Normal-phase TLC separation of enantiomers of 1.4-dihydropyridine derivatives

A new TLC-based method was proposed for the separation of enantiomers and mixtures of racemic DHP derivatives differing in the kind of substituent in the phenyl ring. The conditions for the effective determination of the substances involved and the mechanism of their sorption were also studied. For the separation of felodipine, nilvadipine, and isradipine enantiomers, thin-layer chromatography was used, with a chiral stationary phase of the ligand exchange type, and developing phases of a different concentration of methanol (phi) as an organic modifier. The retention coefficient values k' were used to make the plots log k' = f(log phi) and log k' = f(phi). The processes taking place in the chromatographic systems were shown to be described by the Snyder-Soczewiński equation.     

Solubility Determination and Modeling and Dissolution Thermodynamic Properties of Raspberry Ketone in Binary Solvent Mixtures of Ethanol and Water

The solubility and dissolution thermodynamic properties of raspberry ketone in a set of binary solvent mixtures (ethanol + water) with different compositions were experimentally determined by static gravimetrical method in the temperature range of 283.15–313.15 K at 0.10 MPa. The solubility of raspberry ketone in this series of ethanol/water binary solvent mixtures was found to increase with a rise in temperature and the rising mole fraction of ethanol in binary solvent mixtures. The van’t Hoff, modified Apelblat and 3D Jouyban–Acree–van’t Hoff equations were increasingly applied to correlate the solubility in ethanol/water binary solvent mixtures. The former two models could reach better fitting results with the solubility data, while the 3D model can be comprehensively used to estimate the solubility data in all the ratios of ethanol and water in binary solvent mixtures at random temperature. Furthermore, the changes of dissolution thermodynamic properties of raspberry ketone in experimental ethanol/water solvent mixtures were obtained by van’t Hoff equation. For all the above experiments, these dissolution processes of raspberry ketone in experimental ethanol/water binary solvent mixtures were estimated to be endothermic and enthalpy-driven.     

Electrochemically modulated liquid chromatography and the Gibbs adsorption equation

Electrochemically modulated liquid chromatography uses a conductive material like porous graphitic carbon (PGC) as a chromatographic stationary phase and a working electrode. This dual functionality enables manipulation of separations by changes in the potential applied (E(app)) to the packing. Thus, by monitoring the retention factor (k') with respect to E(app), a chromatographic tool for examination of electrosorption processes can be devised. This novel capability is developed herein by examining the retention of charged aromatic compounds at PGC. The Gibbs adsorption equation and related formulations (e.g., the Lippmann equation) are used to determine interfacial excesses (Gamma) of these solutes in different supporting electrolytes, changes in interfacial tension (dgamma), the charge on the electrode (q(M)), and the potential of zero charge (PZC). Values of the PZC were also determined from plots of ln k' versus E(app). In this case, the dependence of ln k' reveals a shift in the PZC to more cathodic values as the strength of specific adsorption by anions as the electrolyte concentration increases. Together, these results provide insights into the retention mechanism and, more generally, to adsorption at electrified carbon electrodes. Extensions of this strategy as a probe of electrified interfaces with respect to mobile-phase composition, temperature, and pressure are briefly described.     

Characterization of methanol-water and acetonitrile-water association using multivariate curve resolution methods

The solvatochromic comparison method has been used to probe the interactions of solutes with binary solvent mixtures of methanol-water and acetonitrile-water. The solute spectra recorded in these mixtures are composed of the additive spectral contributions of the different solvated species of the solute, i.e., the water-solvated species, the cosolvent-solvated species, and the species solvated by water-solvent complexes. Multivariate curve resolution-alternating least squares has been used to model the solvation of the solutes as a function of the composition of the binary solvent mixture. Spectra and concentration profiles of the dye surrounded by the different solvation environments have been isolated. For the first time, solute spectra solvated exclusively by methanol-water and acetonitrile-water complexes have been obtained, and the solvatochromic parameters of dipolarity/polarizability and hydrogen-bonding acidity have been estimated for these complex species.     

A thermodynamic study of the temperature-dependent elution order of cyclic alpha-amino acid enantiomers on a copper(II)-D-penicillamine chiral stationary phase

The reversal of the elution order of cyclic alpha-amino acid enantiomers as a function of the temperature on a copper(II)-N,S-dioctyl-D-penicillamine ligand-exchange column is described. The thermodynamic parameters accounting for the retention and the separation of analytes were determined by means of van't Hoff plots. The influence of different chromatographic conditions on these parameters was investigated, showing little effect of the Cu(II) concentration in the eluent but strong influence of the organic modifier content on the separation. Further, the pH of the mobile phase was found to be a determining factor for the retention of the analytes. Based on these findings, a separation mechanism is postulated comprising the importance of complex formation for primary docking at the stationary phase, while hydrophobic interactions are crucial for chiral discrimination.     

Reproducibility of temperature-selected mass spectra in matrix-assisted laser desorption ionization of peptides

Matrix-assisted laser desorption ionization of peptides was investigated using α-cyano-4-hydroxycinnamic acid as the matrix. In each experiment, a set of mass spectra was collected by repetitive irradiation of a spot on a sample. Even though shot-to-shot variation in spectral pattern was significant, it was reproducible for different spots and samples. Each spectrum was tagged with the temperature in the early plume (T(early)) estimated through kinetic analysis of the peptide ion survival probability. T(early) decreased as the shot continued because the thermal conduction got more efficient as the sample got thinner. From each spectral set collected under various experimental conditions, a spectrum tagged with a particular T(early) was selected. Then, patterns of the spectra thus selected were the same. The reaction quotient for the matrix-to-peptide proton transfer determined at a specified T(early) was independent of the sample composition, indicating quasi-thermal equilibrium for this reaction. Furthermore, the van't Hoff plots were linear, also indicating quasi-thermal equilibrium. This, together with the thermal kinetics for the fragmentation of peptide and matrix ions, is responsible for the reproducibility of the mass spectral pattern at a specified T(early).     

Temperature dependence of retention in reversed-phase liquid chromatography. 2. Mobile-phase considerations.

The retention mechanism in reversed-phase liquid chromatography (RPLC) has been examined over a wide temperature range with emphasis on the role of the mobile phase. van't Hoff plot shapes were used to assess the retention mechanism, and the data showed evidence of the hydrophobic effect when water-rich and/or hydrogen-bonded mobile phases such as methanol/water were used. However, different van't Hoff plot shape was observed with acetonitrile/water mobile phases, indicating a change in the retention mechanism. These data showed that the hydrophobic effect, which had previously been proposed as the driving force for retention, is not a satisfactory explanation for the retention process in all RPLC systems.     

Retention of ionizable compounds on HPLC. 5. pH scales and the retention of acids and bases with acetonitrile-water mobile phases

The pH calibration procedures that lead to the different pH scales in acetonitrile-water mixtures used as mobile phases in reversed-phase liquid chromatography are discussed. Appropriate buffers of known pH value in acetonitrile-water mixtures are selected and used to establish the relationship (delta values) between the two rigorous acetonitrile-water pH scales: sspH and wspH (pH measured in acetonitrile-water mixtures and referred to acetonitrile-water or water, respectively, as standard state). These delta values allow one to convert pH values measured in acetonitrile-water with electrode systems calibrated with aqueous buffers (wspH scale) to sspH values, which are directly related to the thermodynamic acid-base constants. This offers an easy way to measure the pH of acetonitrile-water mobile phases and to relate this pH to the chromatographic retention of acids and bases through the thermodynamic acid-base constants. The relationships are tested for the variation of the retention of acids and bases with the pH of the mobile phase at several mobile-phase compositions and favorably compared with the relationships obtained with the common wwpH scale (pH measured in the aqueous buffer before mixing it with the organic modifier). The use of the rigorous sspH and wspH scales allows one to explain the retention behavior of bases, which in many instances cannot be justified from the pH measurement in the ill-founded wwpH scale.     

Mechanistic aspects of chiral discrimination on modified cellulose

Cellulose and cellulose derivatives are biopolymers which are often used as stationary phases for the separation of enantiomers. Describing the mechanism of such separations is a difficult task due to the complexity of these phases. In the present study, we attempt to elucidate the types of interactions occurring between a diol intermediate for a LTD(4) antagonist and a tris(4-methylbenzoate)-derivatized cellulose stationary phase. Thermodynamic studies indicate that, at low temperatures, the enantioselectivity is entropy driven. At higher temperatures, the separation is enthalpy driven. DSC and IR experiments reveal that the transitions between the enthalpic and the entropic regions of the van't Hoff plots are a result of a change in conformation of the stationary phase. Investigation of chromatographic kinetic parameters reveals that, at low temperature, the second eluted enantiomer undergoes sluggish inclusion interactions. Subtle changes in the structure of the analyte indicates that π-π interactions do not contribute to enantioselectivity. Finally, molecular modeling of (R)- and (S)-diol and the stationary phase suggests that hydrogen bonding is a primary factor in the separation, and the calculated energy values obtained from the molecular modeling correlate well with the chromatographic elution order.     

Characterization and thermodynamic studies of the interactions of two chiral polymeric surfactants with model substances: phenylthiohydantoin amino acids.

Analytical ultracentrifugation is used for determination of the molecular weights and the sedimentation coefficients of poly(sodium undecanoyl-L-valinate) (PSUV) and poly(sodium undecanoyl-L-threoninate) (PSUT) at different temperatures. Plots of absorbance as a function of radius indicates that both PSUV and PSUT are highly monodispersed. A method for evaluating the partial specific volumes using density measurements is presented. The partial specific volumes of PSUV are slightly higher than those of PSUT. In addition, the temperature dependence of the retention factor in electrokinetic chromatography was used to estimate the enthalpy, the entropy, and the Gibbs free energy of the surfactant/analyte complexes. Five phenylthiohydantoin-DL-amino acids were separated and each enantiomeric pair was completely resolved. Comparison of the thermodynamic values obtained with PSUV vs PSUT using a van't Hoff relationship suggests that PSUT, with a less favorable free energy change (i.e., less negative delta (delta G)), generates a more positive entropy change, hence slightly less chiral resolution.     

Supercritical fluid generated stationary phases for liquid chromatography and capillary electrochromatography

Chromatographic silica-bonded stationary phases have been prepared using supercritical CO(2) as the reaction medium. (29)Si solid-state NMR spectra of the generated bonded silica phases show unreacted silica species Q(3) and Q(4), alongside important resonances for surface-bound ligands, T(1), T(2), and T(3). Initially, a fluorinated octyl silica (C(8)) phase was produced, by reacting (1)H,(1)H,(2)H,(2)H-perfluorooctyltriethoxysilane with silica particles (3 microm) in sc-CO(2) at 60 degrees C and 450 atm for 3 h. In-house-packed LC columns of this fluorinated sc-C(8) silica phase yielded typical reversed-phase behavior when a standard test mix of benzamide (k' = 1.03), benzophenone (k' = 8.11), and biphenyl (k' = 14.92) was eluted. When packed into fused-silica capillaries for CEC, this fluorinated sc-C(8) silica phase gave linear plots of log k' versus percentage MeOH for benzophenone and biphenyl and, in contrast to octyl or octadecyl silica phases, displayed selectivity for aromatic thioureas when chromatographed among a series of synthetic organic thiourea test solutes. Similarily, an octadecyl silica phase (sc-C(18) silica) was prepared by reaction of n-octadecyltriethoxysilane in sc-CO(2), packed at 9500 psi and examined by LC. The sc-C(18) silica LC column yielded high column efficiency (up to 141 000 N/m (fluorene)) and excellent asymmetry factors (1.06, fluorene) without resource to end-capping. Following a second silylating or end-capping step using hexamethyldisilazane in sc-CO(2), sc-end-capped sc-C(18) silica phases elute N,N-DMA before toluene and the toluidine isomers as a single peak, indicating lowered silanol activity according to the Engelhardt test. A rapid separation of the important pharmaceutical substances, ketoprofen, naproxen, fentoprofen, and ibuprofen, on an sc-end-capped sc-C(18) silica phase is also shown.     

Theory for electrostatic interaction chromatography of proteins.

A simple theoretical framework for the effect of the eluting salt concentration on the retention factor of proteins in ion-exchange chromatography under conditions of linear elution is described. It is based on the solution of the linearized Poisson-Boltzmann equation for two oppositely charged planar surfaces in contact with a salt solution. The theory predicts a linear relation between the logarithmic retention factor and the reciprocal square root of the ionic strength of the eluent in the salt concentration range used in linear elution chromatography. A large body of retention data obtained in ion-exchange chromatography of proteins over a wide range of experimental conditions was plotted as ln k' vs 1/square root of I, where k' and I are the retention factor and ionic strength, respectively. The plots are linear or nearly so, as predicted for a moderate salt concentration range by the theory. From the slope of such plots the characteristic charges of the proteins were estimated by using only fundamental physicochemical constants. The chromatographically measured protein charges compare well to those obtained from titrimetric experiments at the same pH, although certain deviations are noted. The theoretical approach presented here offers a more realistic treatment of the ion-exchange chromatography of proteins than the stoichiometric displacement model and can serve as a convenient framework for the analysis of retention data.     

Exploring the possibilities of cryogenic cooling in liquid chromatography for biological applications: a proof of principle

The possibilities to use cryogenic cooling to trap components in liquid chromatography was investigated. In a first step, van 't Hoff plots were measured with a reversed-phase column using the temperature control unit of a conventional high performance liquid chromatography (HPLC) system to gain insight in the retention behavior of proteins at low temperatures. It was estimated that retention factors in the range of k = 10(4) could be achieved at T = -20 °C for lysozyme, indicating that temperature is a usable parameter to trap components in LC. In a next step, trapping experiments were carried out on a nano-LC system, equipped with a UV-detector, using a commercial reversed-phase column. An in-house built setup, allowing cooling of a segment of the column down to temperatures below T = -20 °C, was used to trap components. Experiments were conducted under isocratic and gradient conditions with methanol as organic solvent. It is demonstrated that, by thermally trapping and elution of components, an enhanced S/N ratio and decreased peak widths can be obtained. At the same time, a significant increase in pressure drop occurs during the cooling process. Limitations and benefits of the technique are further discussed.     

Geometrical model for the retention of fullerenes in high-performance liquid chromatography

In high-performance liquid chromatography (HPLC) using a poly(octadecylsiloxane) as a stationary phase, methanol as a mobile phase, C(60) and C(70) fullerenes as solutes, and water as a mobile phase modifier, a study on the surface tension effect of water on fullerene retention was carried out by varying the water concentration [W] and the column temperature T. The thermodynamic parameters for fullerene transfer from the mobile to the stationary phase were determined from linear van't Hoff plots. An enthalpy-entropy compensation revealed that the types of interactions between fullerenes and the stationary phase were independent of the fullerene structure and the mobile phase composition. An analysis of the experimental variations of the retention factor and the selectivity values with [W] was performed using a novel geometrical model. It was shown that the increase in fullerene retention accompanying the water concentration was due to the increased effects of surface tension. This brought about an increase in the interactions between fullerene and the stationary phase, explaining the observed thermodynamic parameter trends over the water concentration range. The theoretical model provided an estimation of the radius of fullerene which was found for C(60) to be equal to 3.3 ? and an activation energy during the transfer equal to 9.8 kJ/mol.     

Thermal unfolding of proteins studied by coupled reversed-phase HPLC-electrospray ionization mass spectrometry techniques based on isotope exchange effects

In this study, a deuterium exchange procedure has been employed to evaluate the thermal stability of globular proteins under conditions that replicate their interactive behavior in reversed-phase high performance chromatographic (RP-HPLC) systems. In particular, this investigation has permitted the conformational stability of two proteins, hen egg white lysozyme (HEWL) and horse heart myoglobin (HMYO) to be examined under different temperature and low-pH solvent regimes. The results confirm that this experimental approach provides an efficient strategy to explore fundamental conformational features of polypeptides or proteins in their folded and partial unfolded states under these interactive conditions. In particular, this analytical procedure permits insight to be readily gained into the processes that occur when polypeptides and globular proteins interact with lipophilic liquid/ solid interfaces in the presence of water-organic solvent mixtures at different temperatures.     

Diverging Thermodynamic Derivatives Associated with Heterogeneous Chemical Equilibrium in a Binary Liquid Mixture with a Consolute Point

The solubilities of tin(II) oxide, copper(II) oxide, and cobalt(II) oxide have been determined in the liquid mixture, isobutyric acid + water, along the critical isopleth. When plotted in van’t Hoff form with \(\ln s\) versus \(1/T\) , the solubility measurements, \(s\) , lie on a straight line for values of the temperature, \(T\) , which are sufficiently in excess of the critical solution temperature, \(T_\mathrm{c}.\) In the case of SnO, the dissolution reaction is exothermic, and the slope of the van’t Hoff plot diverges toward positive infinity as \(T\rightarrow T_\mathrm{c} .\) In the case of both CuO and CoO, the dissolution reaction is endothermic, and the slope of the van’t Hoff plot diverges toward negative infinity as \(T\rightarrow T_\mathrm{c} .\) Analysis of these ternary, heterogeneous equilibria using finite dimensional vector space stoichiometry theory shows that each contains two linearly independent components. According to the Gibbs phase rule, two-phase equilibria of this type can be described by two fixed, intensive variables, which are accounted for by the temperature and the pressure, respectively. The Gibbs–Helmholtz equation and the principle of critical-point universality can be combined to predict under conditions of fixed temperature and pressure that when dissolution is exothermic, \((\partial \ln s/\partial (1/T))\) should diverge toward positive infinity in the critical region, while when dissolution is endothermic, \((\partial \ln s/\partial (1/T))\) should diverge toward negative infinity. Our experiments include examples confirming both these predictions.     

Supramolecular method for the determination of absolute configuration of chiral compounds: theoretical derivatization and a demonstration for a phenolic crown ether-2-amino-1-ethanol system

A combination of anisotropic shielding effect and temperature dependence of NMR chemical shifts makes it possible to develop a supramolecular method for determination of absolute configuration of chiral compounds. The concept, theoretical derivatization, and first demonstration of this daedal noncovalent method using a host-guest system is described. This noncovalent method requires only three substrate solutions and can in principle be applied to any host-guest systems that follow the van't Hoff relation and have a clear anisotropic shielding effect based on a well-defined complex structure regardless of the extent of enantiomer selectivity.     

Fluorous affinity chromatography for enrichment and determination of perfluoroalkyl substances

The adsorption behavior of four perfluoroalkyl acids, including the environmentally relevant perfluorooctanoic acid, has been investigated on a straight-chain perfluorohexyl adsorbent. The aim of this study was to probe the potential of perfluorinated materials for the analysis and enrichment of perfluoroalkyl analytes. Water/acetonitrile mixtures, to which formic acid had been added (generally, 0.1%), were employed as mobile phases. For all perfluorinated acids, a U-shaped retention profile was observed by changing the amount of acetonitrile in the mobile phase. This behavior has been correlated to the excess adsorption of the organic component on the adsorbent surface. The concept of perfluoromethylene selectivity has been defined as the ability of a chromatographic system to discriminate between molecules that differ by a single perfluoromethylene group. The contribution to the Gibbs free energy of phase transfer for the passage of a perfluoroalkyl carbon from the mobile to the stationary phase has been evaluated. This information, in addition to the traditional van't Hoff analysis, has also been used to estimate the analogous contribution for the transfer of a carboxylic unit. Finally, insights into the retention mechanisms of perfluoroalkyl acids on straight-chain perfluorohexyl sorbents are discussed.     

Phase shifts that accompany total internal reflection at a dielectric-dielectric interface

The absolute, average, and differential phase shifts that p- and s-polarized light experience in total internal reflection (TIR) at the planar interface between two transparent media are considered as functions of the angle of incidence phi. Special angles at which quarter-wave phase shifts are achieved are determined as functions of the relative refractive index N. When the average phase shift equals pi/2, the differential reflection phase shift delta is maximum, and the reflection Jones matrix assumes a simple form. For N > square root 3, the average and differential phase shifts are equal (hence deltap = 3 deltas) at a certain angle phi that is determined as a function of N. All phase shifts rise with infinite slope at the critical angle. The limiting slope of the delta-versus-phi curve at grazing incidence (partial partial differential delta/partial partial differential phi)phi=90 degrees = -(2/N)(N2 - 1)1/2 = -2 cos phic, where phic is the critical angle and (partial partial differential2 delta/partial partial differential phi 2)phi=90 degrees = 0. Therefore delta is proportional to the grazing incidence angle theta = 90 degrees - phi (for small theta) with a slope that depends on N. The largest separation between the angle of maximum delta and the critical angle is 9.88 degrees and occurs when N = 1.55377. Finally, several techniques are presented for determining the relative refractive index N by using TIR ellipsometry.     

Viscosities of nonelectrolyte liquid mixtures. I. binary mixtures containing p-dioxane

Viscosity measurements are reported for p-dioxans with cyclohexane, n-hexane, benzene, toluene, carbon tetrachloride, tetrachloroethane, chloroform, pentachloroethane, and ethyl acetate at 303.15 K. Excess Gibbs energies of activation G ^*E of viscous flow have been calculated with Eyring's theory of absolute reaction rates. The deviations of the viscosities from a linear dependence on the mole fraction and values of G ^*E for binary mixtures have been explained in terms of molecular interactions between unlike pairs. The Prigogine-Flory-Patterson theory has been used to estimate the excess viscosity, ln , and corresponding enthalpy ln _H, entropy ln _S, and free volume ln _v terms for binary mixtures of p-dioxane with cyclohexane, n-hexane, benzene, toluene, carbon tetrachloride, and chloroform. Estimates of excess viscosities from this theory for p-dioxane with benzene, toluene, and carbon tetrachloride are good, while for the other three mixtures they are poor. The local-composition thermodynamic model of Wei and Rowley estimates the excess viscosity quite well even for p-dioxane mixtures with cyclohexane and n-hexane.