Cyclodextrin-modified Capillary Zone Electrophoresis Research Articles

Analysis of lipid-oligonucleotide conjugates by cyclodextrin-modified capillary zone electrophoresis

Lipid-oligonucleotide (LONs) based bioconjugates represent an emerging class of therapeutic agents, allowing the delivery of therapeutic oligonucleotide sequences. The LON development requests accurate and efficient analytical methods. In this contribution, LON analysis methods were developed in cyclodextrin-modified capillary zone electrophoresis (CD-CZE). The LONs selected in this study feature different structures, including i) the oligonucleotide length (from 10 to 20 nucleotides), ii) the inter-nucleotide linkage chemistry (phosphodiester PDE or phosphorothioate PTO), and iii) the lipidic part: single- (LONsc) or double-chain (LONdc) lipids. In CD-CZE, the effect of several parameters on the electrophoretic peaks was investigated (buffer, CD, and capillary temperature). The binding interaction between LON and Me-β-CD was studied in affinity capillary electrophoresis and revealed a 1:1 LON:CD complex. Non-linear regression and three usual linearization methods (y-reciprocal, x-reciprocal, and double-reciprocal) were used to determine the binding constants (K values of 2.5.104 M−1 and 2.0.104 M−1 for LON PDE and LON PTO, respectively). Quantitative methods with good performances and analysis time lower than 5 min were achieved. Importantly, the developed analysis allows a separation between the i) full-length sequence LONs and their truncated sequences, (n-1), (n-2), and (n-4)-mers and ii) LONsc, LONdc and their corresponding unconjugated oligonucleotides. This work highlights the interest of CD-CZE methods for LON analysis.

Separation and Migration Behavior of Dichlorophenols in β‐Cyclodextrin‐Modified Capillary Zone Electrophoresis

AbstractThe separation and migration behavior of six isomeric dichlorophenols (DCPs) in cyclodextrin‐modified capillary zone electrophoresis (CD‐CZE) using a phosphate‐borate buffer at alkaline pH with β‐CD and hydroxypropyl‐β‐CD (HP‐β‐CD) as electrolyte modifiers were investigated. The influence of buffer pH and the concentration of β‐cyclodextrins were examined. The results indicate that baseline separation of six isomeric DCPs can be achieved with addition of β‐CD concentration in the range of 2.0‐10 mM or HP‐β‐CD concentration in the range of 4.0‐10 mM at pH 10.0. Binding constants of DCPs to β‐CDs were evaluated for a better understanding of the interaction of DCPs with β‐CDs.

A new example of reversal of the order of migration of enantiomers, as a function of cyclodextrin concentration and pH, by cyclodextrin-modified capillary zone electrophoresis: enantioseparation of 6,6′-dibromo-1,1′-binaphthyl-2,2′-diol

Enantioseparation of 6,6'-dibromo-1,1'-binaphthyl-2,2'-diol (DBBD) by cyclodextrin-modified capillary zone electrophoresis (CD-CZE) was studied using the three native alpha, beta, and gamma cyclodextrins, the three hydroxypropylated cyclodextrins (2-hydroxypropyl-alpha, beta, and gamma), heptakis-2,6-di-O-methyl-beta-CD (DM-beta-CD), and heptakis-2,3,6-tri-O-methyl-beta-cyclodextrin (TM-beta-CD). First, the acidity constants of DBBD were determined using capillary electrophoresis, before performing enantioseparation. The influence of the concentrations of the studied cyclodextrins on the enantioseparation was explored and the experimental optimal concentrations were determined and compared to the theoretical optimal concentrations. Moreover, the apparent complexation constants between each studied cyclodextrin and the two DBBD enantiomers were evaluated using a non-linear curve fitting method and three linear plotting methods (x-reciprocal, y-reciprocal and double reciprocal). For TM-beta-CD, the order of migration of the enantiomers of DBBD reversed as a function of TM-beta-CD concentration. The influence of the nature of methylated cyclodextrin derivatives (methyl-beta-CD (M-beta-CD) and DM-beta-CD) was then studied. Inversion of the order of migration of the enantiomers of DBBD was observed for DM-beta-CD, whereas the S enantiomer of DBBD always migrated first for M-beta-CD.

Enantioseparation of binaphthol and its monoderivatives by cyclodextrin-modified capillary zone electrophoresis: A mathematical approach

A simple model for the separation of atropisomers of binaphthol and its monoderivatives by means of cyclodextrin-modified capillary zone electrophoresis (CD-CZE) was used to describe the migration behavior of poly charged enantiomers in a chiral separation system. This mathematical approach allowed for the determination of the optimal cyclodextrin concentrations for the enantioseparation of binaphthols by taking into account the influence of the formed complex mobilities. Moreover, using this theoretical approach, the reversal of the enantiomers’ migration order as a function of cyclodextrin concentration was predicated. The apparent complexation constants between the cyclodextrins and the binaphthol and its monoderivatives could be calculated using a non-linear curve fitting method and three linear plotting methods ( x-reciprocal, y-reciprocal and double reciprocal). Good agreements between the theoretical and experimental cyclodextrin concentrations were obtained.

Enhanced analysis of triterpenes, flavonoids and phenolic compounds in Prunella vulgaris L. by capillary zone electrophoresis with the addition of running buffer modifiers

A cyclodextrin-modified capillary zone electrophoresis method was developed for the separation and determination of three isomeric compounds (ursolic acid, oleanolic acid and betulinic acid), caffeic acid, p-coumaric acid, rosmarinic acid, rutin and quercetin. Without the addition of β-cyclodextrin (β-CD) and methanol, the separation of these analytes was poorly resolved. These eight compounds, however, were well separated from each other within 20 min with a borax running buffer (40 mM of borax, pH 9.4) containing 2 mM β-CD and 4% (v/v) methanol at the voltage of 25 kV, temperature of 25 °C and detection wavelength of 210 nm. The relative standard deviations (RSDs) of migration time ranged from 0.16 to 0.74% while those of the peak area ratios ranged from 2.17 to 4.61% for six determinations of the analytes at concentration of 10 and 25 μg mL −1. The correlation coefficients of the calibration curves of the analytes were all >0.998, and the recoveries were from 96.8 to 103.6%. The method was successfully applied to determine these bioactive components in the samples of Prunella vulgaris L. and its beverage drink products. Our results reveal that only the isomeric compounds and rosmarinic acid could be detected in the spikes of P. vulgaris L.; other components were either too low to be detected or not present while only rosmarinic acid was detected in the beverage products.

Enantioseparation of binaphthol and its mono derivatives by cyclodextrin-modified capillary zone electrophoresis

Chiral binaphthols belong to the group of most effective ligands for asymmetrical catalysis. In this context, various binaphthols presenting original substituents have been synthesized. Their study through capillary electrophoresis is the object of this work. The literature dedicated to the separation of atropisomers by capillary electrophoresis, corresponding only to binaphthol, reveals that its enantioseparation is always delicate because of the influence of many factors and the resolutions obtained are weak. Therefore, for a structured optimization, we first successfully evaluated the acidity constants of different binaphthols by means of capillary electrophoresis. With these known physicochemical characteristics, we could successfully carry out enantiomeric separations of the different binaphthols at pH 11.5, practically in completely ionized form, in phosphate medium, and in the presence of cyclodextrin (CD), with analysis times lower than 8 min. The nature of CDs (α-CD, β-CD, γ-CD, hydroxypropyl-α-cyclodextrin (HP-α-CD), HP-β-CD, HP-γ-CD and trimethyl-β-CD (TM-β-CD)) and other factors in relation to enantiomeric resolution (applied voltage, nature and concentration of the electrolyte, and concentration of cyclodextrin) were optimized. These studies allowed us to determine the optimal conditions of separation (concentration and nature of CD) for each of the studied binaphthols. It is necessary to mention that, for the 1,1′-binaphthyl-2,2′-diol (Binol) at pH 11.5, the S atropisomer always migrated first, regardless of the nature and concentration of the cyclodextrin used. Moreover, an inversion in elution order of the two atropisomers as a function of pH was observed with γ-CD (pH range: 10–11.5). The R atropisomer migrated first at pH 10. At pH 10.8 the migration order of the two atropisomers of Binol was reversed as a function of γ-CD concentration. Finally, the addition of chiral ionic liquids ( R(−)-1-hydroxy- N, N, N-trimethylbutan-2-aminium bis(trifluoromethylsulfonyl)imide and S(+)-tetrabutylammonium camphorsulfonate) was conducted. In the case of S(+)-tetrabutylammonium camphorsulfonate, a weak antagonistic effect was observed with modeling the evolution of enantiomeric resolution by means of the experimental design, while in the case of R(−)-1-hydroxy- N, N, N-trimethylbutan-2-aminium bis(trifluoromethylsulfonyl)imide the effect was neutral.

Chiral separation of hydroxyflavanones in cyclodextrin-modified capillary zone electrophoresis using sulfated cyclodextrins as chiral selectors

Chiral separations of three hydroxyflavanone aglycones, including 2′-, 3′-, and 4′-hydroxyflavanone, in capillary zone electrophoresis (CZE) using randomly sulfate-substituted β-cyclodextrin (S-β-CD) or dual cyclodextrin (CD) systems consisting of S-β-CD and a neutral CD at low pH were investigated. The results indicate that S-β-CD is an excellent chiral selector for enantioseparation of 2′-hydroxyflavanone and is a good chiral selector for 3′-hydroxyflavanone. Depending on the concentration of S-β-CD ranging from 2.0 to 0.75% (w/v), the enantioresolution values were 10.5–19.5 and 1.8–3.4 for 2′- and 3′-hydroxyflavanone, respectively. The enantiomers of 4′-hydroxyflavanone could be effectively separated with S-β-CD at a concentration of 2.0% (w/v) within 20 min. The enantioselectivity and enantioresolution follow the order 2′-hydroxyflavanone ≫ 3′-hydroxyflavanone > 4′-hydroxyflavanone. Alternatively, with the addition of sodium dodecyl sulfate (SDS) monomers at low concentrations in the electrophoretic system, enantioselectivity of these hydroxyflavanone aglycones could be enhanced with dual CD systems. In this case, SDS monomer acted as a complexing agent probably first with S-β-CD and then subsequently with the analytes for increasing the effective electrophoretic mobility of the analytes towards the anode and as a selectivity controller for affecting the selectivity of hydroxyflavanones. Better enantioseparation between 2′-hydroxyflavanone and 3′-hydroxyflavanone could be achieved with a dual CD system consisting of S-β-CD and γ-CD than that with S-β-CD and β-CD. In addition, possible chiral recognition mechanisms of hydroxyflavanones are discussed.

Studies on degradation of imazalil enantiomers in soil using capillary electrophoresis

A scheme was demonstrated to elucidate the degradation behaviors of the two enantiomers of the fungicide imazalil in soil using cyclodextrin-modified capillary zone electrophoresis. The separation buffer was 50 mmol/L NaH2PO4, 5 mmol/L (NH4)H2PO4, and 5 mmol/L beta-cyclodextrin (pH 3.0). The limits of detection of this CE method were 0.24 and 0.26 microg/mL for (-)- and (+)-imazalil, respectively. Five different soil conditions were investigated in laboratory microcosms: under sunlight; in darkness; under UV irradiation; in sterilized soil; and in soil with wheat planted in it. Radiation, microorganisms, and uptake by wheat benefited the degradation of imazalil in this study. The half-lives (t1/2) of imazalil in soil under the above conditions were 20, 30.5, 11, 27.5, and 21.5 days, respectively. The degradation rate of imazalil in soil under the five different sets of conditions decreased in the order: UV irradiation > sunlight > soil with wheat planted in it > sterilized soil > soil kept in darkness. Imazalil in soil (pH 8.2, slightly alkaline) collected in the suburbs showed non-enantioselective degradation.

Chiral separation of ephedrine alkaloids from Ephedra sinica extract and its medicinal preparation using cyclodextrin-modified capillary zone electrophoresis

A fast capillary zone electrophoresis (CZE) method using dimethyl-β-cyclodextrin (DM-β-CD) as modifier with tetrabutylammonium chloride (TBAC) as addition has been developed for the chiral separation of (±)-ephedrine ((±)-EP), (±)-pseudoephedrine ((±)-PP), (±)-N-methylephedrine ((±)-MP), and (±)-norephedrine ((±)-NP). The electrophoretic separation was performed using a 35-cm × 50 μm I.D. (30-cm effective length) fused silica capillary. Samples were introduced under electromigrated injection at 5 kV for 4 s, and the running voltage was 15 kV at the injector end of the capillary. Within 19 min, eight ephedrine compounds were separated and detected at 210 nm. The method was successively applied to the determination of the ephedrine enantiomers in the Chinese herbal extract from Ephedra sinica and its medicinal preparation (Xiaoerqing feiwan). Parameters affecting the resolution between (+) and (-)-enantiomers, such as pH, cyclodextrin type and concentration, organic modifier, and tetraalkylammonium reagents, were reported.

Enantioseparation of phenothiazines in cyclodextrin-modified capillary zone electrophoresis using sulfated cyclodextrins as chiral selectors

In this study, enantioseparations of five phenothiazines, including promethazine, ethopropazine, trimeprazine, methotrimeprazine, and thioridazine, in CD-modified CZE using dual CD systems consisting of randomly sulfate-substituted CD (MI-S-beta-CD) and a neutral CD as chiral selectors in a citrate buffer (100 mM) at pH 3.0 were investigated. The results indicate that MI-S-beta-CD is an excellent chiral selector for enantioseparation of ethopropazine. The enantiomers of promethazine can also be baseline-resolved with MI-S-beta-CD at concentrations in the range of 0.5-1.0% w/v. On the other hand, thioridazine and trimeprazine interact strongly with neutral CDs. As a result, the enantioselectivity of these two phenothiazines is remarkably and synergistically enhanced with increasing the concentration of neutral CDs in the presence of MI-S-beta-CD and simultaneous enantioseparations of these phenothiazines, except for methotrimeprazine, could favorably be achieved with the use of dual CD systems. Moreover, by varying the concentration of beta-CD or gamma-CD at a fixed concentration of MI-S-beta-CD (0.75% w/v) reversal of the enantiomer migration order of promethazine occurred. This may be attributable to the opposite effects of charged and neutral CDs on the mobility of the enantiomers of promethazine.

Method for simultaneous determination of eight cyclic antidepressants by cyclodextrin-modified capillary zone electrophoresis: applications in pharmaceuticals

Our research team established a selective capillary zone electrophoresis method for the simultaneous determination of amitriptyline HCl, clomipramine HCl, trimipramine maleate (racemate, including R/S, 1:1), nortriptyline HCl, protriptyline HCl, maprotiline HCl, doxepin HCl (including 16.75% of Z-form and 83.25% of E-form), and desipramine HCl. The background buffer was Tris (500 mM, pH 3.50) containing β-cyclodextrin (1.5 mM), hydroxypropyl-β-cyclodextrin (1.5 mM) and 13% (v/v) of acetonitrile in an untreated fused-silica capillary. Analysis was run at a voltage of 20 kV and a detection wavelength of 200 nm. In the method validation, the calibration curves are linear over a range of 125.0–500.0 μM ( r > 0.9974). The R.S.D. and R.E. were all less than 3% for the intra- and inter-day assays. All recoveries were greater than 95%. The limit of detection was Z-doxepine, 4 μM; protriptyline, trimipramine-1 or trimipramine-2, 10 μM; E-doxepine, nortriptyline, maprotiline, amitriptyline, clomipramine, despramine, 5 μM (S/N = 5, hydrodynamic injection 3 s). Our method was applied to commercial tablets (maprotiline, amitriptyline, clomipramine) and capsules (doxepine). All analytical values were within the labeled amount of 90–110% required by the USP XXV.

Enantiomeric analysis of rivastigmine in pharmaceuticals by cyclodextrin-modified capillary zone electrophoresis

Chiral separation method development is usually very time-consuming due to the diversity in chemical structures of pharmaceutical drug substances as well as the suitable separation conditions and the problem to choose the appropriate chiral selector. This paper shows capillary zone electrophoresis (CZE) which was developed for chiral separation of a basic compound – rivastigmine (RIV) using 30 cm × 50 μm i.d. polyacrylamide (PAA)-coated fused-silica capillary (effective length 20 cm), amine-modified phosphate buffer of pH 2.5 and sulfated-β-CD (S-β-CD) as chiral selector. Other selected native or derivatized cyclodextrins (CDs) were also tested: β-CD (5, 30 mM), carboxymethyl-β-CD (5, 30 mM), dimethyl-β-CD (15 mM), hydroxypropyl-β-CD (5, 30 mM), hydroxypropyl-α-CD (5, 30 mM) and hydroxypropyl-γ-CD (5, 30 mM). Complete enantiomeric separation of RIV was achieved at 20 kV, 18 °C and detection at 200 nm within 8 min with R.S.D. for the absolute migration time reproducibility of less than 2.1%. Rectilinear calibration range was 5.0–500.0 μM of each enantiomer ( r = 0.9994–0.9995). The CZE method proposed was used for the control of chiral purity of pharmaceutically active S-RIV and for the analysis of Exelon caps preparation.

Separation and migration behavior of structurally related phenothiazines in cyclodextrin-modified capillary zone electrophoresis.

The influences of buffer pH and the concentration of beta-cyclodextrins (beta-CDs) on the separation and migration behavior of 13 structurally related phenothiazines in CD-modified capillary zone electrophoresis (CD-CZE) using a phosphate background electrolyte at low pH were investigated. We focused on the separation of these phenothiazines, including the enantiomers of chiral analytes, with the use of beta-CD and hydroxypropyl-beta-CD (HP-beta-CD) as electrolyte modifiers or chiral selectors at concentrations less than 8 mM. The results indicate that the interactions of phenothiazines with beta-CDs are very strong and that effective separations of 13 analytes can be achieved with addition of 0.3 mM beta-CD or 0.5 mM HP-beta-CD in a phosphate buffer at pH 3.0. Binding constants of phenothiazines to beta-CDs were evaluated for a better understanding of the interactions of phenothiazines with beta-CDs.

Determination of Active Components in Rhubarb by Cyclodextrin-modified Capillary Zone Electrophoresis

A cyclodextrin-modified capillary zone electrophoresis was developed for the determination of four bioactive components in Rhubarb. Effects of pH, b-CD concentration and organic modifier content on the migration and separation of the components were studied. The four components were separated in the running buffer of 50 mmol/L NaOHH3BO3 (pH 10.7) containing 10 mmol/L b-CD and 12.5% v/v ethanol. The analytical performance of the method was tested with respect to linearity response, precision and recovery. The determination of two commercial Rhubarb samples under the optimized conditions showed a satisfactory result.

ChemInform Abstract: Enantiomeric Separation by Cyclodextrin Modified Capillary Zone Electrophoresis (CD‐CZE) of Quaternary Tetrahydroprotoberberine Alkaloids.

AbstractChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 100 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a “Full Text” option. The original article is trackable via the “References” option.

Enantiomeric separation by cyclodextrin modified capillary zone electrophoresis (CD-CZE) of quaternary tetrahydroprotoberberine alkaloids.

A cyclodextrin modified capillary zone electrophoresis (CD-CZE) for the enantiomeric separation of tetrahydroprotoberberine N-metho salts was established. The resolution was optimized by changing the concentration of the electrolyte solution, hydroxypropyl-beta-CD (0.02 M, 0.07 M, or 0.14 M) or dimethyl-beta-CD (0.05 M or 0.15 M) in phosphate buffer (pH 2.5 or 3) containing 10% acetonitrile with an applied voltage of 20 kV. This method was applied toward the enantioselective bio-conversion of quaternary tetrahydroprotoberberine N-metho salts in cultured cells of Corydalis species.

Chiral Separation of Bencynonate and Econazole by Cyclodextrin-Modified Capillary Zone Electrophoresis

Chiral Separation of Bencynonate and Econazole by Cyclodextrin-Modified Capillary Zone Electrophoresis

Enantiomeric and isomeric separation of herbicides using cyclodextrin-modified capillary zone electrophoresis

Cyclodextrin-modified capillary zone electrophoresis (CD-CZE) was applied successfully to the enantiomeric and isomeric separation of three herbicides (imazaquin, diclofop and imazamethabenz). Commercially available cyclodextrins were evaluated for separation of the enantiomers and isomers of the three herbicides having varied molecular structures. The enantiomers of imazaquin and diclofop, and the isomers of imazamethabenz could be resolved with a resolution of ≥1.5. The resolution was found to depend on pH of the run buffer, cyclodextrin type and cyclodextrin concentration. By employing mixed cyclodextrins in the running buffer, the three herbicides were simultaneously separated in a single run. In addition, rapid (less than 3 min) enantiomeric separation is demonstrated using imazaquin as a model herbicide. The reported capillary electrophoresis (CE) methods are simple, rapid, efficient and reproducible and our results demonstrate that CE provides a powerful analytical tool for enantiomeric and isomeric separation of herbicides.

Migration order reversal of enantiomers in capillary electrophoretic separation

Optical resolution of (R,S)-1-benzyl-4-[(5,6-dimethoxy-1-indanon)-2-yl]methylpiperidine (E2020) was achieved by cyclodextrin-modified capillary zone electrophoresis (CD-CZE) and cyclodextrin-modified micellar electrokinetic chromatography (CD-MEKC). We intend to use the CD-MEKC mode to reverse the migration order of an enantiomer in the CD-CZE mode. For the (S)-enantiomer sample, including a varying amount of the (R)-enantiomer, the limit of quantitation (LOQ) was 3.0% in the CD-CZE mode, where the (S)-enantiomer migrated faster than the (R)-enantiomer, while it was 0.8% in the CD-MEKC mode, where the migration order of these enantiomers was reversed. It is quite useful to choose these two modes properly in order to obtain the precise optical purity. This technique is expected to be applied to optical purity tests of other chiral pharmaceuticals.

Enantioselective determination of oxprenolol and its metabolites in human urine by cyclodextrin-modified capillary zone electrophoresis

A stereospecific capillary electrophoresis assay for oxprenolol enantiomers and their basic metabolites in human urine has been developed using hydroxypropyl-β-CD as a chiral selector in the mobile phase. The bioassay method has been validated and the detection limit from spiked urine samples is 0.2μg/ml. The calibration curves are linear from 0.4 to 16 μg/ml. Extraction recovery ranged from 84.7 to 96.4% for all the compounds studied. The influence of various parameters on the chiral separation of oxprenolol and its basic metabolites have been investigated. Urinary excretion profiles of oxprenolol enantiomers and those of two metabolites have also been studied, following a single oral dose of racemic oxprenolol.