β-Isocryptoxanthin (BICX2), which is naturally occurring carotenoid, was isolated from sea urchins as a racemate. In the present study we analyzed the CD spectra observed for (4R)- and (4S)-BICX2 (BICX2R and BICX2S, respectively) and the corresponding benzoates (BICX3R and BICX3S, respectively) by time-dependent density functional theory (TD-DFT). The separation of the two enantiomers of β-isocryptoxanthin from the racemate was achieved by HPLC on a Chiralcel OD column after conversion to the corresponding monobenzoates. Saponification of each separated monobenzoate gave optically pure BICX2R and BICX2S. First, CD spectra of BICX2 and BICX3 were measured. Next the simulation of CD spectra for BICX2R based on TD-DFT calculations was performed by considering the following three types of conformers: 1) rotational isomers around R1 and R2 caused by steric hindrance between the methyl and methine groups, 2) stereoisomers based on rotation (R3) of thd hydroxy group, 3) conformational isomers of the cyclohexane rings base on puckerin near P1 and P2. The following points were clarified by TD-DFT calculations: 1) the effect of conformation of CD spectrum of BICX2R decreases in the order of R1, R2≫P1, P2>R3, 2) the contrast between the observed CD bands of BICX2R and BICX3R reflects a populaton of each conformer in equilibrium, 3) the intense positive CD band at around 230 nm is insensitive to the conformational changes, indicating that the absolute configuration of compounds similar to β-isocryptoxanthin can be suitably determined by using this CD band.
