The study employed batch shake flasks to evaluate the impact of various nitrogen sources, phosphate levels, and sodium acetate (Na-acetate) on the Rhodotorula mucilaginosa growth and metabolite production. Adding Na-acetate to the medium resulted in significant improvements in critical metabolites. In shake flask experiments, this led to a cell dry weight (CDW) of 1.65 ± 0.94 g L-1, with lipids comprising 66.53% of the biomass. While β-carotene and carotenoid were 5.84 ± 0.05 and 37.66 ± 2.13 µg g-1, respectively. Subsequent experiments in a batch reactor with Na-acetate supplementation further improved these metrics. CDW increased to 5.02 ± 0.83 g L-1, and lipid content to 65.73 ± 0.81%. Carotenoid production rose to 40.33 ± 1.84 µg g-1, with β-carotene reaching 17.63 ± 0.32 µg g-1. The most promising results were obtained using a fed-batch bioreactor strategy with Na-acetate. R. mucilaginosa achieved the highest yields across all parameters: 48.36 ± 1.14 µg g-1 of total carotenoids, 21.38 ± 1.14 µg g-1 of β-carotene, and a lipid content of 68.58 ± 1.95%.