[Petroleum ether fraction of

objectivesTo investigate the effect of the petroleum ether fraction of

methodsThe chemical profile of PEFM was analyzed using GC-MS. Forty Kunming mice were randomized into 5 groups for daily gavage with solvent, dicyclomine, or low-, medium-, and high-dose PEFM for 5 consecutive days, and their effects against neostigmine-induced spasmodic abdominal pain were evaluated. Serum cGMP levels and ileal PKG and p-VASP expressions of the mice were measured. Acute toxicity of PEFM gavage at different doses was assessed in another 32 Kunming mice by monitoring behavioral changes and organ indices. An isolated ileal model of ACh-induced spasm was used to observe the antispasmodic effect of PEFM and its main constituent, stigmast-4-en-3-one (ST), and the mediating roles of the NO/sGC/cGMP/PKG pathway, potassium channels, and calcium channels were investigated using specific inhibitors and a high-potassium-induced spasm model. Molecular docking was used to predict the interactions between compounds and targets.

resultsPEFM contained mainly steroidal compounds, with ST as the most abundant component (31.51%). PEFM at the medium and high doses significantly reduced abdominal pain frequency, prolonged pain latency, and increased serum cGMP and ileal p-VASP expression without producing significant acute toxicity. Both PEFM and ST exhibited concentration-dependent antispasmodic effects against ACh-induced contractions (EC

conclusionsPEFM has spasmolytic and analgesic effects in mice with neostigmine-induced spastic abdominal pain, mediated primarily by its main active component ST, which upregulates cGMP, enhances ileal VASP phosphorylation, activates NO/sGC/cGMP/PKG pathway, promotes opening of voltage-gated potassium channels, and antagonizes L-type calcium channels.