Chromatographic systems normal phase

For HPLC-FTIR, GPC-FTIR, or SFC-FTIR, the design of the interface is more challenging since the mobile phases used for these chromatographic systems normally have strong infrared absorbencies thus, it is important to remove the mobile phase prior to measuring the spectrum. For the interface between the two systems flow-cells or mobile-phase elimination techniques may be used. Some recent developments point toward the elimination of mobile-phase techniques. A microbore column can help to reduce the mobile-phase volume in the system. ... 

As described above, the mobile phase carrying mixture components along a gas chromatographic column is a gas, usually nitrogen or helium. This gas flows at or near atmospheric pressure at a rate generally about 0,5 to 3.0 ml/min and evenmally flows out of the end of the capillary column into the ion source of the mass spectrometer. The ion sources in GC/MS systems normally operate at about 10 mbar for electron ionization to about 10 mbar for chemical ionization. This large pressure... 

The problem is made more difficult because these different dispersion processes are interactive and the extent to which one process affects the peak shape is modified by the presence of another. It follows if the processes that causes dispersion in mass overload are not random, but interactive, the normal procedures for mathematically analyzing peak dispersion can not be applied. These complex interacting effects can, however, be demonstrated experimentally, if not by rigorous theoretical treatment, and examples of mass overload were included in the work of Scott and Kucera [1]. The authors employed the same chromatographic system that they used to examine volume overload, but they employed two mobile phases of different polarity. In the first experiments, the mobile phase n-heptane was used and the sample volume was kept constant at 200 pi. The masses of naphthalene and anthracene were kept... 

Samples and reference substances should be dissolved in the same solvents to ensure that comparable substance distribution occurs in all the starting zones. In order to keep the size of the starting zones down to a minimum (diameter TLC 2 to 4 mm, HPTLC 0.5 to 1 mm) the application volumes are normally limited to a maximum of 5 xl for TLC and 500 nl for HPTLC when the samples are applied as spots. Particularly in the case of adsorption-chromatographic systems layers with concentrating zones offer another possibility of producing small starting zones. Here the applied zones are compressed to narrow bands at the solvent front before the mobile phase reaches the active chromatographic layer. 

From the general framework of the Snyder and Soczewinski model of the linear adsorption TLC, two very simple relationships were derived, which proved extremely useful for rapid prediction of solute retention in the thin-layer chromatographic systems employing binary mobile phases. One of them (known as the Soczewinski equation) proved successful in the case of the adsorption and the normal phase TLC modes. Another (known as the Snyder equation) proved similarly successful in the case of the reversed-phase TLC mode. 

The choice of the chromatographic system depends on the chemical character of the extracts being separated. The mobile phase should accomplish all requirements for PLC determined by volatility and low viscosity, because nonvolatile components (e.g., ion association reagents and most buffers) should be avoided. It means that, for PLC of plant extracts, normal phase chromatography is much more preferable than reversed-phase systems. In the latter situation, mixtures such as methanol-ace-tonitrile-water are mostly used. If buffers and acids have to be added to either the... 

Radke et al. [28] described an automated medium-pressure liquid chromatograph, now commonly called the Kohnen-Willsch instrument. At present, the method is widely used to isolate different fractions of soluble organic matter (for instance, as described in Reference 29 to Reference 31). A combination of normal phase and reversed-phase liquid chromatography has been used by Garrigues et al. [32] to discriminate between different aromatic ring systems and degrees of methylamine in order to characterize thermal maturity of organic matter. 

The PRISMA model was developed by Nyiredy for solvent optimization in TLC and HPLC [142,168-171]. The PRISMA model consists of three parts the selection of the chromatographic system, optimization of the selected mobile phases, and the selection of the development method. Since silica is the most widely used stationary phase in TLC, the optimization procedure always starts with this phase, although the method is equally applicable to all chemically bonded phases in the normal or reversed-phase mode. For the selection of suitable solvents the first experiments are carried out on TLC plates in unsaturated... 

Jandera, P., Holcapek, M., Theodoridis, G. (1998). Investigation of chromatographic behavior of alcohol ethoxylate surfactants in normal-phase and reversed-phase systems using high-performance liquid chromatography-mass spectrometry. J. Chromatogr. A 813(2), 299-311. 

This expresses tR as a function of the fundamental column parameters t0 and k tR can vary between t0 (for k = 0) and any larger value (for k > 0). Since to varies inversely with solvent velocity u, so does tR. For a given column, mobile phase, temperature, and sample component X, k is normally constant for sufficiently small samples. Thus, tR is defined for a given compound X by the chromatographic system, and tR can be used to identify a compound tentatively by comparison with a tR value of a known compound. 

Aliphatic AEOs, considered as environmentally safe surfactants, are the most extensively used non-ionic surfactants. The commercial mixtures consist of homologues with an even number of carbon atoms ranging typically from 12 to 18 or of a mixture of even-odd linear and a-substituted alkyl chains with 11—15 carbons. Furthermore, each homologue shows an ethoxymer distribution accounting typically for 1—30 ethoxy units with an average ethoxylation number in the range 5—15. The separation of the AEO complex mixtures was achieved by reversed-phase and normal-phase chromatographic systems [74—76]. 

Over the past 10 years, liquid chromatography coupled with ultraviolet detection appears to have become the method of choice for the determination of corticosteroids, offering tlie analyst both satisfactory selectivity and sensitivity. Both reversed-phase (544-547) and normal-phase (548) chromatography have been applied to the determination of dexamethasone in plasma, coupled with ultraviolet (UV) detection generally at 254 nm, and in bovine tissues (528, 535). A series of both reversed- and normal phase LC systems have also been used for the simultaneous determination of dexamethasone and other steroids. Two different liquid chromatographic separations have been described for the isolation and simultaneous separation of steroids in serum (549). 

The ELS detector was previously also referred to as a mass detector, pointing to the fact that the response is (mainly) determined by the mass of the sample rather than by its chemical structure. Van der Meeren et al., though, demonstrated that the ELSD calibration curves of phospholipid classes were also dependent on the fatty acid composition (52). The dependence on the fatty acid composition is, however, completely different in nature and much less pronounced than for UV detection. The reason for this behavior is to be found in the partial resolution of molecular species, even during normal-phase chromatography. Thus, the peak shape depends not only on the chromatographic system but also on the fatty acid composition and molecular species distribution of the PL sample (47). Because it was shown before, based on both theoretical considerations and practical experiments, that the ELS detector response is generally inversely proportional to peak width (62,104), it follows that the molecular species distribution of the PL standards used should be similar to the sample components to be quantified. It was shown that up to 20% error may be induced if an inappropriate standard is used (52). 

Another property that is useful in selecting the proper chromatographic conditions (column, mobile phase, etc.) is water solubility or distribution coefficient between a polar (e.g., acetonitrile) and a nonpolar (e.g., -heptane) solvent. Data on water solubility and UV absorption maxima for a large number of NOC can be obtained from Druckrey et al. (25). Eisenbrand et al. (26) reported the distribution coefficients between acetonitrile and n-heptane for several NOC. Those for V-nitroso-dioctylamine, NDMA, and V-nitrosomethyl-2-hydroxyethylamine were reported to be 0.5,17.3, and 32.0, respectively, in this system. This would suggest that in a normal-phase system, using a silica column, these compounds would elute in the same order in which they are mentioned, but the elution order would be reversed in a reversed-phase (C18 column) system. 

In principle, h.p.Lc. arose from conventional liquid column chromatography, following the development of g.l.c. and realisation that it was a rapid and accurate analytical method. This led to a reappraisal of the liquid column chromatographic system, which in turn resulted in research developments in instrument design and in the manufacture of column-packing materials. These now have precise specifications to make them suitable for adsorption, normal and reversed phase partition, ion exchange, gel permeation, and more recently affinity chromatography. 

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