Stationary phase polarity

Trying to determine which column is ideal for a specific analysis can be difficult with over 1000 different columns on the market [74]. A proper choice implies a definition of parameters such as column material, stationary phase (polarity), i.d., film thickness and column length. Guides to column selection are available [74,75]. The most important consideration is the stationary phase. When selecting an i.d., sample concentration and instrumentation must be considered. If the concentration of the sample exceeds the column s capacity, then loss of resolution, poor reproducibility and peak distortion will result. Film thickness has a direct effect on retention and the elution temperature for each sample compound. Longer columns provide more resolving probe, increase analysis times and cost. 

Because of the similarities in the theory and practice of these two procedures, they will be considered together. Both are examples of partition chromatography. In paper chromatography, the cellulose support is extensively hydrated, so distribution of the solutes occurs between the immobilized water (stationary phase) and the mobile developing solvent. The initial stationary liquid phase in thin-layer chromatography (TLC) is the solvent used to prepare the thin layer of adsorbent. However, as developing solvent molecules move through the stationary phase, polar solvent molecules may bind to the immobilized support and become the stationary phase. 

Finally, the sum of the first five McReynolds values has been used to compare the polarities of silicone polymers on OT columns. It has already been stated that the polarity of the silicone polymers can be increased by increasing the percentage of phenyl groups in the polymer. Figure 8.17 shows a plot for five polymers on bonded fused silica WCOT columns (except for SP-2250, which is from packed column data). The increasing McReynolds values clearly show the validity of this method of specifying stationary phase polarity. 

Figure 8.17. Effect of number of phenyl groups on stationary phase polarity as measured by McReynolds values. Reprinted from the Supelco Reporter, Vol. IV, No. 3, May 1985 with the permission of Supelco Inc., Bellefonte, PA.

The stationary phases play an important part in Liquid Chromatography using micellar mobile phases. They interact with both the surfactant and with solutes. To study the interactions with surfactants, adsorption isotherms were determined with two ionic surfactants on five stationary phases an unbonded silica and four monomeric bonded ones. It seems that the surfactant adsorption closely approaches the bonded monolayer (4.5 pmol/m2) whatever the bonded stationary phase-polarity or that of the surfactant. The interaction of the stationary phase and solutes of various polarity has been studied by using the K values of the Armstrong model. The KgW value is the partition coefficient of a solute between the... 

Kg values. Kg values give information about the affinity of the solute for the surfactant covered stationary phases. The first observation indicates that, in spite of this surfactant-coverage, the polar nature of the bonded stationary phases is preserved the order of increasing KgW values of toluene is the same order as the decreasing stationary phase-polarity i.e. silica CN Cl C8 C18. The K values of toluene, with SDS mobile phases, were slighty lower than those with CTAB mobile phases (Table VI). 

The K values of BTAB in SDS mobile phases were similar to the one of toluene. It seems that BTAB was retained as an apolar ion-pair. The order of increasing Kgw values of BTAB corresponds to the decreasing stationary phase-polarity. Benzoic acid in CTAB mobile phases seems to be also retained as an ion-pair. 5% methanol slightly decreased the Kg values. The effect of NaCl was much more significant the decrease was 50% (BTAB with SDS mobile phases) up to 80% (benzoic acid with CTAB mobile phases). Sodium and/or chloride ions decrease the ionic interactions and the ion-exchange capacity. 

Group Stationary phases Polarity 1 Classes of compounds effectively separated by columns with stationary phases of the same group... 

Adsorption TLC selection of the mobile phase is conditioned by sample and stationary-phase polarities. The following polarity scale is valid for various compound classes in NPTLC in decreasing order of K values carboxylic acids>amides>amines>alcohols>aldehydes > ketones > esthers > nitro compounds > ethers > hal-ogenated compounds > aromatics >olefins > saturated hydrocarbons > fluorocarbons. For example, retention on silica gel is controlled by the number and functional groups present in the sample and their spatial locations. Proton donor/acceptor functional groups show the greatest retention, followed by dipolar molecules, and, finally, nonpolar groups. 

Purnell and co-workers [1] pioneered an alternative approach to stationary-phase polarity control for GC. These workers demonstrated that for a wide range of stationary phases made up of binary mixtures, the corrected retention volume of a solute was linearly related to the volume fraction of either one of the two phases. At the time of discovery, this was quite an unexpected relationship, as it was generally accepted that the expression for the retention volume would take the form of the exponent of the stationary-phase composition. The results of Purnell and his co-workers can be summarized by... 

In general, the nature of the analyte determines the choice of stationary phase. For example, for the separation of organochlorine and pyrethroid pesticides, a nonpolar stationary phase such as DB-1 (or OV-1) is recommended. For the separation of somewhat more polar compounds, such as organophosphorus compounds, OV-17 (or DB-1701) can be applied. In addition, for confirmation purposes, the use of two columns with distinct stationary-phase polarities (e.g., DB-1 and DB-1701) is certainly required. A polar stationary phase (e.g., DB-wax) is suitable for the more polar compounds such as methamidofos, but its application to some detection modes is limited due to stationary-phase bleeding. 

Systematic method development guidelines akin to those available for HPLC have not been developed. However, details beyond the scope of this chapter are available.Separation conditions should be evaluated based on the polarity of the solute and the polarity of the stationary phase. Stationary-phase polarity increases in the order C18phenylnonpolar solutes on nonpolar stationary phases, separation may be achieved using pure carbon dioxide. As solute or stationary-phase polarity increases, carbon dioxide modified with methanol (or isopropanol, ethanol, or acetonitrile) or carbon dioxide modified with solvent and an additive such as TFA, acetic acid, triethylamine, or isopropylamine (0.5% or less) is required. 

Gas chromatograph, p. 575 GC columns— packed, capillary, p. 578 Stationary phases—polar to nonpolar, p. 581... 

Partition retention forces intermolecular forces that result in attraction of solute molecules to the stationary phase. Polar retention forces include dipole-dipole attractions, van der Waal s forces and hydrogen bonding. Non-polar retention forces consist of London s dispersion forces arising from induced polarity in non-polar molecules, remember that like attracts like . 

Fig. 6.4. Diagramatic illustration of the effect of temperature in NPC. Chromatographic conditions stationary phases, polar (silica), less polar (cyanopropyl), least polar (n-butyl) (250x2.4 mm I.D.) mobile phase, n-hexane with the addition of water flow rate, 1.6 ml/min solutes, anihne, p-chloroaniline, 2,6-dimethylaniline, m-chloroaniline, n-methylaniline, 2,6-dimethylphenol, N, N-dimethylanihne, o-chloroaniline and benzene. The figure is divided into three different temperature areas (a,b,c). Reproduced from Sisco and Gilpin (1980).

A nonpolar mobile phase passing through a packed column that contains a polar stationary phase defines normal-phase HPLC (NP-HPLC). For example, if -hexane comprises the mobile-phase and silica gel is used for the stationary phase, separations of nonpolar organic analytes as shown in Fig. 4.1 is accomplished. With respect to neutral organic compounds, the polar and ionic domains cannot be reached by NP- HPLC. NP-HPLC was the first high-pressure form of liquid chromatography to be developed. If the stationary phase could be made hydrophobic by chemical treatment and the mobile phase made more polar, a reversal of mobile/stationary-phase polarities could be achieved. Like it or not, we are stuck with this nomenclature RP-HPLC has certainly extended the range of analyte polarity that... 

Rp-HPLC Nonpolar stationary phase/polar mobile phase... 

The cmc is indicated by an arrow. The three nonpolar phases, SAS, MOS and ODS Hypersil , show a Langmuir-like adsorption isotherm with a concave shape or L type in the Giles classification [12]. The polar CPS phase shows a convex shape or S type cooperative adsorption. The amount of adsorbed surfactant increases in the order bare silica < CPS < SAS < MOS < ODS. This is exactly the decreasing order of the stationary phase polarity. No adsorption of SDS on the bare silica was observed in this low concentration range [11]. Above 0.004 M, the SDS adsorption on SAS Hypersil passes the one on MOS Hypersil . Hydrophobic interactions are mainly responsible for SDS adsorption in sub micellar solutions. 

The curvature in the log k vs. n plots, observal with micellar and hybrid mobile phases, was first attributed to the different locations (with different microenvironment polarities) in the micelle, for different members of a homologous series [6, 7], Methylene selectivity decreases as the difference between mobile and stationary phase polarities is reduced. For a given mobile phase composition, the larger and more hydrophobic homologous compounds are located in a less polar environment of micelles, it is then conceivable to assume that these compounds experience a smaller change in their microenvironment polarity upon being transferred from the micellar pseudo-phase to the bonded alkyl stationary phase. The a(CH2) value between n-pentylbenzene and n-butylbenzene is smaller than it is between ethylbenzene and toluene (Table 9.1), because the former pair is located in a more nonpolar environment than the latter. 

The adsorption of surfactant monomers changes the stationary phase polarity, structure, surface, and pore volume. Surfactant molecules coat the interior walls of the pores without completely filling them. 

F1g.4A Effect of stationary phase polarity on a 4-component separation a) Carbowax-20M (polar) (b) DC-200 (nonpolar). Samples and their boiling points (1) /i-heptane (98) (2) tetrahydrofuran (64) (3) 2-butanone (80) (4) /t-propanol (97). Reprinted with permission of the GOW-MAC Instrument Co., Bethlehem, PA, U.S.A. 

Table 2 McReynolds constants and their sum, —y + / + + m + s, used for characterization of liquid stationary phase polarity.

The system of McReynolds constants is a usefiil tool for characterization of the selectivity of stationary phases in GC. The founding principle of this approach is that inter-molecular forces are additive and their individual contributions to retention can be evaluated from the difference in retention index values of selected test probes measured on a liquid phase to be characterized, and on the non-polar reference phase, i.e., squalane. To characterize the stationary phase polarity, the concept uses five special solutes (benzene, n-butanol, 2-pentanone, 1-nitropropane, and pyridine) that are considered to represent typical chemical interactions. 

RP-HPLC (i.e., hydrophobic stationary phase/polar aqueous mobile phase) is the more usual mode to separate and analyze opium alkaloids. The most attractive advantage of RP HPLC is the ability to determine a wide variety of compounds, which have different molecular structures, polarity, and acidity/basicity. Analysis of opium alkaloids by NP-HPLC (i.e., polar stationary phase) is not common and no references have been found in the scientific literature. As it is reflected in Table 33.2, octadecyl colunms (C18) are the most commonly employed. When monolithic columns are used, high flow rates are allowed, which gives rise to very... 

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