Integrated Metabolomic and Biochemical Profiling of Benzodioxane‐Derived Hydrazones Targeting Nrf2 to Mitigate Cadmium‐Induced Metabolic Dysregulation

Benzodioxane-derived hydrazones (BDHs) have emerged as promising antioxidant agents due to their redox-modulatory properties, structural versatility, and ability to activate cellular defense pathways. This study evaluated the protective potential of two BDH derivatives-MBDH (methoxy-substituted) and TBDH (thiophene-substituted)-against Cd-induced toxicity, a major environmental health concern associated with multi-organ dysfunction via oxidative stress and mitochondrial impairment. Using LC-MS/MS-based metabolomic profiling together with biochemical assays in a mouse model, we compared the efficacy of BDHs with that of ascorbic acid. Cd exposure resulted in significant metabolic disruption, including reduced levels of amino acids (serine, methionine), elevated lipid peroxidation products (ceramides, phospholipid fragments), and mitochondrial dysfunction. Ascorbic acid (AA) provided partial metabolic and histological protection. MBDH exhibited moderate effects by reducing lipid peroxidation markers and partially restoring some amino acid levels, with a hydroxylated metabolite detected at m/z 328. TBDH, however, demonstrated superior protection, with near-complete normalization of metabolic profiles, formation of a stable metabolite (m/z 304), and marked attenuation in lipid peroxidation. Additionally, TBDH activated the Nrf2 signaling pathway, increased intracellular glutathione levels, and significantly improved histopathology in the liver, brain, and pancreas. It also reduced the levels of systemic inflammatory markers such as CRP, ESR, and procalcitonin. The enhanced performance of TBDH is attributed to its thiophene moiety, which facilitates improved electron delocalization and redox potential. This study highlights TBDH as a potent Nrf2 activator and antioxidant agent capable of mitigating Cd-induced metabolic and oxidative damage, providing a strong basis for the development of BDH-based therapeutics targeting heavy metal toxicity and related oxidative stress disorders.