Individuals with a diagnosis of adult separation anxiety (ASAD) have extreme anxiety about separations, actual or imagined, from major attachment figures. ASAD might represent a psychological/behavioral model for research probably involving a dysregulation of those neurobiological mechanisms of attachment, in particular central oxytocin (OT), described in numerous animal studies. As experimental strategy, we chose the nucleotidic sequencing of the human OT gene of patients with ASAD to evaluate whether OT mutations were related to potential alteration of its production. With this aim, mutation scanning of proximal promoter and untranslated and coding regions of the OT gene was carried out in 36 patients with ASAD, 14 patients without ASAD, and 26 controls. No mutations were found in promoter and coding regions of the OT gene in our population. One rare 3'UTR single nucleotide variant (rs17339677) and one intron 2 molecular variant (rs34097556), which showed a high frequency, were evidenced. There was no significant difference in the genotype distribution of this intron 2 polymorphism between patients and healthy individuals. Further research is needed to investigate the association between ASAD and OT peptide and receptor polymorphisms.