Blood Leukocyte DNA Methylation of Circadian Genes in Relation to Environmental Phthalate Exposure in Mexican Adolescents

Background Endocrine-disrupting chemicals have been linked to circadian disruption, but their associations with epigenetic regulation of circadian genes during adolescence, a key developmental period, remain unclear. Methods We analyzed 488 adolescents (mean age = 15.4 years; 53.1% female) from the ELEMENT cohort. Urinary phthalate metabolites and blood leukocyte DNA methylation (DNAm) were collected in 2015. Metabolites were grouped into summary phthalate mixtures, and DNAm at 334 CpG sites across nine circadian genes was quantified using the Infinium MethylationEPIC BeadChip. Multivariable linear regression models adjusted for specific gravity, sociodemographic factors, and immune cell composition. Associations with p < 0.05 were evaluated, and false discovery rate (FDR) correction was applied across all CpGs (q < 0.10 denoting statistical significance). Sex-stratified analyses were also performed. Results Several phthalate mixtures were associated with DNAm of circadian genes after FDR correction. For BMAL1 , ΣDEHP, and Σ Plastic were associated with hypomethylation at cg09404790 (q = 0.0419 and 0.0650), and Σ AA was positively associated with cg14666553 (q = 0.0667). In males, multiple CpGs were statistically significant, including CRY1 hypomethylation with Σ AA and Σ DBP (cg06205108 and cg03853227; q = 0.0058–0.0600) and additional loci in BMAL1 (cg05239841; q = 0.0389) and CRY2 (cg26742859; q = 0.0650). In females, only one CpG remained significant after FDR adjustment, with higher methylation at a CRY1 locus observed in relation to Σ Plastic exposure (q = 0.0067). Conclusions Adolescent exposure to phthalate mixtures was significantly associated with DNAm at select circadian gene loci, with stronger and more frequent associations in males.