Bonded phase Alkyl-type phases

Alkyl-Type Phases (C1-C18, C30). Probably 90% of all reversed-phase columns are alkyl-type bonded phases. An enormous amount of publications are devoted to the classiflcation, standardization, and comparison of these phases. In their book Practical HPLC Method Development, Snyder and Kirkland [57] indicate that reversed-phase retention for nonpolar and nonionic compounds generally follows the retention pattern Cl < C4 < C8 = Cl 8. At the same time, they refer to the comparison of Cl 8-type columns from different manufacturers and find dramatic variation in the retention of both polar and nonpolar compounds at the same conditions on different columns. 

Besides the above differentiation, restricted-access media can be further subdivided on the basis of the topochemistry of the bonded phase. Packings with a uniform surface topochemistry show a homogenous ligand coverage, whereas packings with a dual topochemistry show a different chemical modification of the pore internal surface and the particle external surface (114). Restricted-access media of the former type are divided into mixed-mode and mixed-function phases, bonded-micellar phases, biomatrix, binary-layered phases, shielded hydrophobic phases, and polymer-coated mixed-function phases. Restricted-access media of the latter type include the Pinkerton s internal surface reversed-phase, Haginaka s internal surface reversed-phase diol, alkyl-diol silica, Kimata s restricted-access media, dual-zone phase, tris-modified Styrosorb, Svec s restricted-access media, diphil sorbents, Ultrabiosep phases. Bio Trap phases, and semipermeable surface phases. 

C-Alkylations have been performed with both support-bound carbon nucleophiles and support-bound carbon electrophiles. Benzyl, allyl, and aryl halides or triflates have generally been used as the carbon electrophiles. Suitable carbon nucleophiles are boranes, organozinc and organomagnesium compounds. C-Alkylations have also been accomplished by the addition of radicals to alkenes. Polystyrene can also be alkylated under harsh conditions, e.g. by Friedel-Crafts alkylation [11-16] in the presence of strong acids. This type of reaction is incompatible with most linkers and is generally only suitable for the preparation of functionalized supports. Few examples have been reported of the preparation of alkanes by C-C bond formation on solid phase, and general methodologies for such preparations are still scarce. 

Typical heterogeneous Ziegler catalysts operate at temperatures of 70 100°C and pressures of 0.1 2 MPa (15 300 psi). The polymerization reactions are carried out in an inert liquid medium (e.g, hexane, isobutane) or in the gas phase. Molecular weights of LLDPE resins are controlled by using hydrogen as a chain-transfer agent. Reactivities of a-olefins in copolymerization with ethylene depend on two factors the size of the alkyl groups attached to their double bonds and the type of catalyst,... 

The use of reversed-phase liquid chromatography is growing in applications for separating mixtures of peptides. In this type of chromatography, the stationary phase is nonpolar, whereas the mobile phase is polar. The stationary phase is normally porous silica with bonded n-alkyl chains, mainly octadecyl but also octyl, hexyl, butyl, and propyl chains. 

More recently, pyrolysis GC, ESCA, and FTIR have been used to characterize the surface of bonded layers. The reason for the intense interest is the fact that there are significant differences between bonded phases manufactured by different companies, and these investigators hope to find out why. One recent study15 analyzed the decomposition products produced by the reaction of bonded reverse phases with HF, and it was able to determine the type of reaction (monofunctional or polyfunctional), the extent of end capping, and the distribution of lengths of alkyl groups. Some of the results of the study are summarized in Table 7. Such results help explain the differences between bonded phases manufactured by different companies. 

Nonpolar, porous polystyrene-divinylbenzene (PS-DVB) copolymers have also been used (4 /, 45a) as re versed-phase HPLC packings for peptide separations. Compared to the chemically bonded n-alkyl silicas, these supports show generally much lower efficiencies. As they can, however, be used over a wide pH range, e.g., Amberlite XAD-4 can be used over ca. pH 1.5-12.5, these types of supports could find limited use in situations where pH control of the selectivity is mandatory. 

Determinations of the adsorption isotherms for a number of organic solvent-water systems in contact with hydrocarbonaceous stationary phases have shown that a layer of solvent molecules forms on the bonded-phase surface and that the extent of the layer increases with the concentration of the solvent in the mobile phase. For example, methanol shows a Langmuir-type isotherm when distributed between water and Partisil ODS (56). This effect can be exploited to enhance the resolution and the recoveries of hydrophobic peptides by the use of low concentrations, i.e., <5% v/v, of medium-chain alkyl alcohols such as tm-butanol or tert-pentanol or other polar, but nonionic solvents added to aquo-methanol or acetonitrile eluents. It also highlights the cautionary requirement that adequate equilibration of a reversed-phase system is mandatory if reproducible chromatography is to be obtained with surface-active components in the mobile phase. 

Phenyl-Type Phases. Phenyl-type phases have been studied for a long time [58,59]. The presence of a phenyl ring on the surface of a bonded phase introduces so-called n-n interactions with some analytes that are capable of these types of interactions. This introduces an additional specificity for HPLC separations on these stationary phases. Compared to common alkyl-type phases, phenyl columns show lower methylene selectivity in other words, the separation of members of homologous series will be less selective on phenyl columns than on alkyl-modified phases. 

In the contrast to the irreversible adsorption of amphiphilic ions on the reversed-phase surface, the liophiUc ions shows relatively weak interactions with the alkyl chains of the bonded phase. Liophilic means oil-loving. These liophilic ions are usually small inorganic ions and they possess an important ability for dispersive type interactions. They are (a) characterized by significant delocalization of the charge, (b) primarily symmetrical, (c) usually spherical in shape, and (d) absence in surfactant properties. 

Traditional reversed-phase columns appear to offer the most inert surface since the alkyl-type bonded ligands at high bonding density can only participate in weak dispersive interactions, which could be suppressed by practically any organic solvent. An example of the calibration curve made on a commercial reversed-phase column is shown in Figure 6-7. 

The separation of chiral compounds will be discussed in Chapter 22. However, the separation of diastereomers can be accomplished using achiral stationary phases. Another alternative is the use of chiral columns for the separation of diastereomers in either the reversed-phase or normal-phase mode. The use of achiral bonded phases without chiral additives, such as phenyl and alkyl bonded phases for the separation of diastereomeric pharmaceutical compounds, is acceptable. Different selectivities can be obtained by employing stationary phases containing varying functionalities (phenyl, polar embedded moieties). The effect of aqueous mobile-phase pH, temperature, and type of organic eluent (acetonitrile versus methanol) can also play a dramatic role on the separation selectivity of diastereomeric compounds. 

The phase that is most commercially available contains a weak TT-electron acceptor mononitrophenyl bonded to the sUica and usually contains a propyl spacer arm. This packing interacts with analytes through rr-donor interactions. Other nitrophases exist and, as the number of nitro groups increases on these acceptor-type phases, an increased retention of aromatic donor compounds occurs. The phases of 3-(2,4-dinitroanilino)-propyl and (2,4,6-trinitronanilino)-propyl have shown preferential adsorption of PAHs, alkyl aromatic hydrocarbons, and polyarylalkanes, in comparison to aminopropyl silica [3]. 

It is assumed that this is also a rate determining step for the overall reaction. The activation energy of reaction (4) and the site density of oxygen active centers were the only adjustable parameters of the model. In general, a C-H bond scission for reactants and products of the methane dimerization process occurs by an Eley-Rideal (E-R) type mechanism to form a gas-phase alkyl radical and a hydroxyl surface site (HO ) ... 

---