Characterization of various isomeric photoproducts of ergosterol and vitamin D2 generated by UV irradiation

Vitamin D-2 is produced from its precursor ergosterol under the impact of ultraviolet (UV) light which is also commercially carried out to increase vitamin D-2 contents in mushrooms ('Novel Food'). However, this process is accompanied by the formation of various isomers that partly co-elute with the target compound and are currently difficult to analyze. For this reason, vitamin D-2 and ergosterol were irradiated with the goal to generate and characterize various isomeric photoproducts with three analytical methods. High-performance liquid chromatography with ultraviolet detection (HPLC-UV) was accompanied by using a chiral detector (CD) which was serially linked with the UV detector. Applied for the first time in this research area, HPLC-CD chromatograms provided complementary information which was crucial for the identification of several co-elutions that would have been overlooked without this approach. Additional information was derived from gas chromatography with mass spectrometry analysis. Diagnostic fragment ions in the GC/MS spectra allowed to distinguish four classes of tri- (n = 2), tetra-, and pentacyclic isomer groups. Despite several drawbacks of each of the applied methods, the shared evaluation allowed to characterize more than ten isomeric photoproducts of vitamin D-2 including previtamin D-2, lumisterol(2,) tachysterol(2,)trans-vitamin D-2 isomers, and two pentacyclic isomers (suprasterols(2) I and II), which were isolated and characterized by proton magnetic resonance spectroscopy (H-1 NMR).