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Prenatal exposure to per- and polyfluoroalkyl substances (PFAS) may damage newborn mitochondrial function indicated by lower mitochondrial DNA copy number (mtDNAcn), which may help explain the mechanisms underlying adverse health outcomes in offspring. Adequate maternal folate levels may offer protection. We investigated associations between maternal PFAS exposures and newborn mtDNAcn and examined effect modification by maternal folate levels in 572 mother-newborn dyads from the Boston Birth Cohort. We measured eight PFAS in maternal plasma collected 24-72 h postpartum using HPLC-MS/MS and mtDNAcn in cord blood using targeted sequencing. We used multivariable linear regression and Bayesian kernel machine regression models to estimate associations between PFAS and mtDNAcn -score, overall and stratified by newborn sex and maternal folate level. We observed that associations varied by PFAS species, including an inverse association with PFOS and nonlinear associations with Me-PFOSA-AcOH, PFDeA, and PFNA. Associations of PFHxS and PFOS with mtDNAcn differed by sex. Notably, we found inverse associations of Me-PFOSA-AcOH, PFOS, and the PFAS mixture only among newborns whose mothers had low folate levels. In conclusion, prenatal exposures to specific PFAS and the PFAS mixture were associated with altered cord blood mtDNAcn, with adequate maternal folate levels potentially mitigating the associations.