Preparation and Evaluation of Poly-l-Lysine Stationary Phase for Hydrophilic Interaction/Reversed-Phase Mixed-Mode Chromatography

In this work, a poly-l-lysine-grafted stationary phase was synthesized by polymerization of N-carboxyanhydride of l-lysine initiated by 3-aminopropylated silica. The resulting material was characterized by FT-IR spectra, elemental analysis and thermogravimetric analysis, which clearly indicated that the new phase had been prepared successfully. The retention of polar solutes depending on acetonitrile content in mobile phase exhibited ??U-shaped?? curves, which was an indication of hydrophilic interaction liquid chromatography (HILIC)/reversed-phase liquid chromatography (RPLC) mixed-mode retention behavior. The retention mechanisms in HILIC and RPLC modes also were investigated. Phenol compounds, aniline compounds and hydrophilic compounds were separated in RPLC or HILIC mode on the new stationary phase, respectively. This result shows that the new phase could be used for both RPLC and HILIC applications, providing greater flexibility for real sample analysis.     

Enthalpy-Entropy Compensation of Octylsilica Stationary Phase in Reversed-Phase HPLC

Abstract

Enthalpy-entropy compensation has been investigated in reversed-phase high performance liquid chromatography with octylsilica stationary phase. The compensation temperatures were determined for this system, and the results show that their change with the composition of the mobile phase was almost similar to that with octadecylsilica stationary phase. It is concluded that the retention mechanism of the separation of alkylbenzenes in both systems with the mobile phase exceed 20% water content is the same.     

Silica-Based Phenyl and Octyl Bifunctional Imidazolium as a New Mixed-Mode Stationary Phase for Reversed-Phase and Anion-Exchange Chromatography

In this study, octylbenzimidazolium-modified silica (BeImC8-Sil) was prepared by covalent attachment of 1-octylbenzimidazole to γ-chloropropyl silica. The synthesized materials were characterized by the elemental analysis, IR spectrum, and thermogravimetric analysis. Due to the introduction of phenyl and octyl groups on the quaternary imidazolium, the developed BeImC8-Sil column can function via both reversed-phase and anion-exchange retention mechanisms. The chromatographic properties of the synthesized material were investigated by the separations of polycyclic aromatic hydrocarbons, mono-substituted derivatives of benzene, anilines, and phenols, revealing the existence of multiple interactions, including hydrogen bonding, π–π stacking, electrostatic forces, and hydrophobic interactions in reversed-phase mode; inorganic and organic anions were also separated mainly through anion-exchange interaction. The proposed BeImC8-Sil is a promising mixed-mode stationary phase for the separation of complex samples in high-performance liquid chromatography.

     

混合固定相电中溶质的输运特征

 以离子交换和反相固定相构成的混合固定相电中 ,溶质迁移在受到疏水、离子交换作用的同时 ,对于带电溶质而言 ,还受到电泳迁移的影响。根据离子独立迁移原理 ,结合过程中的多种相互作用 ,得到了描述溶质表观迁移速率与其各形态迁移速率、各种相互作用之间相互关系的理论表达式 ;讨论了混合模式电中流动相的 pH及其中的有机调节剂浓度、混合固定相配比等对电渗流的影响及不同形态溶质在柱内的输运特征。结果表明 ,在电中采用混合固定相可以在较大的 pH和有机调节剂浓度范围内得到较强且稳定的电渗流。     

Contribution of Steric Repulsion to Retention on an Octadecylsiloxane-Bonded Silica Stationary Phase in Reversed-Phase Liquid Chromatography

Plots of the retention factor against mobile phase composition were used to organize a varied group of solutes into three categories according to their retention mechanism on an octadecylsilioxane-bonded silica stationary phase, Ascentis ^TM C18, with acetonitrile-water and methanol-water mobile phase compositions containing 10–70% (v/v) organic solvent. The solutes in category 1 could be fit to a general retention model, Eq. (1), and exhibited normal retention behavior for the full composition range. The solutes in category 2 exhibited normal retention behavior at high organic solvent compositions with a discontinuity at low organic solvent mobile phase compositions. The solutes in category 3 exhibited a pronounced step or plateau in the middle region of the retention plots with a retention mechanism similar to category 1 at mobile phase compositions after the discontinuity and a different retention mechanism before the discontinuity. Selecting solutes and appropriate composition ranges from the three categories where a single retention mechanism was operative allowed modeling of the experimental retention factors using the solvation parameter model. These models were then used to predict retention factors for solutes excluded from the models. The overwhelming number of residual values, here defined as the difference between experimental and model predicted retention factors for the excluded solutes, could be explained by contributions from steric repulsion. The latter defined as the inability of solutes to fully insert themselves into the solvated stationary phase because of their size or conformation. Steric repulsion resulted in a systematic reduction in retention compared with predicted values for the fully inserted solute. The bonding density of the stationary phase; the type and composition of the mobile phase; and the size, conformation, type and number of functional groups on the solute are shown to affect the contribution of steric repulsion to the retention mechanism.     

量子化学计算预测高效液相色谱键合相的保留机理

本文用Gaussian98计算软件中的Hartree—Fock方法对高效液相色谱中溶质与固定相发生作用后产生的能量进行计算,并对溶质与C18键合相和苯胺甲基键合相相互作用的计算结果与实验结论进行了比较。结果表明,量化计算结果与色谱保留行为之间具有相关性,即分子间相互作用能量的高低,决定了保留值的大小,表现为能量越负,保留值越大,而且双倍键合量的能量低于单倍键合量。量化计算的方法将为色谱保留性能的研究和键合相的研制提供参考。     

二氢麦角碱在极性嵌入反相色谱固定相上的分离行为

二氢麦角碱的4种组分具有不同的药效特性,需适当的分析方法对其在不同剂型药品中的含量进行分析.传统液相色谱方法使用强碱性流动相,严重腐蚀硅胶基质色谱填料,影响色谱柱寿命.合成了极性嵌入反相固定相--C18酰胺固定相,并在中性流动相条件下分离4种组分.考察了流动相组成和pH对二氢麦角碱在C18酰胺固定相上保留行为的影响.在150 mm×4.6 mm I.D.的C18酰胺色谱柱上,流动相为乙腈-20 mmol/L Na2HPO4(30:70,V/V,pH 7.0)的条件下实现了4种组分的基线分离.与传统方法相比,极性嵌入反相固定相可以有效的改善碱性化合物的分离特性和大幅度地延长色谱柱的使用寿命.     

System Maps for Retention of Small Neutral Compounds on a Superficially Porous Ethyl-Bridged,Octadecylsiloxane-Bonded Silica Stationary Phase in Reversed-Phase Liquid Chromatography

The system constants of the solvation parameter model are used to prepare system maps for the retention of small neutral compounds on an ethyl-bridged, ocatadecylsiloxane-bonded superficially porous silica stationary phase (Kinetex EVO C18) for aqueous mobile phases containing 10–70% (v/v) methanol or acetonitrile. Electrostatic interactions (cation-exchange) are important for the retention of weak bases with acetonitrile–water but not methanol–water mobile phase compositions. Compared with a superficially porous octadecylsiloxane-bonded silica stationary phase (Kinetex C18) with a similar morphology but different topology statistically significant differences in selectivity at the 95% confidence level are observed for neutral compounds that vary by size and hydrogen-bond basicity with other intermolecular interactions roughly similar. These selectivity differences are dampened with acetonitrile–water mobile phases, but are significant for methanol–water mobile phase compositions containing <30% (v/v) methanol. A comparison of a totally porous ethyl-bridged, octadecylsiloxane-bonded silica stationary phase (XBridge C18) with Kinetex EVO C18 indicated that they are effectively selectivity equivalent.     

关于混合模式色谱固定相对蛋白保留特征的研究

为使混合机理色谱(MMC)得到广泛地应用, 合成、表征和评价MMC固定相就成了首先要解决的问题. 依据离子交换色谱柱也具有疏水色谱(HIC)保留机理的特征, 选了4种弱阳离子交换(WCX)柱和一根二维[2D(WCX,HIC)]色谱柱, 研究了标准蛋白在这两类色谱柱上的保留行为. 这四种WCX色谱柱中的两种能在WCX和HIC两种分离模式下分离蛋白, 虽不如2D色谱柱效果好, 但有可能当成“准2D柱”来使用. 发现蛋白在这四种WCX柱上所显示的HIC分离特征各不相同, 且保留值随盐浓度变化呈现出的“U型”曲线也有大的差异. 实验结果显示, “U型”曲线的宽度和临界点分别与色谱动力学和热力学因素相关. 还对这两类色谱固定相的峰容量表征方法和命名提出了建议和说明.     

Novel Monolithic Stationary Phase with Surface-Grafted Triphenyl Selector for Reversed-Phase Capillary Electrochromatography

A triphenylmethylamine-functionalized monolithic capillary column was newly designed for reversed-phase capillary electrochromatographic applications. Incorporation of the three phenyl rings-containing selector (also referred to as trityl selector) was achieved through post-polymerization functionalization of a generic monolithic matrix bearing nucleophilic-sensitive hydroxysuccinimide moieties. Such a 3D polymer matrix was obtained through UV-induced in situ free radical copolymerization of N-acryloxysuccinimide and ethylene dimethacrylate. The separation properties of the trityl monolithic capillary column were initially evaluated vis-à-vis polycyclic aromatic hydrocarbons, as model hydrophobic compounds, and compared to the separation ability of a benzylamine-functionalized monolithic capillary column prepared using the same generic monolithic matrix. Electrochromatographic separation of phenols and anilines was also considered, and our preliminary results suggest the occurrence of hydrophobic interactions due to the aromatic and non-polar nature of the surface-grafted trityl selector. The triphenyl monolithic capillary column exhibited relative standard deviation values (% RSD) below 4.1 % for the here-studied chromatographic parameters, namely, retention factor, selectivity, resolution, and efficiency.

     

十四烷基胺键合固定相的反相色谱性能

制备了一种新型反相液相色谱固定相──十四烷基胺硅胶键合固定相。以甲醇和水为二元流动相,评价了该固定相的疏水性、选择性和残留硅醇基活性。同时分离了包括碱性、酸性和中性有机化合物在内的混合物。结果表明,在该固定相上,碱性化合物的色谱峰对称性较好,硅酸基活性受到抑制,并具有良好的选择性。     

特殊结构新型液相色谱填料的研究进展

本文对最近报道的几种特殊结构液相色谱填料的制备及应用进行评述,包括新型载碳的核壳结构硅胶色谱填料的制备与应用;新型硅胶基质耐酸超桥联固定相的制备及应用和β-环糊精杂化硅胶整体柱的制备及其在手型分离的应用。     

氮杂冠醚聚硅氧烷固定液的色谱保留机理

氨杂冠醚聚硅氧烷是一类呈中等极性的新颖气相色谱固定液．本文通过测定醇、酯、卤代烃和苯系列化合物的色谱保留值和相应势力学参数，研究它对上述不同化合物的保留机理及其分子识别的热力学性质     

Evaluation of a Doubly Zirconized Silica-Based Stationary Phase for HPLC

This paper describes the preparation, characterization, and application of a chemically bonded and endcapped C18 stationary phase having a doubly zirconized silica support. The stationary phase was characterized using infrared and nuclear magnetic resonance (¹³C and ²⁹Si) spectroscopies, elemental analysis, and surface and thermogravimetric determinations, and evaluated chromatographically using several test mixtures, indicating acceptable efficiency, and asymmetry. The new phase was used for several different applications including the determination of the possible presence of six pesticides in orange juice, using a modified QuEChERS procedure for extraction.

     

Evaluation of a Doubly Zirconized Silica-Based Stationary Phase for HPLC

This paper describes the preparation, characterization, and application of a chemically bonded and endcapped C18 stationary phase having a doubly zirconized silica support. The stationary phase was characterized using infrared and nuclear magnetic resonance (¹³C and ²⁹Si) spectroscopies, elemental analysis, and surface and thermogravimetric determinations, and evaluated chromatographically using several test mixtures, indicating acceptable efficiency, and asymmetry. The new phase was used for several different applications including the determination of the possible presence of six pesticides in orange juice, using a modified QuEChERS procedure for extraction.     

A New Definition of the Stationary Phase Volume in Mixed-Mode Chromatographic Columns in Hydrophilic Liquid Chromatography

Polar columns used in the HILIC (Hydrophilic Interaction Liquid Chromatography) systems take up water from the mixed aqueous–organic mobile phases in excess of the water concentration in the bulk mobile phase. The adsorbed water forms a diffuse layer, which becomes a part of the HILIC stationary phase and plays dominant role in the retention of polar compounds. It is difficult to fix the exact boundary between the diffuse stationary and the bulk mobile phase, hence determining the column hold-up volume is subject to errors. Adopting a convention that presumes that the volume of the adsorbed water can be understood as the column stationary phase volume enables unambiguous determination of the volumes of the stationary and of the mobile phases in the column, which is necessary for obtaining thermodynamically correct chromatographic data in HILIC systems. The volume of the aqueous stationary phase, Vex, can be determined experimentally by frontal analysis combined with Karl Fischer titration method, yielding isotherms of water adsorbed on polar columns, which allow direct prediction of the effects of the composition of aqueous–organic mobile phase on the retention in HILIC systems, and more accurate determination of phase volumes in columns and consistent retention data for any mobile phase composition. The n phase volume ratios of 18 columns calculated according to the new phase convention strongly depend on the type of the polar column. Zwitterionic and TSK gel amide and amine columns show especially strong water adsorption.