Separation and determination of flavone and xanthone glycosides in Tibetan folk medicinal species Swertia mussotii and S. franchetiana by capillary electrophoresis

The contents of five pharmacologically active flavone and xanthone glycosides, namely, swertianolin, swertisin, isoorientin, mangiferin, and 7-O-[α-L-rhamnopyranosyl-(1 → 2)-β-D-xylopyranosyl]-1,8-dihydroxy-3-methoxyxanthone, extracted from Tibetan folk medicinal species Swertia mussotii and S. franchetiana were determined by capillary electrophoresis with diode-array detection. The separation of five components has been optimized with a capillary column with a total length of 48.5 cm and effective length of 40 cm (50 μm i.d). The influence of the running buffer, the sodium dodecyl sulfonate (SDS) concentration, organic modifier, etc. on the resolution was evaluated. The background electrolyte contained 30 mM borate buffer, 28 mM SDS, 1.0% (v/v) acetonitrile, and was adjusted to pH 9.0 with 0.1 M NaOH. A good baseline resolution was obtained for the separation of five components within 5 min with the working voltage of 24 kV and a column temperature of 25°C. The established method was rapid and reproducible for the separation and determination of five flavone and xanthone glycosides from the extracts of S. mussotii and franchetiana plant samples. The text was submitted by the authors in English.     

Simultaneous Separation and Determination of Benzoic Acid Compounds in the Plant Medicine by High Performance Capillary Electrophoresis

A simple and inexpensive high performance capillary electrophoresis (HPCE) was applied to separate five benzoic acid compounds simultaneously. The investigation was carried out by micellar electrokinetic capillary chromatography (MECC). To avoid a time‐consuming and tedious procedure, orthogonal experimental design OA₉ (3⁴) for separation experiments was applied to find the optimal conditions in terms of the resolution and analytical time. The best conditions for separation were obtained using a 20 mM borax and 30 mM sodium dodecyl sulfate (SDS) buffer (pH 9.8) containing 2 mM β‐CD and 4% methanol (v/v). Online UV detection was performed at 250 nm. A voltage of 16 kV was applied and the temperature was controlled at 25 °C. Injection was performed for 5 s. The method was validated for the quantification of benzoic acid, salicylic acid and ortho‐aminobenzoic acid in Radix Isatidis, a traditional plant medicine with removal of endotoxin. The separation and determination were satisfactory and quick.     

Rapid Separation of Antioxidants in Food Samples by Coelectroosmotic CE

Rapid separation of a group of eight antioxidants by coelectroosmotic capillary electrophoresis and their preliminary determination in foods (cereal, wine, and beer) is described. The compounds studied were protocatechuic acid, salicylic acid, p-hydroxybenzoic acid, vanillic acid, syringic acid, p-coumaric acid, ferulic acid, and sinapic acid. The best separation was achieved by use of a running buffer consisting of 125 mM boric acid, 49 mM disodium hydrogen phosphate, 0.002% (w/v) hexadimethrine bromide, and 2.5 mM α-cyclodextrin, at pH 7.5; the analysis time was less than 3.5 min. Ligration time and peak area reproducibility, studied on the same day (n=4) and on three different days (n=12), showed the method is reproducible. A study of the antioxidant content of cereal, wine, and beer samples was conducted to evaluate the usefulness of the method. Antioxidants were extracted from cereal samples by sonication with methanol and from wine and beer samples by solid-phase extraction with C₁₈ cartridges. This preliminary study showed that seven of the eight compounds could be detected in wine and cereal samples whereas only five were detected in beer samples.     

Determination of Protocatechuic Aldehyde, Danshensu, Salvianolic Acid B and Gallic Acid in Chinese Medicine 'SHUANGDAN' Granule by MEKC

A simple, accurate and effective micellar electrokinetic capillary chromatography (MEKC) method for the separation and determination of protocatechuic aldehyde (PA), danshensu (DSS), salvianolic acid B (SAB) and gallic acid (GA) in Chinese medicine 'SHUANGDAN' granule was developed in this work. The separation was carried out in a 50.0 cm (42.0 cm to the detector) × 75 μm i.d. fused-silica capillary using the optimum buffer solution containing 5.0 mM borate, 15 mM phosphate, 35 mM SDS and 10% (v/v) acetonitrile. The linear relationships between the concentration of the analytes and the corresponding peak areas were investigated, and excellent linear behavior over the investigated concentration ranges was revealed (R²: 0.9985 for PA, 0.9994 for DSS, 0.9989 for SAB and 0.9991 for GA, respectively). The proposed method was successfully applied to the determination of these four active components contained in 'SHUANGDAN' granule. The recoveries of these four components were 97.6–101.2%, 97.6–100.5%, 96.9–101.6% and 97.6–102.9%, respectively.     

Optimisation of a Headspace Solid-Phase Micro- Extraction Procedure for the Determination of 2,4,6-Trichloroanisole and Various Related Compounds in Cork Washing Waste Water by Use of Gas Chromatography-Mass Spectrometry

This paper reports the use of an anionic cyclodextrin, heptakis(2,3-di-O-methyl-6-O-sulfato)-β-cyclodextrin (HDMS-β-CD), for chiral separations of pharmaceutical enantiomers by nonaqueous capillary electrophoresis (NACE). Enantiomer resolution was affected mainly by HDMS-β-CD concentration and the acidity of the background electrolyte (BGE). The effects of capillary length and applied voltage on enantiomer resolution were also investigated. Results showed that in a methanol solution of 20 mM phosphoric acid, 10 mM sodium hydroxide, and 10 mM HDMS-β-CD, seven anticholinergic drugs were separated to baseline but no chiral separation was obtained for three other similar drugs. NACE is suitable for routine, rapid separation of the enantiomers of pharmaceutical compounds.     

Determination of Phenolic Glucosides in Gentiana piasezkii by Capillary Zone Electrophoresis

A capillary zone electrophoretic method has been developed for the quantitative analysis of five phenolic glucosides, 6′-O-vanilloylarbutin (VA), 7-O-feruloylorientin (FE), lutonarin (LN), isoorient (IO) and luteolin (LL), in Gentiana piasezkii with UV detection at 270 nm. 7-O-β-D-glucosyl-coumarin was selected as the internal standard. The applied voltage was 15 kV and the capillary temperature was kept constant at 25 °C. The effect of pH, the concentration of methanol and boric acid on migration were studied systematically. Optimum separation was achieved with 200 mM boric acid buffer at pH 9.50 containing 10% (v/v) methanol. Regression equations revealed good linear relationship between the peak area ratio of each compound and internal standard and its concentration. The correlation coefficients were 0.9975, 0.9997, 0.9998, 0.9998 and 0.9988 for VA, FE, LN, IO and LL, respectively. The relative standard deviations of migration time and the peak area ratio of each analyte and internal standard were <1.78% and 4.93%, respectively. The contents of the five compounds in Gentiana piasezkii were successfully determined with satisfactory repeatability and recovery.     

Cyclodextrin‐modified capillary electrophoresis for achiral and chiral separation of ergostane and lanostane compounds extracted from the fruiting body of Antrodia camphorata

A CD‐modified capillary electrophoretic method has been developed for achiral and chiral analysis of seven bioactive compounds isolated from the fruiting body of Antrodia camphorata. Such important target analytes exhibit similar chemical structures and are known for their diverse properties including antioxidant and anticancer effects. The analytes were separated in 25 min using a pH 9.3, 20 mM sodium borate buffer containing 20 mM methyl‐β‐CD and 30 mM sulfobutylether‐β‐CD. With the exception of the optical isomer pairs (antcin B or zhankuic acid A, zhankuic acid C, and antcin A), the remaining bioactive compounds including the chiral pair antcin C were baseline‐separated. Analysis time was noticeably longer to baseline separate all of the above chiral pairs (～38 min) by adding 5% DMF to the running buffer. The migration order was reversed compared with the HPLC elution. More hydrophobic compounds complexed favorably with methyl‐β‐CD and emerged earlier in the electropherogram than their more hydrophilic counterparts which were strongly associated with sulfobutylether‐β‐CD. The simple capillary electrophoretic method developed was applicable for rapid separation and characterization of several important bioactive compounds isolated from the fruiting body of A. camphorata.     

Novel Cell Membrane Capillary Chromatography for Screening Active Compounds from Natural Products

Cell membrane chromatography (CMC) is a useful method for the simultaneous isolation and identification of active compounds from natural products. However, it suffers from high cell membrane consumption and is time-consuming to operate. In this study, CMC was performed for the first time with a silica capillary, termed cell membrane capillary chromatography (CMCC). Pancreatic islet cell membranes from a mouse were immobilized onto the capillary inner wall functionalized with aldehyde groups. Scanning electron microscopy observation of the prepared column showed that the cell membrane was evenly coated onto the capillary inner wall. Three model analytes with the pharmacological property of hypoglycemic activity including glibenclamide, glipizide and berberine were tested. They were all retained by the prepared column. Furthermore, the retention factors of the analytes in CMCC correlated well with their pharmacological action. The analytical procedure including washing (to obtain a flat baseline), injection and separation was accomplished within 10 min. The CMCC column was also used for screening active compounds from a natural plant (Coptis chinensis). The hypoglycemia activity of active components such as berberine was verified using the method. The results indicated that CMCC is a viable alternative method for screening active compounds from natural products.     

Determination of N-Methylpyrrolidine in Cefepime for Injection by Capillary Electrophoresis with Indirect UV Detection

N-Methylpyrrolidine in cefepime for injection was determined by capillary electrophoresis with indirect UV detection. Best results were achieved with background electrolyte consisting of 10 mM creatinine adjusted to pH 3.8 with formic acid and an applied voltage of 30 kV in a bare fused-silica capillary. Indirect UV detection was performed at a wavelength of 225 nm. The application of a small amount of inlet pressure during the separation assisted the attainment of a stable baseline. The optimized method was validated regarding selectivity, linearity, accuracy, precision, ruggedness, repeatability and detection limits. Careful control of capillary conditioning enabled migration time precision values of <0.2% RSD. The use of an internal standard enabled precision values of <1% RSD to be obtained for peak area ratios.     

Development of a non-aqueous capillary electrophoresis method with UV-visible and fluorescence detection for phenolics compounds in olive oil

Response surface methodology has been applied to the optimization of a simple and rapid non-aqueous capillary electrophoresis method for the separation and determination of several phenolic compounds belonging to the different families present in olive oil. A Box–Behnken design was employed and a total of 27 experiments were performed using olive oil samples spiked with the phenols and injected directly in the capillary after dilution 1:1 with 1-propanol. Finally, the background electrolyte (BGE) was constituted of 25 mM boric acid and 18 mM KOH in a mixture of 74:26 (v/v) 1-propanol/methanol. The hydrophobicity of the BGE allows its miscibility with the olive oil and, as a consequence, the possibility of characterizing and determining these kinds of compounds in this sample without any pretreatment. A hydrodynamic injection (6 s, −30 mbar) was applied and the separation was carried out using 35 °C and +20 kV of separation temperature and voltage, respectively. A capillary with two detection windows for serial online UV and fluorescence detection was satisfactorily employed. The validation of the method was carried out by setting the calibration curves, and the figures of merit were finally obtained. A lineal relationship between the corrected peak area and concentration and limits of detection in the order of micrograms per milliliter were found.     

Direct Determination of Barbiturates in Urine by Capillary Electrophoresis Using a Capillary Coated Dynamically with Polycationic Polymers

A new, simple and rapid capillary electrophoresis method, using hexadimethrine bromide (HDB) as electro-osmotic flow modifier, has been developed for the identification and determination of nine barbiturates, barbital acid, barbital, phenobarbital, pentobarbital, amobarbital, thiobarbituric acid, butobarbital, N-methyl-5-phenyl-ethyl barbital acid and 5-cyclohexenyl-5-ethyl barbital acid in urine with UV detection at 200 nm. The applied voltage was ?25 kV and the capillary temperature was kept constant at 25 °C. The effects of buffer pH, the concentration of HDB and the concentration of α-cyclodextrin were studied systematically. Optimum separation was achieved with 20 mM borate buffer at pH 10.00 containing 0.04% (w/v) HDB and 2.06 mM α-cyclodextrin. Regression equations revealed good linear relationship between the peak area of each compound and its concentration. The correlation coefficients were from 0.9990 to 0.9997. The relative standard deviations of migration times and peak areas were <3.84 and 5.45% (intra-day). The nine barbiturates in urine were successfully determined within 7 min, without a prior preparation step and the method is useful for the investigation of intoxication.     

Rapid CE‐UV binding tests of environmentally hazardous compounds with polymer‐modified magnetic nanoparticles

Hazardous compounds and bacteria in water have an adverse impact on human health and environmental ecology. Polydopamine (or polypyrrole)‐coated magnetic nanoparticles and polymethacrylic acid‐co‐ethylene glycol dimethacrylate submicron particles were investigated for their fast binding kinetics with bisphenol A, proflavine, naphthalene acetic acid, and Escherichia coli. A new method was developed for the rapid determination of % binding by sequential injection of particles first and compounds (or E. coli) next into a fused‐silica capillary for overlap binding during electrophoretic migration. Only nanolitre volumes of compounds and particles were sufficient to complete a rapid binding test. After heterogeneous binding, separation of the compounds from the particles was afforded by capillary electrophoresis. % binding was influenced by applied voltage but not current flow. In‐capillary coating of particles affected the % binding of compounds.     

Simultaneous Determination of Anthraquinones in Hedyotis diffusa by LC Coupled with UV Detection

A simple and reliable liquid chromatography method coupled with UV detection was developed and validated for the determination of the pharmacologically important anthraquinones in the total plants of Hedyotis diffusa. The separation of four anthraquinones was performed on a C₁₈ column using an isocratic mobile phase consisted of methanol-0.5% glacial acetic acid (58:42, v/v). The detection wavelength was set at 265 nm. The limits of detection obtained for the analytes were in the range of 0.004–0.012 μg mL^?1. The recovery of the developed method for the analysis of the four compounds was determined and ranged from 95.8 to 103.2%. According to the literatures, this is the first LC method reported for the simultaneous determination of the four anthraquinones and the method can be applied to determine the amounts of the bioactive compounds in Hedyotis diffusa.     

Simultaneous identification of natural dyes in the collection of drawings and maps from The Royal Chancellery Archives in Granada (Spain) by CE

A simple and rapid capillary electrophoretic method with UV detection (CE-UV) has been developed for the identification of five natural dyes namely, carmine, indigo, saffron, gamboge and Rubia tinctoria root. The separation was performed in a fused-silica capillary of 64.5 cm length and 50 microm id. The running buffer was 40 mM sodium tetraborate buffer solution (pH 9.25). The applied potential was 30 kV, the temperature was 25 degrees C and detections were performed at 196, 232, 252, 300 and 356 nm. The injections were under pressure of 50 mbar during 13 s. The method was applied to the identification of carminic acid, gambogic acid, crocetin, indigotin, alizarin and purpurin in the collection of drawings and maps at the Royal Chancellery Archives in Granada (Spain). The method was validated by using HPLC as a reference method.     

Method development and validation for the simultaneous determination of five selective serotonin reuptake inhibitors by capillary zone electrophoresis

Summary A capillary zone electrophoresis method is proposed for the separation of five antidepressants. Optimum conditions for the separation were investigated. A background electrolyte solution consisting of 40 mM phosphate buffer adjusted to pH 2.5, hydrodynamic injection, and a 28 kV separation voltage were used. Relative standard deviations (RSD) were <0.9% and <1.7% for migration time and corrected peak area (n=21), respectively. The detection limits for the five antidepressants ranged from 0.3 to 0.7 mg L⁻¹. The stability of the solutions, linearity, accuracy, and precision were examined during validation of the method. The method is rapid and sensitive, when it was tested for the analysis of pharmaceutical formulations the recoveries obtained were between 98 and 103% of the nominal content.     

Determination of adduct forms of antitumor ruthenium(III) complex with cytosolic components by capillary electrophoresis with mass spectrometry

Using capillary electrophoresis with inductively coupled plasma mass spectrometry, the transformation of adduct forms of indazolium trans-[tetrachloridobis(1H-indazole)ruthenate(III)] with apo-transferrin and albumin under the effect of active intracellular reducing agents, glutathione and ascorbic acid, and citric acid as a complexant was studied under conditions simulating a cytosolic environment. These adducts of ruthenium with transport proteins are forms in which the anticancer drug exists after intravenous administration. Two modes of interaction of adducts with glutathione, ascorbic acid, and citric acid were studied: in a capillary using a background electrolyte containing a cytosolic active ingredient and upon incubation of the reaction components in the off-line mode, followed by their separation and determination by capillary electrophoresis. Incubation with intracellular components and separation were carried out in a 10 mM phosphate buffer solution (pH 6.0) containing 4 mM NaCl. The effect 1–10 mM of glutathione, 10 mM of ascorbic acid, and 50 mM of citric acid on the adducts was studied. It is shown that under the selected model conditions, new forms of ruthenium do not emerge.     

Stable physically adsorbed coating of poly-vinyl alcohol for the separation of basic proteins

In aqueous capillary electrophoresis, the electroosmotic flow can be strongly suppressed by coating the inner surface of the capillary. In the present work hydrophilic coating of 4% polyvinyl alcohol (PVA) has been used for the analysis of basic proteins. The coating is simple and easy to obtain. The separation of ribonuclease and α-chymotrypsin has been uniquely done with other three basic proteins (lysozyme, cytochrome-c and trypsin) using a buffer 11.60 mM sodium acetate and 18.40 mM acetic acid at pH 4.5 in addition to positive power supply of 20 kV at 25°C. Detection was performed using UV detector at 230 nm. The proposed PVA coated capillary provides reproducible separation of five basic proteins within 10 min with RSD values for mobility bellow 1.4% (n = 6) for all the five basic proteins. The stability of coated capillary has been checked up to 40 runs. The viscosity measurement for 4% PVA have been studied and scanning electron microscope (SEM) images obtained to make it compatible with future micro-chip applications.     

Separation and Determination of Stereoisomeric Impurity of Folinic Acid Diastereomers by CE with Vancomycin as a Selector

A highly sensitive and rapid method was developed that involves capillary electrophoresis for separation and determination of the stereoisomeric impurity of folinic acid diastereomers. In this method, vancomycin was used as the chiral selector, and a solution of poly(dimethylacrylamide) (PDMA) was prepared for dynamic coating of the capillary wall to minimize the adsorption of vancomycin. This method was optimized for six factors including concentrations of the organic modifier and vancomycin, pH and concentration of the background electrolyte, column temperature, and separation voltage. The following conditions were established: 100 mM Tris-phosphate buffer (pH 6.0) containing 1.0 mM vancomycin and 5 % acetonitrile at 30 °C, and −15 kV applied voltage on the PDMA dynamically coated capillary. Preliminary validation was performed with the determination of limit of quantification and detection, accuracy, precision, and linearity. Under our optimized method, the folinic acid diastereomers were baseline-separated within 7.5 min, and a (6S,2′S)-calcium folinate sample with 0.08 % stereoisomeric impurity was determined.

     

Quick and Sensitive Determination of Fluoroquinolones by Capillary Electrophoresis–Potential Gradient Detection

Abstract

A rapid, capillary electrophoresis–potential gradient detection method was developed for determining three fluoroquinolones: rufloxacin, enoxacin, and moxifloxacin. The fluoroquinolones were baseline separated within 3.5 min with background electrolyte composed of 50 mM acetic acid and 6 mM potassium hydroxide at pH 3.7. For the first time, electrokinetic injection was applied in capillary electrophoretic separation of fluoroquinolones, and an average of ca. 10-fold improvement in detection limits was observed. The method was applied to a fortified chicken tissue sample with detection limits (S/N = 3) of 22–24 ng g^?1, suggesting the potential of the method for determining fluoroquinolone residues in real samples.     

Analysis of anabolic androgenic steroids in urine by full-capillary sample injection combined with a sweeping CE stacking method

This study describes an on-line stacking CE approach by sweeping with whole capillary sample filling for analyzing five anabolic androgenic steroids in urine samples. The five anabolic steroids for detection were androstenedione, testosterone, epitestosterone, boldenone, and clostebol. Anabolic androgenic steroids are abused in sport doping because they can promote muscle growth. Therefore, a sensitive detection method is imperatively required for monitoring the urine samples of athletes. In this research, an interesting and reliable stacking capillary electrophoresis method was established for analysis of anabolic steroids in urine. After liquid–liquid extraction by n-hexane, the supernatant was dried and reconstituted with 30 mM phosphate buffer (pH 5.00) and loaded into the capillary by hydrodynamic injection (10 psi, 99.9 s). The stacking and separation were simultaneously accomplished at ?20 kV in phosphate buffer (30 mM, pH 5.0) containing 100 mM sodium dodecyl sulfate and 40 % methanol. During the method validation, calibration curves were linear (r?≥?0.990) over a range of 50–1,000 ng/mL for the five analytes. In the evaluation of precision and accuracy for this method, the absolute values of the RSD and the RE in the intra-day (n?=?3) and inter-day (n?=?5) analyses were all less than 6.6 %. The limit of detection for the five analytes was 30 ng/mL (S/N?=?5, sampling 99.9 s at 10 psi). Compared with simple MECK, this stacking method possessed a 108- to 175-fold increase in sensitivity. This simple and sensitive stacking method could be used as a powerful tool for monitoring the illegal use of doping.