Microbial consortia in riverbed substrates and their extracellular matrix (biofilms) play a key role in phosphorus (P) entrapment. When P entrapment saturates, the benthic compartment changes from a P sink to a P source thus increasing eutrophication risk. P entrapment saturation is expected to differ between intracellular and extracellular P entrapment and between different magnitudes and durations of P inputs. We studied biofilm P-entrapment following short (48 h) and long (14 days) P loading events in stream bypass flumes supplied with a gradient of dissolved P concentrations. This allowed us to link local biofilm processes in sediments to potential effects on river self-purification, via quantifying the P removal efficiency in the flumes. We found that in short-term events, biofilms develop intracellular mechanisms to cope with P inputs, while long-term events and high P inputs suppress the intracellular uptake mechanisms and increase the prevalence of extracellular entrapment. Specifically, long-term events lowered the threshold for intracellular P entrapment saturation, and decreased the ratio between intracellular and extracellular entrapment resulting in lower removal efficiency for dissolved phosphorus. Our results highlight the risk that aquatic ecosystems may face as the ratio of intracellular to extracellular P entrapment decreases, which may reduce their ability to deal with P inputs, thereby increasing risks of eutrophication.