and semipreparative high-performance liquid chromatography. Phytochemical profiling by HPLC demonstrated a 3.75-fold increase in TSG content, from 22,770.0 ± 815 ppm to 85,387.5 ± 192 ppm, and the identity of the enriched TSG was confirmed by LC-MS and ¹H NMR spectroscopy. PME exhibited potent dose-dependent inhibition of 5α-reductase activity, reaching 92.7% inhibition at 1 μg/mL, comparable to that of finasteride. In a testosterone (TES)-induced androgenetic alopecia (AGA) mouse model, PME markedly accelerated hair regrowth and increased both the number and size of hair follicles. Mechanistically, PME suppressed the conversion of TES to dihydrotestosterone (DHT) through 5α-reductase inhibition and promoted GSK3β inactivation, β-catenin stabilization, and nuclear translocation, indicating activation of the Wnt/β-catenin signaling pathway. Consequently, the expression of hair growth-related proteins, including Ki67 (1.98-fold), epidermal growth factor (EGF, 1.54-fold), insulin-like growth factor-1 (IGF-1, 1.21-fold), and vascular endothelial growth factor (VEGF, 1.34-fold), was significantly upregulated. These findings demonstrate, for the first time, that TSG-enriched PME obtained through optimized MAE and chromatographic purification promotes hair regeneration through a dual mechanism involving 5α-reductase inhibition and Wnt/β-catenin activation, highlighting its potential as a natural therapeutic candidate for androgenetic alopecia.