The culture characteristics, carotenoid production, and associated biosynthetic pathway of strain T-1 were examined. As a result of examining the culture temperature and light irradiation, an increase of neurosporaxanthin and neurosporaxanthin beta-D-glucopyranoside was observed at a low temperature and 0 lx. It was suggested that highly polar carotenoids, such as neurosporaxanthin, and carotenoid glycosides were involved in the stabilization of membrane during nutrition storage other than the defense function of fungus bodies. Strain T-1 produced lycopene, beta-carotene, gamma-carotene, torulene, neurosporaxanthin, and neurosporaxanthin beta-D-glucopyranoside, as assessed by HPLC, LC-MS, and NMR analysis. Carotenoid biosynthesis begins with neurosporene, passing to lycopene and gamma-carotene through cyclization, and produces beta-carotene. In addition, it is saturated, gamma-carotene is converted to torulene, and neurosporaxanthin is produced. Thus, the carotenoid biosynthetic pathway in strain T-1 was estimated.