Freshwater ecosystems play an important role in transporting and accumulating microplastics. Spatial and temporal variability in microplastic pollution can create critical spots and moments of elevated pollution, however, the consequences of their interaction are still poorly understood. This study aimed to assess the interaction between urbanization and flood episodes on river microplastic pollution. The water surface was sampled in two sites of the Garonne River, upstream and downstream a large urban area, during two flood episodes. Samples were chemically digested to facilitate particles isolation, and microplastics (700 μm-5 mm) were characterized through infrared spectroscopy (ATR-FTIR). Microplastic concentration increased by 5-8 fold during flood episodes, driven by river discharge. This increase was more significant in the downstream site. During the flood, there was an overall increase of larger particles on water surface, but only in the downstream site microplastic colours and polymeric compositions significantly varied. Principal component analysis of infrared spectra from polyethylene microplastics revealed that the main variance in the spectral region corresponded to hydroxyl and carbonyl groups. The carbonyl content in microplastics was significantly higher for particles collected during the flood, likely indicating a higher level of degradation. Urbanization modulates freshwater microplastic pollution during floods, and changes in microplastic physicochemical profile should be further integrated within toxicity studies to evaluate risks potentially elevated to animal and human health.