Page 393 - Tandem Techniques

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Figure 9.34. Although the chromatographic separation was not complete, the single ion monitoring of
the cobalt complex ions clearly shows the presence of each carbohydrate.

The use of different reagents to enhance the production of ions in electrospray ionization has been the
subject of a number of investigations, and was employed by Van Breeman [30] in the analysis of
carotenoids. Carotenoids are the metabolic precursors of vitamin A, and are also thought to have
anticancer activity, and to act as in vivo antioxidants. Van Breeman used an LC/MS tandem instrument
in conjunction with the electrospray interface, together with the post-column injection of halogenated
compounds, to enhance the ionization efficiency. The addition of several different halogen compounds
were examined, including chloroform, 2,2,3,3,4,4,4-heptafluoro- 1 -butanol, 2,2,3,3,4,4,4-heptafluoro1-
butyric acid, 1,1,1,3,3,3-hexafluoro-2-propanol and trifluoroacetic acid. The chromatography column
was 12.5 cm long and 4.6 mm I.D., packed with a C18 stationary phase that has been specially prepared
for carotenoid separations. The separation was developed employing a 60 min. linear gradient, from
85:15 to 10:90 methanol/methyl tert-butyl ether, containing 1.0 mM ammonium acetate, at a flow rate
of 1 ml/min. The solute used for testing the different antioxidants was ß-carotene and 0.25 µg was
injected onto the column. It was shown that the presence of the halogen antioxidants did, indeed,
significantly affect the efficiency of ion production. However, in addition, it was evident that an excess
of the additive can also reduce the enhancement significantly. The 1,1,1,3,3,3-hexafluoro-2-propanol
additive provided the greatest sensitivity. The limits of detection were determined for both ß-carotene
(at an m/z value of 536) and lutien (at an m/z value of 568) and the results obtained indicated
sensitivities of about 2 pmol and about 0.6 pmol for ß-carotene and lutein, respectively were easily
obtainable.

An example of the application of the technique to the determination of carotenoids in heat processed
canned sweet potatoes with the post-column addition of heptafluorobutanol is shown in Figure 9.35, (A)
to (E). (A) is the computer reconstructed mass chromatogram of the ß-carotene molecular ion, at an m/z
value of 536, from the injection of ca 20 ng of