Arsenic intoxication affects male reproductive parameters of prepubertal rats. Besides, morphological and functional alterations in their testis and epididymis may remain after withdrawal of arsenic insult, causing potential impairment in male fertility during adulthood. In this study, we aimed at analyzing the effect of prepubertal arsenic exposure on the fecundity of epididymal sperm from sexually mature Wistar rats, assessing fertility indexes, sperm parameters, and sperm phosphoproteins content. Male pups on postnatal day (PND) 21 received filtered water (controls, n = 10) and 10 mg L􀀀 1 arsenite (n = 10) daily for 30 days. From PND52 to PND81, rats from both groups received filtered water. During this period, the males mated with non-exposed females between PND72 and PND75. Our results showed that sexually mature rats presented low sperm production, epididymal sperm count, motility, and quality after prepubertal arsenic exposure. These findings possibly contributed to the low fertility potential and high preimplantation loss. Epididymal sperm proteome detected 268 proteins, which 170 were found in animals from both control and arsenic groups, 27 proteins were detected only in control animals and 71 proteins only in arsenic-exposed rats. In these animals, SPATA 18 and other five proteins were upregulated, whereas keratin type II cytoskeletal 1 was downregulated (q < 0.1). The results of KEGG pathway analysis demonstrated an enrichment of pathways related to dopaminergic response, adrenergic signaling, protein degradation, and oocyte meiosis in arsenic-exposed animals. Moreover, 26 proteins were identified by phosphoproteomic with different phosphorylation pattern in animals from both groups, but SPATA18 was phosphorylated only in arsenic-exposed animals. We concluded that prepubertal exposure to arsenic is deleterious to sperm quality and male fertility, altering the sperm phosphoproteins profile.