Phytochemical Insights into Citrus limon (L.) Osbeck (Kaji Nemu) of Assam as a Potential Natural Modulator of Hyperthyroidism: A Structure-Based and Experimental Approach

Hyperthyroidism is characterized by impaired hormone biosynthesis and dysregulated receptor signaling, and remains a pressing clinical concern as current drugs such as methimazole are often associated with adverse effects and limited efficacy. In this study, we explored the methanolic peel extract of Citrus limon (L.) Osbeck (Kaji Nemu), a distinctive citrus variety from Northeast India, is a potential source of natural therapeutic compounds. LC-MS profiling identified 55 phytochemicals, of which 27 drug-like candidates were shortlisted, alongside ADMET evaluations. Molecular docking revealed three lead compounds including 1-(3,4-Dimethylphenyl)-4-[(4-isopropylphenyl)sulfonyl]piperazine, 2 Benzylcyclohexanol and 2-[(1Z)-2-Methoxy-1-propenyl]-1,3,5-trimethylbenzene showing superior binding affinities with thyroid peroxidase (TPO: –8.2 kcal/mol), thyroid receptor [Formula: see text] (TR[Formula: see text]: –7.6 kcal/mol) and thyroid receptor [Formula: see text]1 (TR[Formula: see text]1: –7.7 kcal/mol), respectively, compared to methimazole. A detailed interaction analysis demonstrated extensive hydrogen bonding, hydrophobic, and [Formula: see text]-interactions, suggesting stable and versatile engagement within binding pockets. Molecular dynamics simulations further confirmed the protein-specific stabilizing effects of candidate ligands, with notable enhancement of backbone stability in TPO and TR[Formula: see text]1. ELISA-based enzyme quantification further supported the therapeutic relevance, with Kaji Nemu peel extract significantly reducing TPO levels in hyperthyroid tissue samples. Overall, the study highlights the translational potential of Kaji Nemu phytochemicals as multi-target modulators capable of simultaneously inhibiting thyroid hormone biosynthesis and regulating receptor activity. These findings position this indigenous citrus resource as a promising avenue for developing safer, plant-derived interventions for managing hyperthyroidism, paving the way for further in vitro and in vivo validations.