This study focused on the simultaneous recovery of carbohydrates (CHO) and phosphorus (P) from Desmodesmus sp. biomass cultivated in municipal wastewater, through a sequential pretreatment. The pretreatment consisted first of ultrasound to trigger cell disruption followed by ozonation to recover CHO and P. For ozone pretreatment, three different parameters were considered: ozone concentration (9, 15, 21, 27, 36, and 45 mg O3/L), contact time (15, 25 and 35 min), and pH (8 and 11). The maximum simultaneous release of 84% of CHO and 58% of P was achieved at the experimental parameters of ozone concentration of 45 mg O3/L, contact time of 35 min, and pH of 11. Also, P was concentrated in solution by 8- to 14-fold with respect to municipal wastewater. The sequential pretreatment was conducted at alkaline pH of 11 and atmospheric conditions, which may considerably reduce energy demand and reagents, in comparison to a traditional hydrolysis pretreatment. The results found suggest that the sequential pretreatment could be feasible on a large scale.