Migration order

The use of t.l.c. for separating organophosphorus compounds has been reviewed. In a study of phosphoryl compounds it was shown that / m values i.e. log [(1 - Rv)lRy on silica gel using hexane-acetone as eluant, depend upon the polarity of the P=0 group.The migration order found was... 

Calvo, D. et al., Migration order of wine anthocyanins in capillary zone electrophoresis, Anal. Chint. Chintica Acta, 524, 207, 2004. 

Anions and uncharged analytes tend to spend more time in the buffered solution and as a result their movement relates to this. While these are useful generalizations, various factors contribute to the migration order of the analytes. These include the anionic or cationic nature of the surfactant, the influence of electroendosmosis, the properties of the buffer, the contributions of electrostatic versus hydrophobic interactions and the electrophoretic mobility of the native analyte. In addition, organic modifiers, e.g. methanol, acetonitrile and tetrahydrofuran are used to enhance separations and these increase the affinity of the more hydrophobic analytes for the liquid rather than the micellar phase. The effect of chirality of the analyte on its interaction with the micelles is utilized to separate enantiomers that either are already present in a sample or have been chemically produced. Such pre-capillary derivatization has been used to produce chiral amino acids for capillary electrophoresis. An alternative approach to chiral separations is the incorporation of additives such as cyclodextrins in the buffer solution. 

The migration order of wine anthocyanins in CE has been studied in detail and the results have been compared with those obtained by RP-HPLC-MS. Wines were filtered and used for the analyses without any other pretreatment. Wine samples of 10 ml were freeze-dried, redissolved in methanol and applied for semi-preparative fractionation. CZE measurements were carried out in a fused-silica capillary (46 cm effective length, 75 /an i.d.). The capillary was conditioned with 0.1 M NaOH (2 min), water (2 min) and running buffer (5 min). The buffer consisted of 50 mM sodium teraborate (pH = 8.4) containing 15 per cent (v/v)... 

Newly used chiral surfactants often have a low critical micellar concentration, are highly soluble and can be synthesized both in L- and D-forms. This last feature makes it possible to easily change the migration order of the optical isomers, which is very interesting for the determination of the optical pnrity of drugs, where for quantification purposes it is favorable that the chiral impurity migrates before the main component. 

The chiral recognition of enantiomers can be of three types (i) desionoselective, (ii) ionoselective, or (iii) duoselective, in which only the non-dissociated, the dissociated or both forms (charged and uncharged), respectively, of the enantiomers selectively interact with the chiral selector. In the case of ionoselective and duoselective interactions, a reversal of the migration order of the enantiomers is theoretically possible by the appropriate selection of CD concentration and the pH of the BGE. The addition of organic modifier to the BGE can also change selectivity by modifying the solubility of the chiral selector and/or of the solute, the complex equilibrium, the conductivity of the BGE and the electroendos-motic flow (EOE) level. Several other factors, such as the temperature, the type and the concentration of the BGE, and the level of the EOE can influence the separation. 

The most electron-rich alkyl group (more substituted carbon) migrates first. The general migration order ... 

The existence of the aforementioned difference between the mobilities of transient diastereomeric complexes of the enantiomers with the chiral selector may have some important consequences in chiral CE. For instance, the enantioseparation can, in principle, be possible even in those cases when the binding constants of both enantiomers to a given chiral selector are the same. On the other hand, this may allow, in certain cases, observation of the reversal of the enantiomer migration order, depending on the concentration of the chiral selector (17). 

The possible reversal of the migration order of the 1,1 -binaphthalene-2,2 -diyl hydrogen phosphate enantiomers could also be predicted based on these studies, but it has been not verified experimentally. 

S Sabah, GKE Scriba. Influence of the structure of cyclodextrins and amino acid sequence of dipeptides and tripeptides on the pH-dependent reversal of the migration order in capillary electrophoresis. J Chromatogr A 894 267-272, 2000. 

B Chankvetadze, K Lomsadze, D Bergenthal, J Breitkreutz, K Bergander, G Blaschke. Mechanistic study on the opposite migration order of clenbuterol enantiomers in capillary electrophoresis with /3-cyclodextrin and single-isomer heptakis(2,3-diacetyl-6-sulfo-)-/3-cyclodextrin. Electrophoresis 22 3178-3184. 

B Chankvetadze, G Pintore, N Burjanadze, D Bergenthal, K Bergander, J Breitkreutz, C Muhlenbrock, G Blaschke. Mechanistic study of opposite migration order of dimethindene enantiomers in capillary electrophoresis in the presence of native /3-CD and heptakis-(2,3,6-tri-0-methyl)-/3-CD. J Chromatogr A 875 455-469, 2000. 

Another problem, the potential increase in background noise due to the addition of solvents and modifiers from the sheath liquid (e.g., volatile salts, acids, and bases), has been studied (19). Moreover, because of the different composition of the initial CE buffer reservoir and the sheath liquid, discontinuous and irreproducible conditions may result. These effects can potentially change migration times or even the migration order of the analytes (20). 

Peptide pf-value Molecular weight No. of amino acids Calculated mobility Migration order... 

Figure 24 Schematic Illustration of the Migration Order of Positively Charged, Negatively Charged, and Uncharged (Neutral) Peptides Separated by Capillary Electrochromatography (CEC) with H,0/0rganic Mobile Phases and Sorbents of Appropriate EOF Properties...

Figure 5.4-—Separation of three steroids on six stationary phases using two sets of solvents. The evolution of the response factor R( and reversal of migration order with changes in mobile phase should be noted (reproduced by permission of Merck).

The migration order of groups in the Baeyer-Villiger oxidation is valid also for chlorofluoro-alkyl ketones, in which the isopropyl group specifically migrates to give the isopropyl ester of chlorofluoroacetic acid (Tabic 11).195... 

The capillary gel electrophoresis separation of a three-component, 3 P-labeled, nucleotide mixture is illustrated in Figure 10. It is interesting to note that the migration order of ATP and CTP is reversed with respect to the free solution separations presented in Figures 8 and 9. This is caused by the absence of electroosmotic flow in the gel-filled capillary. 

In a study presented by Jinno et al. [124], packed column capillary electrochromatography, open-tubular CEC, and microcolumn liquid chromatography using a cholesteryl silica bonded phase have been studied to compare the retention behavior for benzodiazepines. The results indicated that CEC was a promising method, as it yielded better resolution and faster analysis than microcolumn LC for benzodiazepines. Similar selectivity to HPLC was noted, except for a few solutes that were charged under the separation conditions. Columns packed with the ODS and cholesteryl phases were compared and showed totally different migration orders of the analytes. The retention on the cholesteryl silica sta-... 

In addition to CD-modified MEKC, the use of mixed micelles incorporating CDs, CDs together with other chiral selectors (bile salts, ligand exchangers, crown ethers, etc. [5]), or mixed CD systems, neutral with neutral or neutral with charged CDs can also be extremely useful. A very attractive feature of the latter approach is the ability to control the enantiomeric migration order, an aspect that may be extremely useful if quantitation at low levels is required. Details of this technique are available in the review of Chankvetadze et al. [12]. 

Chiral surfactants of amino acid derivatives, such as (5)- and (R)-V-dodecoxycarbonylvaline (DDCV) and V-dodecoxycarbonylproline (DDCP) are available for enantiomer separation by MEKC Several pharmaceutical amines, benzoylated amino acid methyl ester derivatives, piperidine-2,6-dione enantiomers, and aldose enantiomers were successfully resolved. Because both enantiomeric forms of DDCV or (5)- and R) forms are available, we can expect that the migration order of an enantiomeric pair would be reversed. 

In addition to reducing the adsorption, the dynamic additives also play important roles in selectivity enhancement. pH is an important option for protein and peptide separation. An increase in the buffer ionic strength can increase the resolution. Any ion that displays a preferential affinity with the peptides has a potential modifying effect on the selectivity. The addition of organic modifiers, e.g., methanol, ethanol, and acetonitrile, in the BGE, can induce different solvation of the peptide chains and modify the migration order and selectivity of peptides. For separation of very hydrophobic proteins, e.g., lipoproteins, surfactants can act as buffer additives to improve solubilization. 

In general, the migration order of any lipid class is determined by the overall number of double bonds in the molecule. Thus the retention of common fatty acids (chain lengths of 16-22 carbon atoms, methylene interrupted double bonds) increases with increasing number of double... 

This simplified equation allows one to calculate the optimal concentration of a chiral selector [7], a better design of chiral separation systems containing multiple chiral selectors [6], and the design of the enantiomer migration order [6,8] and indicates why micellar additives are sometimes required in chiral CE with CDs. 

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