Amylose Tris Research Articles

Liquid Chromatographic Enantiomer Separation of N‐Phthaloyl Protected α‐Amino Acids on Coated and Immobilized Chiral Stationary Phases Derived from Polysaccharide Derivatives

Liquid chromatographic enantiomer separation of N‐phthaloyl (PHT) protected α‐amino acids on several coated and immobilized chiral stationary phases (CSPs) derived from polysaccharide derivatives was performed. The coated CSP of Chiralpak AD showed more or less enantioseparation than the covalently bonded CSP of Chiralpak IA with the same chiral selector of amylose tris(3,5‐dimethylphenylcarbamate). However, the coated Chiralcel OD showed greater enantioseparation than the covalently bonded Chiralpak IB with the same chiral selector of cellulose tris(3,5‐dimethylphenylcarbamate). Among all examined CSPs, Chiralcel OD afforded the greatest performance for enantiomer resolution of N‐PHT α‐amino acids and, therefore, all analytes enantiomers were baseline separated on Chiralcel OD. The chromatographic method developed in this study was usefully applied for determination of the enantiomeric purity of commercially available N‐PHT α‐amino acids analytes.

Enantiomeric Determination of Tramadol and O-Desmethyltramadol by Liquid Chromatography-Mass Spectrometry and Application to Postoperative Patients Receiving Tramadol

A liquid chromatographic-mass spectrometric assay with atmospheric pressure chemical ionization is presented for quantification (selected-ion mode) of tramadol (T) and O-desmethyltramadol (ODT) in blood plasma after liquid-liquid extraction. The enantiomeric separation was achieved on a Chiralpak AD column containing amylose tris-(3,5-dimethylphenylcarbamate) as chiral selector. The validation data were within acceptable limits. The assay was successfully applied to authentic plasma samples allowing confirmation of diagnosis of overdose or intoxication as well as monitoring of patients' compliance. In postoperative patients receiving T, mean therapeutic plasma concentrations were determined as follows: 422.8 and 440.8 ng/mL for (+)-T and (-)-T, respectively, and 23.7 and 40.7 ng/mL for (+)-ODT and (-)-ODT, respectively. Quantitative results demonstrated a great interindividual variability in postoperative plasma analgesic drug concentrations and, therefore, a wide therapeutic concentration range. The 7.7% of the samples that revealed negative results for ODT are likely explained by genetic polymorphisms resulting in absent enzyme activity of CYP2D6.

Interpretation of chromatographic retentions of simple solutes with an amylose-based sorbent using infrared spectroscopy and DFT modeling

The interactions of amylose tris(3,5-dime- thylphenylcarbamate) (ADMPC, commercially “Chiralpak AD”) with 10 simple solutes—1-propanol, heptane, heptanol, benzene, propylbenzene, benzyl alcohol, pyridine, tetrahydrofuran, diethylamine, and aniline—are studied using attenuated total reflection infrared spectroscopy (ATR-IR) of thin polymer films, DFT modeling, and high performance liquid chromatography (HPLC). ATR-IR is used to determine the changes in the hydrogen bonding states of the C=O and NH groups of the polymer amide I and II bands upon absorption of each of the solutes at 25∘C. DFT modeling with B3LYP/6-311+g(d,p) level of theory is used to predict the IR wavenumbers, the H-bonding interaction energies, and the hydrogen bonding distances of the polymer side-chains with certain solute molecules. The capacity factors of these solutes with ADMPC have been measured at 25∘C in hexane/isopropanol (95/5, v/v) solvent. From IR data and DFT modeling, we conclude that the C=O and NH are key binding sites of the polymer and interact with the functional groups of various solutes. The capacity factors are understood on the basis of hydrogen bonding, hydrophobic, and dipole-dipole interactions of the C=O, NH, and phenyl groups of the sorbent with OH, NH, NH2, O, phenyl, and N functional groups of the solutes.

Direct Probing of Sorbent−Solvent Interactions for Amylose Tris(3,5-dimethylphenylcarbamate) Using Infrared Spectroscopy, X-ray Diffraction, Solid-state NMR, and DFT Modeling

The sorbent-solvent interactions for amylose tris(3, 5-dimethylphenylcarbamate) (ADMPC) with five commonly used solvents, hexane, methanol, ethanol, 2-propanol (IPA), and acetonitrile (ACN), are studied using attenuated total reflection infrared spectroscopy (ATR-IR) of thin sorbent films, X-ray diffraction (XRD) of thin films, (13)C cross polarization/magic angle spinning (CP/MAS) and MAS solid state NMR of polymer-coated silica beads (commercially termed "Chiralpak AD"), and DFT modeling. The ADMPC-polymer-coated silica beads are used commercially for analytical and preparative scale separations of chiral enantiomers. The polymer forms helical rods with intra- and inter-rod hydrogen bonds (H-bonds). There are various nm-sized cavities formed between the polymer side-chains and rods. The changes in the H-bonding states of the C=O and NH groups of the polymer upon absorption of each of the five solvents at 25 degrees C are determined with ATR-IR. The IR wavenumbers, the H-bonding interaction energies, and the H-bonding distances of the polymer side-chains with each of the solvent molecules are predicted using the DFT/B3LYP/6-311+g(d,p) level of theory. The changes in the polymer crystallinity upon absorption of each solvent are characterized with XRD. The changes in the polymer crystallinity and the H-bonding states of C=O groups are also probed with (13)C CP/MAS solid-state NMR. The changes in the polymer side-chain mobility are detected using (13)C MAS solid-state NMR. The H-bonding states of the polymer change upon absorption of each polar solvent and usually result in an increase in the polymer crystallinity and the side-chain mobility. The polymer rods are reorganized upon solvent absorption, and the distance between the rods increases with the increase in the solvent molecular size. These results have implications for understanding the role of the solvent in modifying the structure and behavior of the polymer sorbents.

An improved and validated LC method for resolution of bicalutamide enantiomers using amylose tris-(3,5-dimethylphenylcarbamate) as a chiral stationary phase

An improved HPLC method for determination of enantiomeric purity of bicalutamide in drugs and pharmaceuticals was developed and validated. Baseline separation with resolution ≥6.0 was achieved within 10 min on Chiralpak AD-H (250 mm × 4.6 mm; particle size 5 μm) column using n-hexane:2-propanol (65:35 v/v) as mobile phase at a flow rate of 1.0 ml/min at 25 °C. The detection was made at 270 nm using UV detector while a polarimetric detector connected in series was used for identification of enantiomers. The effects of 2-propanol, ethanol and temperature on enantioselectivity and resolution of enantiomers were evaluated. The method was validated in terms of accuracy, precision and linearity in the range of 10–250 μg/ml and the r 2 was >0.9999. The recoveries were 99.68–100.25% with <1% R.S.D. The limits of detection (LOD) and quantification (LOQ) of enantiomers were (2.4, 3.0 and 7.6, 9.3) × 10 −8 g/ml for ( S)-(+)-BCT and ( R)-(−)-BCT enantiomers, respectively. The method was found to be suitable for rapid determination of enantiomeric purity of bicalutamide in bulk drugs and pharmaceutical formulations.

The role of molecular interaction fields on enantioselective and nonselective separation of chiral sulfoxides

The separation of a series of 23 asymmetric sulfoxides, including the three proton pump inhibitors (PPI) omeprazole, lansoprazole and pantoprazole was investigated by HPLC, under reversed-phase elution with amylose tris(3,5-dimethylphenylcarbamate), amylose tris[( S)-1-phenylethylcarbamate] and amylose tris(3,5-dimethoxyphenylcarbamate) chiral stationary phases, CSP1-3, respectively. The whole set of sulfoxides showed better enantioselectivity and enantioresolution on CSP1. However, the three PPI were enantioseparated only when using CSP1 and CSP3. It was observed an improved enantioselectivity and enantioresolution on CSP3. The mechanisms of retention were evaluated by molecular interaction fields (MIF) generated via GRID force field, which yielded the geometric reasons leading to the scenario outlined. The enantioselective and nonselective interactions are discussed in terms of the reported selectivity. The steric structural outline of the CSP nonselective interaction sites is of major importance to deliver the sulfoxides to the chiral selective sites where the enantioselective interactions take place.

Chiral separation of glycidol enantiomers by normal-phase high-performance liquid chromatography coupled to atmospheric pressure chemical ionization mass spectrometry

A chiral separation method for glycidol enantiomers determination by normal-phase high-performance liquid chromatography coupled to atmospheric pressure chemical ionization mass spectrometry was developed. Two chiral stationary phases, amylose tris–(3,5-dimethylphenylcarbamate) (Chiralpak AD-H) and ( S)-indoline-2-carboxylic acid and ( R)-1-(α-naphthyl) ethylamine (SUMICHIRAL OA-4900) have been investigated. The effects of the mobile phase composition, elution program and column temperature were also studied. Under the best conditions: Chiralpak AD-H column, mobile phase composition n-hexane:ethanol (70:30, v/v), flow rate of 0.8 mL/min and 40 °C column temperature, a good resolution (Rs = 1.6) for both enantiomers has been achieved with an analysis time of 16 min. The method was found to be linear in the range from 100 to 500 ppm for both glycidol enantiomers with a good determination coefficient ( r 2 higher than 0.99) and good precision. Limits of detection of 31 and 50 ppm for ( R)-(+)-glycidol and ( S)-(−)-glycidol, respectively, were obtained. The method was applied to the determination of the enantiomeric excess and yield obtained in a asymmetric epoxidation process of allyl alcohol with a chiral titanium–tartrate complex as catalyst.

Enantiospecific resolution of rabeprazole by liquid chromatography on amylose-derived chiral stationary phase using photo diode array and polarimetric detectors in series

A simple and rapid liquid chromatographic method for enantioselective separation and determination of R-(+) and S-(−) enantiomers of rabeprazole in drugs and pharmaceuticals using photo diode array (PDA) and polarimetric detectors connected in series was developed. Chiralpak AD-H (250 mm × 4.6 mm) 5 μm column packed with amylose tris(3,5-dimethylphenyl carbamate) as a stationary phase and the mobile phase containing n-hexane:ethanol:2-propanol(75:15:10, v/v/v) in an isocratic mode has yielded baseline separation with resolution greater than 3.0 at 40 °C. Effects of ethanol, 2-propanol and temperature on separation were studied for optimum resolution. Lansoprazole sulphone was used as an internal standard (IS) for quantitative determination of individual enantiomers in bulk drugs as well as pharmaceutical formulations. The method was validated in terms of accuracy, precision and linearity according to ICH guidelines. The linearity of the method was studied in the range of 0.5–50 μg/ml and the r 2 was >0.9997. The inter- and intra-day precision of assay were determined (R.S.D. < 1%) and the recoveries were in the range of 99.63–100.22% with <1% R.S.D. The limits of detection (LOD) and quantification (LOQ) were 0.02 μg/ml and 0.07 μg/ml for both the enantiomers, respectively.

Enantiomeric resolution of doxazosin mesylate and its process-related substances on polysaccharide chiral stationary phases

High-performance liquid chromatographic methods for separation of racemic doxazosin mesylate and its synthetic precursors on polysaccharide based stationary phases viz., amylose tris-(3,5-dimethylphenylcarbamate) (Chiralpak AD-H) and cellulose tris-(3,5-dimethylphenylcarbamate) (Chiralcel OD-H) were developed. The base line separation with R s > 1.50 was obtained using a mobile phase containing n-hexane–alcohol–0.1% diethylamine (ethanol, 1-propanol and 2-propanol) in various proportions. The effect of concentration of the alcoholic modifiers on the resolution was studied. A good separation was achieved on amylose based Chiralpak AD-H column when compared with cellulose based Chiralcel OD-H. The effects of structural features of the solutes and solvents on discrimination between the enantiomers were examined. The detection was carried out at 240 nm with UV detector while identification by polarimetric detector connected in series. The method was suitable not only for process development of doxazosin mesylate but also determination of enantiomeric purity of bulk drugs and pharmaceuticals.

High-performance liquid chromatographic enantioseparations on capillary columns containing monolithic silica modified with amylose tris(3,5-dimethylphenylcarbamate)

Monolithic capillary columns containing native silica were modified by in situ coating with amylose tris(3,5-dimethylphenylcarbamate) and applied for enantioseparations in capillary liquid chromatography. Capillary columns were examined for 10 standard racemic compounds in order to compare the performance of monolithic silica columns with the common, 4.6 mm I.D. high-performance liquid chromatographic columns packed with particulate silica. The effects of polysaccharide coating and of the linear velocity of the mobile phase on peak performance were studied. Enantioseparations with an analysis time below 1 min were achieved for some chiral analytes.

Enantiomeric separation of chiral pesticides by high-performance liquid chromatography on an amylose tris-(S)-1-phenylethylcarbamate chiral stationary phase

Amylose tris-(S)-1-phenylethylcarbamate chiral stationary phase (CSP) was prepared. The direct enantiomeric separation of chiral pesticides on this CSP had been studied by HPLC. The mobile phase was n-hexane-isopropanol at a flow rate of 1.0 mL/min. The effects of isopropanol content and column temperature on retention and enantioselectivity were investigated. Thirty-two samples were tested, of which ten interacted enantioselectively with the CSP. Five samples were completely resolved and another five underwent near-baseline or partial resolution. The enantiomers were identified by a circular dichroism detector. Linear van't Hoff plots were established and the thermodynamic parameters were thus calculated.

Application of cellulose and amylose arylcarbamates as chiral selectors in counter-current chromatography

Here we studied the applicability of cellulose and amylose tris(3,5-dimethylphenylcarbamate) as chiral selectors for the separation of enantiomers by counter-current chromatography (CCC). A variety of organic/aqueous biphasic solvent systems and eight racemic analytes of acidic, neutral, and basic nature were tested for this purpose. Classical elution mode and pH-zone-refining displacement conditions were used. Partial enantioseparation of pindolol and warfarin was achieved in methyl isobutyl ketone (MIBK)/aqueous solution and methyl tert-butyl ether (MTBE)/aqueous solution, respectively. For these two racemates enantiomeric excess values from 84% to 97% were achieved under the best conditions tested. However, resolution by CCC was not achieved for propranolol, naproxen and N-(3,5-dinitrobenzoyl)-(±)-leucine, easily separated in HPLC by the same selectors.

Enantioselective chromatography and absolute configuration of N,N‐dimethyl‐3‐(naphthalen‐2‐yl)‐butan‐1‐amines: Potential sigma1 ligands

We describe the preparation of racemic N,N-dimethyl-3-(naphthalen-2-yl)-butan-1-amines, potential sigma1 ligands, and their resolution via chiral HPLC. In order to obtain enantiopure compounds, direct chromatographic methods of separation using chiral stationary phases were investigated. Different methods suitable for both analytical and semipreparative purposes are proposed. The best resolutions were achieved using cellulose tris (3,5-dimethylphenyl carbamate) (Chiralcel OD and OD-H) and amylose tris (3,5-dimethylphenyl carbamate) (Chiralpak AD). On the basis of the preliminary chromatographic results, the resolution of compound 1 was transferred onto a Chiralcel OD semipreparative column. The enantiomers were obtained in high enantiomeric excess. The configurational assignment was performed by circular dichroism. Computational analysis was used to explore the enantioselective recognition process of compound 1 with the Chiralcel OD stationary phase.

Enantioselective determination of a gastroprokinetic drug using amylose tris-(3,5-dimethylphenylcarbamate) as a stationary phase by HPLC

An enantioselective high-performance liquid chromatographic method for determination of enantiomers of mosapride citrate in bulk drugs and pharmaceuticals using UV–vis and polarimetric detectors in series has been developed. Baseline separation with resolution >2.0 was achieved on a column containing amylose tris-(3,5-dimethylphenylcarbamate) as stationary phase using a mobile phase consisting of n-hexane:ethanol:triethylamine (80:20:0.3, v/v/v) at 40 °C. The detection was carried out at UV-276 nm and enantiomers were identified by polarimetric detector. The effect of ethanol, 2-propanol, TEA, temperature and mobile phase flow rate on separation of MSP enantiomers was studied and the method was validated with respect to accuracy, precision, linearity and limits of detection and quantification. The linearity of the method was studied between 6.25 and 50 μg/ml and r 2 was >0.9997. The recoveries were in the range 99.63–100.22%, the method was suitable not only for process development of mosapride citrate but also for quality assurance of the individual enantiomers in bulk drugs and pharmaceuticals.

Immobilized versus coated amylose tris(3,5-dimethylphenylcarbamate) chiral stationary phases for the enantioselective separation of cyclopropane derivatives by liquid chromatography

The solvent versatility of Chiralpak IA, a new chiral stationary phase (CSP) containing amylose tris(3,5-dimethylphenylcarabamate) immobilized onto silica gel, is investigated for the enantioselective separation of a set of cyclopropane derivatives using ethyl acetate or dichloromethane (DCM) as non-standard mobile phase eluent and diluent, respectively in high-performance liquid chromatography (HPLC). A comparison of the separation of cyclopropanes on both immobilized and coated amylose tris(3,5-dimethylphenylcarbamate) chiral stationary phases (Chiralpak IA and Chiralpak AD, respectively) in HPLC using a mixture of n-hexane/2-propanol (90/10 and 99/1, v/v) as mobile phase with a flow rate of 0.5 ml/min and UV detection at 254 nm, is demonstrated. The optimized method of separation is used for an online HPLC monitoring for the Rh(II)-catalyzed asymmetric intermolecular cyclopropanations in dichloromethane. Direct analysis techniques without further purification, workup or removal of dichloromethane were summarized. The method provides an easy and direct determination of the enantiomeric excess of the cyclopropanes and selectivity of the catalyst used without any further work up.

Comparison, Applications, Advantages, and Limitations of Immobilized and Coated Amylose Tris‐(3,5‐Dimethylphenylcarbamate) Chiral Stationary Phases in HPLC

A direct high performance liquid chromatographic enantioselective separation of a set of racemic acidic drugs on the new immobilized and conventional coated amylose tris‐(3,5‐dimethylphenylcarbamate) chiral stationary phases (Chiralpak IA and Chiralpak AD, respectively) was studied using n‐hexane and 2‐propanaol (80∶20 v/v), containing TFA (0.1%) as mobile phase. The separation and elution order of the enantiomers on both columns under the same conditions were compared. The effect of the immobilization of the amylose tris‐(3,5‐dimethylphenylcarbamate) chiral stationary phase on silica (Chiralpak IA) on the chiral recognition ability was noted, as the coated phase (Chiralpak AD) possesses a higher resolving power than the immobilized one (Chiralpak IA). A few racemates, which were not or poorly resolved on the immobilized Chiralpak IA were most efficiently resolved on the coated Chiralpak AD. However, the immobilized phase withstands the prohibited HPLC solvents such as dichloromethane, ethyl acetate, tetrahydrofuran, and others when used as eluents or as a dissolving agent for the analyte itself. The versatility of the immobilized Chiralpak IA in monitoring reactions performed in dichloromethane using direct analysis techniques without further purification, workup, or removal of dichloromethane, was studied on a representative example consisting of the lipase‐catalyzed enantioselective esterification of flurbiprofen, with n‐butanol in dichloromethane as organic solvent.

Comparison, Applications, Advantages, and Limitations of Immobilized and Coated Amylose Tris‐(3,5‐Dimethylphenylcarbamate) Chiral Stationary Phases in HPLC

A direct high performance liquid chromatographic enantioselective separation of a set of racemic acidic drugs on the new immobilized and conventionally coated amylose tris‐(3,5‐dimethylphenylcarbamate) chiral stationary phases (Chiralpak IA and Chiralpak AD, respectively) was studied using n‐hexane and 2‐propanaol (80∶20 v/v) containing TFA (0.1%) as mobile phase. The separation and elution order of the enantiomers on both columns under the same conditions were compared. The effect of the immobilization of the amylose tris‐(3,5‐dimethylphenylcarbamate) chiral stationary phase on silica (Chiralpak IA), on the chiral recognition ability was noted, as the coated phase (Chiralpak AD) possesses a higher resolving power than the immobilized one (Chiralpak IA). A few racemates, which were not resolved or were poorly resolved on the immobilized Chiralpak IA were most efficiently resolved on the coated Chiralpak AD. However, the immobilized phase withstands the prohibited HPLC solvents such as dichloromethane, ethyl acetate, tetrahydrofuran, and others, when used as eluents or as a dissolving agent for the analyte itself. The versatility of the immobilized Chiralpak IA in monitoring reactions performed in dichloromethane using direct analysis techniques without further purification, workup, or removal of dichloromethane, was studied on a representative example consisting of the lipase‐catalyzed enantioselective esterification of flurbiprofen with n‐butanol in dichloromethane as organic solvent.

On the solvent versatility in immobilized amylose tris(3,5-dimethylphenylcarbamate) chiral stationary phase in high performance liquid chromatography: Application to the asymmetric cyclopropanation of olefins

The enantioseparation of a set of cyclopropanes derived from Meldrum's acid and 3,3,3-trifluoro-2-diazopropionate has been achieved on the new immobilized amylose tris(3,5-dimethylphenylcarbamate) chiral stationary phase (Chiralpak IA) in high performance liquid chromatography (HPLC) using a mixture of n-hexane–2-propanol (90:10, or 99:1, v/v) as mobile phase with a flow-rate of 0.5 ml/min and UV detection at 254 nm. Due to its ability to withstand prohibited HPLC solvents like dichloromethane, an online HPLC equipped with a sensitive optical rotation detector for monitoring the optical activity of the racemic and enantioenriched cyclopropanes prepared via the Rh(II)-catalyzed asymmetric inter- and intramolecular cyclopropanations in dichloromethane is demonstrated. Direct analysis techniques without further purification, workup or removal of dichloromethane were summarized. The method provides an easy and direct determination of the enantiomeric excess of the cyclopropanes and selectivity of the catalyst used without any further work up.

Comparative Liquid Chromatographic Enantiomer Resolution on Two Chiral Stationary Phases Derived from Amylose Tris(3,5-dimethylphenylcarbamate)

Polysaccharide-derived chiral stationary phases (CSPs) are known to show high chiral recognition ability in HPLC and have been extensively used to separate a broad range of racemic compounds. These type CSPs are usually prepared coating or adsorbing the polysaccharide derivatives on silica gel. Therefore, the solvents such as chloroform, methylene chloride, and tetrahydrofuran which dissolve or swell the chiral selectors of the polysaccharide derivatives cannot be used as mobile phases. For example, in case of Chiralpak AD prepared by coating amylose tris(3,5dimethylphenylcarbamate) derivative, one of the widely used polysaccharide type CSPs, suitable mobile phases like hexane, 2-propanol and ethanol should be used for the column safety. It may be damaged in case of even the use of only a little amount of inappropriate solvents used as mobile phases and/or sample solvents. These limitations represent a disadvantage for new applications using these CSPs and, especially, preparative separation due to solubilization of analytes. To overcome these problems, therefore, the development of polysaccharide-derived covalently bonded CSPs has been of great interest and various results of different attempts have been reported. Recently, Chiralpak IA prepared by chemically bonding amylose tris(3,5-dimethylphenylcarbamate) derivative on silica gel, which is used as the same chiral selector of coated type Chiralpak AD has been introduced. In this study, we present the comparative liquid chromatographic enantiomer resolution of N-fluorenylmethoxycarbonyl (FMOC) protected αamino acids ethyl ester derivatives on two polysaccharidederived CSPs, covalently bonded type Chiralpak IA and coated type Chiralpak AD. This is the first report concerning the comparison of the chiral separations on Chiralpak IA and Chiralpak AD using normal mobile phases. For enantiomer separation of N-FMOC α-amino acids ester derivatives, very a few results on CSPs have been reported. Rizzi has separated three FMOC α-amino acids methyl esters enantiomers on cellulose triacetate type column. Miyazawa et al. have reported on the resolution of only one analyte of FMOC 2-aminobutanoic acid methyl ester enantiomers among many different N-protected amino acid derivatives. Kusters et al. have reported on the resolution of fifteen N-FMOC α-amino acids methyl and isopropyl esters enantiomers on a polysaccharide-derived CSP. To our knowledge, our results are the first reported for the enantiomer resolution of N-FMOC α-amino acids ethyl ester derivatives. Table 1 shows the effect of mobile phase on the enantiomer separation of some N-FMOC α-amino acids ethyl esters on Chiralpak IA. The enantioselectivities and retention times are greatly influenced by the nature of mobile phase. As shown in Table 1, in general, 5% 2propanol in hexane as a mobile phase afforded the greatest enantioselectivity with the highest resolution factor, whereas 20% chloroform in hexane afforded the lowest enantioselectivity. It is notable that the elution orders of three analytes using 10% tetrahydrofuran or 10% ethyl acetate or 20% chloroform in hexane are different from those using 5% 2-propanol in hexane. Table 2 shows the comparative data of enantiomer separation of N-FMOC α-amino acids ethyl esters on Chiralpak IA and Chiralpak AD using 2-propanol in hexane as a mobile phase. In general, Chiralpak IA showed slightly lower enantioseparation than Chiralpak AD for enantioresolution of N-FMOC α-amino acids ethyl esters. Several results have reported that a certain decrease in the enantio-

Development and validation of HPLC methods for enantioseparation of mirtazapine enantiomers at analytical and semipreparative scale using polysaccharide chiral stationary phases

Novel HPLC methods were developed for the analytical and semipreparative resolution of new antidepressant drug mirtazapine enantiomers. At analytical scale, the separation of the mirtazapine enantiomers was investigated using both cellulose and amylose tris(3,5-dimethylphenylcarbamate) (CDMPC and ADMPC) chiral stationary phases under normal-phases and polar organic modes. Good baseline enantioseparation was achieved using cellulose tris(3,5-dimethylphenylcarbamate) chiral stationary phases under both normal-phases and polar organic modes. Furthermore, the elution order of mirtazapine enantiomic pairs was found reversed by changing the stationary phase from the amylose-based ADMPC–CSPs to its cellulose-based counterpart, CDMPC–CSPs. The validation of the analytical methods including linearity, limit of detection (LOD), limit of quantification (LOQ), recovery and precision, together with the semipreparative resolution of mirtazapine racemate were carried out using cellulose tris(3,5-dimethylphenylcarbamate) chiral stationary phases and methanol as mobile phase without any basic additives under polar organic mode. At analytical scale, the elution times of both enantiomers were less than 6 min at normal temperature and 1.0 ml/min, with the separation factor ( α) 1.99 and the resolution factor (Rs) 3.56. Then, the analytical methods were scaled up to semipreparative loading to obtain small quantities of both mirtazapine enantiomers. At semipreparative scale, about 16 mg/h enantiomers could be isolated and elution times of both enantiomers were less than 10 min at 2.0 ml/min. To increase the throughput, the technique of boxcar injections was used. One enantiomer ((−)-( R)-mirtazapine) was isolated with purity of >99.9% e.e. and >98.0% yield and another ((+)-( S)-mirtazapine) was isolated with purity of >97.0% e.e. and >99.0% yield. In addition, optical rotation and circular dichroism (CD) spectroscopy of both mirtazapine enantiomers isolated were also investigated.