In recent years, there has been an increasing interest in the use of gaseous ozone (O3) to promote the germination of edible seeds. While its ability to improve seedling vigor and stimulate germination is acknowledged, there has been limited research on the impact of gaseous O3 on the biochemical profile, including phenolic compounds (TPC) and antioxidant activity, of cereals, seeds, and their sprouts. The lack of information has led to the initiation of this study, which aims to assess the impact of ozone treatment duration at a concentration of 50 ppm 1 L min−1, ranging from 1 to 5 h, on the biochemical attributes of broccoli (Brassica oleracea), radish (Raphanus sativus), alfalfa (Medicago sativa) seeds, and sprouts, as well as wheat (Triticum aestivum) grains and sprouts. By optimizing O3 exposure parameters, including duration, this approach has the potential to serve as a valuable tool for enhancing the microbiological and nutritional quality of seeds and cereals. The findings revealed that O3 treatment generally had an adverse impact on TPC in seeds, cereals, and sprouts, resulting in a significant reduction in TPC post O3 treatment. Wheat grains, in particular, displayed the lowest TPC following ozone exposure, with an average decrease of 39.4% compared to the untreated sample. However, it is noteworthy that alfalfa seeds exhibited a positive response to 4 and 5 h O3 treatment, manifesting an average increase in TPC of 13.0% and 27.7%, respectively. In turn, broccoli, radish, and wheat sprouts displayed the lowest TPC, with values of 47.7%, 20.2%, and 18.0% lower than the control samples, respectively. This study revealed that plant responses to O3 exposure varied, and the effects of O3 treatment on TPC levels depended on O3 exposure time. Furthermore, the effect of O3 on the sugar content of the seeds, cereals, and sprouts varied among different plant types, with some showing an increase in content and others showing no substantial changes. This suggests that, depending on the type of seed, O3 may have both positive and neutral effects.