Urinary volatile organic compound metabolites and premature menopause: Population-based, network toxicology, and experimental evidence with a focus on HEMA

Using nationally representative data from female participants in NHANES 2005-2018, this study evaluated the association between urinary volatile organic compound (VOC) metabolites and premature menopause and explored potential mechanisms using epidemiological analyses, mixture models, network toxicology, and experimental validation. Multivariable logistic regression and restricted cubic spline models were applied to assess the relationships between 18 urinary VOC metabolites and the risk of premature menopause. Multiple metabolites were associated with increased risk, among which N-acetyl-S-(2-hydroxyethyl)-L-cysteine (HEMA) showed the largest effect estimate in the fully adjusted model (OR = 1.47, 95% CI: 1.15-1.88) and a nonlinear exposure-response relationship. Mixture analyses showed that the overall VOC mixture was positively associated with premature menopause risk in WQS regression (OR = 1.58, 95% CI: 1.08-2.33) and quantile g-computation (OR = 1.51, 95% CI: 1.32-1.72), while BKMR showed a positive joint exposure-response pattern. HEMA had the largest WQS weight (0.388) and the largest positive weight in quantile g-computation. Network toxicology focusing on HEMA-related parent compounds identified targets enriched in DNA damage response and apoptosis-related pathways, with OGG1, CASP3, BCL2, and JUN emerging as hub genes. In a premature menopause-like mouse model, immunohistochemistry showed increased ovarian expression of OGG1, CASP3, and JUN and decreased BCL2 expression. Molecular docking provided preliminary structural compatibility evidence for possible HEMA-hub protein interactions. Together, this multi-level and multi-method evidence links environmental VOC exposure, represented by HEMA, to female reproductive functional decline and provides new support for environmental risk identification in premature menopause.