In vivo, in vitro and in silico evaluation of Rumex nepalensis Spreng. and its active phytoconstituent (Chrysophanol) in gastrointestinal disorders.

ETHNOPHARMACOLOGICAL RELEVANCE: Rumex nepalensis Spreng, is widely used in traditional medicine for management of gastrointestinal (GIT) disorder, including ulcer, diarrhea and inflammation. However, limited scientific evidence exists to validate its traditional use in GIT disorders.

AIM OF THE STUDY: This study aims to investigate the anti-Helicobacter pylori and gastroprotective activities of the crude extract of Rumex nepalensis (Rn.Cr), its fractions [aqueous (Rn.Aq), n-hexane (Rn.n-Hex), ethyl acetate (Rn.ETAC)] and its major phytoconstituent, Chrysophanol.

METHODS: Pharmacological activities of Rumex nepalensis crude extract, its fractions and chrysophanol were assessed using ethanol-induced gastric ulcer and castor oil induced diarrheal in rodents. Antioxidant activity was assessed by measuring levels of reduced glutathione (GSH), glutathione S-transferase (GST), catalase (CAT), and lipid peroxidation (LPO) in gastric tissue homogenates and anti-inflammatory effects were evaluated by quantifying pro-inflammatory biomarkers, including interleukin-8 (IL-8), tumor necrosis factor-alpha (TNF-α), and nuclear factor kappa B (NF-κB). Anti-H. pylori activity was assessed through a disk diffusion assay, while molecular docking and simulation studies elucidated the interaction of chrysophanol with H+/K+-ATPase and TLR4 pathways. Acute toxicity was assessed by OECD Guideline 425, using a single-dose oral toxicity test in rats to determine the safety profile. Hematological (e.g., RBC, WBC, Hb, and platelet count) and biochemical parameters (e.g., ALT, AST, ALP, urea, and creatinine) were analyzed using standard automated hematology analyzers and biochemical kits to evaluate systemic toxicity and organ function.

RESULTS: The plant extracts exhibited a dose-dependent antidiarrheal effect and significantly decreased intestinal fluid secretion in mice, while also reducing the distance traversed by charcoal in a gastrointestinal transit model in rats. Both spontaneous and K+ (80 mM)-induced contractions in rabbit jejunum preparations were relaxed by extracts and in a concentration-dependent manner. Rn.Cr, Rn.ETAC and verapamil were relatively effective against K+-induced contractions and shifted the Ca2+ concentration-response curves (CRCs) to the right like that of verapamil. The isoprenaline-induced inhibitory CRCs were moved to the left by Rn.n-Hex similar to papaverine. Rn.ETAC and Rn.n-Hex were effective against H.pylori. Chrysophanol and extracts have an inhibitory impact on H+/K+-ATPase. ELISA shows decreased expression of inflammatory markers, including COX-2, TNF-α, IL-8 and p-NFƙB. H+/K+-ATPase mRNA levels decline confirmed by RT-PCR. Biochemical and hematological tests, along with kidney, heart, liver, and brain damage, suggest to a safer profile for Rumex nepalensis. Chrysophanol has E-values of -7.7 and -7.6 kcal/mol when docked against the H+/K+-ATPase pump and the voltage-gated L-type calcium channel, respectively. The results from MD simulations and MMPBSA/GBSA agree with those from in-vitro, in-vivo, and docking studies.

CONCLUSION: The findings suggest that Rumex nepalensis and chrysophanol possess gastroprotective effect, mediating through its anti-H. Pylori, antispasmodic, antioxidants and anti-inflammatory properties. This, provide scientific validation for its traditional use in GIT disorders and highlight its potential as a natural therapeutic candidate.