Polar Stationary Phases Research Articles

Preparation and evaluation of a neutral methacrylate-based monolithic column for hydrophilic interaction stationary phase by pressurized capillary electrochromatography

A polar and neutral polymethacrylate-based monolithic column was evaluated as a hydrophilic interaction capillary electrochromatography (HI-CEC) stationary phase with small polar–neutral or charged solutes. The polar sites on the surface of the monolithic solid phase responsible for hydrophilic interactions were provided from the hydroxy and ester groups on the surface of the monolithic stationary phase. These polar functionalities also attract ions from the mobile phase and impart the monolithic solid phase with a given zeta potential to generate electro-osmotic flow (EOF). The monolith was prepared by in situ copolymerization of a neutral monomer 2-hydroxyethyl methacrylate (HEMA) and a polar cross-linker with hydroxy group, pentaerythritol triacrylate (PETA), in the presence of a binary porogenic solvent consisting cyclohexanol and dodecanol. A typical HI-CEC mechanism was observed on the neutral polar stationary phase for both neutral and charged analytes. The composition of the polymerization mixture was systematically altered and optimized by altering the amount of HEMA in the polymerization solution as well as the composition of the porogenic solvent. The monoliths were tested in the pCEC mode. The resulting monoliths had different characteristics of hydrophilicity, column permeability, and efficiency. The effects of pH, salt concentration, and organic solvent content on the EOF velocity and the separation of nucleic acids and nucleosides on the optimized monolithic column were investigated. The optimized monolithic column resulted in good separation and with greater than 140,000 theoretical plates/m for pCEC.

Single fiber-in-capillary annular column for gas chromatographic separation

A method for preparation of a stationary phase-adjustable column with in-column stationary phase-coated fused-silica fiber annular column was successfully developed. The surface of a 0.12 mm o.d. bare optical fiber was first coated with a stationary phase and then inserted into a fused-silica capillary (non-coated or coated) as an annular column for gas chromatographic study. The optical fiber and capillary were coated with polydimethylsiloxane (SE-30) and polyethylene glycol 20M (PEG-20M) as nonpolar and polar stationary phases, respectively. Among the investigated annular and open tubular columns, the PEG-20M-coated fiber-in-PEG-20M-coated capillary annular column showed the highest column efficiency with a minimum plate height of 0.35 mm and an optimum gas velocity of 25 cm/s. When a SE-30/PEG-20M-coated fiber-in-uncoated capillary annular column was applied to separate a 9-component complex mixture, the total analysis time was 5.3 min and the column length was 12 m. By contrast, when a SE-30-coated fiber-in-PEG-20M-coated capillary annular column was used to separate the same 9-component mixture, the analysis time was reduced to 3.5 min and the column length was shortened by half to 6 m. Our results show that the stationary phase-coated fiber-in-stationary phase-coated capillary annular column is a better choice for gas chromatographic separation as it is more efficient and flexible. In addition, the proposed annular column design provides flexibility in using two or even more types of stationary phases to achieve optimal analytical separation.

Effect of salt in the mobile phase on the critical conditions of poly(ethylene glycol) in liquid chromatography-mass spectrometry coupling

The nature of salts introduced in the chromatographic mobile phase to promote on-line electrospray ionization was shown to be a key parameter in the optimization of LCCC-MS couplings, both using non-polar (Si-C18) and polar (Si-NH2) stationary phases. The critical conditions of poly(ethylene glycol), which reflect a compensation process between exclusion and interaction effects, were strongly modified when changing the size of the cation in the eluent. This phenomenon could be attributed to interactions between the cation and water molecules from the mobile phase, in the case of the non-polar stationary phase, giving rise to a salting out effect due to a lowered solvent quality of the eluent; or between the cation and amino-modified silanols of the polar stationary phase, inducing a decrease of the surface adsorptivity. Accordingly, increasing the size of the cation in the mobile phase caused the polymer molecules to be eluted according to the adsorption mode using non-polar adsorbent and according to the exclusion mode using polar stationary phase.

Recent progress of polar stationary phases in CEC and capillary liquid chromatography

The stationary phases including the nonpolar and polar stationary phases in microscale separations have been greatly developed. This review describes the recent progress of the polar stationary phases (PSPs) with the focus on their preparation and the interesting applications in the field of CEC and capillary liquid chromatography (CLC) covering the literatures from the middle of 2006 to the middle of 2008. The PSPs described herein were summarized into three types as open-tubular, particle-packed and monolithic stationary phases for either CEC or CLC, and the separation mechanisms are of hydrophilic interaction, chiral selection, ion-exchange and/or electrostatic interaction, etc. After overviewing the literatures published in the last 2 years, the research efforts on PSPs for CEC and CLC have been remaining in the fabrication of open-tubular and monolithic capillary columns; whereas, the research endeavor of PSPs on particulate-packed format seemed to decline most likely due to the commercial availability of various packing particles for CEC and CLC.

Authentication of fattening diet of goat kid according to their fatty acid profile from perirenal fat

Fatty acids of forty-two samples of perirenal fat of goat kids reared on three different feeding systems: goat milk (B), milk replacer (R) and milk-based starter (F) have been analyzed by Gas Chromatography flame ionization detector. The lipids were extracted by melting of perirenal fat in a microwave oven. The fat was then filtered and dissolved in hexane. This analysis was performed on a column (100 m × 0.25 mm i.d. and 0.25 μm film thickness) coated with a polar stationary phase HP-88 and flame ionization detector was used. Hydrogen (25 psi inlet constant pressure) was used as carrier gas. Programmed temperature was kept at 175 °C and held isothermally for 10 min, and was then raised to 205 °C at a rate of 3 °C/min and held isothermally for 10 min. By using the fatty acids as chemical descriptors, pattern recognition techniques were applied to differentiate between goat milk, milk replacer and milk-based starter fattening diet of goat kid. C18:2 and C18:3 acids were found to be the most differentiating variables.

Modeling of retention of adrenoreceptor agonists and antagonists on polar stationary phases in hydrophilic interaction chromatography: a review

Retention prediction models for reversed-phase liquid chromatography (RPLC) have been extensively studied owing to the fact that RPLC remains the most widely used chromatographic technique especially in the field of pharmaceutical and biomedical analyses. However, RPLC is not always the method of choice for the analysis of some compounds that have high polarity. Hydrophilic interaction chromatography (HILIC) has been gaining interest in the last few years as an alternative option to RPLC for the analysis of polar and hydrophilic analytes. HILIC is a variant of normal-phase liquid chromatography, but utilizes water in a water-miscible organic solvent as the eluent in conjunction with a hydrophilic stationary phase. The present review aims to summarize recent contributions on the development of retention prediction models for a group of basic analytes, namely, the adrenoreceptor agonists and antagonists, on different polar stationary phases. The use of multiple linear regression and artificial neural networks in model building is highlighted.

Preparation and evaluation of a lysine‐bonded silica monolith as polar stationary phase for hydrophilic interaction pressurized capillary electrochromatography

A silica-based monolith as polar stationary phase was described for hydrophilic interaction pressurized capillary electrochromatography (HI-pCEC). The polar monolithic column was prepared by on-column reaction of lysine with epoxy groups on a gamma-glycidoxypropyltrimethosysilane-modified silica monolith. The stationary phase yielded strong hydrophilic interaction due to the slightly polar hydroxyl groups, and the strong polar lysine ligand with amino groups and carboxylic groups contained on the surface of the monolith. In order to evaluate the hydrophilic character of lysine ligand, the chromatographic behaviors of epoxy monolith (before lysine bonded) and diol monolith (hydroxyl groups contained) were also investigated. Two groups of comparative experiment were developed in terms of the separation of typical neutral non-polar and polar compounds performed in a mobile phase of aqueous-acetonitrile solution. Results showed that the lysine monolith was much more hydrophilic than the diol monolith, which presented less hydrophobic than the epoxy monolith. For further study on its hydrophilic character, the lysine monolith was demonstrated in the HI-pCEC mode for the separations of various polar compounds such as phenols, nucleic acid bases and nucleosides.

Investigation of retention behavior of drug molecules in supercritical fluid chromatography using linear solvation energy relationships

In this study, the retention behaviors of pharmaceutical and drug-like compounds in supercritical fluid chromatography (SFC) with polar stationary phases were evaluated using linear solvation energy relationships (LSERs). We used more than 200 compounds, including mono-, di-substituted benzenes, and other marketed drugs with a wide range of solvation descriptors. The dominant contribution to positive retention was the hydrogen bond donor acidity of the solutes, particularly for pyridine and amino columns. That factor is determined by the difference in basicity between the mobile and stationary phases. Another significant contributor to retention behavior was the hydrogen bond acceptor basicity of the solutes made particularly for diol and amino columns. The hydrogen bond donating ability is small for cyano and pyridine stationary phases (or should be zero) since those two phases lack hydrogen bond donor groups. The LSER results showed that the SFC retention behavior of compounds using carbon dioxide–methanol with polar stationary phases was close to that reported for normal phase LC using hexane–methanol. As expected, mobile phases containing high percentages of modifier resulted in decreased retention times for most compounds. A greater knowledge of the nature of solute/stationary phase and solute/mobile phase allows a more rapid and efficient choice in high-throughput screening of compound libraries.

A gas-chromatographic study of the retention of reaction components in the catalytic oxidation of β-picoline to nicotinic acid

The gas-chromatographic retention of pyridine, β-picoline, 3-pyridinecarbaldehyde, 3-pyridinenitrile, nicotinic acid, and nicotinamide on polar stationary phases was studied. A scheme was proposed and a procedure was developed for the determination of the reaction components of the catalytic oxidation of β-picoline to nicotinic acid on a column (3 m × 3 mm) packed with Chromosorb WAW + 10 wt % FFAP. The separation time was 28 min. The detection limits were 0.01 and 0.37 ppm for 3-pyridinenitrile and nicotinic acid, respectively.

Gas Chromatographic Retention Indices of Fentanyl and Analogues

Narcotic analgesics of the fentanyl class are characterized by high potency and relatively short duration of action. These compounds nowadays have become a substitute for heroine and are highly addictive for abusers. Herein, we report retention indices of fentanyl and its eighteen analogues relative to the homologous n-alkane series. These values are determined on a moderately polar BP-5 capillary column under programmed temperature and isothermal chromatographic conditions. The analogues differ in the substituent attached to the piperidine ring nitrogen, and retention indices are found to vary according to the nature of the substituent. The effects of chromatographic conditions like temperature programming rate, carrier gas flow rate, and oven temperature are studied. Retention indices are also determined on a non-polar BP-1 column to study the influence of stationary phase polarity. Standard deviation of all the RI values is less than one index unit.

Application of hydrophilic interaction chromatography for simultaneous separation of six impurities of mildronate substance

The possibility of separating the impurities of mildronate, an antiischemic drug, by hydrophilic interaction chromatography (HILIC) was investigated on different polar stationary phases (silica, amino, cyano and zwitterionic sulfobetaine). The investigations have shown that HILIC is a useful alternative to reversed phase and ion-pair chromatography. The impact of HILIC separation conditions (acetonitrile content, buffer pH in mobile phase) on retention and selectivity has been systematically studied. Importance of these factors was found to be dependent on the structural properties of solutes. A HILIC method using a zwitterionic sulfobetaine stationary phase was developed and validated to determine six impurities in the drug substance. The method was validated in terms of specificity, limit of quantitation, limit of detection, linearity, accuracy and precision.

Optimisation of the GC‐MS conditions for the determination of the 15 EU foodstuff priority polycyclic aromatic hydrocarbons

European legislation has recently established a list of 15 priority polycyclic aromatic hydrocarbons (PAHs) to be monitored in foodstuff. Thus, the accurate determination of these compounds has become a highly relevant issue. The fact that some of these European Union (EU) PAHs differ from those typically analysed, requires the re-evaluation of the instrumental conditions for the proper determination of the new target compounds. In this study, the influence of the stationary phase and dimensions of the GC capillary column on the chromatographic resolution of the 15 EU PAHs has been investigated. Apolar (DB-5 type) and medium polar (DB-17 type) stationary phases with different lengths and film thickness have been evaluated for the separation of the target compounds, with special emphasis on those coelutions involving isomers such as the three benzofluoranthenes included in the EU PAHs. In addition, the influence of the injection technique and the column dimensions on the recovery of the high molecular mass PAHs has been studied. A programmable temperature vaporising (PTV) injector has been used in three different operational modes and the results were compared to those obtained using on-column injection. The experimental parameters involved in the injection step were optimised by using experimental design.

Comparative Study of Cyperus rotundus Essential Oil by a Modified GC/MS Analysis Method. Evaluation of Its Antioxidant, Cytotoxic, and Apoptotic Effects

Gas chromatography coupled with mass spectrometry (GC/MS), using both electron impact (EI) and chemical ionization (CI) detection modes on apolar and polar stationary phases, led to the determination of the volatile composition of the essential oil obtained from tubers of Cyperus rotundus (Cyperaceae). In this study, more than 33 compounds were identified and then compared with the results obtained in our previous work. Cyperene, alpha-cyperone, isolongifolen-5-one, rotundene, and cyperorotundene were the principal compounds comprising 62% of the oil. An in vitro cytotoxicity assay with MTT indicated that this oil was very effective against L1210 leukaemia cells line. This result correlates with significantly increased apoptotic DNA fragmentation. The oxidative effects of the essential oil were evaluated using the 1,1-diphenyl-2-picrylhydrazyl (DPPH), xanthine/xanthine oxidase assays, and the scavenging of superoxide radical assay generated by photo-reduction of riboflavin. The antimutagenic activity of essential oil has been examined by following the inhibition of H(2)O(2) UV photolysis which induced strand-break formation in pBS plasmid DNA scission assay. Based on all these results, it is concluded that C. rotundus essential-oil composition established by GC/MS analysis, in EI- and CI-MS modes, presents a variety of a chemical composition we were not able to detect with only GC/MS analysis in our previous work. This essential oil exhibited antioxidant, cytotoxic, and apoptotic properties.

Separation properties of novel and commercial polar stationary phases in hydrophilic interaction and reversed‐phase liquid chromatography mode

Four novel nonionic polar stationary phases were synthesised by anchoring first 2-mercaptoethanol and 1-thioglycerol, respectively, onto vinylised silica (ME and TG packings) followed by an on-phase oxidation with excess hydrogen peroxide in aqueous medium which yielded sulphoxide analogues of the embedded sulphide groups, i. e. oxidised 2-mercaptoethanol (MEO) and oxidised 1-thioglycerol (TGO) packings. Chromatographic characteristics of these stationary phases were evaluated comparatively to three commercial so-called 'diol' columns. U-shaped response curves of retention factors of adenosine and guanosine with hydro-organic eluents containing 5-95% v/v ACN as well as noticeable CH(2)-increment selectivity demonstrated multimodal separation capabilities of the developed amphiphilic materials, i. e. columns can be operated both in hydrophilic interaction chromatography (HILIC) and in RP mode. Although the selector ligands were physico-chemically related, considerably differing retention and selectivity patterns were observed in the HILIC mode. Thereby the introduction of additional hydroxyl groups in the chromatographic ligand resulted in selectivity increments that were different from those obtained by sulphur oxidation. For example, a set of five vitamins delivered five different elution orders with the overall seven columns. A close examination of HILIC separations of nucleobases and nucleosides on the developed packings revealed that (i) the amount of ACN in the eluent adopts a pivotal role in adjusting retention, (ii) the linearity of the relationship log (retention factor) versus log (volume fraction of water in the eluent) increases with phase polarity in the range of 5-40% v/v water, (iii) the slopes are higher with solutes having more polar interactive sites, (iv) the van't Hoff plots are linear (range 15-45 degrees C) with negative retention enthalpy values DeltaH (-4.5 to -14.5 kJ/mol) and (v) the -DeltaH values tend to be higher with more polar phases and more polar analytes. Based on these data the HILIC retention mechanism is described to be composed of both partitioning and adsorption processes. Distinct types of polar interactive sites in the chromatographic ligands may generate mixed-mode HILIC separation conditions that may additionally be superimposed by surface silanol contributions.

Overview of the retention in subcritical fluid chromatography with varied polarity stationary phases

Retention and separation of achiral compounds in supercritical fluid chromatography (SFC) depend on numerous parameters: some of these parameters are identical to those encountered in HPLC, such as the mobile phase polarity, while others are specific to SFC, as the density changes of the fluid, due to temperature and/or pressure variations. Additional effects are also related to the fluid compressibility, leading to unusual retention changes in SFC, for instance when flow rate or column length is varied. These additional effects can be minimised by working at lower temperatures in the subcritical domain, simplifying the understanding of retention behaviours. In these subcritical conditions, varied modifiers can be mixed to carbon dioxide, from hexane to methanol, allowing tuning the mobile phase polarity. With nonpolar modifiers, polar stationary phases are classically used. These chromatographic conditions are close to the ones of normal-phase LC. The addition of polar modifiers such as methanol or ACN increases the mobile phase polarity, allowing working with less polar stationary phases. In this case, despite the absence of water, retention behaviours generally follow the rules of RP LC. Moreover, because identical mobile phases can be used with all stationary phase types, from polar silica to nonpolar C18-bonded silica, the classical domains, RP and normal-phase, are easily brought together in SFC. A unified classification method based on the solvation parameter model is proposed to compare the stationary phase properties used with the same subcritical mobile phase.

The orthogonal character of stationary phases for gas chromatography

A database of system constants for 32 open-tubular columns at 100 degrees C is used to identify stationary phases for obtaining a wide selectivity space in comprehensive GC. Three parameters based on the Euclidean distance (D-parameter) or vectors (d-parameter and costheta) in hyperspace are used to establish the chemical similarity and retention correlation as an inverse scale of selectivity differences. It is shown that the poly(methyloctylsiloxane) stationary phase is the best candidate for a low-selectivity stationary phase and affords a wider selectivity space when combined with a selective polar stationary phase than poly(dimethylsiloxanes). The most suitable polar stationary phases are poly(ethylene glycols) or bis(cyanopropylsiloxane-co-silarylenes and to a lesser extent poly(methyltrifluoropropylsiloxanes). No systems are truly orthogonal but angles between individual stationary phase vectors of about 75 degrees are possible by choosing the correct combination of stationary phases.

A unified classification of stationary phases for packed column supercritical fluid chromatography

The use of supercritical fluids as chromatographic mobile phases allows to obtain rapid separations with high efficiency on packed columns, which could favour the replacement of numerous HPLC methods by supercritical fluid chromatography (SFC) ones. Moreover, despite some unexpected chromatographic behaviours, general retention rules are now well understood, and mainly depend on the nature of the stationary phase. The use of polar stationary phases improves the retention of polar compounds, when C18-bonded silica favours the retention of hydrocarbonaceous compounds. In this sense, reversed-phase and normal-phase chromatography can be achieved in SFC, as in HPLC. However, these two domains are clearly separated in HPLC due to the opposite polarity of the mobile phases used for each method. In SFC, the same mobile phase can be used with both polar and non-polar stationary phases. Consequently, the need for a novel classification of stationary phases in SFC appears, allowing a unification of the classical reversed- and normal-phase domains. In this objective, the paper presents the development of a five-dimensional classification based on retention data for 94–111 solutes, using 28 commercially available columns representative of three major types of stationary phases. This classification diagram is based on a linear solvation energy relationship, on the use of solvation vectors and the calculation of similarity factors between the different chromatographic systems. This classification will be of great help in the choice of the well-suited stationary phase, either in regards of a particular separation or to improve the coupling of columns with complementary properties.

Electrochromatographic characterization of methacrylate-based monolith with mixed mode of hydrophilic and weak electrostatic interactions by pressurized capillary electrochromatography

A monolithic stationary phase was prepared by in situ copolymerization of 2-hydroxyethyl methacrylate (HEMA), ethylene dimethacrylate (EDMA), and methacrylic acid (MAA), in a binary porogenic solvent consisting of toluene and 1-dodecanol. The resulting monolith was evaluated as a hydrophilic interaction-capillary electrochromatography (HI-CEC) stationary phase under the mode of pressurized capillary electrochromatography (pCEC). Effects of the buffer pH, salt concentration and the mobile phase composition on the electroosmotic flow (EOF) velocity and the retention factors of the compounds were investigated. The generation of cathodic EOF under a broad pH range was attributed to the presence of the carboxyl groups on the surface of the polar stationary phase. The carboxyl groups offered at the same time the possibility of weak electrostatic interaction with analytes. The separation mechanism of the monolithic column was discussed in detail. It was found that the separation mechanism of charged solutes could be attributed to a mixed mode of HI and weak electrostatic interaction, as well as the effect of electrophoresis, while the separation of neutral solutes was based on the hydrophilic interaction at high acetonitrile (ACN) content.

A Simple, Convenient, and Efficient Preparative GC System that Uses a Short Megabore Capillary Column as a Trap

A simple, convenient, and highly efficient preparative GC system has been developed that uses short sections of megabore capillary columns as sample collection (sorbent) traps. The performance of this system with various types of capillary column traps and under various collection conditions was systematically investigated with model compounds, including C4 to C20 normal alkanes, esters, and alcohols. The thickness and polarity of the sorptive stationary phase and the temperature of the collection trap affected trap performance. Each group of compounds was efficiently trapped above a critical Kovat's index, and the type of trap (deactivated, methyl polysiloxane, polyethylene glycol), film thickness, and whether or not the trap was cooled significantly shifted this threshold index. Above this critical index, recovery efficiencies of traps with methyl polysiloxane films were 80-100% for a wide range of injected sample mass. For example, a DB-1 collection trap with a film thickness of 1.5 microm methyl polysiloxane operated at ambient temperature trapped >84% of the mass of injected compounds of all three chemical classes with Kovat's index >1,100 (determined on a nonpolar column) with injected sample mass ranging from 10 to 1,000 ng of each compound. This preparative GC system is technically and economically feasible for most researchers. Furthermore, it is suitable for the preparation of NMR samples of volatile and semivolatile compounds, especially with sample sizes ranging from several nanograms to several micrograms.

A New Approach to Estimate the In-mouth Release Characteristics of Odorants in Chewing Gum

The objective of this study was to develop a new approach that can be used to estimate the in-mouth release characteristics of odorants in chewing gum. This technique, called the “Retronasal Flavor Impression Screening System (R-FISS),” is based on a modified dynamic headspace gas sampling technique. By application of the R-FISS technique, the in-mouth release characteristics of odorants during the chewing of gum was indicated by the peak area ratio, which was calculated by comparing the peak area for 10 min to the peak area for 1 min, of each odorant exhaled from the human nose. In addition, a good overall regression coefficient was found for the correlation between the peak area ratios of each odorant in the model chewing gum obtained by R-FISS and the retention indices on a polar stationary phase GC column (DB-Wax). Therefore, the in-mouth release of odorants in chewing gum seems to be capable of being predicted by their RIs on a polar stationary phase GC column (DB-Wax), and these results appear to suggest that two parameters (vapor pressure and hydrophobicity/hydrophilicity) are the key factors for determining the in-mouth release of odorants from chewing gum.