This study investigated the seasonal water quality characteristics of two key environmental flow sources for the Gwangju Stream: the Yeongsan River water supply and effluent from a sewage treatment plant. Monitoring data collected between 2019 and 2023 were analyzed for both sources, and field surveys from October 2023 to June 2024 examined confluence points where environmental flows entered the Gwangju Stream, measuring both main-stream and inflow waters. The Yeongsan River supply recorded its highest spring organic matter levels (mean BOD: 5.2 mg L-1; maximum: 8.7 mg L-1), while the sewage treatment plant effluent exhibited pronounced seasonal variation in total nitrogen (T-N), ranging from a summer low of 8.2 mg L-1 to a winter high of 13.8 mg L-1. Upstream water quality remained stable; however, downstream BOD increased annually by 8.2%, and total phosphorus (T-P) peaked sharply in summer (0.567 mg L-1). Field survey results indicated that in spring, T-N increased by up to 495%, BOD by 182%, and T-P by 290%; in winter, T-N rose by 239%, BOD by 164%, and COD by 73%. These findings reveal marked seasonal variability in the influence of environmental flow sources, with T-N showing the most substantial increase in spring. The results highlight the need for targeted nutrient management strategies, such as increasing the proportion of the plant’s effluent in spring to stabilize nutrient loads and improving its biological treatment efficiency in winter to reduce T-N concentrations. Season-specific measures of this kind can improve water quality and help sustain the ecological integrity of the Gwangju Stream.