Rhodotorula yeasts which are known as carotenogenic yeasts have a great industrial value due to their ability to produce carotenoids. In particular, the isolated yeast Rhodotorula sp. (strain ATL72) has been reported to be a promising producer of high concentrations of carotenoids. A combination of central composite design (CCD) and Plackett–Burman (PB) design was used to optimize carotenoids produced by this yeast. The optimum production of carotenoids was completed when the yeast was grown in a production medium composed of 3.7 g/L malt extract, 7.7 g/L fructose, 9 g/L urea, 35 g/L NaCl, and 1 g/L yeast extract at 27.5 °C, pH 6.7, and 180 rpm. Two batch runs in 1 L and 7 L bioreactors were conducted which increased the productivity of carotenoid concentration from 21.5 mg/L after 98 h of incubation at the level of the shake flask to 229.9 mg/L after 47 h of incubation at the level of 7 L bioreactor. The carotenoid pigment was extracted in dimethylsulfoxide (DMSO), acetone, petroleum ether, and sodium chloride, and subsequently identified and characterized using UV-visible scanning, thin layer chromatography, and gas chromatography/mass spectrometry.