The present study aimed to assess dentin hypersensitivity (DH) level and quality of life in patients before and after treatment with ozonated oil compared with extra virgin oil. This is a randomized triple-blind controlled clinical trial, consisting of patients with dentin hypersensitivity which was stimulated by evaporative and thermal stimuli. DH level was measured using a numerical rating scale. The patients were treated with ozonated oil and olive oil. Data were measured at baseline, immediately, and 4 weeks after treatment. The Dentine Hypersensitivity Experience Questionnaire (DHEQ-15) and the Oral Health Impact Profile (OHIP-14) were applied at baseline and 4 weeks after treatment. The sample consisted of 8 women and 3 men, with an average age of 23.82 years. The test group was 40 teeth while the control group consisted of 36 teeth, with the most prevalent tooth being the incisor (52.6%). There was a significant reduction in the DH level and an improvement in the quality of life after treatment. Among the sample, 90.1% reported an improvement in the final result. Both ozonated oil and olive oil were effective in reducing dentin hypersensitivity level and improving oral health-related quality of life 30 days after treatment.