The Use of NMR Detection of LC in Carotenoid Analysis - Carotenoids: Physical, Chemical, and Biological Functions and Properties - page 63

OH

7 11 15 8'

12'

14' 10'

8 10 12 14 15' 11' 7'

HO

All- E lutein

9- Z lutein
9΄- Z lutein

13- Z lutein
13΄- Z lutein

0 5 10 15 20 25 min

All- E lutein

13΄- Z lutein

13- Z lutein

9- Z lutein

9΄- Z lutein

0 5 10 15 20 25 min

FIGURE 4.4 Separation of lutein stereoisomers, comparison between C 18 and C 30 phases. (From Dachtler,
M. et al., J. Chromatogr. B , 211, 1998. With permission.)

explains the retention behavior of different isomers due to their differing abilities to penetrate the
alkyl chain clusters (Albert 1988). Figure 4.4 shows a comparison of the separation of lutein deriva-
tives performed on a C 18 versus a C 30 column (Wise and Sander 1985). It is obvious that the C 18 column
is unable to achieve the resolution necessary for the separation of these different compounds.

4.4 ON-LINE CAPILLARY HPLC-NMR COUPLING
HPLC-NMR analysis in a closed-circuit reveals the stereochemical information for elucidating the
structures of unknown compounds (Albert 2002). In contrast to the technique of off-line separation,
sample collection, and peak identii cation closed-circuit analysis guarantees the absence of isomer-
ization and degradation. Very often only small amounts of sample are available after extraction.