中性溶质在反相毛细管电色谱中的保留行为

分别在以甲醇、乙腈、异丙醇和四氢呋喃为有机改性剂的４种二元流动相体系中对中性溶质在反相毛细管电色谱中的保留行为进行了研究。不仅考察了有机改性剂的种类和浓度对电渗淌度的影响,而且建立了溶质容量因子与有机改性剂在流动相中体积分数间的定量关系,此外还对样品在反相毛细管电色谱和反相毛细管高效液相色谱中的保留行为进行了比较,发现中性溶质在这两种分离模式中的容量因子基本相同。     

Investigation of lipophilicity of anticancer-active thioquinoline derivatives

The lipophilicity of a series of anticancer propargylthioquinoline derivatives has been investigated using chromatographic and computational methods. The parameters of relative lipophilicity (R(MO) and logk0) of the tested compounds were determined experimentally both by reversed-phase thin layer (RP-TLC), and high-performance liquid chromatographic methods (RP-HPLC, LiChrospher RP18 column), with mixtures of acetonitrile and water as mobile phases. Their phospholipophilicity (logk(0IAM)) was determined using immobilized artificial membrane HPLC (IAM. PC DD2 Regis column). Mobile phase acetonitrile concentrations were in the ranges 50-90% (RP-TLC), 55-90% (RP-HPLC) and 35-60% (IAM-HPLC). The R(M), logk and logk(IAM) values of the compounds investigated were linearly dependent on acetonitrile concentration. The analysis led to the calculation of R(MO), logk0 and logk(0IAM) parameter values for each of the tested compounds. Their partition coefficients (logP) were also calculated with the Pallas and CAChe programs. The obtained results indicated that, among experimental methods, both RP-TLC and RP-HPLC gave similar results, and these methods enable the determination of lipophilicity of derivatives of thioquinolines. Using the IAM-HPLC technique a simple method of estimation of phospholipoplilicity was described. The CAChe program might better predict calculated lipophilicity logP values, and therefore is a useful tool for the early stage of design of new propargyl thioquinolines.     

Determination of lipophilicity for antitumor acridinone derivatives supported by gradient high-performance liquid chromatography method

The lipophilicity values of selected acridinone (imidazoacridinone and triazoloacridinone) derivatives were measured by gradient reversed-phase high-performance liquid chromatography (RP-HPLC) using a C18 stationary phase with a water/acetonitrile mixture as a mobile phase. The retention times obtained served as input data and appropriate log k_w values (i.e., the retention factor log k_w extrapolated to 0% organic modifier) as an alternative to log P were calculated using the DryLab program. The relationships between the lipophilicity (log k_w) and the chemical structure of the studied compounds, as well as correlation between experimentally determined lipophilicities (log k_w) and log P data calculated using some commonly available software, are discussed.     

A new type of chemically bonded phase for reversed-phase HPLC.

A new type of ether-bonded packing for reversed-phase HPLC (RP-HPLC) was synthesized by reacting 1-octanol with beta-(3,4-epoxycyclohexyl)ethyltrimethoxysilane, followed by coupling the product onto porous silica. The prepared packing was characterized by elemental analysis, solid-state 13C NMR, and Fourier transform infrared (FT-IR) spectroscopy. Chromatographic evaluations were performed by using a mixture of organic compounds as the analyte and methanol-water as binary mobile phase. The influence of the composition of organic modifier on the retention behavior of basic compounds was studied. The hydrolytic stability of the packing between pH 2.5-7.5 was also investigated. The results showed that the new stationary phase has excellent chromatographic properties and good hydrolytic stability.     

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.     

Retention process in reversed phase TLC systems with polar bonded stationary phases

The retention of a solute in RP chromatography is a very complex process which depends on many factors. Therefore, the study of the influence of a mobile phase modifier concentration on the retention in different reversed phase chromatographic systems is very important for understanding the rules governing retention and mechanisms of substance separation in a chromatographic process. Composition changes and the nature of mobile phases enable tuning of the separated analytes' retention over a wide range of retention parameters and optimization of the chromatographic process as well. Optimization of the chromatographic process can be achieved by several different methods; one of them is the so-called interpretative strategy. The key approach adopted in this strategy is the implementation of adequate retention models that couple the retention of solute with the composition of a mixed mobile phase. The use of chemically bonded stationary phases composed of partially non-bonded silica matrix and organic ligands bonded to its surface in everyday chromatography practice leads to questions of the correct definition of the retention model and the dominant retention mechanism in such chromatographic systems. The retention model for an accurate prediction of retention factor as a function of modifier concentration and the heterogeneity of the adsorbent surface should be taken into consideration. In this work the influence of mobile-phase composition on the retention of sixteen model substances such as phenols, quinolines, and anilines used as test analytes in different RP-TLC systems with CN-, NH2-, and Diol-silica polar bonded stationary phases has been studied. The aim of this study is to compare the performance of three valuable retention models assumed as the partition, adsorption/partition, and adsorption mechanism of retention. All the models were verified for different RP-TLC systems by three statistical criteria. The results of investigations presented in this work demonstrate that the best agreement between the experimental and calculated Rf values was obtained by the use of new-generation retention models, which assume heterogeneity of adsorbent surface. The results reported here show that heterogeneity of the adsorbent surface may be important in analysis of the elution process in liquid chromatography. Consideration of the goodness of fit for the experimental data to the examined retention models is in conformity with the adsorption mechanism of retention on all polar bonded stationary phases in most eluent systems for most investigated compounds.     

Lipophilicity determination of N-( benzothiazol-2-yl )-α-amino alkyl phosphonic diesters by RP-HPLC and RP-HPTLC

Using methanol-water mixtures as the mobile phase,the chro-matographic retention parameters k' and Rf were determined by reversed-phase high-performance liquid chromatography(RP-HPLC)and reversed-phase high-performance thin-layer chromatography(RP-HPTLC)for N-(benzothiazol-2-yl)-o) amino alkyl phosphonic diesters and the correlation with lipophilicity parameter(ClogP)was established.Logkw values obtained from RP-HPLC and R values obtained from RP-HPTLC can be used to evaluate the lipophilicity of this kind of compounds.Chromatographic method is a good alternative for lipophilicity measurement.     

Optical resolution of a series of potential cholecystokinin antagonist 4(3H)-quinazolone derivatives by chiral liquid chromatography on alpha1-acid glycoprotein stationary phase

Optical resolution of the enantiomers of new 4(3H)-quinazolone derivatives is investigated using the alpha1-acid glycoprotein chiral stationary phase (Chiral-AGP). Stereoselective separation of the model compounds can be controlled by varying the pH and adding uncharged organic modifiers (acetonitrile and 2-propanol) to the mobile phase. For the majority of quinazolone derivatives, Chiral-AGP is proved to be an excellent enantioselector, because optimized chromatographic conditions allow for the baseline separation of the enantiomers. Separation factors between 1.19 and 1.85 are obtained. The effects of acetonitrile and 2-propanol on the chromatographic behavior of the model compounds are quite different because of their different hydrophobic- and hydrogen-bonding properties. The eluent pH and organic modifier concentration also contributes to the chiral recognition by altering the protein environment. The analysis of the experimental results leads to new information about the chromatographic mechanism on a Chiral-AGP surface.     

表阿霉素及其相关物质在反相离子对色谱中的色谱行为

以表阿霉素及其6种相关物质为研究对象,系统评价了其在反相离子对色谱模式下的色谱行为.分别考察了流动相中有机相种类、有机相比例、水相中离子对试剂浓度、pH值对表阿霉素及其相关物质的影响.结果表明,使用乙腈作为有机相洗脱能力及分离效果优于甲醇,保留时间随乙腈比例增大而减小;随着离子对试剂十二烷基硫酸钠浓度增加,杂质阿霉素酮及柔红霉酮几乎无影响,其他5种物质保留时间增加.同时,表阿霉素及其杂质的保留行为受流动相pH值影响较大,当pH不高于4时可获得较好的分离效果.通过对表阿霉素及其相关物质反相离子对模式下的保留行为进行了系统的评价和定量描述,研究结果将有助于该类化合物液相色谱分离方法的发展.     

Reversed phase liquid chromatography of some benzimidazole derivatives

Summary The retention behaviour of a series of benzimidazole derivatives has been studied as a function of the water content of aqueous methanol and aqueous acetonitrile eluents. The relationship between the retention constant (log k) and the pH of the aqueous phase was linear, with slope values depending on the composition of the aqueous phase, the molecular structure of the compound, and the type of C-18 bonded stationary phase. The type of organic modifier significantly affected the shape of the relationship between log k and the volume fraction of organic modifier in the mobile phase.     

Lipophilicity determination of some potential photosystem II inhibitors on reversed-phase high-performance thin-layer chromatography

The retention characteristics of 25 2-cyano-3-methylthio-3-substituted amine-acrylates are determined using reversed-phase thin-layer chromatography (RP-TLC) with methanol-water mixtures as eluents. The relationship between Rm values and partition coefficients (C log P) are established. The Rm values decrease linearly with increasing methanol concentration in the eluent. The Rm values extrapolated to zero organic modifier concentration (Rm0) in the eluent are highly related to C log P. The Rm0 value can be used to evaluate the lipophilicity of this kind of compound.     

环孢素A在反相液相色谱中的吸附行为及分离纯化北大核心CSCD

环孢素A(cyclosporine A,CsA)是由11个氨基酸组成的中性环状多肽,是临床拮抗器官和组织移植后排异反应的首选药物。高效液相色谱法广泛应用于CsA的分离分析,开展CsA色谱行为的研究是使用制备高效液相色谱纯化CsA的关键。该文首先在C18固定相上比较了CsA在甲醇-水和乙腈-水两种流动相体系中的保留行为,结果表明其保留时间对有机相比例变化比较敏感。控制甲醇比例在84%~88%,或者乙腈比例在75%~85%,CsA的保留因子(k)在3~7范围内。考察了上样量对CsA峰形的影响。随着上样量增加,在两种流动相体系中,CsA的峰形都由对称开始变得拖尾,保留时间前移。因此在进行CsA纯化时,需要特别注意前杂的去除情况。然后采用吸附等温线描述CsA的保留行为,当流动相中CsA的质量浓度较低时,有机相比例对溶质在固定相上的吸附量影响并不明显。随着溶质的质量浓度增加至0.5 g/L以上,有机相比例降低有助于提高CsA在固定相上的吸附量。和甲醇-水体系相比,在乙腈-水体系中固定相对溶质有更大的吸附容量。用模型对CsA的等温吸附曲线拟合,在甲醇-水体系中符合Langmuir模型,在乙腈-水体系中为Moreau模型。由模型参数可知在两种体系下,CsA在C18固定相上均为单层吸附,区别在于乙腈-水体系中CsA会产生较大的分子间作用力。最后,实验采用0~60 min 65%~75%乙腈、60~80 min 75%乙腈的条件开展了环孢素A纯化的探索实验,可将CsA的杂质控制在0.2%以下。本研究结果对采用制备高效液相色谱纯化CsA具有指导意义。     

Retention of some ring-substituted aniline derivatives by porous graphitized carbon. Dependence on physico-chemical parameters

Summary The retention characteristics of 22 aniline derivatives were determined on a porous, graphitized-carbon column in unbuffered acetonitrile-water and methanol-water mixtures. Each aniline derivative gave symmetrical peaks in each eluent without buffers. Good linear correlations were found between the log k and the concentration of the organic component in the eluent. The slope and intercept values differed according to the type of organic modifier and the charcter, number and position of substituents, indicating the different selectivities of methanol and acetonitrile and the good separating power of the column. Multivariate mathematical-statistical calculations proved that the retention of ring-substituted aniline derivatives is mainly governed by electronic parameters and the hydrogen acceptor capacity of substituents.     

Application of chaotropic effect in reversed-phase liquid chromatography of structurally related phenothiazine and thioxanthene derivatives

Chromatographic behavior of two main classes of neuroleptics, derivatives of phenothiazine and thioxanthene in RP systems modified by anionic additives: sodium perchlorate and sodium hexafluorophosphate possessing chaotropic properties, was examined. Influence of the method of pH lowering (by addition of acids: trifluoroacetic or perchloric or by adding the appropriate concentration of phosphate buffer) and the kind of organic modifier in the mobile phase (methanol, acetonitrile) were estimated. Stability of complexes created between protonated drugs and anions of added salts was evaluated by comparison of their desolvation parameters (K), limiting retention factors for unsolved molecules calculated on the basis of chromatographic data. Experimentally obtained parameters were used in QSAR studies. It appeared that chosen parameters reflect not only physico-chemical properties of analytes but also contain information about the strength of their antipsychotic activity. Multidimensional cluster analysis has been performed. On the basis of the results obtained, it could be concluded that chaotropic systems can generate useful parameters for further QSAR studies.     

Structural modifications of dicationic acetylcholinesterase reactivators studied under ion‐pairing mechanism in reversed‐phase liquid chromatography

A study focused on the chromatographic behavior of several acetylcholinesterase reactivators under ion‐pairing mechanism is reported. Among these reactivators, dicationic oximes and carbamoyl‐based pyridinium congeners were studied, which form ion pairs with alkylsulfonate anions. This mechanism was studied for some major experimental parameters, such as the chain length of the ion‐pairing agent added to the aqueous phase, its concentration, temperature, and nature of the organic modifier from mobile phase. Retention data showed one or two possibilities of forming ion pairs and the tautomerism of the studied reactivators, for different pH values of the aqueous component. Double sigmoid shapes were obtained for the studied compounds for the dependence between retention factor and pH, indicating the possibility of one or two tautomeric equilibria: at pH close to 7 these compounds are not stable as dicationic species and they participate in the retention process as nitroso forms, which are not able to form ion pairs with alkylsulfonates. The dependences of the retention factor on the organic modifier content from mobile phase were linear. Two complementary theoretical models were used to explain the functional dependences for the retention data on the experimental parameters.     

Influence of the mobile phase on salmon calcitonin analysis by reversed-phase liquid chromatography

The retention properties of calcitonins on a reversed-phase column are examined using salmon calcitonin as the model compound. The effect of the concentration of organic modifier, buffer strength, pH of the mobile phase, and ion-pair reagent are studied. In the absence of an ionic modifier in the eluent the calcitonin peak shapes are not symmetrical. The addition of 0.1% trifluoroacetic acid (TFA), however, results in good peak characteristics without the need to add nonvolatile salts. The retention of the calcitonins was found to be very sensitive to the concentration of the organic modifier (acetonitrile) present in the mobile phase. A change of pH between 2 and 5 has only a slight effect of the k' of salmon calcitonin, but the k' increases significantly at higher pH values. The addition of a phosphate buffer to the mobile phase and an increase in the buffer concentration (0-0.2 M) causes a decrease in the retention of salmon calcitonin. Evidence shows that reproducible, quantitatively measurable data can be obtained using reversed-phase chromatography if the ion-pairing reagent and organic modifier concentrations are carefully controlled. The system also shows a good selectivity for the calcitonin series. Therefore, both highly selective methods (qualitative) as well as quantitative methods for analytical, pharmaceutical, and manufacturing use can be developed by adjusting the high-performance liquid chromatography (HPLC) conditions as discussed.     

Relationship between the lipophilicity and specific hydrophobic surface area of some pesticides by RP-HPLC and HPTLC

Summary Using methanol-water mixtures as the mobile phases, the retention behaviors of thirty-seven pesticides were determined in RP-HPLC and RP-HPTLC. Regular retention behavior was observed for all the investigated pesticides: theirR _m and logk values decreasing linearly with increasing concentration of methanol in the mobile phase. The lipophilicity and specific hydrophobic surface area values for each compound were obtained and they have a good linear relationship. Although the chemical structures of these pesticides were different, factor analysis proved that the lipophilicity and specific hydrophobic surface area of these compounds have much in common, and the insecticides, fungicides and herbicides could not be distinguished from each other according to their lipophilicity parameters obtained from chromatography method.     

Comparison of Retention Indices of Some Explosives and Related Compounds Calculated by Different Mathematical Methods in Reversed-Phase Liquid Chromatography

In reversed phase liquid chromatography, the retention indices of some neutral and acidic explosives and related compounds (nitramines, nitroaromatics, aminoaromatics, and nitrophenols) based on the alkan-2-one, alkyl aryl ketone, and 1-nitroalkane retention index standards have been determined by the application of a new mathematical adaptation method, viz. a multiparametric least-squares regression iterative method. This method was applied to two types of columns. The first group includes six octadecyl-C₁₈ columns with different packing materials and obtained from different manufacturers, while the second group comprises one octyl-C₈ column. The retention indices have been extensively studied using either methanol-water or methanol-phosphate buffer mobile phase systems. The calculated multiparametric retention indices values were compared with those obtained by Guardino's, Grobler's, and Kovàts' methods. The influences of the concentration of the organic modifier in the mobile phase, aqueous mobile phase pH, and the column packing material on the retention indices of the explosives were also investigated. Good agreement was observed between the retention indices calculated by the use of the four mathematical methods for both neutral and acidic explosives.     

Insights into the Retention Mechanism on a Pentafluorophenylpropylsiloxane-Bonded Silica Stationary Phase (Discovery HS F5) in RP-LC

The solvation parameter model is used to elucidate the retention mechanism of neutral compounds on the pentafluorophenylpropylsiloxane-bonded silica stationary phase (Discovery HS F5) with methanol-water and acetonitrile-water mobile phases containing from 10 to 70% (v/v) organic solvent. The dominant factors that increase retention are solute size and electron lone pair interactions while polar interactions reduce retention. A comparison of the retention mechanism with an octadecylsiloxane-bonded silica stationary phase based on the same silica substrate and with a similar bonding density (Discovery HS C18) provides additional insights into selectivity differences for the two types of stationary phase. The methanol-water solvated pentafluorophenylpropylsiloxane-bonded silica stationary phase is more cohesive and/or has weaker dispersion interactions and is more dipolar/polarizable than the octadecylsiloxane-bonded silica stationary phase. Differences in hydrogen-bonding interactions contribute little to relative retention differences. For mobile phases containing more than 30% (v/v) acetonitrile selectivity differences for the pentafluorophenylpropylsiloxane-bonded and octadecylsiloxane-bonded silica stationary phases are no more than modest with differences in hydrogen-bond acidity of greater importance than observed for methanol-water. Below 30% (v/v) acetonitrile selectivity differences are more marked owing to incomplete wetting of the octadecylsiloxane-bonded silica stationary phase at low volume fractions of acetonitrile that are not apparent for the pentafluorophenylpropylsiloxane-bonded silica stationary phase. Steric repulsion affects a wider range of compounds on the octadecylsiloxane-bonded than pentafluorophenylpropylsiloxane-bonded silica stationary phase with methanol mobile phases resulting in additional selectivity differences than predicted by the solvation parameter model. Electrostatic interactions with weak bases were unimportant for methanol-water mobile phase compositions in contrast to acetonitrile-water where ion-exchange behavior is enhanced, especially for the pentafluorophenylpropylsiloxane-bonded silica stationary phase. The above results are compatible with a phenomenological interpretation of stationary phase conformations using the haystack, surface accessibility, and hydro-linked proton conduit models.