Phytochemical characterization and multi-target protective effects of Lycopodium serratum var. longipetiolatum...

Primary dysmenorrhea is associated with aberrant uterine contractility, inflammatory activation, and oxidative stress, resulting in nociceptive hypersensitivity and impaired quality of life. Lycopodium serratum Thunb. var. longipetiolatum Spring. (LS), a fern endemic to Taiwan, has been traditionally used to alleviate menstrual disorders; however, its mechanistic basis remains undefined.

This study aimed to investigate the chemical constituents, uterine relaxant, and antinociceptive effects of LS extracts, and to elucidate their molecular mechanisms in dysmenorrhea.

Ethanolic extracts of LS and their solvent-partitioned fractions, ethyl acetate (LSE-EA), n-butanol (LSE-BuOH), and aqueous (LSE-H₂O) were characterized by LC-MS/MS for phenolic constituents. The relaxant and antinociceptive effects were assessed in ex vivo uterine contraction assays induced by prostaglandin F₂α (PGF₂α), oxytocin, acetylcholine, and carbachol, and in acetic acid- and oxytocin-induced pain models in ICR mice. Western blot, biochemical, and histopathological analyses were performed to delineate molecular and oxidative pathways.

LSE-EA exhibited the strongest inhibition of uterine contraction and pain responses. LC-MS/MS identified ferulic acid, caffeic acid, and chlorogenic acid as major metabolites. Mechanistically, LSE-EA downregulated oxytocin receptor (OTR) and myosin light chain kinase (MLCK), suppressed TLR-4/NF-κB/COX-2 and ERK activation, reduced uterine IL-6 expression, and attenuated oxidative stress, as evidenced by decreased malondialdehyde levels and restoration of redox balance.

Lycopodium serratum extract confers protection against dysmenorrhea through concurrent suppression of Ca²⁺-dependent uterine contraction, inflammatory signaling, and oxidative stress. These findings identify LSE-EA as a novel bioactive fraction with therapeutic potential in redox-mediated uterine dysfunction.