We isolated mutants of Phaffia rhodozyma strain NRRL Y-17269 that overproduced astaxanthin when grown on corn-based fuel ethanol stillage (thin stillage, TS, or fuel ethanol byproducts). Ten mi cultures of mutant strain JB2 produced 1.54+/-0.21 mu g carotenoid/mg dry weight when grown on 70% thin stillage at pH 5.2, compared with 0.38+/-0.04 mu g/mg produced by the parental strain. Furthermore, JB2 produced similar astaxanthin concentrations when grown in either thin stillage or yeast malt broth. By comparison, previously described astaxanthin overproducing strain NRRL Y-17811 yielded 1.08+/-0.07 mu g/mg in yeast malt broth but only 0.67+/-0.03 mu g/mg in thin stillage. Five liter fermentation experiments using JB2 grown on 70% thin stillage at pH 5.2 yielded 2.01+/-0.17 mu g/mg astaxanthin. Thus, JB2 is uniquely suited for astaxanthin production from low cost thin stillage.