Chiral separation of flurbiprofen enantiomers by preparative and simulated moving bed chromatography

This study presents the chiral resolution of flurbiprofen enantiomers by preparative liquid chromatography using the simulated moving bed (SMB) technology. Flurbiprofen enantiomers are widely used as nonsteroidal anti‐inflammatory drugs, and although demonstrate different therapeutic actions, they are still marketed as a racemic mixture. The results presented here clearly show the importance of the selection of the proper solvent composition for the preparative separation of flurbiprofen enantiomers. Chiral SMB separation is carried out using a laboratory‐scale unit (the FlexSMB‐LSRE®) with six columns, packed with the Chiralpak AD® stationary phase (20 μm). Results presented include the experimental measurement of equilibrium and kinetic data for two very different solvent compositions, a traditional high hydrocarbon content [10%ethanol/90%n‐hexane/0.01% trifluoroacetic acid (TFA)] and a strong polar organic composition (100%ethanol/0.01%TFA). Experimental data, obtained using the two mobile phase compositions, are used to predict and optimize the SMB operation. After selecting 10%ethanol/90%n‐hexane/0.01%TFA as the most appropriate solvent composition, three feed concentrations of racemic flurbiprofen were considered. Using 40 g/l of racemic flurbiprofen feed solution, the purities for both outlet streams were above 99.4%, the productivity was 13.1 g_feed/(L_bed h), and a solvent consumption of 0.41 L_solvent/g_feed was achieved. Chirality, 2011. © 2011 Wiley‐Liss, Inc.     

Separation of Nadolol Stereoisomers Using Chiralpak IA Chiral Stationary Phase

Chiralpak IA adsorbent is used for both analytical and preparative chromatographic separation of nadolol stereoisomers. The results include a complete screening of the mobile phase composition for both the baseline resolution of all four nadolol stereoisomers (analytical separation) and the simulated moving bed (SMB) pseudo‐binary separation of the most retained stereoisomer. The experimental results show that analytical baseline resolution of nadolol stereoisomers can be achieved using alcohol/hydrocarbon and alcohol/acetonitrile solvent mixtures. The 10%ethanol/90%acetonitrile mixture is presented as the one that presents baseline resolution with lower retention. For the preparative pseudo‐binary separation, pure ethanol, pure methanol, alcohol/acetonitrile, and alcohol/tetrahydrofuran mixtures proved to allow good separation results. The 100%methanol/0.1%diethylamine solvent composition was selected to perform the experimental SMB separation. Using a 10 g/L total feed concentration, the more retained stereoisomer was recovered at the extract outlet stream with 99.5% purity, obtaining a system productivity of 1.98 gL^?1 h^?1 and requiring a solvent consumption of 3.13 L/g of product. Comparing these results with the ones recently presented by Ribeiro et al. (2013), this work shows that the Chiralpak IA chiral adsorbent is an interesting alternative to Chiralpak AD for the separation of nadolol stereoisomers at both analytical and preparative scales. Chirality 28:399–408, 2016. © 2016 Wiley Periodicals, Inc.     

The application of preparative batch HPLC, supercritical fluid chromatography, steady-state recycling, and simulated moving bed for the resolution of a racemic pharmaceutical intermediate

This article discusses the chromatographic resolution of a racemic pharmaceutical intermediate. Preparative batch high performance liquid chromatography (HPLC), supercritical fluid chromatography (SFC), steady-state recycling (SSR), and simulated moving bed (SMB) were used to resolve a total of 12.2 kg of a racemic pharmaceutical intermediate. In this study, a first batch of 0.8 kg of racemate was separated on the preparative batch HPLC and SFC, and subsequently another 5.9 kg of racemate was separated on the SSR. Lastly, a third batch of 5.5 kg was separated on the SMB. The separation conditions and results of these techniques are discussed. The productivities and solvent costs of SFC versus HPLC are compared. The productivities and solvent costs of SMB, SSR, and HPLC are also compared. The analytical method development and process optimization of these processes are also discussed in this article.     

Preparative scale enantioseparation of a chiral epoxide: Comparison of liquid chromatography and simulated moving bed adsorption technology

The feasibility of using simulated moving bed technology (SMB) for chiral separation on cellulose triacetate is demonstrated on the preparative scale: 1 kg of a chiral epoxide has been separated. On comparing SMB technology with conventional liquid chromatography it turns out that the main advantage of SMB lies in the significant reduction of mobile phase consumption. The process design for SMB is made theoretically and the predictions are confirmed by our pilot study. © 1993 Wiley-Liss, Inc.     

Improving the performance of nadolol stereoisomers' preparative separation using Chiralpak IA by SMB chromatography

The pseudobinary preparative separation of nadolol stereoisomers is performed by simulated moving bed chromatography (SMB). Using the Chiralpak IA adsorbent, a new 25:75:0.1 (v/v/v) methanol‐acetonitrile‐diethylamine solvent composition was selected to perform the experimental SMB separation and compare it with the previous results obtained using pure methanol. Using a 2 g L^?1 total feed concentration of an equimolar mixture of the four stereoisomers of nadolol, the more retained component was fully recovered (100% purity and 100% recovery), with a system productivity of 0.77 g L^?1 hour^?1 and a solvent consumption of 9.62 L g^?1. Comparing these results with the ones previously reported using 100:0.1 methanol‐diethylamine solvent composition, this work shows that the 25:75:0.1 methanol‐acetonitrile‐diethylamine is a better alternative for the preparative separation of nadolol stereoisomers by SMB chromatography. These results are confirmed by simulation of the SMB operation for higher feed concentrations, by comparing the performances of the two solvent compositions using the data obtained experimentally through the measurement of the adsorption equilibrium isotherms and the kinetic data obtained for both solvents. The new experimental and simulation results stress out that the performance of the preparative separation can be improved by a careful selection of the solvent composition.     

Separation of Tryptophan Enantiomers by Ligand‐Exchange Chromatography With Novel Chiral Ionic Liquids Ligand

Chiral ionic liquids (CILs) with amino acids as cations have been applied as novel chiral ligands coordinated with Cu²⁺ to separate tryptophan enantiomers in ligand exchange chromatography. Four kinds of amino acid ionic liquids, including [L‐Pro][CF₃COO], [L‐Pro][NO₃], [L‐Pro]₂[SO₄], and [L‐Phe][CF₃COO] were successfully synthesized and used for separation of tryptophan enantiomers. To optimize the separation conditions, [L‐Pro][CF₃COO] was selected as the model ligand. Some factors influencing the efficiency of chiral separation, such as copper ion concentration, CILs concentration, methanol ratio (methanol/H₂O, v/v), and pH, were investigated. The obtained optimal separation conditions were as follows: 8.0 mmol/L Cu(OAc)₂, 4.0 mmol/L [L‐Pro][CF₃COO] ,and 20% (v/v) methanol at pH 3.6. Under the optimum conditions, acceptable enantioseparation of tryptophan enantiomers could be observed with a resolution of 1.89. The results demonstrate the good applicability of CILs with amino acids as cations for chiral separation. Furthermore, a comparative study was also conducted for exploring the mechanism of the CILs as new ligands in ligand exchange chromatography. Chirality 26:160–165, 2014. © 2014 Wiley Periodicals, Inc.     

Enantiomeric Separations of Pyriproxyfen and its Six Chiral Metabolites by High‐Performance Liquid Chromatography

Pyriproxyfen is a chiral insecticide, and over 10 metabolites have been identified in the environment. In this work the separations of the enantiomers of pyriproxyfen and its six chiral metabolites were studied by high‐performance liquid chromatography (HPLC). Both normal phase and reverse phase were applied using the chiral columns Chiralpak IA, Chiralpak IB, Chiralpak IC, Chiralcel OD, Chiralcel OD‐RH, Chiralpak AY‐H, Chiralpak AD‐H, Chiracel OJ‐H, (R,R)‐Whelk‐O 1, and Lux Cellulose‐3. The effects of the chromatographic parameters such as mobile phase composition and temperature on the separations were investigated and the enantiomers were identified with an optical rotation detector. The enantiomers of these targets could obtain complete separations (resolution factor Rs > 1.5) on Chiralpak IA, Chiralpak IB, Chiralcel OD, Chiralpak AY‐H, or Chiracel OJ‐H under normal conditions. Chiralcel OJ‐H showed the best chiral separation results with n‐hexane as mobile phase and isopropanol (IPA) as modifier. The simultaneous enantiomeric separation of pyriproxyfen and four chiral metabolites was achieved on Chiralcel OJ‐H under optimized condition: n‐hexane/isopropanol = 80/20, 15°C, flow rate of 0.8 ml/min, and UV detection at 230 nm. The enantiomers of pyriproxyfen and the metabolites A , C , and D obtained complete separations on Chiralpak IA, Chiralpak IC, and Lux Cellulose‐3 under reverse phase using acetonitrile/water as the mobile phase. The retention factors (k) and selectivity factors (α) decreased with increasing temperature, and the separations were better under low temperature in most cases. The work is of significance for the investigation of the environmental behaviors of pyriproxyfen on an enantiomeric level. Chirality 28:245–252, 2016. © 2016 Wiley Periodicals, Inc.     

Separation of Ofloxacin and Its Six Related Substances Enantiomers by Chiral Ligand‐Exchange Chromatography

A chiral ligand‐exchange high‐performance liquid chromatography method was developed for the enantioseparation of ofloxacin and its six related substances termed impurities A, B, C, D, E, and F. The separation was performed on a conventional C₁₈ column. Different organic modifiers, copper salts, amino acids, the ratio of Cu²⁺ to amino acid, pH of aqueous phase, and column temperature were optimized. The optimal mobile phase conditions were methanol‐water systems consisting of 5 mmol/L copper sulfate and 10 mmol/L L‐isoleucine (L‐Ile). Under such conditions, good enantioseparation of ofloxacin and impurities A, C, E, and F could be observed with resolutions (R_S) of 3.54, 1.97, 3.21, 3.50, and 2.12, respectively. On the relationship between the thermodynamic parameters and structures of analytes, the mechanism of chiral recognition was investigated. It was concluded that ofloxacin and impurities A, C, E, and F were all enthalpically driven enantioseparation and that low column temperature was beneficial to enantioseparation. Furthermore, the structure–separation relationship of these analytes is also discussed. Chirality 27:843–849, 2015. © 2015 Wiley Periodicals, Inc.     

Chiral Separation of Four Stereoisomers of Ketoconazole Drugs Using Capillary Electrophoresis

This work aimed to develop a chiral separation method of ketoconazole enantiomers using electrokinetic chromatography. The separation was achieved using heptakis (2, 3, 6‐tri‐O‐methyl)‐β‐cyclodextrin (TMβCD), a commonly used chiral selector (CS), as it is relatively inexpensive and has a low UV absorbance in addition to an anionic surfactant, sodium dodecyl sulfate (SDS). The influence of TMβCD concentration, phosphate buffer concentration, SDS concentration, buffer pH, and applied voltage were investigated. The optimum conditions for chiral separation of ketoconazole was achieved using 10 mM phosphate buffer at pH 2.5 containing 20 mM TMβCD, 5 mM SDS, and 1.0% (v/v) methanol with an applied voltage of 25 kV at 25 °C with a 5‐s injection time (hydrodynamic injection). The four ketoconazole stereoisomers were successfully resolved for the first time within 17 min (total analysis time was 28 min including capillary conditioning). The migration time precision of this method was examined to give repeatability and reproducibility with RSDs ≤5.80% (n =3) and RSDs ≤8.88% (n =9), respectively. Chirality 27:223–227, 2015. © 2014 Wiley Periodicals, Inc.     

Separation of Glycopeptides by High Performance Liquid Chromatography

A method is presented for separation of tryptic glycopeptides-containing oligosaccharides of the N-asparagine-linked type. High performance liquid Chromatography (HPLC) of glycopeptides on a C18 reverse-phase system eluted with a gradient of 0%–50% acetonitrile in 0.1 M NaPO₄ pH 2.2 resolves the two major glycosylation sites from the envelope glycoprotein (G) of vesicular stomatitis virus. Glycopeptides containing N-linked oligosaccharides of the complex type coelute with those containing N-linked oligosaccharides of the neutral, high mannose type, indicating that separation is based upon peptide rather than carbohydrate composition. The contribution of the carbohydrate component to glycopeptide elution, as determined by cleavage of the high mannose oligosaccharides with endo-β-Nacetylglucosaminidase H, is that of a significant, but minor, decrease in peptide retention time. Comparison of the tryptic glycopeptide profiles of G isolated from both wild type and mutant strains of VSV illustrates the rapid, reproducible, and quantitative nature of the technique. Through HPLC analysis of appropriately treated glycopeptides, it is possible to explore both the nature and extent of glycosylation at individual sites in glycoproteins in a single step.     

Preparative resolution of the enantiomers of tert-leucine derivatives by simulated moving bed chromatography

The enantiomers of two different derivatives of tert-leucine were separated by continuous chromatography on chiral stationary phases applying the simulated moving bed technique. About 1 kg of racemic N-carbobenzoxy-tert-leucine was resolved on the cellulose-based phase Chiralcel OD using a mixture of heptane/ethanol and 0.1% of trifluoroacetic acid modifier as the mobile phase, while 520 g of the N-Boc-tert-leucine-benzylester was resolved on the amylose-based phase Chiralpak AD with a mixture of heptane/2-propanol as the mobile phase. In both instances the corresponding enantiomers were obtained in high yield and high optical purity.     

高速逆流色谱结合半制备液相色谱制备甘青青兰中3种活性成分

采用高速逆流色谱（HSCCC）结合液相色谱法制备甘青青兰（Dracocephalum tanguticum Maxim.）中绿原酸、胡麻甙-6″-乙酯、迷迭香酸,建立快速分离制备甘青青兰中活性成分的方法。采用半制备型高效液相色谱（SP-HPLC）富集甘青青兰乙酸乙酯萃取物中绿原酸、胡麻甙-6″-乙酯、迷迭香酸,再用制备液相（Pre-HPLC）和HSCCC对富集物进行分离纯化,获得54 mg化合物Ⅰ、130 mg化合物Ⅱ和200 mg化合物Ⅲ,纯度分别为96.9%、97.9%和95.1%,经核磁共振碳谱（¹³CNMR）和氢谱（¹HNMR）分别鉴定为绿原酸、胡麻甙-6″-乙酯和迷迭香酸。本实验方法适用于甘青青兰乙酸乙酯萃取物中绿原酸、胡麻甙-6″-乙酯和迷迭香酸的分离制备,并避免了传统分离方法操作繁多、试剂消耗量大、不可回收等弊端,为分离甘青青兰活性成分、制备对照品等研究提供了参考依据。     

高速逆流色谱分离制备甘草中的甘草苷和芒柄花苷

应用高速逆流色谱分离制备甘草中的甘草苷和芒柄花苷。将甘草乙酸乙酯提取物经聚酰胺柱粗分后,30%乙醇洗脱物用高速逆流色谱进一步分离,所用两相溶剂系统为乙酸乙酯-水(5∶5,v/v),转速850 rpm,流速2.0 mL/min,检测波长254 nm,从50 mg30%乙醇洗脱物中得到甘草苷8.7 mg、芒柄花苷4.2 mg,纯度分别为99.5%和97.3%。所得产物的结构经核磁共振谱(NMR)鉴定。利用该方法可以对甘草中的甘草苷和芒柄花苷进行快速的分离和纯化。     

Use of Sulfated Cyclofructan 6 and Sulfated Cyclodextrins for the Chiral Separation of Four Basic Pharmaceuticals by Capillary Electrophoresis

Sulfated cyclofructan 6 (S‐CF6) and sulfated cyclodextrins (S‐α‐, β‐, γ‐CDs) are highly selective chiral selectors for the enantioseparation of basic solutes. In this study, S‐CF6 was introduced for the enantiomeric separation of four basic pharmaceuticals (including tamsulosin, tiropramide, bupivacaine, and norephedrine) by capillary electrophoresis (CE), and the enantiomeric separation performance was compared with S‐α‐, β‐, γ‐CDs. The effects of the chiral selector type, chiral selector concentration, operating voltage, and column temperature were examined and optimized. Excellent resolutions were obtained for all solutes on these chiral selectors. Chirality 25:735–742, 2013. © 2013 Wiley Periodicals, Inc.     

Liquid Chromatographic Resolution of Mexiletine and Its Analogs on Crown Ether‐Based Chiral Stationary Phases

Mexiletine, an effective class IB antiarrhythmic agent, and its analogs were resolved on three different crown ether‐based chiral stationary phases (CSPs), one (CSP 1 ) of which is based on (+)‐(18‐crown‐6)‐2,3,11,12‐tetracarboxylic acid and the other two (CSP 2 and CSP 3 ) are based on (3,3’‐diphenyl‐1,1’‐binaphthyl)‐20‐crown‐6. Mexiletine was resolved with a resolution (R_S) of greater than 1.00 on CSP 1 and CSP 3 containing residual silanol group‐protecting n‐octyl groups on the silica surface, but with a resolution (R_S) of less than 1.00 on CSP 2 . The chromatographic behaviors for the resolution of mexiletine analogs containing a substituted phenyl group at the chiral center on the three CSPs were quite dependent on the phenoxy group of analytes. Namely, mexiletine analogs containing 2,6‐dimethylphenoxy, 3,4‐dimethylphenoxy, 3‐methylphenoxy, 4‐methylphenoxy, and a simple phenoxy group were resolved very well on the three CSPs even though the chiral recognition efficiencies vary with the CSPs. However, mexiletine analogs containing 2‐methylphenoxy group were not resolved at all or only slightly resolved. Among the three CSPs, CSP 3 was found to show the highest chiral recognition efficiencies for the resolution of mexiletine and its analogs, especially in terms of resolution (R_S). Chirality 26:272–278, 2014. © 2014 Wiley Periodicals, Inc.     

Studies of the Heterogeneity of a Candida cylindracea (rugosa) Lipase: Monitoring of Esterolytic Activity and Enantioselectivity by Chiral Liquid Chromatography

A method for the determination of lipase activity in terms of rate and enantioselectivity of hydrolysis of a chiral ester substrate has been developed. When this method was applied to fractions, isolated from preparative, column chromatographic separations (anion-exchange, molecular sieve) of the lipase, significant differences in enantioselectivity (E) was found between the fractions. The highest enantioselectivity was found in the first main peak obtained on DEAE-Sepharose chromatography, meaning that the enzyme with the highest isoelectric point shows the highest esterolytic enantioselectivity.

The experimental results are discussed in the light of some earlier reported results and with respect to the possible existence of subunit aggregates and isoenzymes.     

Stereoselective Determination of a Novel Chiral Insecticide,Sulfoxaflor, in Brown Rice,Cucumber and Apple by Normal‐Phase High‐Performance Liquid Chromatography

An effective high‐performance liquid chromatography method was developed for the stereoselective determination of a new sulfoximines insecticide, sulfoxaflor, in brown rice, cucumber and apple. Target compounds were extracted with acetonitrile and an aliquot cleaned with Cleanert PestiCarb/PSA (primary and secondary amine) cartridge. Five polysaccharide‐based columns were investigated on the separation of sulfoxaflor stereoisomers and the best was achieved on a ChromegaChiral CCA column with n‐hexane/ethanol/methanol (90:2:8, v/v/v) as mobile phase by UV detection at 220 nm at 20ºC. The resolutions of the four stereoisomers were 1.85, 1.54 and 3.08, and the elution order was identified by optical rotation and stereoisomers ratio. The mean recoveries of sulfoxaflor stereoisomers ranged from 77.1% to 99.3%, with relative standard deviations less than 8.9% at three concentration levels in all matrices. The limits of detection for all stereoisomers varied from 0.05 mg/kg to 0.07 mg/kg, while the limit of quantification did not exceed 0.22 mg/kg. The method was then successfully applied to determine the sulfoxaflor stereoisomers in authentic samples, confirming that it is convenient and reliable for stereoselective determination of sulfoxaflor stereoisomers in food. Chirality 26:114–120, 2014. © 2014 Wiley Periodicals, Inc.     

Xterra RP18柱高效液相色谱法快速分离测定氨基酸

建立了一种用XterraRP1 8色谱柱快速分离测定水解氨基酸的方法。所采用的色谱条件是 :WatersAlliance系统 ,柱温 5 6℃ ,流速 1 .8ml/min ,检测波长 2 4 8nm ,梯度分离 ,运行周期 2 5min,柱反压低于 2 0 0 0Psi。在 1 7.5min内分离了包括AMQ、NH3 和牛磺酸在内的 2 1种氨基化合物 ,适应于复合氨基酸注射液、含牛磺酸的氨基酸口服液及水解氨基酸样品的分析测定