Separation of Structurally Related Anthocyanins by MEKC

Summary A method based on micellar electrokinetic chromatography has been developed for the simultaneous separation of six anthocyanins (malvidin-3,5-diglucoside, malvidin-3-glucoside, malvidin-3-galactoside, pelargonidin-3-glucoside, cyanidin-3,5-diglucoside, cyanidin-3-galactoside). Optimum selectivity was achieved in the buffer 30 mM phosphate + 400 mM borate-TRIS, pH=7.0 supported with 50 mM sodium dodecylsulphate. High content of borate was essential mainly for the separation of diastereomeric pair malvidin-3-glucoside-malvidin-3-galactoside. The calibration dependencies exhibit good linearity in the ranges of concentration 10–100 g mL^–1 for diglycosylated and 25–100 g mL^–1 for monoglycosylated derivatives (R² = 0.9711–0.9989). The optimized method was applied to a sample of wine grape skin extract. Malvidin-3-glucoside was identified as main anthocyanin colorant in this sample.     

Separation of Di-dansyl amino acids by MEKC with a co-surfactant

Summary A method has been developed for qualitative determination, in some natural products, of those amino acids which form di-dansyl derivatives. The capillary electrophoresis methods so far developed have not completely solved the problem of separation of di-dansyl amino acids. These amino acids are highly non-polar, because they contain two dansyl groups linked to the amino acid part of the molecule, and are poorly separated from each other, or not separated at all, by micellar electrokinetic chromatography. A new separation mechanism has been tested using a different co-surfactant. Preliminary data have led to 2-methyl-1-propanol being chosen for further investigation of the capabilities of the method of separation.     

Separation of Fluoroquinolones by MEKC Modified with Hydrophobic Ionic Liquid as a Modifier

The hydrophobic ionic liquid [BMIM]PF₆ (1-butyl-3-methylimidazolium hexafluorophosphate) can interact with sodium dodecyl sulfate (SDS) micelles in aqueous solution and modify their physicochemical properties to produce a unique separation efficiency in micellar electrokinetic chromatography (MEKC). An MEKC method was developed using [BMIM]PF₆ as a modifier for separating eight fluoroquinolone compounds (ciprofloxacin, enrofloxacin, gatifloxacin, ofloxacin, norfloxacin, enoxacin, pazufloxacin, and tosufloxacin). The effects of several parameters on the separation selectivity, e.g., pH, concentration of background electrolyte, concentration ratio and amount of [BMIM]PF₆ and SDS, were investigated. Under the optimal conditions of 10 mmol L⁻¹ sodium borate, pH 7.1, 1.7% (w/w) SDS, 1.5% (w/w) [BMIM]PF₆ with 18 kV as running voltage, the eight investigated quinolone compounds were baseline separated within 15 min. The selectivity of the developed method differed from that of the simple SDS micelles system containing no ionic liquid. The results suggest that hydrophobic ionic liquids should be promising modifiers in capillary electrophoresis, especially in MEKC analysis.

     

Separation of Biphenyl Nitrile Compounds by Microemulsion Electrokinetic Chromatography with Mixed Surfactants

A mixture of nine biphenyl nitrile compounds with high hydrophobicity and similar structures was successfully separated by microemulsion electrokinetic chromatography (MEEKC) within 30 rain. The buffer system contained 100 mmol/L sodium dodecyl sulfate (SDS), 80 mlnol/L sodium cholate (SC), 0.81% heptane, 7.5% n-butanol, 10% acetonitrile and 10 mmol/L borate. The addition of SC, organic modifiers, sample preparation and temperature all showedremarkable effect on the separation. Meanwhile, the MEEKC method was briefly compared with micellar electrokinetic chromatography (MEKC) method.     

RNA analysis by MEKC with LIF detection

We have developed and validated a procedure of high sensitivity for the analysis of RNA. The procedure is based on the separation and detection of the 5'-monophosphates of ribonucleosides selectively conjugated with 4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacene-3-propionyl ethylene diamine hydrochloride (BODIPY FL EDA) at the 5'-phosphate group using CE with LIF. BODIPY conjugates of the four common ribonucleoside-5'-monophosphates were prepared and subjected to CE-LIF to serve as standard compounds for peak assignment and to develop separation conditions. After digestion of RNA or oligoribonucleotides to 5'-monophosphates by nuclease P1 and fluorescence labeling BODIPY conjugates were detected and resolved by CE-LIF without further purification steps. Comparative CE-LIF analyses with DNA digested to deoxyribonucleoside-5'-monophosphates showed that the assay is equally efficient and sensitive for RNA analysis. Conditions to determine the modified ribonucleosides inosine, xanthosine, pseudouridine and 2'-O-methyladenosine were also established. The limits of detection were in the range of 80-200 pM. After calibrating the assay with oligoribonucleotides, pseudouridine was quantified in total RNA of Drosophila, human liver, human kidney and t-RNA of Saccharomyces cerevisiae. These studies demonstrate good potential of fluorescence labeling of ribonucleoside-5'-monophosphates with BODIPY FL EDA and detection by CE-LIF to determine RNA composition with high accuracy and sensitivity.     

Study of Spontaneous Imbibition of Water by Oil-Wet Sandstone Cores Using Different Surfactants

The mechanism of spontaneous imbibition of water by sandstone cores and the relationship between reservoir wettability and imbibition recovery were studied by investigating factors influencing the spontaneous imbibition of different surfactants by oil-wet sandstone cores. Ultimate oil recovery of cores using the cationic surfactant CTAB was higher than that of the cores using the nonionic surfactant TX-100 and the anionic surfactant POE (1) at the same concentration. For CTAB and TX-100, the ultimate oil recovery by spontaneous imbibition increased with increase in surfactant concentration. In regard to imbibition recovery, TX-100 and POE(1) at high temperatures were superior to those at low temperatures. Ultimate oil recovery of the high-permeability core was higher than that of the low-permeability core at room temperature. According to changes in the driving force during the imbibition process, the imbibition curve could be divided into three regions: (1) mainly capillary force, (2) both capillary and gravity forces, and (3) mainly gravity force. The stronger the hydrophilicity of the rock surface, the higher the spontaneous imbibition recovery.     

三正辛胺络合萃取法分离提取糖果中8种合成色素

以三正辛胺为络合剂,四氯化碳为稀释剂,采用络合萃取法对糖果中的柠檬黄、新红、苋菜红、胭脂红、日落黄、诱惑红、亮蓝和赤藓红8种合成色素进行了预处理研究。考察了样品溶液p H值、稀释剂种类、三正辛胺浓度、反萃相组成等对预处理效果的影响。结果表明,当样品溶液p H值为6.0,以四氯化碳为稀释剂,三正辛胺浓度为5%,无水乙醇-15%氨水(体积比1∶1)为反萃相时,8种合成色素的回收率为90%~98%。该预处理方法操作简单、快速、有机溶剂用量少、回收率高,适用于糖果中多种合成色素的分离提取。     

烷基苯磺酸盐型表面活性剂的HPLC及HPCE分离分析进展*

作为一种常见的表面活性剂,烷基苯碘酸盐（ABS）被广泛用于许多领域。本文结合作者最近的研究结果,对烷基苯磺酸盐表面活性剂的HPLC及HPCE分离分析现状及其应用前景进行了综述。     

Study of recombinant cytochrome P450 2C9 activity with diclofenac by MEKC

Cytochrome P450 2C9 (CYP2C9) is one of the most important isoforms in human liver involved in the metabolism of a large number of therapeutic agents. The aim of this paper is to demonstrate the applicability of CE for the determination of the enzymatic activity of CYP2C9 with diclofenac as a probe substrate. MEKC with SDS as a pseudostationary phase was used for this purpose. Compared to other assays, the MEKC-based method is rapid, can be automated and requires only a small quantity of enzymes and substrate. Moreover, the enzymatic reaction can be monitored with high sensitivity and repeatability even when the reaction mixture is used for the analysis without any pretreatment. The kinetic study on the given enzymatic reaction was also performed since the basic characterization of drug biotransformation generally begins with the enzyme kinetic analysis of metabolite formation. As a result, the Michaelis constant and maximum reaction velocity were evaluated, the values 3.44 +/- 0.45 microM and 19.78 +/- 0.76 nmol min(-1) nmol(-1), respectively, were in agreement with the literature data. On the other hand, a slight deviation from typical Michaelis-Menten kinetics with a weak positive cooperativity was found at diclofenac concentrations below 2 microM. The same atypical kinetic behavior of CYP2C9 was also observed by other authors.     

Chromatographic Separation of Synthetic Isoflavones on Stannic Molybdate Papers

Abstract

Chromatographic separation of 24 synthetic isoflavones on stannic molybdate papers has been described. Various analytically and biochemically important separations, such as separation of isomers and that of having same molecular weights have been achieved. Out of different solvent systems used aqueous methanol was found to be the best eluent.     

季铵盐表面活性剂引起非水混合溶剂的分相

当季铵盐表面活性剂在极性溶剂中具有合适溶解度时, 它可使该极性溶剂与另一种非极性溶剂形成的混合溶剂自发分相, 形成稳定的两相界面. 借鉴表面活性剂双水相现象的概念, 这种现象被称为表面活性剂非水双相(NSTP), 也可简称为双油相. 产生此种现象的原因被归结为季铵盐表面活性剂在极性溶剂中达到合适的溶解度时, 从而逐出与之不亲合的非极性溶剂.     

Separation and determination of biogenic amines in fish using MEKC with novel multiphoton excitation fluorescence detection

Biogenic amines are a group of biological molecules derived from the enzymatic decarboxylation of natural amino acids. They can be found in a variety of foods and some of them are involved in essential cellular pathways regulating cellular functions. To address the issues raised by conventional detection methods for biogenic amines, such as laborious sample preparation and limited sensitivity, a new micellar electrokinetic chromatography scheme was developed based on multiphoton excitation fluorescence (MPEF) detection. Six FITC-labeled biogenic amine species were used for the evaluation of this MEKC-MPEF method in comparison to single photon excitation fluorescence detection. The results indicated that MEKC-MPEF had superior resolution with a detection volume as low as aL. Quantitative analysis of varying concentrations of biogenic amine species has also been achieved suggesting a ymole mass detection limit, a linear dynamic range of about two orders of magnitude, and 95-105% recoveries. Furthermore, the biogenic amine profile of decayed oriental crucian carps was successfully determined and quantified using this new method.     

Protein Binding Study of Isoflavones by High-Performance Frontal Analysis

High-performance frontal analysis was used for protein binding study of isoflavones (daidzein, genistin, and genistein) to human serum albumin. The analysis was performed on a Develosil 100-Diol-5 column (10 cm × 4.6 mm). Sodium phosphate solution (pH 7.4, ionic strength 0.17) was used as the mobile phase at a flow rate of 1 mL min^–1. UV wavelength was set at 260 nm. To ensure the drug to be eluted as a trapezoidal peak with a plateau, injection volumes of 700 L for daidzein and genistin, 900 L for genistein were chosen, respectively. Experimental data were fitted by Scatchard equation. The binding constants (K) and binding affinities (nK) of isoflavones to HSA were: K=1.581 × 10⁵ (L mol^–1), nK=0.77 × 10⁴ (L mol^–1) for daidzein, K=1.082 × 10⁵ (L mol^–1), nK=0.32 × 10⁴ (L mol^–1) for genistein, and K=3.533 × 10⁵ (L mol^–1), nK=0.70 × 10⁴ (L mol^–1) for genistin, respectively.     

Determination of triptonide by cloud point extraction combined with MEKC

A method has been developed for the determination of triptonide in the traditional Chinese herb Tripterygium wilfordii Hook F. by micellar electrokinetic capillary chromatography combined with cloud point extraction. The analyte was extracted at pH 3.0 by micelles of the nonionic surfactant polyoxyethylene 7,5-octylphenyl ether (Triton X-114). A 250-muL aliquot from the extracted surfactant-rich phase was diluted to 400 muL with ethanol to reduce its viscosity before separation by MEKC. Under optimum conditions, an enrichment factor of 25 is obtained and the determination limit of triptonide is found to be 3.15 x 10(-7) mol/L. The proposed method has been successfully applied to the determination of triptonide in T. wilfordii tablet and spiked urine matrix, demonstrating the feasibility and reliability of the proposed method.     

Separation of cold medicine ingredients using a precise MEKC method at elevated pH

An MEKC method was developed in order to separate a cold medicine formulation containing acetaminophen, ephedrine sulfate, doxylamine succinate, and dextromethorphan hydrobromide as active pharmaceutical ingredients. Because of their similar physical and chemical properties, it was a challenge to separate the basic compounds without sample pretreatment. In addition, the high content of alcohol and sucrose together with the variety of further excipients had to be considered. Thus, the complex matrix required several optimization steps. These included the search for the optimum pH and for a suitable sodium dodecyl sulfate concentration to avoid matrix-capillary wall interaction and to ensure precision. As a second developing step, an internal standard (benzocaine) was chosen to guarantee a high level of quantitative performance. An RSD% value of the peak areas between 1.0 and 2.0 was reached. The employed method development strategy can be generalized to similar separation approaches in the future.     

1-Hexadecyl-3-methylimidazolium Ionic Liquid as a New Cationic Surfactant for Separation of Phenolic Compounds by MEKC

The ionic liquid 1-hexadecyl-3-methylimidazolium bromide ([C₁₆MIm]Br) has been used as a novel cationic surfactant for separation of phenolic compounds, including quinol, phloroglucinol, resorcinol, phenol, p-cresol, and m-nitrophenol, by micellar electrokinetic capillary chromatography (MEKC). The effects of buffer concentration and pH, concentration of [C₁₆MIm]Br, and applied potential were studied. Use of the optimized buffer (25 mmol L^?1 NaH₂PO₄), 10 mmol L^?1 [C₁₆MIm]Br, and an applied potential of ?15 kV enables optimum separation with regard to resolution and migration time. The phenolic compounds were detected at 214 nm. The micelle of this long-alkyl-chain imidazolium ionic liquid acts as a pseudo-stationary phase in this MEKC separation.     

Determination of Dexamethasone in Cosmetics by MEKC

A rapid and reliable method based on micellar electrokinetic capillary chromatography has been developed for the determination of dexamethasone in cosmetics. Effects of buffer composition, concentration and pH, the detection wavelength, separation voltage, and injection time were systematically investigated. The optimum conditions were: 30 mM borax buffer containing 20 mM sodium dodecyl sulfate at pH 9.0, detection at 254 nm, injection time 10 s at a height of 10 cm, and a separation voltage of 15 kV. Under these conditions, the analysis of dexamethasone in cosmetics was carried out within 6 min. The method was validated for stability, precision, linearity and accuracy. Excellent linearity was obtained in the range of 50–1,000 μg mL⁻¹, and acceptable precision, in intra-day and inter-day analysis, was also obtained with relative standard deviation in the range of 0.19–0.86 and 2.50–4.90% for migration time and peak area ratio, respectively. The method was used to analyse eight cosmetic samples purchased locally.     

Analytical characterization of PEG polymers by MEKC

Characterization of PEGs with average molecular masses of up to 2000 has been achieved using MEKC with UV detection. A rapid derivatization procedure with phenyl isocyanate using microwave radiation, in order to introduce chromophore groups in PEGs, has been developed involving a reaction time of 60 s. Different optimized conditions in accordance with the molecular weight have been studied to obtain the oligomer separation. The weight‐average molecular mass the number‐average molecular mass and the degree of polydispersity (molecular mass distribution) were calculated for the different PEGs obtaining similar results with those certified for standards. A good precision was obtained for characterizing the different oligomers. Ethylene glycol was used as the internal standard for the analysis of low‐molecular‐weight PEGs. The developed method was satisfactorily applied to the characterization of these polymers in several real samples, such as lubricant eye drops, toothpaste, tap water and eye make‐up remover.