The response of periphyton biofilm and the submerged macrophyte tape grass (Vallisneria natans) to internal loading from eutrophic lake sediments were evaluated in microcosms. The sediments from the littoral zone and center of a lake were selected to carry out the microcosm experiment. To determine how the differences in the periphyton biofilm and V. natans growth alone or in combination, we measured changes in water quality, growth, and TP in the periphyton biofilm and V. natans in microcosms containing these sediments. The results showed that the average daily TN and TP removal rates were 32.6 and 35.4%, respectively, in the microcosms containing the lake center sediments by V. natans and the periphyton biofilm. The presence of the periphyton biofilm and V. natans increased the pH, dissolved oxygen, and redox potential and decreased the conductivity in the overlying water in all treatments. Compared to the state before the treatments, V. natans grew well, with a significant increase in biomass (3.1- to 5.5-fold growth) and TP amount (5.1- to 8.8-fold) in all treatments after 48 days. However, the growth of V. natans that combined with the periphyton biofilm was better than that of V. natans alone, as reflected by the dry weight, chlorophyll a content, malondialdehyde content, and TP amount. In conclusion, the periphyton biofilm was beneficial for the growth of V. natans, and the appropriate combination of V. natans and periphyton biofilm would be a potential method for the ecological restoration of eutrophic lakes.