The carotenoids lutein and capsanthin, widely used as colorants, mainly occur as esters of fatty acids in their producer plants marigold and paprika. The enzymatic hydrolysis of these esters is reported here. Out of 26 commercial lipases, fastest ester hydrolysis under conditions was obtained using an enzyme from Candida antarctica (69% release of capsanthin, 44% of lutein from the respective oleoresins). Bile salts were essential for the activity of all commercial enzymes. A novel hydrolase was discovered in the mycelium-free supernatant of submerged cultures of the edible basidiomycete Pleurotus sapidus. The fungal enzyme cleaved several carotenoid esters almost completely and in the absence of bile salts.