Effect of Temperature on the Chromatographic Behavior of Epirubicin and its Analogues on High Purity Silica Using Reversed-Phase Solvents

The influence of temperature on the retention behavior of epirubicin and its analogues on high purity silica with reversed-phase solvents has been systematically investigated. It was found that temperature effects on retention are highly dependent on the type and concentration of organic modifier, as well as the pH of the mobile phase. In organic-rich mobile phases, the type of organic modifier plays an important role. With an aprotic solvent as modifier, retention times show anomalous increases with elevated temperature. At the same time, both efficiency and resolution are significantly improved but this is not the situation with a protic solvent as modifier. In addition, temperature shows different effects on retention time and selectivity when the pH is changed, and temperature-dependent selectivity reversal is found at higher pHs. In aqueous-rich mobile phases, regardless of the nature of the organic solvent and pH, retention of solutes drops as temperature is raised. It seems that the effect of temperature on chromatographic behavior of the solutes on bare silica using mobile phases containing various organic modifiers or pHs, results from a number of different retention mechanisms.     

A fundamental study of the effects of modifiers in supercritical fluid chromatography

The effects of organic modifiers on retention and peak shape in packed-column supercritical fluid chromatography were studied. The adsorption behavior of different modifiers was investigated on stationary phases consisting of hydrocarbon monolayers chemically bonded on silica. Adsorption isotherms for several modifiers were recorded using breakthrough measurements. The results were compared with those obtained by a simple method based on the injection of different sample sizes. Modifiers were selected to reflect various types of interactions with the silica support. Isotherms were found to be approximately Langmuirean. Experiments were performed to elucidate the influence of adsorbed molecules on the retention of selected test solutes. The introduction of a solvent modifier can lead to a substantial change in the mobile phase density, the effect of which is comparable with that of an increase in the density of pure carbon dioxide. Our results confirm that the effects of low concentrations of modifiers (between 0 and 2 percent) in packed-column SFC are largely due to deactivation of residual silanol groups on the silica support. The accessibility of the active sites was found to depend strongly on the size and structure of the modifier molecules. The decrease of retention due to the addition of low concentrations of modifiers could be described accurately by a model derived from Langmuir adsorption behavior. Some semi-quantitative rules for the selection of modifiers and the required concentrations for optimum deactivation of the support are discussed.     

Influence of preferential adsorption of mobile phase on retention behavior of amino acids on the teicoplanin chiral selector

The adsorption behavior of two amino acids, i.e., l,d-threonine and l,d-methionine has been investigated on the chiral stationary phase (CSP)column packed with teicoplanin bonded to a silica support. The study has been performed under non-linear conditions of adsorption isotherm for various types of organic modifiers (methanol, ethanol, propan-2-ol and acetonitrile) in the reversed-phase mode. A heterogeneous adsorption mechanism of amino acids has been identified that was strongly affected by the nature of organic modifier. Generally, isotherm non-linearity and retention decreased with decrease of the modifier content in the mobile phase exhibiting a minimum at water-rich mobile phases. These trends were suggested to result from a combined effect of the mobile as well as the adsorbed phase composition. To determine the composition of the adsorbed phase the excess adsorption of modifiers in aqueous solutions has been measured and their binary adsorption equilibria have been quantified and compared. Strongly non-ideal behavior of solvents in the mobile phase and the adsorbed phase has been accounted for by activity coefficients. The fraction of the modifiers in the adsorbed phase decreased in the sequence: methanol, ethanol, propan-2-ol and acetonitrile.     

Unusual Retention Behavior of Some Cationic-Type Aldoximes Used as AChE Reactivators Under Ion-Pairing Liquid Chromatographic Mechanism

Retention of some cationic-type aldoximes used as AChE reactivators was studied under ion-pairing (IP) liquid chromatographic conditions, using alkyl sulphonates as IP agents. An unusual chromatographic retention behavior was observed: the functional dependencies between the logarithm of the capacity factor and the methanol content in the mobile phase followed a binomial pattern (U-shaped), with a minimum positioned within the interval 50–80% methanol. The tautomerism of the investigated structures and the stationary phase morphological modification caused by the adsorption of the ion-pairing agent may explain the experimental findings. The study focused on the influences brought by the pH, the alkyl chain length of the ion-pair agent, and the organic modifiers on the retention behavior of these compounds.     

Separation of imidacloprid and its degradation products using reversed phase liquid chromatography with water rich mobile phases

Water rich mobile phases in RPLC are not generally used because of the longer retention times involved when organic modifiers such as methanol or acetonitrile are used. The problem of longer retention times can be addressed using hydrophobic alcohols such as pentanol in low quantities (less than 1%) as organic modifiers. The advantages of using these mobile phases in RPLC for the separation of water soluble and weakly retained congeners is demonstrated through the separation of imidacloprid and its degradation products using a 0.4% pentanol in water mobile phase with 0.2% acetic acid.     

Packed column supercritical fluid chromatography with light-scattering detection. II. Retention behaviour of squalane and glucose with mixed mobile phases

Summary Evaporative light scattering detectors can be used to detect organic substances without chromophoric groups in packed column supercritical fluid chromatography (SFC). A detector of this type has been used to detect squalane and glucose after SFC with various packed columns and binary mobile phases. In this study, the amount of organic modifier in carbon dioxide/modifier mixtures was varied. The results give further insight into the mechanisms that influence retention behaviour in packed column separations with super- and subcritical mobile phases. Squalane is an ideal non-polar test solute which shows long retention times on non-polar columns while its elution can be accelerated by non-polar modifiers in carbon dioxide. Glucose is an extremely polar solute containing hydroxyl groups. Elution of this sugar can be improved with polar modifiers. Column packings with polar end groups lead to high capacity ratios and long retention times for glucose. Most columns used in this study contained silica-based packing materials. For purposes of comparison, a polymeric packing (HEMA RP-18) was also employed.     

几种手性化合物在直链淀粉-三(环己基氨基甲酸酯)手性固定相上高压液相中的拆分

正相条件下, 在自制的涂敷型直链淀粉-三(环己基氨基甲酸酯)手性固定相上, 直接拆分了九种不同类型的手性化合物, 考察了流动相中极性添加剂对手性拆分的影响, 并与直链淀粉-三(3,5-二甲基苯基氨基甲酸酯)进行了比较. 研究发现, 此种手性固定相不仅具有较高的手性识别能力, 而且对某些类型手性化合物的选择性与后者明显不同; 探讨了此种固定相可能的拆分机理.     

Effect of class I and II organic modifiers on retention and selectivity in vesicle electrokinetic chromatography

Vesicles are large aggregates of surfactant monomers consisting of a spherical bilayer surrounding an internal cavity of solvent. The bilayer structure allows vesicles to be attractive models for the study of various transmembrane and binding processes. The use of thermodynamically stable vesicles (TSV) formed from oppositely charged surfactants for use as a pseudostationary phase in electrokinetic chromatography (EKC) was first accomplished using dodecyltrimethylammonium bromide and sodium dodecyl sulfate (DTAB/SDS). Surfactant vesicles have demonstrated enhanced separation characteristics compared to conventional micelles in EKC, although only investigated in aqueous media. Organic modifiers have been widely studied and used in EKC to enhance separation conditions. In this study, vesicles formed from cetyltrimethylammonium bromide and sodium octyl sulfate (CTAB/SOS) were investigated in the presence of "class I and II" organic modifiers. Electrophoretic and chromatographic parameters were examined as well as linear solvation energy relationship analysis (LSER) to characterize the effects of the modifiers on retention and selectivity in EKC. LSER analysis is a useful way to quantitatively investigate solute/solvent interactions responsible for retention and selectivity.     

Thermodynamic and kinetic study of chiral separations of coumarin-based anticoagulants on derivatized amylose stationary phase

Thermodynamic and kinetic studies are performed on amylose derivatized with tris-(3,5-dimethylphenyl carbamate) stationary phase for the chiral separation of coumarin-based anticoagulants. Polar-organic eluents that contain acetonitrile as bulk solvent with modifiers such as methanol, i-butanol, t-butanol, and tetrahydrofuran are used in the study. Temperature is varied from 5 to 45 °C at constant pressure of 1500 psi. In general, both retention and enantioselectivity decrease as the temperature increases and as hydrogen bond donating ability of the modifiers increases. The van’t Hoff plots are found to show both linear and non-linear behavior. The non-linear plots are believed to be the result of conformational changes in the derivatized amylose phase and are observed around room temperature. The retention behavior in acetonitrile mobile phase provides a linear enthalpy–entropy compensation plot, indicating that all coumarins may have a similar retention mechanism. In contrast, enthalpy–entropy compensation is not observed for warfarin and coumatetralyl enantiomers when separated with different organic modifiers in the mobile phase. The kinetic data indicate that the rate of sorption is always greater than the rate of desorption. An increase in the concentration of alcohol modifiers causes an increase in the desorption rate constant. In contrast, an increase in the concentration of tetrahydrofuran causes a decrease in the desorption rate constant. This effect is most significant for the second eluted enantiomer of coumatetralyl, for which the desorption rate is 36 times slower than the first eluted enantiomer.     

Effect of the organic modifier concentration on the retention in reversed-phase liquid chromatography II. Tests using various simplified models

Ten simplified expressions for the retention factor, k', that arise from either the adsorption or partition mechanism for retention in reversed-phase chromatographic columns are examined in what concerns the model they express and their performance to fit experimental data. In order to test the simplified expressions, which describe the variation of the retention of a solute with the organic modifier content in the mobile phase, a wide range of solutes in mobile phases modified with three different organic modifiers was used. It is shown that a new three-parameter expression of ln k' works more satisfactorily, since it combines simplicity, high applicability and good numerical behavior. It is also shown that the applicability of a simplified equation does not entail the validity of its model and thus no molecular information can be gained from its use.     

Effects of stationary-phase polarity on retention in reversed bonded phase HPLC columns

Summary Variations in retention and selectivity have been studied in cyano, phenyl and octyl reversed bonded phase HPLC columns. The retention of toluene, phenol, aniline and nitrobenzene in these columns has been measured using binary mixtures of water and methanol, acetonitrile or tetrahydrofuran mobile phases in order to determine the relative contributions of proton donor-proton acceptor and dipole-dipole interactions in the retention process. Retention and selectivity in these columns was correlated with polar group selectivities of mobile phase organic modifiers and the polarity of the bonded stationary phases. In spite of the prominent role of bonded phase volume and residual silanols in the retention process, each column exhibited some unique selectivities when used with different organic modifiers.     

Effect of the eluent pH and acidic modifiers in high-performance liquid chromatography retention of basic analytes

The retention of ionogenic bases in liquid chromatography is strongly dependent upon the pH of the mobile phase. Chromatographic behavior of a series of substituted aniline and pyridine basic compounds has been studied on C18 bonded silica using acetonitrile-water (10:90) as the eluent with different pHs and at various concentrations of the acidic modifier counter anions. The effect of different acidic modifiers on solute retention over a pH range from 1.3 to 8.6 was studied. Ionized basic compounds showed increased retention with a decrease of the mobile phase pH. This increase in retention was attributed to the interaction with counter anions of the acidic modifiers. The increase in retention is dependent on the nature of the counter anion and its concentration in the mobile phase. It was shown that altering the concentration of counter anion of the acidic modifier allows the optimization of the selectivity between basic compounds as well as for neutral and acidic compounds.     

Selectivity patterns in micellar electrokinetic chromatography: Characterization of fluorinated and aliphatic alcohol modifiers by micellar selectivity triangle

The usefulness of the micellar selectivity triangle (MST) for prediction and interpretation of separation patterns in micellar electrokinetic chromatography (MEKC) separations is presented. In addition, we demonstrate the capability of controlling selectivity properties of micelles through addition of organic modifiers with known solvation properties as predicted by MST. The examples are modification of the hydrogen bond donor (HBD) micelle of lithium perfluorooctanesulfonate, the hydrogen bond acceptor (HBA) micelle of tetradecyltrimethylammonium bromide, and the sodium dodecyl sulfate micelles with intermediate hydrogen bonding properties with two hydrophobic organic modifiers. One is an aliphatic alcohol, n-pentanol that can act as both a HBA and a HBD; by contrast, the other organic modifier is a fluorinated alcohol, hexafluoroisopropanol that is a strong HBD modifier and would enhance the hydrogen bond donor strength of micelles. A test sample composed of 20 small organic solutes representing HBA, HBD, and non-hydrogen bond aromatic compounds was carefully selected. The trends in retention behavior of these compounds in different micelles are consistent with the selectivity patterns predicted by the MST scheme. To the best of our knowledge, this is the first report on the unique selectivity of fluorinated alcohols as modifiers in MEKC. These results demonstrate the usefulness of the MST scheme for identifying pseudo-phases with highly similar or different selectivities and can serve as a guide for judicious selection of modifiers to create pseudo-phases with desired selectivity behavior on a rational basis.     

Retention prediction in ternary solvent reversed-phase liquid chromatography systems based on the variation of retention with binary mobile phase composition

An extension of the treatment adopted in a recent paper [P. Nikitas, A. Pappa-Louisi, P. Agrafiotou, J. Chromatogr. A 946 (2002) 33] was used to derive expressions describing the variation of solute retention k with composition in ternary reversed phase liquid chromatography, RP-LC, solvent systems. The equation of the partition model obtained in this way for a ternary mobile phase was identical to that previously derived using the solubility parameter concept. This equation as well as two new expressions of In k versus organic modifiers content were tested in a variety of ternary solvent systems in order to examine the possibility of predicting retention behavior of solutes under ternary solvent mixture elution conditions from known retention characteristics in binary mobile phases. It was demonstrated the superiority of both new equations derived in this paper to that previously proposed and applied to date in ternary solvent mixtures.     

An unexpected observation concerning the effect of anionic additives on the retention behavior of basic drugs and peptides in reversed-phase liquid chromatography

Anionic species with ion pair forming ability are commonly used to enhance the retention and efficiency of basic analytes in RPLC separations. However, little is known about the interactions between organic mobile phase modifiers and such ion pairing anions. In this work, we measured the magnitude of the retention increase of basic drugs and peptides upon addition of strong inorganic ion pairing anions (e.g. perchlorate) as a function of the volume fraction of modifier in acidic water-acetonitrile mobile phases on two different stationary phases. We found that the increase in retention upon addition of various salts depended strongly on the eluent strength. In general, larger retention increases upon addition of the anion were observed at higher organic fractions. Regression of retention against the volume fraction of organic modifier indicated that the ion pair forming anions substantially decreased S values while only slightly changing ln k'w values. The decrease in S is the major cause of the retention increase of basic drugs and peptides when such anions are added to the mobile phase.     

Effects of organic modifiers on solute retention and electrokinetic migrations in micellar electrokinetic capillary chromatography.

Influences of seven organic modifiers, including urea, methanol (MeOH), dioxane (DIO), tetrahydrofuran (THF), acetonitrile (ACN), 1-propanol (1-PrOH) and 2-propanol (2-PrOH), on the solute retention and the electrokinetic migrations in micellar electrokinetic capillary chromatography (MEKC) are investigated with sodium dodecyl sulfate (SDS) micelle as pseudostationary phase. It is observed that in the limited concentration ranges used in the MEKC systems the effect of organic modifier concentration on the retention can be described by the equation logk1=logk1w-SC for most binary aqueous-organic buffer, but deviations from this retention equation are observed at ACN and particularly THF as organic modifiers. With parameter S as a measure of the elutropic strength, the elutropic strength of the organic modifiers is found to follow a general order urea 相似文献   

Ion-exchange properties of silica gel with covalently bonded histidine

The ion-exchange and acid-base properties of Silasorb 600 (10 μm) with immobilized histidine (His-SiO₂) were investigated. The acid-base properties of the sorbent were estimated from the dependence sof the retention of organic acids and bases on the eluent pH. The effect of various organic modifiers (benzoic, citric, oxalic, and sulfuric acids) and the eluent pH on the retention of inorganic ions was studied. It was shown that His-SiO₂ exhibits anion-exchange properties in acidic and weakly acidic media at pH < 5.7. Eight anions were separated under optimal conditions on a 150 × 4.6-mm column in 21 min using a 5 mM oxialic acid solution as an eluent.     

Mixture-designed electrolytes for the MEKC separation of natural and synthetic steroids

In this work, the separation of 11 natural and synthetic steroids was studied using MEKC electrolytes modified by property-selected organic solvents: ethanol, ACN, and THF. The interplay between electrophoretic behavior and structural features was disclosed and the effects of organic modifiers to modulate retention and to alter selectivity were discussed in terms of system linear solvation energy relationships (LSER). The LSER indicated the total organic solvent percentage in the electrolyte as a major parameter to control retention and as a minor contribution, the hydrogen bond acidity. By evaluating the electropherograms obtained from mixture-designed electrolytes, a favorable separation condition for all solutes was achieved in ca. 25 min with an electrolyte composed of 20 mmol/L sodium tetraborate at pH 9.4, 20 mmol/L SDS and 20% EtOH (0.80% CV for migration time and 2.5% CV for peak area, n = five consecutive injections). The applicability of the proposed separation condition was demonstrated by the inspection of estrogens in urine sample (puberty stage).