Evaluation of Eupatorium perfoliatum loaded chitosan nanocarriers in silico, in vitro and in vivo for anti-inflammatory, antipyretic and analgesic potential

This study explores the development of cost-effective, cross-linked chitosan nanoparticles(CSNPs) for anti-inflammatory and toxicity evaluations, using Eupatorium perfoliatum at ultra-dilution concentrations of 3C and 1000C. In silico studies revealed strong binding affinities and dynamic stability of Eupatorium perfoliatum bioactive compounds within the COX-2 protein–ligand complex, highlighting their potential antipyretic activity. Molecular dynamics simulations further confirmed stable interactions, providing mechanistic insights at the molecular level. Eupatorium perfoliatum was successfully encapsulated into low-molecular-weight (LM) and high-molecular-weight (HM) chitosan nanocarriers. The synthesis and characterization of these nanocarriers confirmed the particle size distribution, surface morphology, and functional group interactions, validating the successful formation of nanoparticles. Zero-order, first-order, Higuchi, and Korsmeyer-Peppas kinetic models were used to assess in vitro drug-release experiments using dialysis membrane diffusion, demonstrating sustained release profiles of 62–88% over 48 h. The nanoparticles exhibited enhanced anti-inflammatory activity, with 56–60% inhibition of lipoxygenase (LOX) and heat induced hemolysis, along with notable antimicrobial activity against Staphylococcus aureus and Escherichia coli. Toxicity assessments, including hemolytic assays and zebrafish embryo toxicity, indicated a significantly lower toxicity profile for drug-loaded nanoparticles compared to the free drug. In vivo studies were conducted using Wistar rats (n = 6 per group; total 8 groups), where EP-loaded chitosan nanocarriers (0.5% HM, 200 μL dose) demonstrated significant antipyretic and analgesic activities compared to the control confirming the therapeutic efficacy of the formulation. Overall, this study highlights the promising potential of homeopathic drug-loaded chitosan nanocarriers as sustainable nanotherapeutic systems for anti-inflammatory and antipyretic applications.