Environmental conditions resulting from climate change may have diverse impacts on the safety and performance of infrastructure. Bridges, as pivotal structural components of transportation systems, are subject to various natural and environmental factors. Therefore, periodic definition of their dynamic characteristics, such as natural frequencies through experimental measurements, is crucial for promptly and accurately assessing their current state. In this study, experimental analysis is employed for this purpose, involving the measurement of structural responses under ambient conditions. This paper presents an investigation into the environmental effects on the dynamic characteristics of reinforced concrete (RC) frame bridges. The study focuses on two overpasses with similar geometries. Three sets of measurements were conducted for each overpass: in October 2017, March 2020, and May 2022. The identified dynamic characteristics were compared across different time points and correlated with environmental effects. The analysis results indicate that the identified natural frequencies effectively reflect changes in the dynamic characteristics of the overpasses due to environmental effects. A significant difference in identified natural frequencies is observed in the longitudinal direction, while minimal variation occurs in the vertical direction.