Isoniazid (INH), a cornerstone of tuberculosis therapy, is plagued by dose-limiting hepatotoxicity with limited therapeutic options. While Schisandra chinensis lignans exhibit hepatoprotective potential, their mechanistic interplay with ferroptosis—a newly recognized driver of INH-induced liver injury—remains unexplored. Here, we delineate a novel molecular axis by which Schisandrin A (SinA) and Schisandrin B (SinB) mitigate INH-induced hepatotoxicity through ferroptosis suppression. Using human hepatocyte models (L02, WRL68) and C57BL/6J mice, we demonstrate that SinA/SinB significantly attenuate hepatic injury markers (ALT/AST), iron overload, lipid peroxidation, and glutathione depletion. Crucially, both compounds rescued GPX4 expression by blocking its ubiquitin-proteasomal degradation, as evidenced by CETSA, DARTS, and Co-IP. Strikingly, GPX4 knockout abolished their protective effects, underscoring GPX4 as the pivotal target. ITC and SPR revealed high-affinity binding of SinA/SinB to GPX4, while molecular docking identified Lys31 and Lys90 as critical residues for SinB-GPX4 interaction. Mutagenesis studies confirmed that K31/K90 substitutions abolished SinB’s efficacy, highlighting a structure-dependent mechanism. Notably, in vivo administration of SinA/SinB restored hepatic GPX4 levels, suppressed ferroptotic markers (MDA, GSH), and ameliorated histopathological damage. This work not only uncovers GPX4 stabilization as a druggable strategy against ferroptosis but also positions SinA/SinB as first-in-class natural compounds targeting this pathway for INH-induced liver injury. Our findings bridge traditional medicine with modern mechanistic insights, offering a blueprint for developing standardized, multitargeted therapies to address drug-induced hepatotoxicity.