Polar compounds retention

The phases were tested for polar compound retention by running a gradient method using a set of probe compounds made up of samples and standards that were effectively unretained on the generic LC/MS method. To compare the different stationary phases... 

Polar compounds present the most problems because of their low breakthrough volumes with common sorbents. In the last few years, highly crosslinked polymers have become commercially available which involve higher retention capacities for the more polar analytes (37, 38). Polymers have also been chemically modified with polar groups in order to increase the retention of the compounds previously mentioned (35, 37). 

When analytes lack the selectivity in the new polar organic mode or reversed-phase mode, typical normal phase (hexane with ethanol or isopropanol) can also be tested. Normally, 20 % ethanol will give a reasonable retention time for most analytes on vancomycin and teicoplanin, while 40 % ethanol is more appropriate for ristocetin A CSP. The hexane/alcohol composition is favored on many occasions (preparative scale, for example) and offers better selectivity for some less polar compounds. Those compounds with a carbonyl group in the a or (3 position to the chiral center have an excellent chance to be resolved in this mode. The simplified method development protocols are illustrated in Fig. 2-6. The optimization will be discussed in detail later in this chapter. 

This technique is based on the same separation mechanisms as found in liquid chromatography (LC). In LC, the solubility and the functional group interaction of sample, sorbent, and solvent are optimized to effect separation. In SPE, these interactions are optimized to effect retention or elution. Polar stationary phases, such as silica gel, Florisil and alumina, retain compounds with polar functional group (e.g., phenols, humic acids, and amines). A nonpolar organic solvent (e.g. hexane, dichloromethane) is used to remove nonpolar inferences where the target analyte is a polar compound. Conversely, the same nonpolar solvent may be used to elute a nonpolar analyte, leaving polar inferences adsorbed on the column. 

Reversed-phase extraction of nonpolar compounds. Provides less retention of hydrophobic compounds. Normal phase extraction of polar compounds. Adsorption of polar coeqsounds. 

The instrument used to generate the data shown in Figures 1 and 2 (LECO Pegasus III GC x GC-ToF-MS) has a modulator at the end of the first 30 mx 0.25 mm non-polar column (HP-5MS, 0.25 pm film thickness). As compounds elute from this column, the modulator concentrates them over a short period to focus them and then sends them down the second, shorter and narrower 2 m x 0.10 mm, polar column (BPX-50, 0.10 pm film thickness) situated in its own oven compartment within the main oven. This operation is repeated throughout the analytical run. Having the two columns coupled in this way allows compounds to be separated by volatility on the first analytical column and by polarity on the second column. Hence for complex mixtures, peaks with a similar (or identical) retention on the first column can be separated by the second column. Non-polar compounds emerge before polar components. 

Retention of polar compounds is greater than in RP-HPLC... 

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