Amino phase

The reagent can be employed on silica gel, kieselguhr, polyamide and cellulose layers. Only dipping solution I can be employed on amino phases. 

Note The reagent can be employed for qualitative and quantitative analysis on silica gel and RP layers. Ammonia, amine and acid-containing mobile phases should be completely removed beforehand. Amino phases cannot be employed. The NBD-chloride reagent is not as sensitive as the DOOB reagent (qv.) on RP phases. 

Note The dipping solution may also be used as spray solution [1]. The reagent may be applied to RP layers it is not suitable for amino phases. 

The last example for thermal activation to be discussed involves amino phases. Table 2.3 lists the publications concerning the specific detection of sugars and creatine derivatives by means of the fluorescence obtained on heating mobile phase-free amino layer chromatograms . 

The reagent can be used on silica gel, kieselguhr. Si 50000, aluminium oxide and RP layers amino phases are unsuitable. 

Amino-3-nitrotoluene 1 b 415,419 Amino phases la 3 Aminophenazone lb312,314,354 2-Aminophenol lb 309, 310, 381 4-Aminophenol lb 309, 310, 381 Aminophenols lb 309,381,383,401 9-(p-Aminophenoxy)acridine lb 145 1-Aminopyrene la 61 4-Amino salicylic acid lb 309,310 Amino sugars lb 47,232-235,354 Aminotrimethylenephosphonic acid la 172... 

Additional modes of HPTC include normal phase, where the stationary phase is relatively polar and the mobile phase is relatively nonpolar. Silica, diol, cyano, or amino bonded phases are typically used as the stationary phase and hexane (weak solvent) in combination with ethyl acetate, propanol, or butanol (strong solvent) as the mobile phase. The retention and separation of solutes are achieved through adsorp-tion/desorption. Normal phase systems usually show better selectivity for positional isomers and can provide orthogonal selectivity compared with classical RPLC. Hydrophilic interaction chromatography (HILIC), first reported by Alpert in 1990, is potentially another viable approach for developing separations that are orthogonal to RPLC. In the HILIC mode, an aqueous-organic mobile phase is used with a polar stationary phase to provide normal phase retention behavior. Typical stationary phases include silica, diol, or amino phases. Diluted acid or a buffer usually is needed in the mobile phase to control the pH and ensure the reproducibility of retention times. The use of HILIC is currently limited to the separation of very polar small molecules. Examples of applications... 

A mixture of acetonitrile and water is capable of separating sugars on amino phase. Problems with the determination of reducing sugars can occur, since the reaction of the keto group with the amino moiety is well known. In a similar way, an amino phase in normal-phase mode should not be used with acetone. NH2 packing can be used as a weak anion exchanger. The diol... 

Finally, other proposals include the introduction of an amine into the eluent so that, unlike in bonded phases, preparation of the amino phase takes place in situ. This competes advantageously in terms of degradation of the column, since with this procedure columns may be immediately repaired by recoating and therefore losses in performance are minimized and even avoided. 

Equation (8) is a fundamental relationship for retention in LSC as a function of the solvent strength of the mobile phase. It states that log A values for different solutes will yield linear plots against values of for different mobile phases, and the slopes of these plots will be proportional to the molecular size A, of the solute. Numerous data are summarized or referenced in Ref. /. showing the validity of Eq. (8) when applied to LSC systems where the solute and solvent molecules are nonlocalizing (nonpolar or moderately polar compounds—see Section II,B below). Similar data showing the applicability of Eq. (8) to amino-phase polar-bonded-phase columns are given in Ref. 17. 

The linearity of plots of log k versus e° has been demonstrated in many studies referenced in Ref. /. For example, Fig. 4 (from Ref. 1) shows plots for 9 different solutes, using 7 different solvents (pentane, CCI4, benzene, CH2CI2) and/or mixtures thereof. Similar plots for an amino-phase column (/7) are shown in Fig. 5 for 17 different solutes and hexane/tetra-hydrofuran binary mobile phases. 

There are two reasons to suspect that restricted-access delocalization is not responsible for the rise in cb at small 0b in Fig. 10. First, an apparent function can be derived from these latter data and is plotted in Fig. 11. A single curve (solid line) fits the data points for different B-solvents, as expected for restricted-access delocalization. However, this solid curve in Fig. 11 differs dramatically from the dashed curve of Fig. 11, which is the plot of 7cversus 0b for localizing solvents (C) on alumina and silica [Eq. (40)]. Whereas the latter curve suggests localized adsorption of the B-solvent for 0b as large as 0.75 (% p = 0.5), the corresponding value of 0b for the localized monolayer on an amino-phase surface is only 0.18. It is reasonable to assume that localized solvent molecules might occupy as much as 75% of the adsorbent surface, but unreasonable to conclude that a localized monolayer exceed 18% of a monolayer. Furthermore,... 

A second reason to doubt the occurrence of restricted-access delocalization on amino-phase columns is that this increase in eu (Fig. 11) is observed for both localizing and nonlocalizing solvents ethyl acetate and tetrahydrofuran (localizing) and CH2CI2 and CHCHs (nonlocalizing). This contrasts with theory, which predicts that delocalization effects are only associated with localizing compounds. 

The data of Fig. 11 thus indicate that restricted-access delocalization does not exist for amino-phase packings. Furthermore, the importance of the variation of eo with 6n, shown in Fig. 11, is insignificant, so far as calculations of e" are concerned. We will therefore assume that cb is constant for a given solvent B and amino-phase packings, in the calculation of ° for multicomponent mobile phases (Appendix). 

Fig. 14. Representation of the surface structure of four LSC adsorbents the active site is circled in each case (a) alumina refers to vacancy-site, (b) Cie-silica, (c) silica, (d) amino phase.

Terminal amino groups are believed to comprise adsorption sites on the surface of amino-phase packings (77). These (1) Eire exposed for direct interaction with adsorbate molecules, (2) are not rigidly positioned on the surface, and (3) have a much lower surface concentration (2-3 /xmol/m ). These characteristics of the various LSC packings so far discussed are summarized in the surface representations shown in Fig. 14 for each adsorbent. The m or adsorption sites in each case are indicated by enclosure within a circle (except the vacancy site for alumina, shown as an asterisk in Fig. 14). 

Site-competition delocalization will be favored by surface sites that are accessible to lateral interaction by solvent molecules that are adjacent to a localized solute or solvent molecule. The relative accessibility of sites for this type of interaction is as follows alumina (least), Qg-silica, and silica or amino phase (most). This is in fact the order of increasing site-competition delocalization noted in Table III for delocalization of either solvent or solute molecules. 

Restricted-access delocalization is favored by a high concentration of surface sites and by the rigid positioning of these sites on the surface. Alumina and silica meet these requirements and exhibit restricted-access delocalization. Amino-phase columns do not meet these requirements and do not exhibit this phenomenon. Qg-silica would be expected to exhibit the effect in lesser degree, due to the lower concentration of surface sites. 

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