Background/aim: Activity of the prestin gene may have a role in the pathogenesis of salicylate-induced ototoxicity. We investigated DNA methylation for prestin gene exon 1 in salicylate-injected guinea pigs.Materials and methods: Fifteen guinea pigs (30 ears) underwent audiological evaluation including 1000 Hz probe-tone tympanometry and a distortion product otoacoustic emission (DPOAE) test. The animals were randomly divided into three groups. Groups 2 (8 ears) and 3 (14 ears) were injected with intramuscular saline and sodium salicylate (200 mg/kg), respectively twice daily for 2 weeks. Group 1 (8 ears) received no injection. DPOAE measurements were performed at baseline; after 1, 2, 4, and 8 h (acute effect); and after 1 and 2 weeks (chronic effect). After audiological measurements, the animals were sacrificed for DNA isolation.Results: While a significant decrease (P < 0.01) was found for the acute effect in all frequencies in Group 3 according to baseline measurements, there was no difference in terms of chronic effect. DNA methylation increased during the acute phase of salicylate administration, whereas it returned to initial levels during the chronic phase.Conclusion: Salicylate-induced changes in DPOAE responses may be related to prestin-gene methylation. These results may have important implications for salicylate ototoxicity.