Zeineb Aturki

Use of cyclodextrins in capillary electrophoresis for the chiral resolution of some 2-arylpropionic acid non-steroidal anti-inflammatory drugs

Abstract The enantiomeric separation of racemic compounds of some 2-arylpropionic acid non-steroidal anti-inflammatory drugs (profens), namely fenoprofen, ibuprofen, flurbiprofen, suprofen, ketoprofen and indoprofen, was performed by capillary zone electrophoresis. The separation was obtained by supporting the background electrolyte with derivatized β-cyclodextrins. The type and concentration of cyclodextrin used and the background electrolyte composition (pH and amount of methanol) influenced the complexation and the chiral resolution. All the modified β-cyclodextrins used (heptakis-2,6-di-O-methyl-β-, heptakis-2,3,6-tri-O-methyl- and 6 A -methylamino-β-cyclodextrin) showed good complexing effects with the profens tested. Tri-O-methyl-β-cyclodextrin proved to be the best stereoselective additive because it allowed the enantiomeric resolution of all the profens studied whereas the dimethylated and methylamino-β-cyclodextrin were able to separate only some of them.

Use of vancomycin silica stationary phase in packed capillary electrochromatography I. Enantiomer separation of basic compounds.

Chiral separation of basic compounds was achieved by using 75 or 100 μm ID fused‐silica capillaries packed with a vancomycin‐modified diol silica stationary phase. The capillary was firstly packed for about 12 cm with a slurry mixture composed of diol‐silica (3:1) then with the vancomycin modified diol‐silica (3:1) (23 cm), and finally with diol‐silica (3:1) for about 2 cm. Frits were prepared by a heating wire at the two ends of the capillary; the detector window was prepared at 8.5 cm from the end of the capillary where vancomycin was not present. The influence of the mobile phase composition (pH and concentration, organic modifier type and concentration) on the velocity of the electroosmotic flow, chiral resolution and enantioselectivity was studied. Good enantiomeric resolution was achieved for atenolol, oxprenolol, propranolol, and venlafaxine using a mobile phase composition of 100 mM ammonium acetate solution (pH 6)/water/acetonitrile (5:5:90 v/v/v) while for terbutaline a mixture of 5:15:80 v/v/v provided the best separations. The use of methanol instead of acetonitrile caused a general increase of enantiomer resolution of the studied compounds together with a reduction of efficiency and detector response. However, the combination of acetonitrile and methanol in the mobile phase (as, e.g., 10% methanol and 80% acetonitrile) allowed to improve the enantiomer resolution with satisfactory detector response.

Enantiomeric resolution study by capillary electrophoresis Selection of the appropriate chiral selector

Abstract The enantiomeric separation of several arylpropionic acids, namely carprofen, cicloprofen, flurbiprofen, ibuprofen, indoprofen, ketoprofen, naproxen and suprofen has been studied by capillary zone electrophoresis using different chiral selectors added to the background electrolyte with the aim to find the optimum experimental conditions for both qualitative and quantitative purposes. The chiral selectors used included two β-cyclodextrin derivatives and a glycosidic antibiotic, namely 2,3,6-tri-O-methyl-β-CD, heptamethylamino-β-CD and vancomycin. When the CDs were used the chiral selector was present in both capillary and electrode compartments while for vancomycin the partial filling method was used (the chiral selector was not present at the detector window in order to improve the sensitivity; vancomycin is strongly absorbing at low wavelengths). Enantiomeric resolution was recorded for all the compounds studied except for ibuprofen when heptamethylamino-β-CD was used; resolution generally increased by increasing the chiral selector concentration. Good sensitivity and good precision for both migration times and corrected peak areas (Asample/migration time) were achieved using the three chiral selectors. Among the three chiral additives employed vancomycin proved to be the most effective for the enantiomeric separation of the studied arylpropionic acids. The optimized method achieved analysis with the shortest time (

Use of β-cyclodextrin polymer as a chiral selector in capillary electrophoresis

Enantiomers of several basic compounds of pharmaceutical interest were successfully separated by capillary electrophoresis using a modified β-cyclodextrin polymer. As the cyclodextrin contained carboxylic groups, the chiral selector could be used in either an unchanged or a charged mode, selecting the appropriate pH of the background electrolyte. The effect of the pH of the background electrolyte on the effective mobility, resolution and selectivity was studied in the range 2.6–6.2 for the enantiomer resolution of β-hydroxyphenylethylamine, norphenylephrine, terbutaline, ephedrine, norephedrine, ketamine, epinephrine and propranolol. Very good enantiomeric resolution was achieved for all the compounds except for ephedrine and norephedrine (R < 0.5). An increase in the pH of the electrolyte caused an inversion of mobility for either terbutaline and propranolol owing to strong complexation with the negatively charged polymer.

Current applications of miniaturized chromatographic and electrophoretic techniques in drug analysis.

In the last decade, miniaturized separation techniques have become greatly popular in pharmaceutical analysis. Miniaturized separation methods are increasingly utilized in all processes of drug discovery as well as quality control of pharmaceutical preparation. The great advantages presented by the analytical miniaturized techniques, including high separation efficiency and resolution, rapid analysis and minimal consumption of reagents and samples, make them an attractive alternative to the conventional chromatographic methods for drug analysis. The purpose of this review is to give a general overview of the applicability of capillary electrophoresis (CE), capillary electrochromatography (CEC) and micro/capillary/nano-liquid chromatography (micro-LC/CLC/nano-LC) for the analysis of pharmaceutical formulations, active pharmaceutical ingredients (API), drug impurity testing, chiral drug separation, determination of drugs and metabolites in biological fluids. The results concerning the use of CEC, micro-LC, CLC, and nano-LC in the period 2009-2013, while for CE, those from 2012 up to the review draft are here summarized and some specific examples are discussed.

Analysis of synthetic cannabinoids in herbal blends by means of nano-liquid chromatography

In this study, a rapid and simultaneous separation of 12 synthetic cannabinoids and Δ(9)-tetrahydrocannabinol (Δ(9)-THC) in herbal blends was obtained by means of nano-liquid chromatography (nano-LC). The nano-LC experiments were performed in a 100μm i.d. capillary column packed with Cogent(®) bidentate C(18) silica particles for 25.0cm. All compounds were resolved using an isocratic elution mode in less than 30min. A mobile phase containing ACN/MeOH/H(2)O/formic acid 69/5/25/1 (v/v/v/v) was employed for the chromatographic separation. The developed analytical method was validated in terms of precision, linearity, sensitivity and accuracy. Under optimal nano-LC-UV conditions, the resulting RSD percentages for intra-day and inter-day repeatability, related to retention time and peak area, were below 2.98 and 6.40%, respectively. Limits of detection and quantification were 0.2 and 0.5μg/mL, respectively, for all the studied compounds. Linearity was assessed in the concentration range of interest for all analytes with determination coefficients r(2)≥0.9975. The method was then applied to the determination of synthetic cannabinoids in herbal blends. Quantitative analyses of the cannabimimetic compounds in six products showed that there was a wide difference in the concentration of the studied compounds among different products. Further, the nano-LC system was coupled with a mass spectrometer measuring the MS and MS-MS spectra to unequivocally identify the cannabinoids present in smoking mixtures.

Chiral analysis of UV nonabsorbing compounds by capillary electrophoresis using macrocyclic antibiotics: 1. Separation of aspartic and glutamic acid enantiomers.

Glycopeptide antibiotics, namely vancomycin or teicoplanin, were evaluated in capillary electrophoresis for the analysis of UV nonabsorbing compounds such as aspartic and glutamic acid enantiomers. Electrophoretic runs were performed in laboratory‐made polyacrylamide‐coated capillaries using the partial filling‐counter current method in order to avoid the presence on the detector path of the absorbing chiral selector. The background electrolyte consisted of an aqueous or aqueous‐organic buffer in the pH range of 4.5–6.5 of sorbic acid/histidine and the appropriate concentration of chiral selector. Several experimental parameters such as antibiotic concentration and type, buffer pH, organic modifier, type and concentration of absorbing co‐ion (for the indirect UV detection) were studied in order to find the optimum conditions for the chiral resolution of the two underivatized amino acids in their enantiomers. Among the two investigated chiral selectors, vancomycin resulted to be the most useful chiral selector allowing relatively high chiral resolution of the studied compounds even at low concentration. The optimized method (10 mM sorbic acid/histidine, pH 5, and 10 mM of vancomycin) was used for the analysis of real samples such as teeth dentine and beer.

Analysis of hesperetin enantiomers in human urine after ingestion of blood orange juice by using nano-liquid chromatography

Hesperetin (HT) is a flavanone abundantly found in citrus fruits. It has been reported that HT possesses significant antioxidant, anticancer, anti-inflammatory and analgesic activities. This explains the necessity of developing new methods more powerful and sensitive for analyzing HT in biological fluids. Taking into account the chiral nature of HT, the study of the stereospecific kinetics of in vitro and in vivo metabolism and tissue distribution could be a useful tool for further understanding stereoselective biotransformations in human body. A simple nano-liquid chromatographic method for the determination of the enantiomeric composition of hesperetin in human urine was developed. Chiral separation was achieved using a 100 microm I.D. capillary, packed with phenyl-carbamate-propyl-beta-cyclodextrin stationary phase, employing a mobile phase composed by a mixture of triethylammonium acetate buffer (1%, v/v, pH 4.5) and water/methanol (30:70, v/v) at room temperature. The detection was done by using on-column UV detector at 205 nm. Calibration curves were linear in the studied concentration range from 0.25 to 25 microg/mL (r(2)>0.999). Precision assay was <4.5% and was within 3% at the limit of quantification (0.5 microg/mL). The recovery of 7-ethoxycoumarin (IS), R- and S-hesperetin was greater than 82.48%, utilizing a liquid-liquid extraction procedure. The developed method was successfully applied to the determination of hesperetin enantiomers in urine samples obtained from a male volunteer, after the ingestion of 1L of a commercial blood orange juice.

Analysis of phenolic compounds in extra virgin olive oil by using reversed-phase capillary electrochromatography.

In this work, the simultaneous separation of ten phenolic compounds (protocatechuic, p‐coumaric, o‐coumaric, vanillic, ferulic, caffeic, syringic acids, hydroxytyrosol, tyrosol and oleuropein) in extra virgin olive oils (EVOOs) by isocratic RP CEC is proposed. A CEC method was optimized in order to completely resolve all the analyzed compounds by studying several experimental parameters. The influence of the stationary phase type (C18 and C8 modified silica gel), buffer concentration and pH as well as the organic modifier content of the mobile phase on retention factors, selectivity and efficiency were evaluated in details. A capillary column packed with Cogent® bidentate C18 particles for 23 cm and a mobile phase composed by 100 mM ammonium formate buffer pH 3/H2O/ACN (5:65:30 v/v/v) allowed the baseline resolution of the compounds under study in less than 35 min setting the applied voltage and temperature at 22 kV and 20°C, respectively. A study, evaluating the intra‐ and interday precision as well as LOD and LOQ and method linearity was developed in accordance with the analytical procedures for method validation. LODs were in the range of 0.015–2.5 μg/mL, while calibration curves showed a good linearity (r2 >0.997). The CEC method was applied to the separation and determination of these compounds in EVOO samples after a suitable liquid–liquid extraction procedure. The mean recovery values of the studied compounds ranged between 87 and 99%.

CEC-ESI ion trap MS of multiple drugs of abuse.

This article describes a method for the separation and determination of nine drugs of abuse in human urine, including amphetamines, cocaine, codeine, heroin and morphine. This method was based on SPE on a strong cation exchange cartridge followed by CEC‐MS. The CEC experiments were performed in fused silica capillaries (100 μm×30 cm) packed with a 3 μm cyano derivatized silica stationary phase. A laboratory‐made liquid junction interface was used for CEC‐MS coupling. The outlet capillary column was connected with an emitter tip that was positioned in front of the MS orifice. A stable electrospray was produced at nanoliter per minute flow rates applying a hydrostatic pressure (few kPa) to the interface. The coupling of packed CEC columns with mass spectrometer as detector, using a liquid junction interface, provided several advantages such as better sensitivity, low dead volume and independent control of the conditions used for CEC separation and ESI analysis. For this purpose, preliminary experiments were carried out in CEC‐UV to optimize the proper mobile phase for CEC analysis. Good separation efficiency was achieved for almost all compounds, using a mixture containing ACN and 25 mM ammonium formate buffer at pH 3 (30:70, v/v), as mobile phase and applying a voltage of 12 kV. ESI ion‐trap MS detection was performed in the positive ionization mode. A spray liquid, composed by methanol–water (80:20, v/v) and 1% formic acid, was delivered at a nano‐flow rate of ∼200 nL/min. Under optimized CEC‐ESI‐MS conditions, separation of the investigated drugs was performed within 13 min. CEC‐MS and CEC‐MS2 spectra were obtained by providing the unambiguous confirmation of these drugs in urine samples. Method precision was determined with RSDs values ≤3.3% for retention times and ≤16.3% for peak areas in both intra‐day and day‐to‐day experiments. LODs were established between 0.78 and 3.12 ng/mL for all compounds. Linearity was satisfactory in the concentration range of interest for all compounds (r2≥0.995). The developed CEC‐MS method was then applied to the analysis of drugs of abuse in spiked urine samples, obtaining recovery data in the range 80–95%.