毛细管区带电泳法研究肾上腺素类药物的手性分离

使用β－环糊精（β－ＣＤ）及β－ＣＤ－羧甲基（ＣＭ－β－ＣＤ）作为手性选择剂,采用毛细管区带电泳法（ＣＺＥ）对去甲肾上腺素、肾上腺素和异丙肾上腺素的手性分离进行了研究。对影响这类药物手性分离的主要因素〔手性选择剂、背景电解质（ＢＧＥ）、分离体系的酸度和温度〕进行了讨论,并对手性识别机理进行了探讨。     

Cyclodextrin-mediated micellar electrokinetic chromatography and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry for the enantiomer separation of racemorphan in human urine.

Micellar electrokinetic chromatography (MEKC) was successfully and conveniently applied to the chiral separation with the addition of cyclodextrins (CDs) as chiral selector to the running buffer. Chiral separation depended on the type of CD; in particular, beta-CD was effective for the chiral separation of racemorphan. We investigated the optimal conditions of type and concentration of CD as chiral selector for the routine enantiomeric separation of racemorphan with good reproducibility. The effects of other parameters such as buffer pH and detection wavelength were also investigated to obtain the optimum conditions for the enantiomeric separation of racemorphan. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry was used for confirmation of racemorphan. The optimal conditions for enantiomeric separation of the racemorphan were as follows: 50 mM borate buffer at pH 9.4 with 50 mM SDS, 10 mM beta-CD and 20% 1-propanol, 57 cm x 50 microns fused-silica capillary column, and UV detection at 192 nm. Based on the developed method, racemorphan in human urine was also separated and determined using solid-phase extraction and MEKC.     

用羧甲基聚合环糊精作手性剂的毛细管电泳法分离3种碱性药物

 采用 5种环糊精衍生物对碱性药物硫喷妥钠、盐酸氟桂利嗪、山梗菜碱进行了毛细管区带电泳的手性拆分。结果表明 ,采用含 2 % (质量分数 ,其余相同 )聚合 β 环糊精 (P β CD)或 0 5%羧甲基聚合 β 环糊精 (CM P β CD) 30mmol/L的Tris H3PO4缓冲液可使这 3种药物达到基线分离 ;使用CM P β CD时 ,分离度高达 4～35。     

Separation and control of the elution order of N-t-butyloxycarbonyl amino acids D/L isomers by reversed-phase HPLC using cyclodextrins as chiral selectors for the mobile phase.

The enantiomeric resolution of N-t-butyloxycarbonyl (N-t-Boc) amino acids D/L isomers by reversed-phase HPLC was investigated using cyclodextrins (CD's) as chiral selectors for the mobile phase. The use of a low pH (pH<4) for the mobile phase enabled the enantioseparation of N-t-Boc amino acids. The opposite elution order of D/L isomers was observed when hydroxypropyl-derivatized beta-CD was used instead of native beta-CD. A computer simulation of the enantioseparation showed that the ratio of the retention factors of the chiral selector and the sample determined the elution order and the resolution. When the retention factor of the chiral selector is smaller than that of the sample, an isomer having larger complex formation constant eluted faster. However, when the chiral selector had a larger retention factor than the sample, an opposite elution order of the isomers was obtained. The large difference in the retention factors between the chiral selector and the sample led to good enantiomeric separation.     

Chiral separation of synthetic vicinal diol compounds by capillary zone electrophoresis with borate buffer and beta-cyclodextrin as buffer additive.

The investigation on capillary electrophoretic enantioseparation of six synthetic compounds containing vicinal diol groups has been undertaken to acquire the optimum conditions using native beta-cyclodextrin (beta-CD) as chiral selector and borate as a background electrolyte. The separation was carried out in an uncoated capillary (58.5 cm x 75 microm i.d., effective length 48.5 cm) and the effects of several important factors were investigated in detail. The results showed that beta-CD as a chiral selector exhibited good enantioselectivity and that the enantioseparation was greatly influenced by the structure of the diols, the borate concentration and the buffer pH. The optimum performance was obtained for the chiral vicinal diols under the conditions of 200 mM borate buffer of pH 9.8 containing 1.7% beta-CD at an applied voltage of 15 kV and a capillary temperature of 20 degrees C. Under the conditions, four diols were baseline separated with fast analysis time and the good theoretical plate numbers (above 10 x 10(4)) and favorable migration-time reproducibilities (RSDs below 3.0%) were obtained. The separation results were satisfactory.     

Enatiomeric analysis of simendan by CE with beta-CD as chiral selector compared with CMPA-HPLC

The chiral separation of simendan enantiomers using capillary electrophoresis was studied with beta-cyclodextrin (beta-CD) as chiral selector. The influences of the concentration and pH of borate buffer solution, beta-CD concentration and methanol content in the background electrolyte were investigated. These factors were compared with those in an HPLC with beta-CD as chiral mobile phase additive (CMPA-HPLC). The quantification properties of the developed CE method were examined. A baseline separation of simendan enantiomers was achieved in the background electrolyte of 20 mmol/L borate buffer (pH 11.0) containing 12 mmol/L beta-CD-methanol (50:50 in volume ratio). The CE method is comparable with CMPA-HPLC in chiral resolution, although the optimal pH in CE (11.0) is much higher than that (6.0) in CMPA-HPLC. This chiral CE method is applicable to the quantitative ananlysis and enantiomeric excess value determination of L-simendan.     

Enantiomeric separation of synthetic 2,3-dihydroxy-3-phenylpropionate compounds by beta-cyclodextrin-modified capillary electrophoresis

A new capillary electrophoretic method was developed for enantiomeric separation and optical impurity analysis of three synthetic 2,3-dihydroxy-3-phenylpropionate compounds using native beta-cyclodextrin (beta-CD) as chiral selector and borate as a background electrolyte. The separation was carried out in uncoated capillary (58.5 cm x 75 microm I.D., effective length 48.5 cm). The results showed that beta-CD as the chiral selector exhibited good enantioselectivity and the baseline separation was obtained at pH 9.8, 200 mM borate buffer containing 1.7% beta-CD at applied voltage 15 kV and capillary temperature 20 degrees C within 15 min. The precision of each tested compound was less than 1.0% at migration time and 5.0% in corrected peak area and the accuracy of the method was in the range of 98.7-105%. Furthermore, the developed method was successfully applied to the determination of the undesirable trace (2S,3R)-(+)-form impurity in the synthetic (2R,3S)-(-)-2,3-dihydroxy-3-phenylpropionate samples.     

应用四甲基氢氧化铵控制电渗流的毛细管电泳方法分离多沙唑嗪及其中间体的对映体

建立了毛细管电泳手性分离多沙唑嗪中间体对映体的方法,并同时分离了多沙唑嗪对映体。考察了不同种类季铵盐对电渗流及分离的影响,其中四甲基氢氧化铵(TMB)能有效控制电渗流并提高组分的分离度。实验还考察了其他因素,如pH值、分离电压和磷酸二氢钠浓度对分离的影响。所用的毛细管为40 cm(有效长度30 cm)×50 μm,缓冲液为12 mmol/L β-环糊精、30 mmol/L TMB、60 mmol/L 磷酸二氢钠(pH 2.2),分离电压为20 kV。在此条件下多沙唑嗪及其中间体的对映体均达到了基线分离。实验结果表明,一些用β-环糊精不能完全分离的对映体通过加入TMB控制电渗流能达到满意的分离效果。     

Chiral separability of hydrophobic boron cluster anions with native cyclodextrins in water-methanol background electrolytes

Cluster anions of boron are built up on three-center two-electron bonds in contrast to naturally occurring compounds and their synthetic analogs. Methanol works as a solvent and as a competing agent, which advantageously adjusts reasonable strength of their interaction with native CDs in water-organic BGE. The highest methanol concentration preserving chiral discrimination of atropoisomers of individual anions is approximately 35, 55 and 75% v/v for alpha-, beta- and gamma-CD, respectively. alpha-CD separates anionic 7, 8-nido-dicarbaundecaborate clusters with small exo-skeletal substituents. beta-CD separates anions of all four tested structural types. The efficiency of separation of a compound with alpha- or beta-CD is always markedly lower than the separation efficiency at the absence of a CD in BGE. The efficiency of separation of a compound with beta-CD is always lower than the efficiency of separation of the compound with alpha-CD. gamma-CD was proved to be unsuitable as a chiral selector because in BGEs with gamma-CD, effective mobilities of analytes as well as their differences continuously decrease. The decrease was ascribed to the decomposition of the gamma-CD. The assessment of analytical prospect of alpha- and beta-CDs as chiral selectors for chiral separations of boron cluster anions requires knowledge of stability of individual CDs at the conditions of analyses and recognition of the chance to eliminate low separation efficiency.     

用去氧胆酸盐和β-环糊精的毛细管胶束电动色谱拆分手性药物

 用β－环糊精（β－ＣＤ）和去氧胆酸钠（ＳＤＣ）的环糊精改性毛细管胶束电动色谱，实际拆分了ＥＭＤ－５６４３１和扑尔敏两种手性药物，研究了ＳＤＣ和β－ＣＤ浓度及ｐＨ值对分离的影响。初步讨论了分离的机理。认为ＣＤ－ＳＤＣ体系中胶束单体分子几乎均被ＣＤ包合；ＣＤ可能与部分胶束单体包合而存在于ＳＤＣ的胶束中；该拆分体系中ＳＤＣ与β－ＣＤ的浓度比在４∶１～４∶３时拆分效果最好。并发现ＳＤＣ对β－ＣＤ有显著的增溶作用。     

Polymeric alkenoxy amino acid surfactants: II. Chiral separations of beta-blockers with multiple stereogenic centers

Two amino acid-based (leucine and isoleucine) alkenoxy micelle polymers were employed in this study for the separation of multichiral center-bearing beta-blockers, nadolol and labetalol. These polymers include polysodium N-undecenoxy carbonyl-L-leucinate (poly-L-SUCL) and polysodium N-undecenoxy carbonyl-L-isoleucinate (poly-L-SUCIL). Detailed synthesis and characterization were reported in our previous paper [26]. It was found that poly-L-SUCIL gives better chiral separation than poly-L-SUCL for both nadolol and labetalol isomers. The use of 50-100 mM poly-L-SUCIL as a single chiral selector provided separation of four and three isomers of labetalol and nadolol, respectively. Further optimization in separation of both enantiomeric pairs of nadolol and labetalol was achieved by evaluation of type and concentration of organic solvents, capillary temperature as well type and concentration of cyclodextrins. A synergistic approach, using a combination of poly-L-SUCIL and sulfated beta-CD (S-beta-CD) was evaluated and it showed dramatic separation for enantiomeric pairs of nadolol. On the other hand for labetalol enantiomers, separation was slightly decreased or remain unaffected using the dual chiral selector system. Finally, simultaneous separation of both nadolol and labetalol enantiomers was achieved in a single run using 25 mM poly-L-SUCIL and 5% w/v of S-beta-CD in less then 35 min highlighting the importance of high-throughput chiral analysis.     

Chiral separation of amino acids derivatized with fluoresceine-5isothiocyanate by capillary electrophoresis and laser-induced fluorescence detection using mixed selectors of beta-cyclodextrin and sodium taurocholate

Chiral separation of 20 pairs of amino acids derivatized with fluoresceine-5-isothiocyanate (FITC) by capillary electrophoresis and laser-induced fluorescence detection was studied using the mixture of beta-cyclodextrin (beta-CD) and sodium taurocholate (STC) as selector. Resolution was considerably superior to that obtained by using either beta-CD or STC alone. The molar ratio of beta-CD to STC of about 2:3 was found to be critical to achieve maximum separation. At this beta-CD-to-STC ratio, chiral separation occurred at really low total concentration of beta-CD and STC (<0.1 mM). Other impacting factors were investigated including the total concentration of beta-CD and STC, pH, and capillary conditioning procedure between two successive runs. Using a running buffer of 80 mM borate containing 20 mM beta-CD and 30 mM STC at pH 9.3, all of the 20 pairs of FITC-amino acid enantiomers were baseline resolved. The resolutions of the most pairs of the amino acid enantiomers (17 of 20) were higher than 3.0, only three pairs gave a resolution lower than 3.0 but higher than 1.90 (beta-phenylserine, pSer). The highest resolution reached 14.58 (Glu). Two derivatives of beta-CD, 2-hydroxypropyl-beta-CD (HP-beta-CD) and heptakis(2,6-di-O-methyl)-beta-CD (DM-beta-CD) were also explored. HP-beta-CD showed similar cooperative effect with STC, while DM-beta-CD together with STC led to poorer chiral separation.     

Enantioseparation of cetirizine by sulfated-beta-cyclodextrin-mediated capillary electrophoresis

A sulfated beta-cyclodextrin (sulfated beta-CD)-mediated capillary electrophoresis method is described for the enantioseparation of cetirizine using achiral cefazolin as an internal standard. The enantioseparation of the drug was performed in a borate buffer (5 mM, pH 8.7) with 1% sulfated beta-CD (w/v) as chiral selector at 10 kV. Several parameters affecting the separation were studied, including the pH and the concentration of borate buffer and chiral selector. Under optimized conditions, a baseline separation of two enantiomers was achieved in less than 7 min. Using cefazolin as an internal standard (IS), the linear range of the method for the determination of levocetirizine was over 1.0 to 50.0 microg/mL; the detection limit (signal-to-noise ratio = 3) of levocetirizine was 0.5 microg/mL. The method allowed the enantioseparation of cetirizine in bulk samples and enantiomeric purity evaluation of levocetirizine (R-enantiomer) in pharmaceutical tablets (Xyzal), and it was also found to be suitable for enantioseparation in human plasma.     

Modeling and optimization of the chiral selectivity of basic analytes in chiral capillary electrophoresis with negatively charged cyclodextrins using electrochemical detection

A mathematical model concerning the separation selectivity of basic analytes in chiral capillary electrophoresis (CE) modified with negatively charged cyclodextrins (CDs) has been presented to describe the dependence of chiral selectivity on the buffer pH and the chiral selector concentration. The electrophoretic method to determine the parameters of the model has also been developed. The model has been tested with racemic epinephrine and isoproterenol as target analytes and sulfonated beta-CD as chiral selector. The agreements have been found between the calculated and the measured values. Some significant conclusions to optimize chiral CE separation have been derived from the model and proven by the experiments. Electrochemical detection was used to meet the requirement of the low introduced concentration of analytes.     

Enantiomeric separation of citalopram and its metabolites by capillary electrophoresis

A simple and fast capillary electrophoretic method has been developed for the enantioselective separation of citalopram and its main metabolites, namely N-desmethylcitalopram and N,N-didesmethylcitalopram, using beta-cyclodextrin (beta-CD) sulfate as the chiral selector. For method optimisation several parameters were investigated, such as CD and buffer concentration, buffer pH, and capillary temperature. Baseline enantioseparation of the racemic compounds was achieved in less than 6 min using a fused-silica capillary, filled with a background electrolyte consisting of a 35 mM phosphate buffer at pH 2.5 supplemented with 1% w/v beta-CD sulfate and 0.05% w/v beta-CD at 25 degrees C and applying a voltage of -20 kV. A fast separation method for citalopram was also optimized and applied to the analysis of pharmaceutical formulations. Racemic citalopram was resolved in its enantiomers in less than 1.5 min using short-end injection (8.5 cm, effective length) running the experiments in a background electrolyte composed of a 25 mM citrate buffer at pH 5.5 and 0.04% w/v beta-CD sulfate at a temperature of 10 degrees C.     

Separation of basic drug enantiomers by capillary zone electrophoresis using glucuronyl glucosyl beta-cyclodextrin as a chiral selector.

Separations of basic drug enantiomers have been investigated using glucuronyl glucosyl beta-cyclodextrin (GUG beta-CD) as a chiral selector in the background electrolyte by capillary zone electrophoresis. The effects of GUG beta-CD concentration and running buffer pH on the migration times and resolution of 16 basic drug enantiomers were precisely examined using a linear polyacrylamide-coated capillary. High resolution of 16 basic drug enantiomers was generally attained with a running buffer pH 2.5 or 3.5 containing 10 mM GUG beta-CD. Next, we compared the chiral resolution abilities of GUG beta-CD with those of beta-CD and maltosyl beta-CD (G2 beta-CD). GUG beta-CD showed higher resolution for basic drug enantiomers tested than beta-CD and G2 beta-CD. This could be due to that hydrogen bonding or ionic interactions of uncharged and charged glucuronyl glucosyl groups of GUG beta-CD with an analyte could stabilize the inclusion complex.     

Enantioselective separation of azole compounds by EKC. Reversal of migration order of enantiomers with CD concentration

The enantioselective separation of a group of six weak base azole compounds was achieved in this work using EKC with three neutral beta-CDs as chiral selectors. The native beta-CD and two other beta-CD derivatives with different types and positions of the substituents on the CD rim ((2-hydroxy)propyl-beta-CD (HP-beta-CD) and heptakis-2,3,6-tri-O-methyl-beta-CD (TM-beta-CD)) were employed. Apparent binding constants for each pair compound-CD were determined in order to study analyte-CD interactions. The best enantiomeric resolutions for miconazole, econazole, and sulconazole were observed with HP-beta-CD whereas for the separation of the enantiomers of ketoconazole, terconazole, and bifonazole, TM-beta-CD was the best chiral selector. The enantioseparations obtained were discussed on the basis of the structure of the compounds taking into account that inclusion into the hydrophobic CD cavity occurred through the phenyl ring closer to the azole group. In addition, a change in the migration order for the enantiomers of two of the compounds studied (ketoconazole and terconazole) with the concentration of HP-beta-CD was observed for the first time.     

Separation of multicomponent mixtures of 2,4-dinitrophenyl labelled amino acids and their enantiomers by capillary zone electrophoresis

The use of capillary zone electrophoresis (CZE) for the separation of a group of 33 2,4-dinitrophenyl labeled amino acids (DNP-AA), including DNP-AA racemates, DNP-L-AA enantiomers and achiral DNP-AAs, was investigated. Alpha-, beta- and gamma-cyclodextrins (CDs) and their derivatives (hydroxypropyl derivatives of alpha-, beta- and gamma-CDs, polymeric beta-CD and 6A-methylamino-beta-cyclodextrin (MA-beta-CD)) served as complexing agents and chiral selectors in this investigation. Although native alpha- and gamma-CDs and their derivatives influenced the effective mobilities of the studied DNP-AAs in different ways, they generally failed to resolve enantiomers of the individual DNP-AAs. On the other hand, beta-CD and all of its derivatives were found to be effective in this respect. Of these, the best results were achieved with a positively charged MA-beta-CD and this chiral selector resolved enantiomers of ten DNP-AA racemates available for this study. However, a complete resolution of these enantiomers in one CZE run required that the effect of the chiral selector be complemented by complexing effects of polyvinyl pyrrolidone (PVP) or gamma-CD. Complexing and chiral recognition capabilities of MA-beta-CD combined with complexing effects of gamma-CD and PVP provided separating conditions suitable for the CZE separations of multicomponent mixtures of DNP-AAs with preserved resolutions of the enantiomers. For example, a mixture consisting of 43 DNP-AA constituents was resolved using an MA-beta-CD/gamma-CD combination with three peak overlaps.     

Enantioseparation of linear and cyclic chiral bis(phenethyl)amines by means of cyclodextrin-modified capillary electrophoresis

For two years drugs introduced to the market have had- to be enantiomerically pure. Rapid and cheap methods of high reproducibility must, therefore, be available for evaluation of enantiomeric purity. Within the framework of a larger project dealing with chiral recognition of phenethylamines by means of native and derivatized cyclodextrins it was intended to find capillary electrophoresis methods suitable for separation of the enantiomers of chiral bis(phenethyl)amines and their corresponding cyclic analogues, within 10 min, using small amounts of a chiral selector, to save time and money. Heptakis(2,3-O-diacetyl-6-sulfato)beta-CD was found to be the most promising candidate most often fulfilling these requirements.     

Simultaneous separation and enantioresolution of racemic local anesthetic drugs by capillary zone electrophoresis with Tween 20 and methyl-beta-cyclodextrin as selectors, employing a double plug technique

A new approach for simultaneous chiral and achiral separations by capillary zone electrophoresis is described. Two adjacent selector plugs, consisting of Tween 20 as an achiral and methyl-beta-cyclodextrin (CD) as a chiral selector, are employed and four related local anesthetics are used as model compounds. The principles of the partial filling technique, whereby the capillary is filled with the chiral selector solution followed by the micellar solution at different plug lengths and concentrations, prior to application of the solutes, was employed. During the run both capillary ends were dipped in a simple buffer, i.e., one without additives. The two separation media worked independently without any interaction. Separation of the solutes and their enantiomers was regulated by adjusting both the concentration and plug length (PL) of the micellar solution in the capillary, employing methyl beta-CD as chiral selector either at 38 or 76 mM. The solutes were separated on the basis of their affinity towards the micellar phase before they reached the methyl-beta-CD plug for enantioseparation. In the absence of the micellar plug, the enantiomers of prilocaine overlapped those of bupivacaine. The solutes and their enantiomers were completely separated by employing two adjacent plugs consisting of 100 mM Tween 20 solution (PL approximately 10 cm) and methyl-beta-CD solution at either 38 or 76 mM (PL approximately 30 cm).