Normal-phase liquid chromatography separations

One example of normal-phase liquid chromatography coupled to gas chromatography is the determination of alkylated, oxygenated and nitrated polycyclic aromatic compounds (PACs) in urban air particulate extracts (97). Since such extracts are very complex, LC-GC is the best possible separation technique. A quartz microfibre filter retains the particulate material and supercritical fluid extraction (SPE) with CO2 and a toluene modifier extracts the organic components from the dust particles. The final extract is then dissolved in -hexane and analysed by NPLC. The transfer at 100 p.1 min of different fractions to the GC system by an on-column interface enabled many PACs to be detected by an ion-trap detector. A flame ionization detector (PID) and a 350 p.1 loop interface was used to quantify the identified compounds. The experimental conditions employed are shown in Table 13.2. 

Normal-phase liquid chromatography is thus a steric-selective separation method. The molecular properties of steric isomers are not easily obtained and the molecular properties of optical isomers estimated by computational chemical calculation are the same. Therefore, the development of prediction methods for retention times in normal-phase liquid chromatography is difficult compared with reversed-phase liquid chromatography, where the hydrophobicity of the molecule is the predominant determinant of retention differences. When the molecular structure is known, the separation conditions in normal-phase LC can be estimated from Table 1.1, and from the solvent selectivity. A small-scale thin-layer liquid chromatographic separation is often a good tool to find a suitable eluent. When a silica gel column is used, the formation of a monolayer of water on the surface of the silica gel is an important technique. A water-saturated very non-polar solvent should be used as the base solvent, such as water-saturated w-hexane or isooctane. 

Example 2 Chromatography of nitroaniline isomers. The elution order of the nitroaniline isomers was ortho, meta, and para in normal-phase liquid chromatography using H-butanol-w-hexane mixtures as the eluent, when the stationary phase material was either silica gel, alumina, an ion-exchanger, polystyrene gel, or octadecyl-bonded silica gel. The results indicate that the separation of these compounds can be performed on a range of different types of stationary phase materials if the correct eluent is selected. The best separation will be achieved by the right combination of stationary phase material and eluent.68... 

The qualitative analysis of retention behaviour in liquid chromatography has now become possible. Quantitative retention-prediction is, however, still difficult the prediction of retention time and the optimization of separation conditions based on physicochemical properties have not yet been completely successful. One reason is the lack of an ideal stationary phase material. The stationary phase material has to be stable as part of an instrument, and this is very difficult to achieve in normal-phase liquid chromatography because the moisture in organic solvents ages the silica gel. 

Perrin, C., Vu, V.A., Maftouh, M., Massart, D.L., Vander Heyden, Y. Screening approach for chiral separation of pharmaceuticals Part I. Normal-phase liquid chromatography. J. Chromatogr. A 2002, 947, 69-83. 

Attempts were first made in this study to separate dimers by normal-phase liquid chromatography. Silica gel absorption chromatography was first used as a separation method. Hexane, as the mobile phase, gave very long retention times for the various dimers, and no separation was... 

AA Karlsson, P Michelsen, A Larsen, G Odham. Normal-phase liquid chromatography class separation and species determination of phospholipids utilizing electrospray mass spectrometry/tandem mass spectrometry. Rapid Communications Mass Spectrom 10 775-780, 1996. 

Stenhoff et al. [117] determined enantiomers of omeprazole in blood plasma by normal-phase liquid chromatography and detection by atmospheric-pressure ionization tandem mass spectrometry. The enantioselec-tive assay of omeprazole is using normal-phase liquid chromatography on a Chiralpak AD column and detection by mass spectrometry. Omeprazole is extracted by a mixture of dichloromethane and hexane and, after evaporation, redissolution and injection, separated into its enantiomers on the chiral stationary phase. Detection is made by a triple quadrupole mass spectrometer, using deuterated analogs and internal standards. The method enables determination in plasma down to 10 nmol/1 and shows excellent consistency suited for pharmacokinetic studies in man. 

It also was reported by Lagerstroem et al that pantoprazole and its enantiomers can be separated by using normal-phase liquid chromatography, and detection by atmospheric pressure ionization tandem mass spectrometry [13]. 

Whilst gas chromatography has been used for the analysis of many of the lycoctonine-based alkaloids [52], the larger, less volatile, and more thermally labile MSAL compounds require analytical procedures such as TLC and HPLC for separation and detection. For example, both normal phase liquid chromatography [53] and reversed phase liquid chromatography [54] with UV detection have been used for separation, detection, and quantitation of alkaloids from Delphinium species associated with livestock poisonings in the western US and Canada. The introduction of API techniques has allowed the analysis of all types of diterpene alkaloids by direct MS methods and with MS methods coupled to liquid chromatography. 

In normal-phase liquid chromatography on silica and alumina the hydrogen bonding of fullerenes with surface hydroxyl groups of adsorbent is the most important. On separation of Ceo and C70 on a column packed by alumina Alusorb N 200 from n hexane the selectivity is 1.80 (Fig. 2 a.). The reversed-phase adsorbents such as silica with bonded diphenylsilyl... 

Andersson, J. T., Separations on a mercuric-acetate-substituted phenylsihcia phase in normal-phase liquid chromatography, Fresenius Z. Anal. Chem., 326, 425-433, 1987. 

Usually, different types of solvent mixtures are used in normal-phase liquid chromatography, especially in thin-layer liquid chromatography. For example, sedative medications were separated hy thin-layer liquid chromatography using different types of eluents (developing solvents). 

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