Prediction of retention time of phenols in liquid chromatography

The chromatographic behavior of phenols in reversed-phase mode liquid chromatography differs from that of non-ionic compounds such as alkyl alcohols, alkylbenzenes, halogenated benzenes, polyaromatic hydrocarbons, and aromatic acids. Therefore, the retention times of 61 phenols were measured in a system of an octadecyl bonded silica gel and acetonitrile/water mixtures. The logarithm of the capacity ratio (log k') was found to be a linear function of the hydrophobicity (log P) in acidic acetonitrile/water mixtures. This result was applied to a different octadecyl bonded silica gel. Eight phenols were selected as standard compounds, and their log k' values were measured in 0.05 M phosphoric acid in 10 to 90% acetonitrile/water mixtures. An empirical polynomial relation was obtained between the concentration of acetonitrile and the slope of the log k' vs log P curve. Finally the capacity ratio of all phenols were calculated in given eluents by the equations derived from the measurements of standard compounds and the calculated log P values. The difference between predicted capacity ratios and measured ones was within 10%.     

Liquid Chromatographic Behavior of Nitrogen Compounds

Abstract

Chromatographic behavior of nitrogen compounds differed from others. The column efficiency was poor for the compounds and sometimes solutes were not eluted out from a column. Therefore, the elution volume of alkylamines, anilines, pyridines, pyrazines, quinolines and aminopolyaromatic hydrocarbons was measured on a methacrylate gel and octadecyl bonded silica gels in pH controlled acetonitrile/water mixtures. The solvent effect on the dissociation constant differs from that obtained for aromatic acids. The values in acetonitrile/water mixtures are smaller than those obtained in 100% water. The linear relation between log P and log k′ values is obtained in eluents of pH 7 where the retention of these compounds is maximized. Some hydrophobic fragmental constants are proposed from this result. Prediction of retention time of these compounds from their log P values can be done in the individual groups on octadecyl bonded silica gels in pH controlled acetonitrile/water mixtures.     

Determination of lipophilicity of alpha-(4-phenylpiperazine) derivatives of N-benzylamides using chromatographic and computational methods

This paper describes the evaluation of lipophilicity of alpha-(4-phenylpiperazine) derivatives of N-benzylamides. We employed reversed-phase thin-layer chromatography (RP-TLC) and reversed-phase high performance liquid chromatography (RP-HPLC) as experimental methods, using mixtures of acetonitrile and water as the mobile phases with addition of 0.1%TFA in the HPLC experiments. Retention parameters (R(M)) and capacity factors (log k) determined by applying these methods were linearly dependent on the acetonitrile concentration and enabled us to estimate the relative lipophilicity factors: R(M0) and log k(0). These factors were compared with the calculated partition coefficients C log P obtained using several software packages. The results indicate that both experimental methods (RP-TLC and RP-HPLC) yielded similar results, and these methods enable determining the lipophilicity of alpha-(4-phenylpiperazine) derivatives of N-benzylamides. Significant correlations were found between log P values calculated by Pallas, ALOGPS and C log P Chem3D programs and the experimental data.     

The study of the lipophilicity of alpha-(4-phenylpiperazin-1-yl)-gamma-phthalimidobutyramides using chromatographic and computational methods

The lipophilicity of the series of alpha-(4-phenylpiperazin-1-yl)-gamma-phthalimido-butyramides (1-8) has been investigated. Several methods, like reversed-phase thin-layer chromatography and high-performance liquid chromatography using reversed-phase RP18 and IAM.DD2 columns, were applied to determine RMO, log k0 and log k(0IAM) factors. The RP-TLC investigations were performed in mixtures of acetone-water and acetonitrile-water. For RP-HPLC method mixtures of acetonitrile, water and 0.01% TFA were used as the mobile phases while for IAM.DD2 investigations mixtures of acetonitrile and water were applied. The partition coefficients of compounds (1-8) were also calculated with the Pallas and CAChe programs. All the obtained data, both from experimental methods and computational calculations, were compared and a suitable conclusion was reached.     

Evaluation of a Simple HPLC Correlation Method for the Estimation of the Octanol-Water Partition Coefficients of Organic Compounds

Abstract

A simple reverse-phase high performance liquid chromatographic method is evaluated for the estimation of octanol-water partition coefficients (log P) of organic compounds by correlation with their chromatographic capacity factors (k′). Using an unmodified commercial octadecylsilane column and a mobile phase consisting of methanol and an aqueous buffer, a linear relationship is established between the literature log P values of 68 compounds and the logarithms of their k′ values. For the determination of the partition coefficients of unknowns, one of two sets of standards is used to calibrate the system, the choice being dependent on the hydrogen-bonding character of the compounds being evaluated. The overall method is shown to be rapid and widely adaptable and to give log P data which are comparable to results obtained by classical or other correlation methods.     

The effect of stationary phase on lipophilicity determination of beta-blockers using reverse-phase chromatographic systems

Evaluation of lipophilicity parameters for basic compounds using different chromatographic stationary phases is presented. An HPLC method for determination of lipophilic molecule-stationary phase interactions was based on gradient analysis. Differences in correlation between the lipophilicity of compounds and experimental chromatographic results obtained in pseudo-membrane systems showed a strong influence of stationary phase structure and physico-chemical properties. beta-Blocker drugs with varying lipophilicity and bio-activity were chosen as test compounds. The stationary phases used for the study were monolithic rod-structure C18 and silica gel octadecyl phase SG-C18 as reference material. The second group was silica gel-based polar-embedded alkylamide and cholesterolic phases. The mobile phase was composed of acetonitrile or methanol with ammonium acetate, and a linear gradient of methanol and acetonitrile in mobile phase was performed. A linear correlation of plots of log k(g) = f(log P) was observed, especially for polar-embedded phases, and this allowed log P(HPLC) to be calculated. The behavior of stationary phases in methanol and acetonitrile buffer showed differences between obtained log P(HPLC) values.     

Micellar liquid chromatography: suitable technique for screening analysis

The screening capability of micellar liquid chromatography (MLC) is discussed using the reported chromatographic data of several sets of compounds (amino acids, beta-blockers, diuretics, phenethylamines, phenols, polynuclear aromatic hydrocarbons, steroids and sulfonamides) and new results (sulfonamides and steroids). The chromatographic data are treated with an interpretive optimisation resolution procedure to obtain the best separation conditions. Usually, the pH and the concentration of surfactant (sodium dodecyl sulfate, SDS, or cetyltrimethylammonium bromide) for the optimal mobile phase were 2.5-3 and < 0.12 M, respectively. The nature and concentration of organic solvent depended on the polarity of the eluted compounds: a low volume fraction of propanol (approximately 1%, v/v) was useful to separate the amino acids, with log P(o/w) < -1 (where P(o/w) is the octanol-water partition coefficient). A greater concentration of this solvent (approximately 5-7%) was needed for compounds in the range -1 < log P(o/w) < 2, as with the studied diuretics and sulfonamides, and a high concentration of propanol (approximately 15%) or a low concentration of butanol (< 10%) had to be used for less polar compounds with 1 < log P(o/w) < 3, such as the beta-blockers. Pentanol (< 6%) was more suitable for the even less polar compounds with log P(o/w) > 3, such as the steroids. For basic drugs such as the phenethylamines (0 < log P(o/w) < 1.7), eluted with a micellar eluent of anionic SDS, propanol was too weak. A study is also shown for mixtures of sulfonamides (log P(o/w) = -1.2 to 1.7) and steroids (log P(o/w) = 3.0-8.1) eluted from conventional C18 columns with SDS mobile phases containing acetonitrile and 1-pentanol, respectively, which are compared with classical acetonitrile-water and methanol-water mixtures. The results complement a previous study on beta-blockers (log P(o/w) = -0.03 to 2.8) and reveal that MLC is a very competitive technique for the screening of compounds against conventional RPLC, due to its peculiar behaviour with regard to the selectivity and elution strength. The concentration of organic solvent needed to obtain sufficiently low retention times (even for highly hydrophobic steroids with log P(o/w) = 7-8) is also appreciably smaller for MLC, which reduces the environmental impact of the mobile phases.     

Determination of pKa values of nonsteroidal antiinflammatory drug‐oxicams by RP–HPLC and their analysis in pharmaceutical dosage forms

In this study, pK_a values were determined by using the dependence of the capacity factor on the pH of the mobile phase for four ionizable substances, namely, tenoxicam, piroxicam, meloxicam, and naproxen (I.S.). The effect of the mobile phase composition on the ionization constant was studied by measuring the pK_a at different ACN concentrations, ranging from 30 to 40%. The adequate condition for the chromatographic determination of these compounds in pharmaceutical dosage forms was established based on the different retention behaviors of the species. An octadecylsilica Nucleosil C18 column (150×4.6 mm, 5 μm) was used for all the determinations. The chromatographic separation of oxicams was carried out using acetonitrile (ACN)/water at 35% v/v, containing 65 mM phosphoric acid and UV detection at a wavelength of 355 nm. The method developed was successfully applied to the simultaneous determination of these drug compounds in laboratory‐prepared mixtures and their commercial pharmaceutical dosage forms. Each analysis requires no longer than 12 min.     

Liquid chromatographic retention and separation of phenols and related aromatic compounds on reversed phase columns

Summary Isocratic column liquid chromatographic systems with UV absorbance detection at 280 nm have been developed for the separation of 29 phenolics and related compounds.The selectivity was investigated on silica-, carbon- and polymer-based separation columns for the separation of phenolic type of components. The effects of various acetonitrile/buffer mixtures, and pH of the mobile phase, and their impact on the retention of the phenols was assessed. Tables of retention times on the four columns for the 29 phenols with two different acetonitrile/buffer mixtures, together with the retention times at three pHs from 6.5 to 2.3 with varying levels of organic modifier on the LiChrospher RP 18 column are presented.As an application, the analysis of real river water samples from the Ebro river is described using a solid phase extraction step prior to injection into the chromatographic system.     

Use of solid-phase microextraction for measuring oil-water partition coefficients and correlation with high-performance liquid chromatographic methods for lipophilicity

For flavour compounds, lipophilicity is often estimated by the partition coefficient between oil and water (log Koil-water), which is highly relevant to food. A modification of the shake-flask method is reported here where compounds are quantified in the two phases using solid-phase microextraction (SPME). SPME's highly sensitivity to non-polar compounds facilitates quantification in the water phase. Twelve flavour compounds representing a broad range of lipophilicities and functional groups were analysed by two methods. Their log Koil-water was determined using SPME quantitation and their log k(w) using a reversed-phase HPLC methodology. The isocratic capacity factor at 60% methanol and predicted log P value also showed high correlation factors with other methods. The octadecyl silylated surface of the HPLC column provides a matrix that interacts with lipophilic compounds where the retention time is the indication of lipophilicity. Both methods gave reproducible results (median 3% and 4% RSD) and similar but not identical values for lipophilicity. The relationship between the two methods is log k(w) =0.85 log Koil-water +0.48 with a correlation coefficient of 0.94. The new SPME detection method, with the ability to quantify limonene and 2-pentylfuran at 1 ppm in the water phase, is preferred for flavour compound analysis due to the applicability of oil-water partitioning in food.     

Relationships between LC retention, octanol-water partition coefficient, and fungistatic properties of 2-(2,4-dihydroxyphenyl)benzothiazoles

The retention behavior of newly synthesized compounds with antimycotic activity from the 2-(2,4-dihydroxyphenyl)benzothiazole group by high-performance liquid chromatography has been investigated. RP-18 stationary phase and methanol-acetate buffer aqueous mobile phases at pH 4 and 7.4 have been used. In the case of the mobile phase at pH 7.4, higher concentrations of water can be applied than at pH 4. The studied compounds showed regular retention behavior, their log k values decreasing linearly with an increasing concentration of methanol in the mobile phase. On the basis of these relationships, the lipophilicity (log kw), specific hydrophobic surface area (S), and isocratic chromatographic hydrophobicity index (psi0) were determined. Similar log kw values and sensitivity to changes in the structure of compounds studied for both mobile phases have been found. Moderate correlations between the chromatographic parameters and the calculated octanol-water log P values were found. Finally, the lipophilicity parameters were compared with the fungistatic properties of compounds expressed by log MIC (minimum inhibitory concentration) values to find quantitative structure activity relationship equations.     

Immobilized artificial membrane liquid chromatography: proposed guidelines for technical optimization of retention measurements

The objectives of this study were to establish guidelines for the proper measurement of capacity factors (log k(IAMw) on immobilized artificial membrane (IAM) stationary phases. In this context, some aspects related to the extrapolation of log(kIAMw) values, the stability and properties of IAM.PC.DD2 stationary phases and the column-to-column variability are discussed. No significant difference was observed when using either acetonitrile or methanol for the linear extrapolation of log k(IAM) values. However, methanol seems more appropriate when working with ionized compounds. Plotting isocratic capacity factors against the percentage (v/v) of co-solvent instead of the mole fraction leads to more reliable log k(AMW) values. Furthermore, our results with a YMC ODS-AQ and an IAM.PC.DD2 HPLC column indicate that only small differences arise between extrapolated capacity factors when using the (w(w))pH or the (s(w))pH operational scale and correcting or not the ionic strength for dilution caused by the co-solvent. The use of the (s(w))pH scale is recommended when working with ionized compounds in order to avoid parabolic relationships during linear extrapolation. The pH-dependent retention of three ionizable drugs on an IAM.PC.DD2 phase showed that secondary interactions with the charged moieties of the chromatographic surface affect the retention of ionized compounds around physiological pH. Finally, it was shown that column ageing occurs also with IAM.PC.DD2 stationary phases and that it depends on the column as well as on the investigated analyte. The intra-batch variability for IAM.PC.DD2 phases was small, whereas a marked and solute-dependent batch-to-batch variability was apparent.     

Separation and determination of biphenyl nitrile compounds by microemulsion electrokinetic chromatography with mixed surfactants

A mixture of six biphenyl nitrile compounds and three related substances with high hydrophobicity and similar structures was successfully separated by microemulsion electrokinetic chromatography (MEEKC) within 30 min. The microemulsion system contained 100 mM sodium dodecyl sulfate (SDS), 80 mM sodium cholate (SC), 0.81% v/v heptane, 7.5% v/v n-butanol, 10% v/v acetonitrile, and 10 mM borate. The addition of SC, organic modifiers, sample preparation, and temperature all showed remarkable effects on the separation. The capacity factor (k) was calculated by using dodecyl benzene as the marker for microemulsion, and the calculated partition coefficient log P(o/w) of the solutes was in the range of 3.35-7.38. The log k values matched well with the log P(o/w) with a correlation coefficient of 0.96. In addition, the linear correlation coefficients of each compound between peak area and concentration were from 0.996 to 0.998 with the repeatability RSD value < 1.2% for migration time and < 4.8% for peak area, and the highest theoretic plate number was > 586000. MEEKC was compared with micellar electrokinetic chromatography (MEKC) indicating that the former method is more suitable for this separation and can be used for the quality control of biphenyl nitrile compounds in the synthesis of liquid crystals.     

胆固醇基键合相-反相高效液相色谱法测定正辛醇/水分配系数

通过反相高效液相色谱法系统地考察了中性和弱酸性化合物在新型胆固醇基键合相色谱柱（Cholester柱）上的保留行为。以甲醇和乙腈为有机调节剂,建立了保留因子（k）与有机调节剂比例（φ）间的关系,并外推获取100%水相为流动相时的log k_w;同时进一步建立并验证了不同流动相下正辛醇/水分配系数的对数（log K_ow）和log k_φ（log k_w）间的模型,并预测了部分化合物的log K_ow。结果表明,使用Cholester柱测定log K_ow时,甲醇比乙腈更适合作为有机调节剂;对中性化合物和中性状态的酸性化合物,可以用任意甲醇比例下获取的log k_φ预测log K_ow;对存在离解的酸性化合物,依然用外推方式获取的log k_w预测log K_ow。将采用Cholester柱与文献中采用C₁₈柱、C₈柱建立的log K_ow-log k_φ模型进行对比,结果表明化合物与胆固醇基键合相存在特别的作用。     

Influence of solvent structure on the aquation of <Emphasis Type="BoldItalic">trans</Emphasis>-[Ru(3-MePy)<Subscript>4</Subscript>Cl<Subscript>2</Subscript>] complex in mixed aqueous solvents

The kinetics of the solvolytic aquation of trans-[Ru (3-Me Py)₄Cl₂] was studied spectrophotometrically in water – isopropanol in the range (30–90% v/v), and water acetonitrile in the range (10–70% v/v), and in the temperature range 50–65 °C. Plots of log k versus the reciprocal of the relative permittivity and Grunwald–Winstien gave non-linear plots. This non-linearity is derived from a large differential effect of solvent structure between the initial and transition states. The plot of log k versus water concentration was also non linear; evidence for the presence of a S _N ¹ mechanism. However, extrema in the variation of enthalpy ΔH* and entropy ΔS* of activation correlate well with the extrema in physical properties of the mixtures which are related to changes in solvent structure. Linear plots of ΔH* versus ΔS* were obtained and the iso- kinetic temperature indicates that the reaction is entropy controlled.     

HPTLC and magnetochromatography of new complexes of carboxylates with transition metals or rare earth elements and their ligands – study of lipophilicity

Nineteen new complexes of carboxylates with transition and rare elements as central ions and their ligands were characterized by chromatographic analyses. The parameter of relative lipophilicity (R _M0) of the tested compounds was determined experimentally by the reversed‐phase high‐performance thin layer chromatography method with mixtures of various organic modifiers (acetonitrile, acetone, dioxane) and water as a mobile phase. The extrapolated R _M0 values were compared with the logP values calculated from the molecular structures of tested solutes. Similarities between the lipophilicity indices were analysed by principal component analysis and linear regression. Thin‐layer chromatography combined with a magnetic field has been proposed as a complementary method for determination of lipophilicity of the investigated compounds. The chromatograms in the field and outside it were developed simultaneously in two identical chromatographic chambers. One of them was placed in the external magnetic field of 0.4 T inductivity. We proved that chelation causes a drastic change in compound lipophilicity, but all complexes did not exhibit enhanced activity as compared with the parent ligand. Also in the magnetic field the retention of some complexes changed, which means that the presence of the field influences the physicochemical properties of the compounds and their interactions with the stationary phase.