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In this study, the combined effects of hydrological and chemical stressors on benthic macroinvertebrates were evaluated in order to explore the response of the biological community to multiple stressors. The Adige River, located in the south‐eastern Alps, was selected as a case study because representative of the situation of a large river in which the variety of stressors present in the Alpine region act simultaneously. As expected, streamflow showed a seasonal pattern, with high flows in the spring–summer period; however, locally, the natural hydrological regime was altered by the presence of hydropower systems, which chiefly affected low flows. Multivariate analysis showed seasonal and spatial patterns in both chemical and hydrological parameters with a clear gradient in the concentration of nitrate, personal care, and pharmaceutical products moving from headwaters to the main stem of the river. The macroinvertebrate community composition was significantly different in summer and winter and between up and downstream sites. Streamflow alteration chiefly due to water use by hydropower affected community composition but not richness or diversity. Gammarus sp., Hirudinea, and Psychomyia sp., were positively correlated with flow variability, increasing their densities in the sites with higher streamflow variability because of hydropeaking. The results obtained in this study show that the composition of the macroinvertebrate community responded to seasonality and to changes in the main stressors along the river and highlights the importance of the spatial and temporal variability of stressors in this Alpine river. Taking into account, this variability will help the decision‐making process for improving basin management. [ABSTRACT FROM AUTHOR]