Evaluation of a silicon oxynitride hydrophilic interaction liquid chromatography column in saccharide and glycoside separations

The retention characteristics of a silicon oxynitride stationary phase for carbohydrate separation were studied in hydrophilic interaction chromatography mode. Four saccharides including mono‐, di‐, and trisaccharides were employed to investigate the effects of water content and buffer concentration in the mobile phase on hydrophilic interaction liquid chromatography retention. For the tested saccharides, the silicon oxynitride column demonstrated excellent performance in terms of separation efficiency, hydrophilicity, and interesting separation selectivity for carbohydrates compared to the bare silica stationary phase. Finally, the silicon oxynitride hydrophilic interaction liquid chromatography column was employed in the separation of complex samples of fructooligosaccharides, saponins, and steviol glycoside from natural products. The resulting chromatograms demonstrated good separation efficiency and longer retention compared with silica, which further confirmed the advantages and potential application of silicon oxynitride stationary phase for hydrophilic interaction liquid chromatography separation.     

Elucidation of steroid glycosides in Anemarrhena asphodeloides extract by means of comprehensive two-dimensional reversed-phase/polyamine chromatography with mass spectrometric detection

In the work presented here, a novel approach to comprehensive two-dimensional liquid chromatography is evaluated. Reversed-phase liquid chromatography was employed for the first-dimension separation and polyamine chromatography was chosen for the second-dimension separation mode. The two dimensions are highly orthogonal and the separation efficacy of the developed octadecylsilica × polyamine was tested by separating an extract from Anemarrhena asphodeloides. The steroid glycosides identified by comprehensive two-dimensional liquid chromatography in this experiment were compared to those obtained for monodimensional liquid chromatography. The comprehensive two-dimensional liquid chromatography system, thanks to the complementary separation selectivity and enhanced peak capacity provided by the two columns, allowed to distribute five compounds of low amounts otherwise unachievable by monodimensional liquid chromatography. In addition, four steroid isomers with similar fragmentation characteristics in MS/MS spectra, were newly separated based on their different chemical structures.     

Off‐line comprehensive two‐dimensional reversed‐phase countercurrent chromatography with high‐performance liquid chromatography: Orthogonality in separation of Polygonum cuspidatum Sieb. et Zucc

Off‐line comprehensive two‐dimensional reversed‐phase countercurrent chromatography with high‐performance liquid chromatography was investigated in separation of crude ethanol extract from traditional Chinese medicinal herb Polygonum cuspidatum Sieb. et Zucc. Two‐dimensional contour plots for countercurrent chromatography with high‐performance liquid chromatography was obtained after comprehensive separation was completed. Total peak capacity was evaluated and approximately 810 peaks were obtained through a comprehensive two‐dimensional separation. A highly orthogonality of 52.23% and a large separation space occupancy of 88.86% were achieved. Meanwhile, it was found that several components could be well separated by countercurrent chromatography while they could not be separated by high‐performance liquid chromatography, and vice versa, which further indicated the orthogonality of the two separation methods. The off‐line comprehensive two‐dimensional countercurrent chromatography with high‐performance liquid chromatography provided a promising and powerful method for separation of complex natural products.     

Application of Thin-Layer Chromatography to High-Performance Liquid Chromatographic Separation of Steroidal Hormones and Cephalosporin Antibiotics

Abstract

High-performance liquid chromatographic (HPLC) separation of steroidal hormones and cephalosporin antibiotics was investigated by adsorption chromatography and reversed-phase chromatography, respectively.

Prior to the HPLC separation of these pharmaceuticals, silica gel thin-layer adsorption chromatography of steroidal hormones and reversed-phase thin-layer partition chromatography of cephalosporin antibiotics with chemically bonded dimethylsilyl silica gel were performed in order to obtain suitable HPLC separation systems.

In the separation of steroidal hormones, the same binary mobile phase ratios of TLC did not give satisfactory results in HPLC. For the sharp separation in HPLC, solvent strength in the binary solvent mixture used for TLC had to be decreased.

The difference in solvent strength for efficient separation between TLC and HPLC might be attributed to the fact that in HPLC the solvent elution power acts in an isocratic manner while in TLC it acts in a gradient manner.

On the other hand, a correlation of mobility between TLC and HPLC separation for cephalosporin antibiotics was obtained, and the possibility of direct transfer of chromatographic systems from TLC to HPLC for separation of these antibiotics was confirmed.     

Opportunities and challenges of high-pressure ion exchange chromatography for nuclide separation and enrichment

With the rapid development of the nuclear industry, more-stringent requirements are proposed for high-level radioactive waste liquid treatment and the enrichment of isotope products. High-pressure ion exchange chromatography has been widely accepted for the fine separation of elements and nuclides due to its advantages, such as high efficiency, environmental friendliness, ease of operation, and feasibility for large-scale industrial applications. Here, we summarized the evolution of high-pressure ion exchange chromatography and the relevant research progress in ion exchange equilibrium and related separation technology. The prospects for application of high-pressure ion exchange chromatography to rare earth elements, actinide elements and isotope separation were discussed. High-pressure ion exchange chromatography represents a promising strategy for the extraction of rare earth elements and actinide elements from high-level radioactive waste liquid, as well as being an effective method for the automated production of high purity isotope products with great environmental benefits.     

Two dynamic chromatographies and their application

Hydrodynamic chromatography and slalom chromatography known as dynamic liquid chromatography were introduced and reviewed, mainly for the recent development of separation principle, theoretical model, and applications. Fifty-two references were cited.     

流体动力色谱法和障碍色谱法及其应用

介绍了流体动力色谱(HDC)和障碍色谱(SC)及其在生物、化工分离中的研究进展,着重于它们的分离原理、理论发展及二者之间的联系与转化,引用52篇文献.     

离子色谱-非抑制电导法同时测定铵盐、四乙基铵、甲基三乙基铵

建立了离子色谱非抑制电导法同时分离测定铵根与两种季铵盐四乙基铵、甲基三乙基铵的方法。分别实验了在亲水性和疏水性阳离子交换色谱柱上三种铵类的分离效果,研究了使用不同淋洗液和流速情况下离子的分离情况,结果表明使用SH-Cation-101型疏水性阳离子色谱柱,淋洗液采用甲烷磺酸(5.0mmol/L),其中加入乙腈(7%),于0.8mL/min的流速条件下,三种铵类物质分离良好,其中结构极为相似的两种季铵盐四乙基铵和甲基三乙基铵分离度达到1.5以上,分离时间短,3种物质在13min内实现完全分离。采用国产离子色谱仪非抑制电导法检测,无需使用抑制器,成本低,操作简便可行。检测结果的灵敏度高,线性范围铵根为0.5～50mg/L,四乙基铵和甲基三乙基铵为5～500mg/L,相关系数均高于0.999,相对标准偏差均在3%以内,平均加标回收率在98.5%～101.2%。     

径向展开薄层色谱法分离生物碱及其分离效能

径向展开薄层色谱法是一种将样品由中心沿径向向外展开的简便、快速、高效的色谱方法。该文组装了简单的径向展开薄层色谱装置,并建立了朱砂安神丸的径向展开薄层色谱检测法,对其中的生物碱成分进行分离,研究了径向展开薄层色谱的分离特性。从薄层色谱基础理论出发,对径向展开薄层色谱和一般薄层色谱的分离效能进行了比对研究,设计试验进行计算和求解。证明了径向展开薄层色谱法更快、更高效、更经济,适用于生物碱等高极性样品分离。探索了径向展开薄层色谱法高分离效率的理论根源,这一研究思路也为理论创新提供了新的方法和思路。     

A comparative study of chiral separation of proton pump inhibitors by supercritical fluid chromatography and high-performance liquid chromatography

A comparative study of chiral separation of pantoprazole and rabeprazole is carried out using supercritical fluid chromatography and high-performance liquid chromatography. The columns used were Chiralpak IA and Chiralpak IE. The best mobile phase in supercritical fluid chromatography was carbon dioxide-0.2% triethylamine in methanol (60:40) and 0.1% triethylamine in n-hexane-ethanol (50:50) in high-performance liquid chromatography. For supercritical fluid chromatography, values of the retention factor of pantoprazole enantiomers were 3.97 and 4.88. These values for rabeprazole enantiomers were 6.10 and 7.52. The values of separation and resolution factor for pantoprazole and rabeprazole were 1.23 and 1.23 and 2.20 and 3.36, respectively. Similarly, for high-performance liquid chromatography, the values of retention factor for enantiomers of pantoprazole were 4.02 and 7.32. These values for rabeprazole enantiomers were 5.32 and 7.88, respectively. The values of separation and resolution factor for pantoprazole and rabeprazole were 1.82 and 1.48 and 9.22 and 6.58, respectively. A comparison was carried out, which confirmed supercritical fluid chromatography as the best method due to its fastness, eco-friendly, and inexpensiveness. The reported methods are effective, efficient, and reproducible and may be used to separate and identify pantoprazole and rabeprazole in any unknown samples.     

First separation of four aromatic acids and two analogues with similar structures and polarities from Clematis akebioides by high‐speed counter‐current chromatography

In this paper, we report an efficient method by high‐speed counter‐current chromatography for the first separation of four aromatic acids and two analogs with similar structures and polarities from Clematis akebioides. First, the ethyl acetate extract was treated by silica gel column chromatography to enrich the target compounds. And then the fraction with target compounds were purified by high‐speed counter‐counter chromatography using a two‐phase solvent system consisting of chloroform/acetonitrile/water (10:6:4, v/v). The results showed high‐speed counter‐current chromatography could be a powerful technology for the separation of compounds with similar structures and polarities. Besides, it was found acetonitrile could be a good methanol substitute when a chloroform/methanol/water system could not provide a good separation factor. This study provides a reference for the separation of compounds from Clematis akebioides.     

A practicable strategy for enrichment and separation of four minor flavonoids including two isomers from barley seedlings by macroporous resin column chromatography,medium‐pressure LC,and high‐speed countercurrent chromatography

Separation of minor compounds especially with similar polarities and structures from complex samples is a challenging work. In the present study, an efficient method was successfully established by macroporous resin column chromatography, medium‐pressure liquid chromatography, and high‐speed countercurrent chromatography for separation of four minor flavonoids from barley seedlings. Macroporous resin column chromatography and medium‐pressure liquid chromatography were used for enrichment of these four flavonoids. High‐pressure liquid chromatography analysis showed the total content of these four flavonoids increased from 2.2% in the crude extract to 95.3% in the medium‐pressure liquid chromatography fraction. It was indicated that the combination of macroporous resin column chromatography and medium‐pressure liquid chromatography could be a practicable strategy for enrichment of minor compounds from complex sample. Then, high‐speed countercurrent chromatography was employed for separation of these four flavonoids using ethyl acetate/n‐butanol/water (0.1% glacial acetic acid) (4:1:5, v/v/v) as solvent system. As a result, four flavonoids including two isomers with purities higher than 98% were obtained. Interestingly, two flavonoids existing in one high‐pressure liquid chromatography peak were also successfully separated. All these indicated high‐speed countercurrent chromatography had great potential for separation of compounds with similar structures and polarities. This study provides a reference for efficient enrichment and separation of minor compounds from complex sample.     

Comparison of ultra high performance supercritical fluid chromatography,ultra high performance liquid chromatography,and gas chromatography for the separation of synthetic cathinones

A comparison of ultra high performance supercritical fluid chromatography, ultra high performance liquid chromatography, and gas chromatography for the separation of synthetic cathinones has been conducted. Nine different mixtures of bath salts were analyzed in this study. The three different chromatographic techniques were examined using a general set of controlled synthetic cathinones as well as a variety of other synthetic cathinones that exist as positional isomers. Overall 35 different synthetic cathinones were analyzed. A variety of column types and chromatographic modes were examined for developing each separation. For the ultra high performance supercritical fluid chromatography separations, analyses were performed using a series of Torus and Trefoil columns with either ammonium formate or ammonium hydroxide as additives, and methanol, ethanol or isopropanol organic solvents as modifiers. Ultra high performance liquid chromatographic separations were performed in both reversed phase and hydrophilic interaction chromatographic modes using SPP C18 and SPP HILIC columns. Gas chromatography separations were performed using an Elite‐5MS capillary column. The orthogonality of ultra high performance supercritical fluid chromatography, ultra high performance liquid chromatography, and gas chromatography was examined using principal component analysis. For the best overall separation of synthetic cathinones, the use of ultra high performance supercritical fluid chromatography in combination with gas chromatography is recommended.     

Mixed‐mode chromatography integrated with high‐performance liquid chromatography for protein analysis and separation: Using bovine serum albumin and lysozyme as the model target

A type of mixed‐mode chromatography was integrated with high‐performance liquid chromatography for protein analysis and separation. The chromatographic behavior was tested using bovine serum albumin and lysozyme as model proteins. For the mixed‐mode column, the silica beads were activated with γ‐(2,3‐epoxypropoxy)‐propytrimethoxysilane and coupled with 4‐mercaptopyridine as the functional ligand. The effects of pH, salt, and the organic solvent conditions of the mobile phase on the retention behavior were studied, which provided valuable clues for separation strategy. When eluted with a suitable pH gradient, salt concentration gradient, and acetonitrile content gradient, the separation behavior of bovine serum albumin and lysozyme could be controlled by altering the conditions of the mobile phase. The results indicated this type of chromatography might be a useful method for protein analysis and separation.     

Efficient strategies for preparative separation of iridoid glycosides and flavonoid glycosides from Hedyotis diffusa

Efficient strategies for the preparative separation of iridoid glycosides and flavonoid glycosides from Hedyotis diffusa using preparative high-performance liquid chromatography combined with appropriate pretreatment technologies were developed. Four fractions (Fr.1-1, Fr.1-2, Fr.1-3, and Fr.2-1) were firstly isolated from the crude extract of Hedyotis diffusa by column chromatography with C18, resin, and silica gel materials, respectively. Then, corresponding separation strategies were developed according to the polarity and chemical constituents. High-polar compounds of Fr.1-1 were purified by hydrophilic reversed-phase liquid chromatography and hydrophilic interaction liquid chromatography mode. The combination of C18 and phenyl columns realized the complementary separation of iridoid glycosides in Fr.1-2. Meanwhile, the improved selectivity caused by the change of organic solvent in the mobile phase was utilized to realize the purification of flavonoid glycosides in Fr.1-3 and Fr. 2-1. Finally, 27 compounds (purity > 95%) mainly involving nine iridoid glycosides and five flavonoid glycosides were obtained. A complete strategy was established for the separation of a complex sample with a wide polarity range, to jointly solve the problems of enrichment of target components and separation of structural analogs.     

Analysis of insecticidal proteins from Bacillus thuringiensis and recombinant Escherichia coli by capillary electrokinetic chromatography

Bacillus thuringiensis and recombinant Escherichia coli proteinaceous protoxins were subject to proteolysis and analyzed by capillary electrokinetic chromatography. Three resulting toxins (65 kDa) were baseline-resolved within 22 min using a 10 mM borate, pH 11 separation buffer consisting of 25 mM sodium dodecyl sulfate (SDS) and 30 mM phytic acid. The toxins displayed differential interactions with the SDS and phytic acid phases to effect their separation. The ion-pairing interaction between the analyte and phytic acid was also useful in preventing adsorption to the capillary walls and thus enhanced separation resolution and efficiency. The use of electrokinetic chromatography allows achievement of the separation in a significantly shorter time than conventional high-performance liquid chromatography (HPLC) using a diethylaminoethyl (DEAE) weak-anion exchanger.     

An effective method based on medium‐pressure liquid chromatography and recycling high‐speed counter‐current chromatography for enrichment and separation of three minor components with similar polarity from Dracocephalum tanguticum

The separation of minor compounds, especially those with similar polarities from a complex sample, remains challenging. In the proposed study, an effective method based on medium‐pressure liquid chromatography and recycling high‐speed counter‐current chromatography was developed for the enrichment and separation of three minor components from Dracocephalum tanguticum. The crude extract was directly introduced to medium‐pressure liquid chromatography for the enrichment of the three minor components. Based on high‐performance liquid chromatography analysis, the total content of these three compounds increased from 0.48% in the crude extract to 85.3% in the medium‐pressure liquid chromatography fraction. In addition, high‐speed counter‐current chromatography was employed to separate the enriched compounds using the solvent system hexane/ethyl acetate/methanol/water (1.18:8.82:1.18:8.82, v/v/v/v). As a result, compound 3 and a mixture of compounds 1 and 2 were obtained. In order to improve the resolution of compounds 1 and 2 while saving separation time, a recycling and heart‐cut mode was used. Finally, compounds 1 and 2 were obtained after five cycles. These compounds were identified as 3‐phenylethyl β‐d ‐glucopyranoside ( 1 ), tazettoside E ( 2 ), and cirsiliol‐4′‐glucoside ( 3 ). Compounds 1 and 2 were primarily separated from D. tanguticum. Moreover, the developed method provided a reference for the separation of minor components from the complex sample.     

Hybridation of different chiral separation techniques with ICP-MS detection for the separation and determination of selenomethionine enantiomers: chiral speciation of selenized yeast

Enantioseparation and determination of selenomethionine enantiomers in selenized yeast was investigated using chiral separation techniques based on different principles, coupled on-line to inductively coupled plasma mass spectrometry (ICP-MS) for selenium-specific detection. High performance liquid chromatography (HPLC) on a beta-cyclodestrin (beta-CD) column, cyclodextrin-modified micellar electrokinetic chromatography (CD-MEKC), gas chromatography (GC) on a Chirasil-L-Val column, and HPLC on a Chirobiotic T column have been investigated as the chiral separation techniques. For HPLC separation on the beta-CD column, and also for CD-MEKC, selenomethionine enantiomers were derivatized with NDA/CN(-). For chiral separation by GC, selenomethionine enantiomers were converted into their N-trifluoroacetyl (TFA)-O-alkyl esters. The developed hybridation methodologies are compared with respect to enantioselectivity, sensitivity and analysis time. The usefulness of the best-suited method [HPLC (Chirobiotic T)-ICP-MS] was demonstrated by its application to the successful chiral speciation of selenium and D-and L-selenomethionine content determination in selenized yeast.     

Determination of Antiviral Drugs and Their Metabolites Using Micro-Solid Phase Extraction and UHPLC-MS/MS in Reversed-Phase and Hydrophilic Interaction Chromatography Modes

Two new ultra-high performance liquid chromatography (UHPLC) methods for analyzing 21 selected antivirals and their metabolites were optimized, including sample preparation step, LC separation conditions, and tandem mass spectrometry detection. Micro-solid phase extraction in pipette tips was used to extract antivirals from the biological material of Hanks balanced salt medium of pH 7.4 and 6.5. These media were used in experiments to evaluate the membrane transport of antiviral drugs. Challenging diversity of physicochemical properties was overcome using combined sorbent composed of C₁₈ and ion exchange moiety, which finally allowed to cover the whole range of tested antivirals. For separation, reversed-phase (RP) chromatography and hydrophilic interaction liquid chromatography (HILIC), were optimized using extensive screening of stationary and mobile phase combinations. Optimized RP-UHPLC separation was carried out using BEH Shield RP18 stationary phase and gradient elution with 25 mmol/L formic acid in acetonitrile and in water. HILIC separation was accomplished with a Cortecs HILIC column and gradient elution with 25 mmol/L ammonium formate pH 3 and acetonitrile. Tandem mass spectrometry (MS/MS) conditions were optimized in both chromatographic modes, but obtained results revealed only a little difference in parameters of capillary voltage and cone voltage. While RP-UHPLC-MS/MS exhibited superior separation selectivity, HILIC-UHPLC-MS/MS has shown substantially higher sensitivity of two orders of magnitude for many compounds. Method validation results indicated that HILIC mode was more suitable for multianalyte methods. Despite better separation selectivity achieved in RP-UHPLC-MS/MS, the matrix effects were noticed while using both chromatographic modes leading to signal enhancement in RP and signal suppression in HILIC.     

薄层色谱综合分离程度的定量量度

薄层色谱对多元组分的分离程度,目前常用信息量~[1]来度量.但由于计算信息量时需人为地固定R_f值的分段,且认为同一区段内的R_f值等同,而不考虑每相邻两组分的R_f差值,也忽略了同一R_f区段内不同R_f值的细微差异.同时,由于薄层色谱分离的信息量并非连续函数,它只能取很有限的若干个分立值.因此,导致有时相同的信息量值却对应着相去甚远