1. River regulation and abstraction have dramatically altered the natural flow regime of many rivers world-wide, but experimental investigations of the biological effects are infrequent. In the mid-region of the Murray River, Australia, river regulation has reduced the frequency and duration of spring floods and increased the frequency of summer floods. We used controlled floods (treatments: no floods, spring floods, summer floods and spring + summer floods) to determine how the growth of river red gum Eucalyptus camaldulensis trees, aquatic macrophytes and biofilms varied with the seasonal timing and frequency of flooding.2. After 6 years of controlled flooding, above-ground net production of wood by river red gum trees was equal and greatest in plots receiving spring + summer floods and summer floods (mean 496 g m(-2) year(-1)). Production was significantly lower in plots receiving spring floods or no controlled floods, which had similar rates of production (mean 330 g m(-2) year(-1)).3. During 2 years of measurement in wetlands created by flooding, production and species richness of aquatic macrophyte were both greater in spring than in summer floods. The history of flood frequency at any experimental site did not affect macrophyte production or species richness. The aquatic macrophyte community in shallow regions of wetlands differed significantly with the seasonal timing of floods, but not flood frequency.4. The accumulation of chlorophyll a and total mass of biofilm on wood surfaces in wetlands created by flooding were greater in spring (mean chlorophyll a, 0.88 mug cm(-2); mean mass, 0.066 mg cm(-2)) than in summer floods (mean chlorophyll a, 0.09 mug cm(-2); mean mass, 0.034 mg cm(-2)). The history of flood frequency at any experimental site did not affect accumulation of either the autotrophic or heterotrophic components of biofilms.5. Spring flooding, while not as beneficial for tree growth, is critical for the growth of wetland macrophytes, the maintenance of macrophyte species richness, and favours better development of autotrophic biofilms. Maintenance of both the timber harvest and wetland conservation values of these floodplains will require the return of more natural flood flows in the spring period. Restoration of floodplain rivers requires a thorough understanding of the relationships between ecological functions and the natural flow regime.