ABSTRACT Outcrops play an important role in groundwater recharge. Understanding groundwater origins, dynamics and its correlation with different water sources is essential for effective water resources management and planning in terms of quantity and quality. In the case of the Guarani Aquifer System (GAS) outcrop areas are particularly vulnerable to groundwater pollution due to direct recharge processes. This study focuses on the Alto Jacaré-Pepira sub-basin, a watershed near Brotas, a city in the central region of the state of São Paulo, Brazil, where groundwater is vital for supporting tourism, agriculture, urban water supply, creeks, river and wetlands. The area has a humid tropical climate with periods of both intense rainfall and drought, and the rivers remain perennial throughout the year. Therefore, the aim of this study is to investigate the interconnections between a spring and its potential sources of contribution, namely rain and groundwater, in order to elucidate the relationships between the different water sources. To achieve this, on-site monitoring of groundwater depth, rainfall amount, and stable isotope ratios (deuterium (2H) and oxygen-18 (18O)) from rain, spring discharge, and a monitoring well was carried out from 2013 to 2021. The results indicate that the mean and standard deviations for δ18O in rainwater exhibit higher variability, resulting in −4.49 ± 3.18 ‰ VSMOW, while δ18O values from the well show minor variations, similar to those of the spring, recording −7.25 ± 0.32 ‰ and −6.94 ± 0.28 ‰ VSMOW, respectively. The mixing model’s outcomes reveal seasonal variations in water sources contribution and indicate that groundwater accounts for approximately 80 % of spring discharge throughout the year. Incorporating stable isotopes into hydrological monitoring provides valuable data for complementing watershed analysis. The values obtained support the significance of the aquifer as a primary source, thereby offering critical insights into stream dynamics of the region.