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Enhancement in the production of β-carotene by the hyper producer mutant 32 of Rhodotorula glutinis by manipulation of temperature and illumination.Growth and β-carotene production was investigated in a 1 litre fermenter at different temperature and illumination conditions. The optimum temperature for growth and β-carotene production was 30 and 20°C, respectively. At 30°C, β-carotene production was 125 ± 2 mg l−1 and accounted for 66% of the total carotenoids in 72 h; at 20°C, it was 250 ± 7 mg l−1 and accounted for 92% of total carotenoid content. Continuous illumination of the fermenter by 1000 lx white light hampered growth as well as carotenoid synthesis. At 30°C, illuminating the fermenter in late logarithmic phase resulted in a 58% increase in β-carotene production with a concurrent decrease in torulene; at 20°C, however, it showed no appreciable increase.Proper manipulation of culture conditions enhanced β-carotene production by R. glutinis which makes it a significant source of β-carotene.