E Chromatographic Separations

Hansson, C., Agrup G., Rorsman, H., Rosengren, A.M., and Rosengren, E., Chromatographic separation of catecholic amino acids and catecholamines on immobilized phenylboronic acid, J. Chromatogr., 161, 352, 1978 Chem. Abs., 90, 50771d, 1979. 

While being a most powerful and widely used preparative separation technique, analytical lEC has the disadvantage of being difficult to couple to MS due to the need for a high salt elution buffer. For the same reason, miniaturization is difficult since high salt buffers are intrinsically unsuited for electrochro-matographic applications, i.e., chromatographic separation, where the electro-osmotic flow is used to drive the mobile phase. Pressure-driven p-LC is possible in the lEC mode but is more difficult to perform. 

Fig. 3 (I) Chromatogram of tenfold-diluted sheep urine. (E) Chromatographic separation of standard solutions. Peaks A, allantoin B, uric acid C, hypoxanthine D, allopurinol (IS) E,...

More sensitive detection methods and more objective recording methods (e g the employment of scanners) are constantly been striven for m order to overcome this illusion It IS for this reason too that fluorescent methods have been introduced to an increasing extent on account of their higher detection sensitivity This allows an appreciable reduction in the amount of sample applied, so that possible interfering substances are also present m smaller quantibes This increases the quality of the chromatographic separation and the subsequent m situ analysis... 

The optimization of chromatographic separations can generally be seen as a compromise between speed, i.e., to produce the largest possible amount of data or substance per unit time, and resolution, i.e., to produce the highest possible quality of data or purity of substance. Obviously the goal for optimization differs according to the purpose of the separation and also between scale of operation. Therefore, different parameters are critical for different situations. Still, some basic rules for optimization may be applied. 

The first (inconclusive) work bearing on the synthesis of element 104 was published by the Dubna group in 1964. However, the crucial Dubna evidence (1969-70) for the production of element 104 by bombardment of 94PU with loNe came after the development of a sophisticated method for rapid in situ chlorination of the product atoms followed by their gas-chromatographic separation on an atom-by-atom basis. This was a heroic enterprise which combined cyclotron nuclear physics and chemical separations. As we have seen, the actinide series of elements ends with 103 Lr. The next element should be in Group 4 of the transition elements, i.e. a heavier congenor of Ti, Zr and Hf. As such it would be expected to have a chloride... 

Polysaccharide derivatives for chromatographic separation of heterocyclic enantiomers 98AG(E)1021. 

P. Harmala, E. Botz, O. Sticher and R. Hiltunen, Two-dimensional planar chromatographic separation of a complex mixture of closely related coumarins from the genus Angelica , J. Planar Chromatogr. 3 515-520 (1990). 

Although many attempts have been made to separate or exclusively synthesize one isomer of an unsymmetrically substituted phthalocyanine,72-89-296,297 the product mixture has been separated in only two cases.96 103,104 Besides the chromatographic separation of the statistical product mixture it is also possible to prepare exclusively the D4h isomer by use of steric hindrance of bulky substituents, e.g. 7-ferr-butylnaphthalene-l,2-dicarbonitrile only forms the respective An isomer of the tetra(to -butyl)-substituted 1,2-NcFe by heating in hexan-l-ol.73 Recently, some 1,8,1 5,22-substituted pure isomers have also been synthesized by the use of bulky substituents in 3-substituted phthalonitriles298,299 at low temperature (see Section 2.1.4.).94... 

Factors may be classified as quantitative when they take particular values, e.g. concentration or temperature, or qualitative when their presence or absence is of interest. As mentioned previously, for an LC-MS experiment the factors could include the composition of the mobile phase employed, its pH and flow rate [3], the nature and concentration of any mobile-phase additive, e.g. buffer or ion-pair reagent, the make-up of the solution in which the sample is injected [4], the ionization technique, spray voltage for electrospray, nebulizer temperature for APCI, nebulizing gas pressure, mass spectrometer source temperature, cone voltage in the mass spectrometer source, and the nature and pressure of gas in the collision cell if MS-MS is employed. For quantification, the assessment of results is likely to be on the basis of the selectivity and sensitivity of the analysis, i.e. the chromatographic separation and the maximum production of molecular species or product ions if MS-MS is employed. 

The application of high tension (e.g. 20 kV, 0.5 MHz) in an evacuated system (0.2. .. 8 torr) causes the residual gas to form a highly ionized mixture of positive and negative ions, electrons, photons and neutral gas molecules. In the presence of active sorbents this plasma reacts with the chromatographically separated substances to eld reactive ions and radicals. 

Alkaline hydrolysis (saponification) has been used to remove contaminating lipids from fat-rich samples (e.g., pahn oil) and hydrolyze chlorophyll (e.g., green vegetables) and carotenoid esters (e.g., fruits). Xanthophylls, both free and with different degrees of esterification with a mixture of different fatty acids, are typically found in fruits, and saponification allows easier chromatographic separation, identification, and quantification. For this reason, most methods for quantitative carotenoid analysis include a saponification step. 

The calibration solutions, which typically contain a number of analytes at known concentrations, are useful for validating the chromatographic separation step (e.g. retention times and analyte detector response). 

The most significant differences (i.e. independence) in the analytical methods are provided in the final chromatographic separation and detection step using GC/ MS and LC-FL. GC and reversed-phase LG provide significantly different separation mechanisms for PAHs and thus provide the independence required in the separation. The use of mass spectrometry (MS) for the GC detection and fluorescence spectroscopy for the LG detection provide further independence in the methods, e.g. MS can not differentiate among PAH isomers whereas fluorescence spectroscopy often can. For the GC/MS analyses the 5% phenyl methylpolysiloxane phase has been a commonly used phase for the separation of PAHs however, several important PAH isomers are not completely resolved on this phase, i.e. chrysene and triphenylene, benzo[b]fluoranthene and benzofjjfluoranthene, and diben-z[o,h]anthracene and dibenz[a,c]anthracene. To achieve separation of these isomers, GC/MS analyses were also performed using two other phases with different selectivity, a 50% phenyl methylpolysiloxane phase and a smectic liquid crystalline phase. 

Francotte, E. and Junker-Buchheit, A., Preparative chromatographic separation of enantiomers, /. Chromatogr., 576, 1, 1992. 

E. Forgdcs and T. Cserhdti, Molecular Basis of Chromatographic Separation, CRC Press, Boca Raton, FL (1997). 

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