Effect of Mobile Phase and Modifiers on Carotene Separations

Supercritical CO2 has been the most commonly used mobile phase in SFC. It is nontoxic and also has a low critical temperature (31°C), which enables the separation of thermally labile compounds such as caro-tenoids at low column temperatures. Furthermore, it

Echinenone

Figure 1 Chemical structures of some basic carotenoids.

Table 1 Advances in SFC of carotenoid pigments

Date Development

1968 Separation of a- and ^-carotene by packed-column SFC

(Giddings etal.) 1983 Separation of lycopene and a- and ^-carotene (Gere) 1989 Separation of geometrical isomers of a- and ^-carotene by open tubular SFC (Schmitz et al.) 1991 Understanding of the effect of temperature, pressure, modifier and stationary-phase type on carotene separations (Lesellier et al., Aubert et al.) 1994 Investigation of spectral shifts of carotenoids in supercritical CO2 (Hui etal.)

is compatible with a wide range of HPLC and GC detectors. Nitrous oxide exhibits a polarity similar to CO2 but has only received limited use because of its oxidizing properties. Nevertheless, both of these mobile phases have been investigated with ^-carotene as a probe molecule. Interestingly, the retentive properties of the polar amino stationary phase for ^-caro-tene are largely affected by the CO2 and N2O mobile phases, in comparison to the nonpolar octa-decylsilyl stationary phase, which is not appreciably affected by these mobile phases.

In cases where increased solubilizing power is required to elute components of interest, modifiers have been successfully added to the supercritical fluid. The resulting effect is reduced solute retention, improvement of chromatographic efficiency and, in some cases, altered elution order. One of the advantages of supercritical CO2 is that it is chemically compatible and miscible with a large number of modifiers. The effects of a range of modifiers - methanol, acetonit-rile, tetrahydrofuran, dichloromethane and trich-lorotrifluoroethane - on the SFC of carotenes have been studied using binary and ternary mixtures with CO2 containing 3-20% (v/v) of the modifiers. Addition of each of these modifiers produces a concentration-dependent decrease in the capacity factors (£') of the carotenes. The effectiveness of the modifiers has not been directly correlated with their densities nor with their polarities, and this suggests that specific interactions between the solutes and the modifiers are important.

Minimal retention is obtained with the strongest modifier, dichloromethane, which is closest to the dipole-dipole interaction of Snyder's triangle. Furthermore, increasing the dichloromethane concentration in the CO2 results in an overall decrease in retention of the carotenes and also a decrease in selectivity, probably due to reduced interactions between the solutes and the stationary phase, as all the carotenes have high affinities for this solvent. A plot of 1/k' against modifier concentration is linear for a mixture of tetrahydrofuran-methanol in CO2 whereas, for more polar modifiers, the line curves slightly downwards due to increased solvent polarity, which indicates that solute-solvent interactions are important. On the other hand, an exponential curve is observed for the nonpolar modifiers, suggesting that the solubility of the carotenes is enhanced by an amount independent of the concentration of the modifier.

The selectivity between the trans and cis isomers is unaffected by the modifiers, whereas the selectivity between the a- and ^-all-trans compounds diminishes with increasing percentage of modifier. With a CO2-methanol-acetonitrile mixture, the selectivity between trans and cis isomers falls as the proportion of acetonitrile is increased, for both the a- and ^-carotenes. This phenomenon has been used to advantage in optimizing the separation of the components of a carrot extract, as shown in Figure 2. With methanol-carbon dioxide, the peaks were not well resolved; however, replacing some of the methanol with acetonitrile allowed on additional component in the extract to be resolved and eluted before all-trans a-carotene.

Solar Panel Basics

Solar Panel Basics

Global warming is a huge problem which will significantly affect every country in the world. Many people all over the world are trying to do whatever they can to help combat the effects of global warming. One of the ways that people can fight global warming is to reduce their dependence on non-renewable energy sources like oil and petroleum based products.

Get My Free Ebook


Post a comment