We investigated if prenatal exposures to tobacco smoke lead to changes in mitochondrial DNA content (mtDNA) in cord serum and adversely affect newborns’ health. Umbilical cord serum cotinine levels were used to determine in utero exposure to smoking. Cord serum mtDNA was measured by quantitative polymerase chain reaction analysis of the genes coding for cytochrome c oxidase1 (MT-CO1) and cytochrome c oxidase2 (MT-CO2). Log transformed levels of mtDNA coding for MT-CO1 and MT-CO2 were significantly higher among infants of active smokers with higher serum level of cotinine (p p = 0.08; p = 0.02). Structural equation modeling results confirmed a positive association between cotinine and MT-CO1 and2 (p p = 0.02) and IGF-1 (p MT-CO1 and MT-CO2 associated to increased cord serum cotinine and decreased gestational age.