Indirect detection in reversed-phase liquid chromatography: Ion-pair retention models for anionic analytes and mobile phase components on polystyrene polymers using an alkaline mobile phase

Summary Retention models for anionic analytes and mobile phase compounds in reversed-phase ion-pair chromatographic systems have been studied. The solid phase was a polystyrene-divinylbenzene copolymer. The analytes were mono- and divalent anions monitored by indirect detection technique. The mobile phase was highly alkaline and contained a detectable anion, sulfanilic acid.Expressions for the ion-pair retention of solutes and mobile phase components have been developed assuming Langmuir distribution of ion-pairs to a solid phase with one kind of binding site. The validity of the expressions and the origin of system peaks have been confirmed by determination of distribution constants using linear regression.     

High-performance liquid chromatographic separation and extraction investigation for the simultaneous determination of mefloquine and its carboxylic acid metabolite

Distribution ratios of mefloquine ion pairs with perchlorate and heptanesulphonate as counter-ions, the base distribution for mefloquine, the acid distribution for the carboxylic acid metabolite and the ion-pair distribution with quaternary ammonium ions as counter ions have been determined. Differences in retention characteristics with several commercially available supports bearing the same octadecyl label were found. The effect on the retention time of the pH, the percentage acetonitrile and the addition of an amine to the mobile phase were studied. Other antimalarial drugs present concurrently in the sample showed no chromatographic interference with mefloquine and its metabolite. A suitable internal standard for use in the analytical method was also tested.     

Fluorimetric determinations by ion-pair extraction : Part V. Studies on ion-pair extraction with the fluorescent anion 9,10-dimethoxyanthracene-2-sulphonate

The fluorescent anion 9,10-dimethoxyanthracene-2-sulphonate has been studied in ion-pair extractions. Its ability to extract amines and quaternary ammonium compounds was investigated by fluorimetric determination of extraction and dissociation constants with methylene chloride as organic phase for two amines (amitriptyline and protriptyline) and two quaternary ammonium ions (tetrabutyl-and tetrapropylammonium). The optical properties of the anion and its ion pairs were characterized by quantum yield of fluorescence and a fluorescence sensitivity index. The determination of two of the ion pairs was studied, both after extraction to an organic phase and after a subsequent back-extraction of the anion component to an aqueous phase. Recoveries and standard deviations are given.     

Ion-pair LC of carbohydrates at alkaline pH. Separation and retention behaviour of glycoconjugates

Summary A new technique for the separation of carbohydrates as ion-pairs in strongly alkaline solution is presented. Carbohydrates are weakly acidic and partly present as anions at pH12 [1]. They are retained as ion-pairs on polymeric solid phases (PRP-1 and PLRP-S) with a hydrophobic quaternary ammonium counter ion present in the mobile phase. The effects of nature and concentration of mobile phase components on the retention of carbohydrates have been investigated and an ion-pair distribution model is proposed. The influence of temperature indicated no changes in retention mechanism with high counter ion concentration, but the resolution decreased with increasing temperature. Saccharides added to the mobile phase were shown to increase the retention and the selectivity.     

Effect of the steric accessibility of the exchange site in long-chain quaternary ammonium salts on the anion-exchange extraction of divalent ions

The constants of anion-exchange extraction of divalent ions by quaternary ammonium salts of various structures were determined. The exchange constants depend significantly on steric accessibility of the exchange site. For the exchange of small-size anions, the affinity of the ion exchanger for divalent ions is higher than for monovalent ions. For large divalent anions, the exchange constant may increase or decrease, depending on the size of a monovalent anion. The obtained results are explained by the specifics of the ion-pair association of monovalent and divalent anions with quaternary ammonium cations.     

Effects of Counter Ions in Ion-Pair Liquid Chromatography of Hydrophobic Amines on Non-Polar Bonded Phases

Abstract

The retention behaviour of alprenolol and related hydrophobic amines in ion-pair adsorption systems has been examined with particular emphasis on the influence of different mono-and divalent counter ions (dihydrogenphosphate, bromide, perchlorate, dimethylcyclohexyl sulphate, sulphate and ethylenediaminetetraacetate). N,N-dimethyloctylamine (DMOA) and 1-pentanol were used as modifiers in the aqueous eluent and LiChrosorb RP-8 as stationary phase.

The retention is evaluated according to a two-site adsorption model and equilibrium constants are given for ion pair adsorption of DMOA. The retention of alprenolol has been evaluated in terms of ion exchange with DMOA and the ion-exchange constants are shown to be of the same magnitude and independent of the nature of the counter ion used. The ion-pair adsorption and the ion-exchange approaches are analogous expressions for the distribution process governing the retention.     

Indirect detection in reversed-phase liquid chromatography: Ion-pair retention models for ionic compounds on μ Bondapak phenyl

Summary Retention models for ionic compounds in reversed-phase ion-pair chromatographic systems have been developed on the basis of the capacity ratios of solutes and ionic mobile phase components using indirect detection. The binding to the adsorbent, μBondapak Phenyl, has been expressed by ion-pair distribution constants and tested in the retention models by non-linear regression. All the ionic compounds followed adsorption models of the Langmuir type comprising two kinds of binding sites with widely different adsorption capacities and binding constants. The results indicate certain differences between the two sites regarding the binding of cationic and anionic components, respectively.     

Indirect detection in liquid chromatography. II. Response models for reversed-phase non-ionic systems

Summary Uncharged solutes devoid of inherent detectable properties can be monitored and quantified in reversed-phase chromatographic systems by including a non-ionic detectable component (probe) in the eluent. Response models have been developed, which are based on the fact that the distribution of the solute and the probe are connected to each other by common interaction effects, such as competition for the binding surface. The direction and magnitude of the indirect response can be predicted quantitatively by means of expressions which clearly show the influence of the retention of the solute relative to the probe as well as the effect of the solid phase coverage by the probe. The relatively low response obtained in non-ionic systems may be due to a low loading of the probe on the solid phase and to a simultaneous distribution of the solute in complexed and uncomplexed forms, the two distribution modes giving opposite response patterns.     

Indirect detection in liquid chromatography I. Response models for reversed-phase ion-pairing systems

Summary Ionic compounds without inherent detectable properties show good response in reversed phase ion-pairing systems containing a detectable ionic compound (probe) in the mobile phase. The basis for the indirect detection is the common interaction effect for solutes and probe caused by the limited binding capacity of the adsorbent. Quantitative models for solutes and probe with the same or opposite charges show a simple dependence of the response on the retention of the solute and the coverage of the binding surface by the probe. Applications of the models have shown good agreement between calculated and found response. The binding to sites of different kinds on the adsorbent and possibly ion-pair formation in the mobile phase have an influence on the indirect response. Uncharged solutes give low response in such systems owing to low binding to the competitive sites.     

Ion-pair LC of sugars at alkaline pH and pulsed electrochemical detection

Summary Sugars, sugar acids, amino sugars and oligomers are separated as ion-pairs with hydrophobic counter ions at alkaline pH using PLRP-S and Hypercarb as solid phases. Important parameters for regulation of retention and selectivity are nature and concentration of the counter ion, pH (hydroxide concentration) and temperature. Reversals in the elution order wer obtained in some cases. Oligosaccharides are highly retained in these systems. The addition of organic modifiers to the mobile phase for elution of the solutes were found to interfere with the pulsed electrochemical detection (PED). Anions added to the mobile phase compete with the solutes for ion-pair retention, hence, decreasing the capacity factors, and phosphate could be used for this purpose in the separation of maltooligomers (M2-M10) from corn syrup.     

缓冲盐类型和离子对试剂非对离子对强离解酸性化合物离子对反相液相色谱保留行为的影响

在离子对反相液相色谱(IP-RPLC)分析中,溶质保留受对离子(counter ion)的影响比较受人关注,但鲜有研究流动相中缓冲盐类型和离子对试剂中非对离子(non-counter ion)对溶质保留行为的影响。鉴于此,该文以14种磺酸化合物为研究对象,甲醇为有机调节剂,分别考察了3种缓冲盐体系(磷酸二氢铵、氯化铵和乙酸铵)和5种离子对试剂体系(四丁基溴化铵、四丁基磷酸二氢铵、四丁基硫酸氢铵、四丁基硝酸铵和四丁基乙酸铵)下强离解酸性化合物的IP-RPLC保留行为,通过比较不同流动相条件下得到的溶质log k_w(100%水相作流动相时的保留因子)、S(线型溶剂强度模型线性回归得到的常数),以及CHI(色谱疏水指数,log k_w/S),寻找保留行为规律。研究表明,流动相中的缓冲盐类型和离子对试剂非对离子均会影响化合物的log k_w和S值,所有化合物在氯化铵缓冲盐体系下具有最大的log k_w值。相对于无机阴离子,离子对试剂中弱离解性有机阴离子(乙酸根)的存在有利于增加磺酸化合物的S值。通过对比不同条件下的保留行为,推测磺酸化合物的IP-RPLC保留机理中同时存在着离子对模型和动态离子交换模型。与log k_w和S值不同,化合物的CHI值受缓冲盐类型以及离子对试剂非对离子的影响较弱。此外,研究发现化合物的表观正辛醇/水分配系数(log D)与log k_w、S、CHI之间均具有良好的线性相关性。不同缓冲溶液和不同离子对试剂非对离子条件下获得的log k_w和S值存在着一定的差异,而CHI值相对稳定,因此,CHI更适用于IP-RPLC中定量结构-保留行为关系模型的建立。     

Reversed-Phase Ion-Pair High-Performance Liquid Chromatography of Phosphatidylinositols

Abstract

Reversed-phase ion-pal, high-performance liquid chromatographic (HPLC) separations of molecular species of phosphatidylinositols (PI) were studied. Mobile phases of acetonitrile-methanol-water containing various tetraalkylammonium phosphates (TAAP) were used for optimization. Stationary phases of macroporous polystyrene divinylbenzene (MPD), octylsilica, and octadecylsilica exhibited arying degrees of retentivity toward PI solutes. Without exception capacity factors (k') of PI on either alkylsilica or MPD increased with the alkyl chain length and the concentration of the quaternary ammonium counter ions evaluated. the results can be interpred in terms of an ion-pair retention mechanism. Logarithmic k' values were linearly related to the total number of carbons in TAAP. Perbenzoylation of PI yielded derivatives readily resolvable without the use of any mobile phase additive. Incorporation of TAAP to a mobile phase facilitated component separations of early-eluting perbenzoates. In HPLC with     

Ion-pair chromatography of acidic drug metabolites and endogenic compounds.

Liquid-liquid chromatographic systems based on ion-pair partition with silica microparticles as the support for the stationary phase have been used for the separation of anionic compounds of biochemical and pharmacological interest. A high separating efficiency can be obtained with both aqueous and organic mobile phases and the retention is easily regulated by the nature and the concentration of the quaternary ammonium counter ion, present in the aqueous phase. The influence of the composition of the liquid phases on the selectivity and separating efficiency has been studied, as well as equilibration methods and the stability of the systems. Examples are given of separations of sulphonamides, barbiturates, glucuronic and sulphuric acid conjugates of steroidal compounds and phenols glycine conjugates of carboxylic acids (hippuric, nicotinuric and salicyluric acid) and anionic metabolites of biogenic amines (indoleacetic, benzoic, mandelic and phenylacetic acid derivatives).     

A Mixed Retention Model of Reversed Phase HPLC

The retention behavior of solutes in reversed phase chromatographic system, especially containing hydrophobic ions in mobile phase, has been studied by many authors, but the silanol effect on alkyl-modified silica surface was neglected. Actually, because of stereo hindrance, numerous unreacted (residual) silanol groups, which can interact with some solutes, are left within the bonded phase after the silica surface has be modified. A mixed retention model, which considers adsorbed hydrophobic ions on the bonded phase can decrease the hydrophobicity and mask residual silanol groups, is proposed, based on hydrophobic distribution of neutral solutes, ion-pair distribution of ionic solutes and coulombic attraction between cationic solutes and the dissociated silanols.     

羧基甜菜碱型亲水作用色谱柱的制备及性能研究

以甲基丙烯酰氧乙基二甲基乙酸铵(CBMA)为功能单体,利用表面引发原子转移自由基聚合(Surface-initiated atom transfer radical polymerization, SI-ATRP)技术,将CBMA接枝到硅胶表面,得到接枝聚合物CBMA的亲水作用色谱固定相(Silica-CBMA).通过改变SI-ATRP反应体系中单体的浓度,制备了3种不同接枝量的亲水作用色谱固定相.考察了Silica-CBMA固定相对有机酸类化合物的分离性能以及流动相中pH值、盐浓度、水含量等因素对溶质保留行为的影响.结果表明,在亲水作用色谱模式下,Silica-CBMA固定相对有机酸类化合物的分离是离子交换作用与亲水作用的混合色谱模式.流动相中盐浓度增大,溶质保留减弱,符合离子交换作用特征;固定相和溶质的离子化程度受流动相pH值影响较大,pH值增大,溶质保留增强;而溶质的保留时间随流动相水含量增加而降低则是典型的亲水作用色谱特征.使用自制Silica-CBMA柱,建立了芦丁片中维生素C、芦丁含量的亲水作用色谱测定方法,操作方法简单,为强极性样品的分离测定提供了新方法.     

A chemometric investigation of the selectivity of multisolvent mobile phase systems in RP-HPLC

Summary The selectivity of multisolvent mobile phase systems consisting of water, methanol, acetonitrile and tetrahydrofuran was studied when the solvent strength decreases at a constant ratio of the modifiers. From the retention measurements of six benzene derivatives on an octylsilica column at eleven different mobile phase compositions the relation between the logarithm of the capacity factor of each solute and the mobile phase composition was modeled by a quadratic equation. From the models the capacity factors of the solutes were predicted for 3 binary, 3 ternary and 7 quaternary mobile phase systems when the fraction of water decreases and the ratio of the organic modifiers is kept constant for the tenary and quaternary solvent systems. The selectivity factors, alpha, of five pairs of solutes were calculated from the capacity factors and plotted against the solvent strength of the mobile phase systems. The selectivity remained not constant, but varied with the solvent strength: if the water fraction of a multi-solvent system is changed at a constant ratio of the modifiers, not only the solvent strength, but also the selectivity changes. The consequence of this result for optimization strategies is discussed.     

UPLC a Powerful Tool for the Separation of Imidazolium Ionic Liquid Cations

Ultra-performance liquid chromatography (UPLC) in reversed-phase (RP), ion pair (IP) and hydrophilic interaction chromatography (HILIC) has been investigated for the separation of imidazolium-based ionic liquid (IL) cations. Among the three stationary phases (i.e., C18, C8 and phenyl) studied under RP conditions the phenyl phase provided much stronger retention for the IL cations. Four acids (hydrochloric, methanesulfonic, perchloric and trifluoroacetic) as mobile phase additives were compared in light of their effects on the retention of IL cations. It was shown that the retention of all IL cations decreased upon acidification of the mobile phase, possibly due to suppression of residual silanol ionization. Very fast (~3 min) and efficient RP-UPLC separation of six cations was achieved by gradient elution with acetonitrile?Cwater mobile phase containing 2.5 mmol L^?1 perchloric acid. In IP-UPLC all solutes were well resolved in about 4 min by gradient elution with acetonitrile?Cwater mobile phase containing 1 mmol L^?1 sodium 1-octanesulfonate as ion pairing reagent. Finally, under HILIC conditions by using isocratic elution with acetonitrile?Cwater (85:15, v/v) mobile phase containing 5 mmol L^?1 ammonium formate (pH 3.2) the separation time was reduced to less than 2 min while maintaining excellent peak shapes and sufficient resolution. Compared to current LC systems UPLC allowed considerably faster separations with better peak shapes.     

非极性毛细管电色谱原位柱的阳离子表面活性剂动态修饰

毛细管电色谱是在毛细管中依靠电渗流来驱动流动相 ,同时溶质与固定相发生相互作用的一种色谱分离模式 ,它有高效液相色谱的高选择性 ,同时兼具毛细管电泳的高效性 [1] .传统电色谱柱是将HPLC填料装入毛细管 ,但由于装柱困难且易产生气泡而在一定程度上阻碍了电色谱的发展 [2～ 4 ] .通过柱内合成的方法直接在毛细管中制成连续床毛细管电色谱柱 ,可避免两端烧塞 . 1 995年 Svec等 [5,6]首次将连续床层色谱柱用于毛细管电色谱 ,此后 ,有关毛细管中原位合成连续床电色谱柱的方法得到了应用 [7～ 11] .为了使原位合成电色谱柱能产生电渗流 ,…     

Retention in Reversed-Phase Ion-Pair Chromatography of Amines On Alkyl-Bonded Phases

Abstract

Reversed phase ion-pair chromatography of phenethylamine derivatives (noradrenaline, adrenaline, dopamine, synephrine, tyramine and pholedrine) and lower alkylamines has been performed with octyl sulfate as counter ion in an aqueous eluent with a low content of 1-pentanol. LiChrosorb RP-18 was used as the solid phase. The retention of lithium and potassium in the system has also been studied.

On the basis of adsorption and retention studies a model for the chromatographic behaviour of the amines is proposed that includes interaction with two sites with different binding ability in the stationary phase. The adsorption capacity of the sites has been calculated as well as adsorption constants for the octyl sulfate ion pairs.     

The role of analogue ions in the ion-pair reversed-phase chromatography of quaternary ammonium compounds

The choice of analogue ion of the mobile phase additive is shown to significantly affect the analysis of quaternary ammonium compounds (QACs) in ion-pair reversed-phase high-performance liquid chromatography. A series of bromide-containing and dodecyl-sulfate-containing mobile phase additives are investigated using two QAC probe analytes. In all instances, the quaternary-ammonium-containing mobile phase additives perform better than the corresponding sodium-containing additives for effective QAC elution. These results indicate that the structure of the analogue ion, not just its formal charge, is important in the reversed-phase ion-pair chromatography of these compounds. The relative elution order of the QAC probe analytes is also influenced by the counter ions of the mobile phase additives, with bromide and dodecyl sulfate offering opposite elution orders.