BackgroundAlthough adverse health effects of phthalates have been reported, very few studies have assessed the associations between biomarkers of phthalate exposure and serum folate concentrations in children. ObjectivesWe aimed to examine the association between urinary phthalate metabolites, as biomarkers of exposure to phthalates, and total serum folate concentrations in children using national data from the United States. MethodsWe conducted cross-sectional analyses of 2100 individuals aged 6–18 y enrolled in the National Health and Nutrition Examination Survey, 2011–2016. Multivariable linear regression was applied to examine the relationship between natural logarithm (ln)-transformed urinary phthalate metabolites and serum folate concentrations. The quantile-based g-computation was used to assess the association of urinary phthalate metabolite mixture with serum folate levels. Subgroup analyses were conducted by sex, age, and race/ethnicity, and the interactions were assessed by adding interaction terms of these stratifying variables and phthalates and modeling through the Wald test. ResultsIn multiple linear regression models, for participants in the highest tertile of MEHHP, MEOHP, DEHP, MCPP, and MCOP, total serum folate concentrations were 1.566 [β: −1.566; 95% confidence interval: −2.935, −0.196], 1.423 (−1.423; −2.689, −0.157), 1.309 (−1.309; −2.573, −0.044), 1.530 (−1.530; −2.918, −0.142), and 1.381 (−1.381; −2.641, −0.122) ng/mL lower than those in the lowest tertile. The inverse associations were consistent in different subgroups by sex, age, and race/ethnicity (P for interaction ≥0.083 for all). In addition, the phthalate mixture showed a strong inverse correlation with serum folate; a quartile increase in the phthalate mixture on the ln scale was associated with 0.888 (−0.888; −1.677, −0.099) ng/mL decrease in the serum folate. ConclusionsHigher concentrations of urinary phthalate metabolites were associated with lower serum folate concentrations in children. Although our findings should be validated through additional population and mechanistic studies, they support a potential adverse effect of phthalate exposure on folate metabolism in children.