Jiwon Yoo (Korea Basic Science Institute)

Ju Yeon Lee (Korea Basic Science Institute)

Jiwon Yoo (Korea Basic Science Institute)

Sung Ho Yun (Korea Basic Science Institute)

Geonu Mo (Korea Basic Science Institute)

Ju Hwan Song (Korea Basic Science Institute)

So-Young An (Department of Health Sciences, The Graduate School of Dong-A University, DAU G-LAMP Project Group, Innovation Center for Atomic Science, The Graduate School of Dong-A University)

Wan-Seob Cho (Department of Health Sciences, The Graduate School of Dong-A University)

Jiyoung Jeong (Korea Radioisotope Center for Pharmaceuticals (KRICP), Korea Institute of Radiological & Medical Sciences (KIRAMS),)

Mihye Kwon (Korea Radioisotope Center for Pharmaceuticals (KRICP), Korea Institute of Radiological & Medical Sciences (KIRAMS),)

Dong Hyun Kim (Korea Radioisotope Center for Pharmaceuticals (KRICP), Korea Institute of Radiological & Medical Sciences (KIRAMS),)

Tae Hwan Shin (3Department of Health Sciences, The Graduate School of Dong-A University)

Ju Yeon Lee (Korea Basic Science Institute)

To investigate organ-specific molecular responses to acute inhalation exposure, TMT-based quantitative proteomic analysis was performed on hydramethylnon and methyl chavicol, two chemical constituents commonly found in household products whose inhalation toxicity remains poorly characterized. Male Sprague-Dawley rats received a single intratracheal instillation of hydramethylnon (1 mg/mL) or methyl chavicol (40 μg/mL), and lung, liver, and small intestine tissues were collected 24 h after exposure. Proteins were processed using the S-Trap workflow, labeled with TMT reagents, fractionated by high-pH reversed-phase chromatography, and analyzed by LC-MS/MS. Protein identification and quantification were performed using the Integrated Proteomics Pipeline (IP2). Differentially expressed proteins (DEPs) were identified using Student's t-test (p < 0.05, |log₂FC| ≥ 0.5), followed by functional enrichment analysis using Metascape. A total of 4,335 and 5,069 proteins were quantified in the two TMT sets, respectively. Methyl chavicol exposure induced extensive proteomic alterations in the small intestine (2,199 significant proteins) and liver (374 proteins), whereas hydramethylnon exposure altered 493 proteins in the lung and 116 proteins in the liver. Pathway enrichment analysis revealed significant changes in membrane organization, intracellular transport, protein localization, metabolic processes, and immune- and inflammation-related pathways, demonstrating distinct tissue-specific responses to each chemical. Collectively, these findings demonstrate that quantitative TMT-based proteomics is a powerful approach for providing mechanistic insight into the organ-specific molecular responses and inhalation-induced toxicity of household chemicals, while facilitating the discovery of candidate biomarkers for toxicological assessment.