Simultaneous determination of eight vitamin D2 isomers by proton magnetic resonance spectroscopy

Simultaneous determination of vitamin D2 and its isomers, with the aid of pmr employing a paramagnetic shift reagent Eu (dpm)3, is presented. Thus, a mixture of vitamin D2 isomers is acetylated and its pmr spectra are run with and without the addition of the shift reagent. Relative paramagnetic shift values (gamma rel) of the acetoxyl protons of the acetylated isomers are estimated for solutions in CCl4 against the reference acetoxyl signal of vitamin D2 acetate. Estimated values are as follows: vitamin D2 acetate = 100, 5,6-trans-vitamin D2 acetate = 71 +/- 1, ergosteryl acetate = 86 +/- 1, lumisteryl2 acetate = 123 +/- 3, isovitamin D2 acetate = 82, isotachysteryl2 acetate = 97, tachysteryl2 acetate = 100 (105 +/- 1 in C6D6), and previtamin D2 acetate = 80 +/- 2. On the basis of a dramatic shift of an acetoxyl signal and a constancy of each gamma rel, this method permits immediate identification of the isomers and provides valuable information on their stereochemistry. This technique also facilitates a simultaneous determination of these isomers in a mixture by estimating each signal area.