Brassinosteroids (BRs) are essential plant steroid hormones that regulate a wide range of developmental and physiological processes. In addition, these hormones play a significant role in helping plants cope with stress by modulating responses to various stressors, including heat, cold, drought, and heavy metals. Freezing temperatures during winter and spring pose a serious threat to citrus production, putting the sustainability of this newly emerging citrus industry in north Florida. Based on the protective effects of BRs, we hypothesized that exogenous application of BRs could enhance the basal tolerance of citrus plants exposed to freezing temperatures. To test this hypothesis, we evaluated the photosynthetic performance and cellular damage of cold sensitive grapefruit (cv. Ray Ruby) plants sprayed with BRs (6 ppm) under freezing stress. Plants were treated with BRs (0 and 6 ppm) at 12, 24, and 48 hours before freezing stress (hbfs). Plants maintained at 20°C and sprayed with BRs (0 and 6 ppm) served as controls. Following treatment, the plants were placed in a freezing chamber and subjected to a gradual temperature decrease until reaching -6°C, where they were maintained for 1.5 hours. Freezing stress induced greater cellular damage in exposed plants compared to those kept at 20°C. However, BRs-treated plants at 12, 24, and 48 hbfs showed approximately 30% less cellular damage compared to water-sprayed plants. Photosynthetic parameters were affected by freezing stress, leading to reductions in CO₂ assimilation (
