The substantial discharge of vinasse biogas slurry (VBS) from anaerobic digestion poses significant environmental risks, while microalgae could recover nutrient for biomass production from VBS treatment. This study comprehensively investigated the bio-remediation potential of microalgae via rotating biofilm reactor, focusing on microalgae species and inoculation ratios, as well as VBS dilution ratios. The results revealed that Desmodesmus sp. had a better performance in biomass productivity (0.62 g/L), and total phosphorus (TP) removal efficiency (77.03%) than the other two microalgae (p < 0.05). The low VBS proportion (≤30%) obtained a higher biomass accumulation of 0.87–0.89 g/L than groups of high VBS proportion of 50%. These growth inhibition in high VBS proportion groups were attributed to high ammonium nitrogen (NH₄⁺-N) toxicity and elevated turbidity of biogas slurry. Both the total nitrogen (TN) removal of 34.11–36.48% and NH₄⁺-N removal of 83.63–97.38% revealed ammonium assimilation during the treatment of VBS via microalgae. Inoculum optimization demonstrated that reduced initial inoculation ratio (10% v/v) obtained highest TN removal efficiency of 46.52% and biomass growth rate of 0.04 g/(L·d), likely through alleviated light limitation. In the rotating biofilm reactor, treatment with 50-time diluted VBS achieved the highest biomass growth rate of 1.60 g/(m²·d), and the removal of TN, NH₄⁺-N, and TP was 74.93%, 93.94% and 98.01%, respectively. Moreover, coarse ribbed fiber fabric (CRFF) exhibited higher attached biomass (48.76 g/m²) and NH₄⁺-N removal efficiency (76.30%) than other attached materials. The research offered insights into trade-offs between the nutrient recovery and accumulating microalgae biomass during biogas slurry treatment.