Hydrophilic Chromatography Research Articles

Chromatographic properties of silica modified with 6,10-ionene and sodium lignosulfonate

Adsorbents for hydrophilic chromatography: silica layer-by-layer modified with 6,10-ionene and sodium lignosulfonate (Sorbent-1), and silica modified with chitosan and sodium lignosulfonate (Sorbent-2) have been investigated. The influence of the nature of polycation used for the synthesis of adsorbents and the composition of the mobile phase on the retention and separation of phenol carboxylic acids have been studied. It was found that the retention of organic acids and the selectivity of their separation are higher on Sorbent-1. The possibility of the isocratic separation of a mixture of caffeine, sorbic, benzoic, vanillic, gallic, salicylic, sinapic, ferulic, p-cumaric, and caffeic acids in 30 min on a column filled with Sorbent-1 was demonstrated.

Rapid capillary electrochromatographic profiling of phytohormones on a hydrophilic interaction/strong anion-exchange mixed-mode monolith

An improved hydrophilic interaction/strong anion-exchange (HI-SAX) monolith for rapid capillary electrochromatographic profiling has been developed to detect carboxylic phytohormones with high resolution. The HI-SAX monolith was prepared by copolymerization of 2-(methacryloyloxy)ethyltrimethylammonium methyl sulfate, pentaerythritol triacrylate and porogenic solvents. Detailed polymerization compositions were investigated and improved, and the characteristics in terms of morphology, retention mechanism and column reproducibility were studied. A typical hydrophilic chromatography mechanism was observed when ACN content exceeded 30%, and the HI-SAX mixed-mode was verified with nucleic acid bases and nucleosides. Based on the improved HI-SAX monolith, capillary electrochromatographic profiling of typical phytohormones was evaluated by using indole-3-acetic acid (IAA), abscisic acid (ABA) and gibberellic acid (GA(3)) as the mode analytes. Effects of the test parameters on carboxyl phytohormones were investigated, and a fast analysis with high resolution for the representative phytohormones was achieved within 4.0 min with intraday RSDs (n = 3) less than 2.6% and 6.3% for the retention time and peak area, respectively. When applied to spiked corn samples, mean recoveries between 82.3% and 95.4% were obtained.

Three-dimensional patterning of porous materials using vapor phase polymerization

The ability to pattern porous materials with functional polymeric coatings is important for the fabrication of next-generation microfluidic platforms, membranes, tissue scaffolds, and optical devices. Here, we demonstrate for the first time that solventless initiated chemical vapor deposition (iCVD) can be used for three-dimensional patterning of porous substrates. The individual fibers of hydrophilic chromatography paper were uniformly coated with a thin layer of hydrophobic photoresponsive poly(o-nitrobenzyl methacrylate) (PoNBMA). X-Ray photoelectron spectroscopy and contact angle measurements confirmed that the PoNBMA coating penetrated the entire depth of the paper and scanning electron microscope images confirmed that the porosity and hierarchical structure of the paper were retained during the coating process. The PoNBMA coating was then patterned through the entire depth of the paper by exposure to ultraviolet light followed by rinsing in biologically compatible buffer. We demonstrated the utility of our patterning process by fabricating three-dimensional hydrophilic and hydrophobic regions into the chromatography paper for use as paper-based microfluidic devices. Our patterning process represents an environmentally friendly method to pattern three-dimensional materials since no organic solvents are used during the polymerization process or patterning step.

Simultaneous determination of sixteen metabolites related to neural tube defects in maternal serum by liquid chromatography coupling with electrospray tandem mass spectrometry

Disturbances in maternal folate, homocysteine, and glutathione metabolism have been reported to be associated with neural tube defects (NTDs). However, the role played by specific components in the metabolic pathways leading to NTDs remains unclear. Thus an analytical method for simultaneous measurement of sixteen compounds involved in such three metabolic pathways by high performance liquid chromatography–tandem mass spectrometry was developed. The use of hydrophilic chromatography column improved the separation of polar analytes and the detection mode of multiple-reaction monitoring (MRM) enhanced the specificity and sensitivity so as to achieve simultaneous determination of three class of metabolites which have much variance in polarity and contents. The influence of parameters such as temperature, pH, flow rate on the performance of the analytes were studied to get an optimal condition. The method was validated for its linearity, accuracy, and precision, and also used for the analysis of serum samples of NTDs-affected pregnancies and normal women. The result showed that the present method is sensitive and reliable for simultaneous determination of as many as sixteen interesting metabolites which may provide a new means to study the underlying mechanism of NTDs as well as to discover new potential biomarkers.

Recent Developments in the Enrichment of Glycopeptides for Glycoproteomics

Protein glycosylation has been recognized as one of the most important posttranslational modifications in nature. Although mass spectrometry has been proved as a powerful technique in glycoproteomic and glycomic research, an efficient enrichment procedure of glycopeptides is highly required to reduce the biological sample complexity. We summarize various enrichment approaches for extracting glycopeptides from biological milieus prior to mass spectrometric analysis. These methodologies include lectin‐based affinity chromatography, hydrophilic chromatography, and oxidation‐hydrazine chemistry.

High-Throughput Quantitative Profiling of Serum N-Glycome by MALDI-TOF Mass Spectrometry and N-Glycomic Fingerprint of Liver Fibrosis

The use of MALDI-TOF mass spectrometry (MS) in quantitative glycan profiling has not been reported. In this study, we attempted to establish a high-throughput quantitative assay for profiling serum N-glycome, and we applied the new assay to identifying serum N-glycans for diagnosis of liver fibrosis and cirrhosis. N-glycans from whole serum proteins in 2 microL serum were released by enzymatic digestion, cleaned up by hydrophilic chromatography, and subsequently quantitatively profiled with a linear MALDI-TOF MS system, which was originally designed for quantitative proteomic profiling. Serum N-glycome profiles from 46 patients with chronic hepatitis B infection and with different degrees of liver fibrosis were examined. The intra- and interassay CVs of peak intensities of the standard N-glycans were <8% and <17%, respectively. When the assay was applied to the analysis of serum N-glycome profiles, 17 peaks were found to be potential biomarkers for detection of liver fibrosis/cirrhosis. Linear regression analysis revealed that 4 peaks of 1341.5, 1829.7, 1933.3, and 2130.3 m/z (all P <0.005) had complementary value in detecting liver fibrosis and included them, but not any serological markers, in the diagnostic model. Leave-one-out cross-validation showed the diagnostic model could identify significant fibrosis (Ishak score > or = 3) and cirrhosis (Ishak score > or = 5), both at 85% accuracy. This is the first study to illustrate the quantitative aspect of MALDI-TOF MS in N-glycome profiling and the first study to reveal the potential value of the serum N-glycan profile for identifying liver fibrosis.

Hydrophilic-interaction chromatography for the separation of peptides, nucleic acids and other polar compounds

When a hydrophilic chromatography column is eluted with a hydrophobic (mostly organic) mobile phase, retention increases with hydrophilicity of solutes. The term hydrophilic-interaction chromatography is proposed for this variant of normal-phase chromatography. This mode of chromatography is of general utility. Mixtures of proteins, peptides, amino acids, oligonucleotides, and carbohydrates are all resolved, with selectivity complementary to those of other modes. Typically, the order of elution is the opposite of that obtained with reversed-phase chromatography. A hydrophilic, neutral packing was developed for use in high-performance hydrophilic-interaction chromatography. Hydrophilic-interaction chromatography is particularly promising for such troublesome solutes as histones, membrane proteins, and phosphorylated amino acids and peptides. Hydrophilic-interaction chromatography fractionations resemble those obtained through partitioning mechanisms. The chromatography of DNA, in particular, resembles the partitioning observed with aqueous two-phase systems based on polyethylene glycol and dextran solutions.