Strong Anion-exchange Stationary Phase Research Articles

Separation of phenylpropionic acids both by strong anion exchange stationary phase and strong cation exchange stationary phase in supercritical fluid chromatography, using the same additive

This study demonstrates the expanded application of ion-exchange stationary phases (including strong cation exchange, SCX and strong anion exchange, SAX) in supercritical fluid chromatography (SFC), and more importantly, provides a deeper understanding of the retention mechanisms of these two stationary phases when using the same acidic additive. Phenylpropionic acid compounds (belonging to phenolic acids) were selected as probes. On the SCX column, the π-π and polar interactions originating from the bonded benzenesulfonic acid groups were important foundations for prolonging the retention time of solutes, but they were also the main reason for solutes’ tailing profiles. It was found that adding 0.1% phosphoric acid can generate sufficient strength of electrostatic repulsion to obtain satisfactory peak shapes. Here, phosphoric acid can be adsorbed on the surface of the stationary phase to accumulate more negative charges, and at the same time, the phosphate anion and the phenylpropionic acid in mobile phase might combine into the contact ion pair (CIP)- like through the polar interactions to form the apparently negatively charged solute. On the SAX column, phosphate anions generated by ionization of phosphoric acid, were considered as counterions to effectively mask the electrostatic attraction of quaternary ammonium groups, to significantly reduce the retention of phenylpropionic acids, but improve their resolution. Finally, utilizing two developed SFC methods, i.e., SCX with MeOH + 0.1% phosphoric acid in CO2, or SAX with MeOH + 0.3% phosphoric acid in CO2, the baseline separation of the extract of Lonicerae Japonicae Flos was achieved within 6 min and 10 min, respectively.

Arsenic Speciation in Fish and Rice by HPLC-ICP-MS Using Salt Gradient Elution

This paper presents arsenic speciation studies by high-performance liquid chromatography with inductively coupled plasma-mass spectrometer (HPLC-ICP-MS) using a chromatography column containing a strong anion exchange stationary phase (Agilent Bio SAX G3154A/101) for determination of arsenobetaine (AsB), dimethylarsinic acid (DMA), AsIII, monomethylarsonic acid (MMA), and AsV. The best separation was achieved in 15 min, using a 4 min isocratic step with 1 mM (NH4)2HPO4 pH 10.2, followed by a 2 min salt gradient from 1 to 10 mM, which was kept for 4 min. Compared to the Food and Drug Administration (FDA) method for rice using the PRP-X-100 column, the Bio SAX method was quicker, provided at least equal separation of AsB from the next peak, and led to more symmetrical peaks. The linearity between 0.5 and 60 µg L-1 was shown. The method was tested with 10 µg L-1 fortified rice and fish samples extracted in ultrasound using methanol/water 1:1, and recoveries between 40 and 90% were obtained.

Sulfobutylether-β-cyclodextrin as a chiral selector for separation of amino acids and dipeptides in chromatography

Various amino acids, dipeptides and their isomers were (enantio)separated using sulfobutylether-β-cyclodextrin as a chiral selector. Two different approaches were employed: first, dynamic coating of sulfobutylether-β-cyclodextrin onto a strong anion-exchange stationary phase and, second, use of sulfobutylether-β-cyclodextrin as a mobile phase additive in a separation system with a C18 column. Measurements were carried out using RP-HPLC and hydrophilic interaction liquid chromatography. Mobile phases composed of organic modifier (methanol) and four different aqueous parts: (i) deionized water, (ii) an aqueous solution of formic acid (pH 2.1), (iii) ammonium acetate buffer (pH 4.7), and (iv) ammonium acetate buffer (pH 8.8) in various volume ratios. Under these separation conditions, out of 23 chiral analytes, 9 were baseline enantio-resolved and 7 were partially separated. Of 9 mixtures of dipeptide isomers, 8 were baseline-separated. Sulfobutylether-β-cyclodextrin proved to be suitable for the separation of chiral and also achiral analytes. The use of sulfobutylether-β-cyclodextrin as a dynamic coating agent or as a mobile phase additive depends on the particular chromatographic system and analytes of interest.

Preparation of a dibenzylated 1,4-diazacyclohexane-derived strong anion-exchange stationary phase

This paper reports the preparation of a novel, silica-based, strong anion-exchange stationary phase from a 1,4-diazacyclohexane derivative. To prepare the difunctional strong anion-exchange stationary phase, activated silica beads were first bonded with 3-chloropropyltriethoxysilane and then reacted with 1-methylpiperazine followed by benzyl chloride. The silica beads, the strong anion-exchange stationary phase and its precusors were instrumentally characterized. Aromatic acids were separated with non-aqueous anion-exchange chromatography. After elution with eluant prepared in mixed solvents of water and methanol, the resulting 1,4-diazacyclohexane-derived, difunctional, strong anion-exchange stationary phase exhibited good separation and selectivity for the aromatic acids investigated. The effects of eluant pH, eluant ion concentration and solvent composites on the separations were investigated. Organic acids with different substituents were eluted in order of decreasing dissociation coefficients, with no observable peak shape differences.

MIXED-MODE MONOLITHIC SILICA AS A CHROMATOGRAPHIC SEPARATION MEDIUM

Capillary electro chromatography (CEC) is a separation technique that combines the dual properties of capillary zone electrophoresis (CZE) and micro-liquid chromatography (μ-LC). A hybrid-type silica monolith capillary column was prepared from a mixture of tetramethoxysilanes (TMOS) and methyltrimethoxysilanes (MTMS). Surface modification with octadecyldimethyl-(N-N-diethylamino)silane (ODS-DEA) and N-3[-(dimethylamino)propyl]acrylamide methyl] chloride-quaternary salt, (DMAPAA-Q) was carried out to form a mixed-mode of C18 and a strong anion exchange stationary phase. The synthesized mixed-mode silica monolithic column was then characterized physically with a scanning electron microscope (SEM) and shown to have an average through-pore size and skeleton size of 2.2 µm and 2.0 µm, respectively. This new column with mixed-mode characteristic of reversed-phase (RP) and strong anion exchange (SAX) functionalities was then applied as the stationary phase in CEC for the separation of inorganic and organic ions using a 50 mM phosphate electrolyte of pH 6.9. The inorganic and organic anions were base-line separated within 10 min with column efficiency up to 114 900 plates/min.

Sulfoalkylbetaine‐based monolithic column with mixed‐mode of hydrophilic interaction and strong anion‐exchange stationary phase for capillary electrochromatography

A novel monolithic stationary phase with mixed mode of hydrophilic and strong anion exchange (SAX) interactions based on in situ copolymerization of pentaerythritol triacrylate (PETA), N,N-dimethyl-N-methacryloxyethyl N-(3-sulfopropyl) ammonium betaine (DMMSA) and a selected quaternary amine acrylic monomer was designed as a multifunctional separation column for CEC. Although the zwitterionic functionalities of DMMSA and hydroxy groups of PETA on the surface of the monolithic stationary phase functioned as the hydrophilic interaction (HI) sites, the quaternary amine acrylic monomer was introduced to control the magnitude of the EOF and provide the SAX sites at the same time. Three different quaternary amine acrylic monomers were tested to achieve maximum EOF velocity and highest plate count. The fabrication of the zwitterionic monolith (designated as HI and SAX stationary phase) was carried out when [2-(acryloyloxy)ethyl]trimethylammonium methylsulfate was used as the quaternary amine acrylic monomer. The separation mechanism of the monolithic column was discussed in detail. For charged analytes, a mixed mode of HI and SAX was observed by studying the influence of mobile phase pH and salt concentration on their retentions on the poly(PETA-co-DMMSA-co-[2-(acryloyloxy)ethyl]trimethylammonium methylsulfate) monolithic column. The optimized monolith showed good separation performance for a range of polar analytes including nucleotides, nucleic acid bases and nucleosides, phenols, estrogens and small peptides. The column efficiencies greater than 192 000 theoretical plates/m for estriol and 135 000 theoretical plates/m for charged cytidine were obtained.

Preparation and characterization of silica confined ionic liquids as chromatographic stationary phases through surface radical chain-transfer reaction

Ionic liquids (ILs) have been used in various regions of chemistry. Ionic liquids have designability, thus giving us the ability to manipulate the structure (with respect to the organic cation, inorganic anion and the length of the side chain attached to the organic cation) and consequently their properties. In this manuscript, two new different chromatographic stationary phases based on different ionic liquid-modified silica particles were prepared through surface radical chain-transfer reaction and characterized with elemental analysis, Raman spectroscopy and X-ray photoelectron spectroscopy. 1-Allyl-3-butylimidazolium ionic liquid-modified silica particles can be used as a strong anion-exchange stationary phase. 1-Allyl-3-(butyl-4-sulfonate)imidazolium ionic liquid-modified silica particles were used for the first time as the covalently bonded charge-balanced zwitterionic stationary phase in electrostatic ion chromatography for the simultaneous separation of inorganic anions and cations with pure water as the mobile phase.

Anion exchange silica monolith for capillary liquid chromatography

An anion exchange monolithic silica capillary column was prepared by surface modification of a hybrid monolithic silica capillary column prepared from a mixture of tetramethoxysilane (TMOS) and methyltrimethoxysilane (MTMS). The surface modification was carried out by on-column copolymerization of N-[3-(dimethylamino)propyl]acrylamide methyl chloride-quaternary salt (DMAPAA-Q) with 3-methacryloxypropyl moieties bonded as an anchor to the silica surface to form a strong anion exchange stationary phase. The columns were examined for their performance in liquid chromatography (LC) and capillary electrochromatography (CEC) separations of common anions. The ions were separated using 50 mM phosphate buffer at pH 6.6. Evaluation by LC produced an average of 30,000 theoretical plates (33 cm column length) for the inorganic anions and nucleotides. Evaluation by CEC, using the same buffer, produced enhanced chromatographic performance of up to ca. 90,000 theoretical plates and a theoretical plate height of ca. 4 mum. Although reduced efficiency was observed for inorganic anions that were retained a long time, the results of this study highlight the potential utility of the DMAPAA-Q stationary phase for anion separations.

Monolithic columns with mixed modes of reversed-phase and anion-exchange stationary phase for capillary electrochromatography

A capillary electrochromatography (CEC) monolithic column with mixed modes of reversed-phase and anion-exchange stationary phases was prepared by in situ polymerization of 2-(methacryloxy)ethyltrimethylammonium methyl sulfate (MEAMS) and ethylene dimethacrylate (EDMA) in a binary porogenic solvent consisting 1-propanol and 1,4-butanediol. The ammonium groups on the surface of the stationary phase generate an electroosmotic flow (EOF) from cathode to anode, and serve as a strong anion-exchange stationary phase at the same time. The EOF of the stationary phase can be determined by the amount of MEAMS monomer in reaction mixtures during the polymerization. The monolithic stationary phases exhibited reversed-phase chromatographic behavior toward neutral solutes. For charged solutes, hydrophobic as well as electrostatic interaction/repulsion with the monoliths was observed. Separations of aromatic compounds and basic compounds on the prepared column were performed under the mode of CEC. Peak tailing of basic compounds was avoided and the efficient separation of aromatic acids was achieved using neutral mobile phase due to the same direction of EOF and electrophorestic mobility of negatively charged solutes.

Analysis of carbohydrates using different quaternized polystyrene-divinylbenzene particles and pulsed amperometric detection

This paper reports on the use of a polymer-based, strong anion-exchange stationary phase for rapid, selective and sensitive analysis of physiological important mono-, di- and oligosaccharides by high-performance anion-exchange chromatography (HPAEC) with pulsed amperometric detection (PAD) under alkaline conditions. The adsorbent was obtained by direct nitration of 3 and 5 μm, spherical non-porous highly cross-linked, styrene-divinylbenzene copolymer (PS-DVB) beads, followed by reduction of superficially introduced nitro groups with nascent hydrogen and quaternization of the resultant amino groups with iodomethane. Extended exposure to high pressure and strong alkaline conditions did not have any untoward effect on mechanical stability and chromatographic performance. A comparison of the 3 and 5 μm beads showed, that the synthesized 3 μm highly cross-linked PS-DVB particles are the preferred phases for the separation of monosaccharides and the 5 μm particles are preferable for the separation of oligosaccharides. To demonstrate the suitability for the analysis of complex samples, the optimized and validated system was used for the determination of glucose, fructose and sucrose in apple juice and other soft drinks such as Coca Cola. Finally, analysis within a few minutes without sample pretreatment down to a lower limit of detection of 0.174–0.504 μg mL−1 at a linearity with R2>0.994 and a repooducibility higher than 98% further confirmed the efficiency of these polymeric sorbents.

The Efficient Removal of Endotoxins from Insulin Using Quaternized Polyethyleneimine-Coated Porous Zirconia

The synthesis and use of a zirconia-based, alkali-stable strong anion-exchange stationary phase are described for the removal of pyrogenic lipopolysaccharides (LPS) from insulin. The strong anion-exchange material is produced by deposition of polyethyleneimine (PEI) onto porous zirconia particles, followed by cross-linking with a novel reagent, 1,2-bis-(2-iodoethoxy) ethane, and quaternization with iodomethane. Physical characterization of the chromatographic support shows that it has an ion-exchange capacity of 0.6 mmol/g, and 82% of the amine sites on the surface are in quaternized form. Isocratic elution of small benzoic acid derivatives shows good column efficiency. The two primary virtues of this material are its chemical stability under alkali conditions (up to pH 13) and its lower hydrophobicity compared to previously described alkali-stable PEI-coated zirconia supports cross-linked with 1,10-diiododecane. Using this new zirconia-based phase, a purification protocol is developed for the efficient removal of Escherichia coli 0111:B4 LPS from bovine insulin samples. An endotoxin clearance rate of up to 1.3 × 108 was attained. Endotoxin levels were reduced to less than 5 endotoxin units/ml even at initial contamination levels as high as 5.0 × 106 endotoxin units/ml. Furthermore, endotoxin adsorbed to the porous zirconia column may be easily removed (depyrogenated) using alkali for repeated purification cycles.

Determination of Anionic, Neutral, and Cationic Species of Arsenic by Ion Chromatography with ICPMS Detection in Environmental Samples

Ion chromatographic methods developed to separate either cationic, neutral, and anionic arsenic species or soluble and suspended arsenic species were successfully used in DORM-2 standard reference material and in water samples of environmental interest. The most effective separation of the analytes within 10 min was achieved with a nitric acid gradient elution using a strong anion-exchange stationary phase with additional capacity for hydrophobic interactions (IonPac AS7). The elemental-specific detection mode allows the sensitive determination of the arsenic species in the submicrogram per liter range. The calibration results were compared with those obtained by an alkaline water-methanol mixed eluent combined with a weak anion-exchange column (IonPac AS4A-SC). Differences in sensitivities were eclipsed by the low level of the baseline and the noise when using nitric acid. The gradient method was used to determine arsenic species in highly ferrous/ferric-contaminated leachates of lignite spoil. The companion elements underwent parallel screening to explain the interactions of arsenic species with the major elements.

Synthesis of a non-porous, polystyrene-based strong anion-exchange packing material and its application to fast high-performance liquid chromatography of proteins

Adsorbed coating technology has been used to produce a strong anion-exchange stationary phase on 3-μm non-porous poly(styrene-divinylbenzene) particles. In order to take full advantage of the excellent kinetic properties of the resultant packing material, small columns of 5 mm × 6 mm I.D. were used. These columns were pressure- and pH-stable and allowed protein separations to be made in less than 1 min at ambient temperature.