Methacrylic acid stationary phases

The mixture of deprotected amino acid derivatives in solution was then immobilized onto a polymeric solid support, typically activated 5-)xm macroporous poly(hydroxyethyl methacrylate-co-ethylene dimethacrylate) beads, to afford the chiral stationary phases with a multiplicity of selectors. Although the use of columns... 

There is a wide variety of commercially available chiral stationary phases and mobile phase additives.32 34 Preparative scale separations have been performed on the gram scale.32 Many stationary phases are based on chiral polymers such as cellulose or methacrylate, proteins such as human serum albumin or acid glycoprotein, Pirkle-type phases (often based on amino acids), or cyclodextrins. A typical application of a Pirkle phase column was the use of a N-(3,5-dinitrobenzyl)-a-amino phosphonate to synthesize several functionalized chiral stationary phases to separate enantiomers of... 

In the case of amorphous intermediate 4, the weakly acidic cation exchange resin DIAION WKIOO was selected as the CEC stationary phase. The resin comprises a methacrylate resin matrix and carboxylic acid, which functions as an exchange group with a high ion exchange rate. Figure 9.4 illustrates the concept behind the CEC procedure for 4 ... 

Electro-osmotic flow depends largely on the zeta potential arising at the interface of the stationary phase with the electrolyte. For this potential to develop, the stationary phase has to carry some amount of ionic or ionizable groups at its surface. When MIPs are used as stationary phase in CEC their composition has to be designed with this point in mind besides the usual criteria for an MIP. Some of the more usual MIP compositions, like MIPs made with the functional monomer methacrylic acid, satisfy this criterion quite well. It is also important that the stationary phase is well retained in the capillary. Frits at the ends of the packing are difficult to make in capillaries, and therefore monoliths covalently bound to the (prederivatized) surface of the capillary are advantageous. The preparation of MIP monoliths has been mostly promoted in the CEC field. 

Fig. 6.21. Electrochromatographic separation of benzene derivatives on monolithic capillary column prepared by UV initiated polymerization. Conditions capillary column, 100 pm i.d. x 25 cm active length stationary phase poly(butyl methacrylate-co-ethylene dimethaciylate) with 0.3 wt. % 2-acrylamido-2-methyl-l-propanesulfonic acid pore size, 296 nm mobile phase, 75 25 vol./vol mixture of acetonitrile and 5 mmol/L phosphate buffer pH 7 UV detection at 215 nm 25 kV pressure in vials, 0.2 MPa injection, 5 kV for 3 s. Peaks thiourea (1), benzyl alcohol (2), benzaldehyde (3), benzene (4), toluene (5), ethylbenzene (6), propylbenzene (7), butylbenzene (8), and amylbenzene (9).

The methacrylate-based polymers are stable even under extreme pH conditions such as pH 2 or 12. Fig. 6.24 shows the CEC separations of aromatic acids and anilines at these pH values [14]. The sulfonic acid functionalities of the monolithic polymer remain dissociated over the entire pH range creating a flow velocity sufficient to achieve the separations in a short period of time. In contrast to the stationary phase, the analytes are uncharged, yielding symmetrical peaks. Needless to say that typical silica-based packings may not tolerate such extreme pH conditions. 

Schmid et al. [60] demonstrated the enantiomer separation of underivatized amino acids on a monolithic chiral ligand-exchange phase by rod-CEC. The chiral stationary phase was prepared in situ in the capillary by polymerization of methacrylic acid, piperazine diacrylamide, vinylsulfonic acid and /V-(2-hydroxy-3-alloxypropyl)-L-4-hydroxyproline. The monolithic separation bed was covalently linked to the internal capillary wall and thus no frits were required. Fig. 9.13 shows the enantiomer separation of phenylalanine by (A) pure CEC (30 kV), (B) nano-LC (12 bar) and (C) pressure supported CEC (30 kV, 12 bar at the inlet vial). The shortest elution time was clearly obtained by pressure supported CEC, while the highest resolution was found in the pure CEC mode (CEC Rs = 2.11 nano-LC Rs = 0.98 pressure supported CEC Rs= 1.60). 

Finally, libraries aimed to chiral resolution of racemates will be covered here in particular, the use of chiral stationary phases (CSPs) has recently been reported for the identification of materials to be used for chiral separation of racemates by HPLC. The group of Frechet reported the selection of two macroporous poly methacrylate-supported 4-aryl-1,4-dihydropyrimidines (DHPs) as CSPs for the separation of amino acid, anti-inflammatory drugs, and DHP racemates from an 140-member discrete DHP library (214,215) as well as a deconvolutive approach for the identification of the best selector phase from a 36-member pool library of macroporous polymethacrylate-grafted amino acid anilides (216,217). Welch and co-workers (218,219) reported the selection of the best CSP for the separation of a racemic amino acid amide from a 50-member discrete dipeptide iV-3,5-dinitrobenzoyl amide hbrary and the follow-up, focused 71-member library (220). Wang and Li (221) reported the synthesis and the Circular Dichroism- (CD) based screening of a 16-member library of CSPs for the HPLC resolution of a leucine ester. Welch et al. recentiy reviewed the field of combinatorial libraries for the discovery of novel CSPs (222). Dyer et al. (223) reported an automated synthetic and screening procedure based on Differential Scanning Calorimetry (DSC) for the selection of chiral diastereomeric salts to resolve racemic mixtures by crystallization. Clark Still rejxrrted another example which is discussed in detail in Section 9.5.4. 

Hjerten et al. [124] introduced the monolithic stationary phases based on acrylamides in the late 1980s. The cross-linked polyacrylamide can be directly synthesized within the mold by a one step free-radical chain polymerization. Acrylamide, methacrylamide, or piperazine diacrylamide are often employed as monomers, while V,A -methylene-bis-acrylamide is used as a cross-linker. 2-acrylamido-2-methylpropane sulphonic acid, vinylsulphonic acid, butyl methacrylate, or stearoyl methacrylate are usually added to the polymerization mixture to provide charge and functional groups [114]. 

Xiong, B. etal. Capillary electrochromatography with monolithic poly(styrene-co-divinylbenzene-co-methacrylic acid) as the stationary phase, J. High Resol. Chromatogr., 23, 67, 2000. 

Note 8. Washing and equilibration of the packed column are very important. To obtain the release of the template, a mobile phase able to interfere with the noncova-lent interaction between the template, and the stationary phase should be used. In this tutorial methanol and acetic acid are very efficient, disrupting the hydrogen bond between theophylline and methacrylic acid residues. The column washing could be considered complete when the UV signal falls below 0.001 units of absorbance. 

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