The Water Framework Directive requires all water bodies in EU countries to ensure the ecological integrity of freshwater ecosystems in all water bodies. A “good” status in water bodies leads to their biological elements to reproduce reference conditions, so quantity and timing should be considered in describing environmental flows. However, the availability of daily natural flow data is one of the major drawbacks, especially in highly water-regulated countries, such as Spain, and hydrological models, such as the Soil and Water Assessment Tool (SWAT+) model, can be used to generate them. This work evaluated three different approaches for obtaining daily flows used to characterise the natural regime and to estimate environmental flows in the Piloña River (northern Spain) and Tagus River (centre Spain) basins: Scenario 1) daily flows simulated by the SWAT+ model calibrated on a monthly scale; Scenario 2) daily flows estimated by applying monthly-to-daily disaggregation techniques from the monthly series obtained from the SWAT+ model calibrated on a monthly scale using Water Rights Analysis Package (WRAP) software; and Scenario 3) daily flows simulated by the SWAT+ model calibrated on a daily scale. The Kling–Gupta efficiency (KGE), percent bias (PBIAS), Nash–Sutcliffe efficiency (NSE), root mean square error (RMSE) and standard deviation ratio (RSR) were used to evaluate the model’s performance. Furthermore, the indicators of hydrologic alteration in rivers (IAHRIS) and the basic flow methods were assessed and compared with those obtained directly using the real data observed at the gauging station. Our results indicated that Scenario 2 outperformed the rest of the approaches for all the statistics used. KGE and NSE varied between 0.78 and 0.92 for both basins in Scenario 2 while these values decreased to 0.50–0.60 for KGE and 0.20–0.40 for NSE in Scenarios 1 and 3. Moreover, PBIAS ranged between 3.24 and 4.47% in validation periods in Scenario 2, while it decreased to −14.64% in Scenario 3. Likewise, Scenario 2 was also observed to show better performance related to both the patterns of the natural regime and the basic flow values, which were generally underestimated by the other scenarios. The methodology proposed provides a valuable tool for evaluating and managing the hydrological and environmental status of river basins.