Adrenal glands as secondary target organs of inhaled benzalkonium chlorides

Benzalkonium chloride (BKC) is a quaternary ammonium compound widely used in aerosol disinfectants and sanitizers, raising concerns about its biodistribution and potential systemic toxicity following inhalation exposure. In this study, biodistribution using ¹⁴C-labeled BKC and the toxicity of unlabeled BKC were evaluated in rats following intratracheal and oral administration. Biodistribution after a single intratracheal instillation revealed accumulation of BKC in the adrenal glands, which exhibited the highest accumulation per organ weight among extrapulmonary secondary organs, while oral exposure showed minimal gastrointestinal absorption. Repeated exposures of BKC at doses of 50 and 100 μg/rat (10 times with three-day intervals) induced no pulmonary inflammation but elicited mild systemic responses, including increased white blood cell counts. However, other systemic toxicity-related parameters, such as serum biochemistry, coagulation, and serum cytokine levels, remained unchanged. Among secondary organs, the adrenal glands showed significant toxicity-related changes, including vacuolation in the zona fasciculata and elevated serum corticosterone levels. Transcriptomic analysis using RNA sequencing identified 1214 differentially expressed genes (DEGs) at a dose of 100 μg/rat compared to the control. Gene ontology analysis highlighted mitochondrial-associated processes, and network analysis revealed that alterations in mitochondrial membrane potential and reactive oxygen species (ROS) generation were closely linked to lipid accumulation, oxidative stress, and cell death. These changes formed a distinct dose-dependent pattern indicative of mitochondrial dysfunction. In conclusion, these findings identify the adrenal glands as previously unrecognized extrapulmonary targets of BKC toxicity, underscoring the need for further studies on associated long-term human health risks.