Chiral Mobile Phase Additives Research Articles

Analytical enantioseparation of N-alkyl drugs by reversed-phase liquid chromatography with carboxymethyl-β-cyclodextrin as mobile phase additive.

Carboxymethyl-β-cyclodextrins (CM-β-CDs) with five kinds of degrees of substitution were synthesized and characterized. Analytical enantioseparation of six basic drugs containing N-alkyl groups, including pheniramine, chlorpheniramine, labetalol, propranolol, venlafaxine, and trans-paroxol, was achieved by reversed-phase high-performance liquid chromatography (RP-HPLC) using the synthesized CM-β-CD as chiral mobile phase additives. Key influence factors were optimized, including organic modifier, pH value, CM-β-CD with different degrees of substitution, and concentration of CM-β-CD. The mobile phase was composed of methanol and 10 mmol L-1 of phosphate buffer pH 4.0 containing 10 mmol L-1 of CM-β-CD. Peak resolution for six racemic drugs was gradually increased with an increasing degree of substitution of the synthesized CM-β-CD. The stoichiometric ratio and binding constants for the inclusion complex formed by CM-β-CD and enantiomer were determined, which showed that the stoichiometric ratio for each inclusion complex was 1:1.

Recent Advances on Chiral Mobile Phase Additives: A Critical Review

Recent Advances on Chiral Mobile Phase Additives: A Critical Review

Enantiomeric Separation of 2-Hydroxyglutarate Using Chiral Mobile Phase Additives

Enantiomeric Separation of 2-Hydroxyglutarate Using Chiral Mobile Phase Additives

A simple and cost-effective synthesis of sulfated β-cyclodextrin and its application as chiral mobile phase additive in the separation of cloperastine enantiomers

A simple and cost-effective method for the synthesis of sulfated β-cyclodextrin, one of the most widely used chiral mobile phase additives, using sulfamic acid as a sulfonating agent has been described. The method was optimized, and the synthesized product was characterized by spectroscopic, size-exclusion chromatographic, thermal, and microscopic methods and was compared to the marketed Sigma Aldrich sulfated β-cyclodextrin. β-Cyclodextrin, hydroxypropyl β-cyclodextrin, sulfated β-cyclodextrin (marketed and synthesized) were evaluated as chiral mobile phase additives for the enantiomeric separation of cloperastine, an antitussive agent, using reversed-phase HPLC. Under optimized conditions, a resolution of 3.14 was achieved within 15 min on an achiral Kromasil C8 (150 × 4.6 mm, 5 µm) column, with 5 mM monopotassium phosphate containing 10 mM synthesized sulfated β-cyclodextrin pH 3.0 and 45% methanol as mobile phase. The method utilizing synthesized sulfated β-cyclodextrin as chiral mobile phase additive was validated as per ICH guidelines and applied for the quantitative determination of cloperastine enantiomers in active pharmaceutical ingredients and pharmaceutical formulations. The selectivity changes imparted by sulfated β-cyclodextrin were proven to be beneficial for chiral separation. For the enantiomeric separation of cloperastine, synthesized sulfated β-cyclodextrin afforded better resolution than marketed sulfated β-cyclodextrin.

Quality-by-Design Is a Tool for Quality Assurance in the Assessment of Enantioseparation of a Model Active Pharmaceutical Ingredient.

The design of experiments (DoE) is one of the quality-by-design tools valued in analytical method development, not only for cost reduction and time effectiveness, but also for enabling analytical method control and understanding via a systematic workflow, leading to analytical methods with built-in quality. This work aimed at using DoE to enhance method understanding for a developed UHPLC enantioseparation of terbutaline (TER), a model chiral drug, and to define quality assurance parameters associated with using chiral mobile phase additives (CMPA). Within a response surface methodology workflow, the effect of different factors on both chiral resolution and retention was screened and optimized using Plackett-Burman and central composite designs, respectively, followed by multivariate mathematical modeling. This study was able to delimit method robustness and elucidate enantiorecognition mechanisms involved in interactions of TER with the chiral modifiers. Among many CMPAs, successful TER enantioresolution was achieved using hydroxypropyl β-cyclodextrin (HP-β-CD) added to the mobile phase as 5.4 mM HP-β-CD in 52.25 mM ammonium acetate. Yet, limited method robustness was observed upon switching between the different tested CMPA, concluding that quality can only be assured with specific minimal pre-run conditioning time with the CMPA, namely 16-column volume (60 min at 0.1 mL/min). For enantiorecognition understanding, computational molecular modeling revealed hydrogen bonding as the main binding interaction, in addition to dipole-dipole inside the CD cavity for the R enantiomer, while the S enantiomer was less interactive.

A Review on Chiral Columns/Stationary Phases for HPLC

The chiral separation of pharmaceutical molecules and their precursors is one of the important areas of application of HPLC in pharmaceutical analysis for obtaining enantiomerically pure drug. The latter procedures include the use of so-called chiral selectors to enantio-selectively recognise and isolate the enantiomer. The direct approaches, i.e. those which do not derivate the compound of interest before separation, are addressed in detail, since they are now the most common approaches. The role of stereochemistry in medicinal products is being given greater emphasis to medical practice. For physicians to make conscious choice about the use of single-enantiomered medicinal products, basic knowledge is required. For few treatments single-enantiomer formulations can provide more selectivity than a combination of enantiomers in their biological purposes, enhanced therapeutic indexes and/or better pharmacokinetics. This highlights the possible biological and pharmacological variations between the two drug enantiomers and underlines the clinical experience of individual enantiomers. Particular emphases have been put on chiral separation by HPLC on chiral stationary phases (CSPs). Chiral derivatization reagents (CDRs) are optically pure reagent on reaction with drugs forms a pair of diastereoisomers that can be separated on conventional achiral phase. In Chiral mobile phase additive (CMPA) method, the stationary phase is achiral and the chiral selector is dissolved during the mobile phase. Interaction with the analyte’s enantiomer leads to the formation of transient diastereomeric complexes that are separated by their affinity towards mobile/stationary phase. Separation mechanisms and method development for chiral molecules using these phases are discussed in this review.

Assessment and validation of green stability indicating RP-HPLC method for simultaneous determination of timolol and latanoprost in pharmaceutical dosage forms using eco-friendly chiral mobile phase

Impurity profiling of active pharmaceutical ingredients is a crucial step in assessing their quality. In this study, a green stability indicating method for simultaneous determination of latanoprost (LTN) and timolol (TIM) was validated and assessed for greenness. TIM & LTN are two anti-glaucoma drugs manufactured as combination therapy. Their separation was done in presence of their pharmacopeial impurities including LTN C3-epimer using chiral mobile phase additives. Stress conditions were applied to affirm the stability indicating power of the LC-separation. Gradient elution was employed using mixture of aqueous phase (containing 11.35 g β-cyclodextrin and 1.5 g sodium octane sulfonate/L) and the green organic modifier ethanol. Separation was done on monolithic RP-C18 column and 210 nm UV-detection. Elution time was <18 min with retention times 4.9 and 16.1 min for TIM and LTN, respectively. The proposed method was validated for linearity, accuracy and precision and was found to be acceptable over the range of 50–750 and 1–50 μg/mL for TIM and LTN, respectively. The method was then applied successfully to determine TIM and LTN in marketed pharmaceutical ophthalmic solution. Since claiming greenness is not enough, two assessment tools (Analytical eco-scale and Green Analytical Procedure Index; GAPI) were utilized to compare our proposed method to other reported methods.

Enantioseparation of Cinacalcet, and its Two Related Compounds by HPLC with Self-Made Chiral Stationary Phases and Chiral Mobile Phase Additives

Background:Cinacalcet is one of the second-generation calcimimetics which consists of a chiral center. The pharmacological effect of R-cinacalcet is 1000 times greater than that of the Scinacalcet. As mentioned in many literatures, 1-(1-naphthyl)ethylamine is used as the starting material for the synthesis of cinacalcet. The absolute structure of cinacalcet is influenced by the starting materials.Methods:We present the chiral separation of cinacalcet and its starting material, 1-(1-naphthyl) ethylamine along with one of its intermediates, N-(1-(naphthalen-1-yl) ethyl)-3- (3-(trifluoromethyl) phenyl) propanamide by high-performance liquid chromatography with chiral stationary phase and chiral mobile phase additives.Results:On vancomycin and cellulose tri 3,5-dimethylphenylcarbamate) chiral stationary phase, cinacalcet and 1-(1-naphthyl)ethylamine achieved enantioseparation under normal phase with addition of triethylamine additives, respectively. Meanwhile, 1-(1-naphthyl)ethylamine and N-(1-(naphthalen-1- yl)ethyl)-3-(3-(trifluoromethyl) phenyl) propanamide achieved enantioseparation on 1-napthalene vancomycin chiral stationary phase using D-tartaric acid, diethyl L-tartrate and diethyl D-tartrate as chiral mobile phase additives.Conclusion:The chiral recognition in our experiment was based on the hydrogen-bonding, dipoledipole and π-π interactions among the solutes, chiral stationary phases and chiral mobile phase additives. In addition, the space adaptability of chiral stationary phases also affected the separation efficacy.

Chiral Mobile-Phase Additives in HPLC Enantioseparations.

High-performance liquid chromatography (HPLC) is one of the main separation techniques for chiral drugs. Among the chiral HPLC techniques available, the chiral mobile-phase additive (CMPA) technique is a valuable method for the direct enantioseparation of chiral chemical entities. In the CMPA method, the chiral selector is dissolved in the mobile phase while the stationary phase is achiral. Interaction with the analyte enantiomers results in the formation of transient diastereomeric complexes. These complexes differ in their formation constants and/or distribution between the (achiral) stationary phase and the mobile phase resulting in an enantioseparation. This chapter describes the HPLC separation applying CMPA methods by several most useful types of chiral selectors including chiral ligand exchangers, macrocyclic antibiotics, and cyclodextrins.

Application and Comparison of High-Speed Countercurrent Chromatography and High Performance Liquid Chromatography in Preparative Enantioseparation of α-Substitution Mandelic Acids

This paper is mainly about extending research on application and comparison of preparative high-speed countercurrent chromatography (HSCCC) and preparative high performance liquid chromatography (HPLC) in chiral separations. Preparative enantioseparations of α-cyclopentylmandelic acid and α-methylmandelic acid by HSCCC and HPLC were compared using hydroxypropy-β-cyclodextrin (HP-β-CD) and sulfobutyl ether-β-cyclodextrin (SBE-β-CD) as the chiral mobile phase additives. In preparative HPLC the enantioseparation was achieved on the ODS C18 reverse phase column with the mobile phase composed of a mixture of acetonitrile and 0.10 mol L−1 phosphate buffer at pH 2.68 containing 20 mmol L−1 HP-β-CD for α-cyclopentylmandelic acid and 20 mmol L−1 SBE-β-CD for α-methylmandelic acid. The maximum sample size for α-cyclopentylmandelic acid and α-methylmandelic acid was only about 10 mg and 5 mg, respectively. In preparative HSCCC the enantioseparations of these two racemates were performed with the two-phase solvent system composed of n-hexane-methyl tert.-butyl ether-0.1 molL−1 phosphate buffer solution at pH 2.67 containing 0.1 mol L−1 HP-β-CD for α-cyclopentylmandelic acid (8.5:1.5:10, v/v/v) and 0.1 mol L−1 SBE-β-CD for α-methylmandelic acid (3:7:10, v/v/v). Under the optimum separation conditions, totally 250 mg of racemic α-cyclopentylmandelic acid could be completely enantioseparated by HSCCC with HP-β-CD as a chiral mobile phase additive in a single run, yielding 114-116 mg of enantiomers with 98-99% purity and 89-92% recovery. But, no complete enantioseparation of α-methylmandelic acid was achieved by preparative HSCCC with either of the chiral selectors due to their limited enantioselectivity. In this paper, preparative enantioseparation by HSCCC and HPLC was compared from various aspects.

Rapid chiral separation and impurity determination of ropivacaine and bupivacaine by Densitometry-HPTLC, using mucopolysaccharide as chiral mobile phase additive

Enantio separation of the local anesthetic drugs; ropivacaine ( Rop ), bupivacaine ( Bup ) and potential organic impurities (2,6-Dimethylaniline, 2,6-DMA ) were accomplished on HPTLC using mucopolysaccharide selector (Chondroitin) as chiral mobile phase additive ( CMPA ). The enantioseparation was achieved in acetonitrile:water:methanol (16:3:1, v:v:v ) containing 0.25% chondroitin as chiral mobile phase additive. The influence of separation conditions, including type and concentration of chiral selector, organic modifiers and temperatures on enantioseparation were evaluated. The enantioselective HPTLC method was validated to control the enantiomeric purity of the ( S )-enantiomers ( S-Rop and S-Bup ); the active ingredients contained in drug products. In these conditions, linearity over the concentration range, 1.0-10.0 µg/spot for each ( R )-enantiomer and 1.0-8.0 µg/spot for 2,6-DMA main organic impurity were obtained. The detection limits are less than 0.6 µg/spot of chiral and organic impurities. The intra and inter-day assay precision was less than 3.0% (RSD%).

Thin-layer chromatographic enantioseparation of ofloxacin and zopiclone using hydroxy-propyl-beta-cyclodextrin as chiral selector and thermodynamic studies of complexation

Summary A novel economic thin-layer chromatographic procedure for stereoselective separation of racemic mixtures of each of zopiclone and ofloxacin, and determination of their enantiomers: eszopiclone, (+)(S)-zopiclone, and levofloxacin, (−)-(S)-ofloxacin, was described. The method was based on using normal plates and hydroxy propylβ-cyclodextrin (HP-β-CD) as chiral mobile phase additive (CMPA). The spots were detected under UV lamp 254 nm, followed by densitometric measurements at 304 and 330 nm for (+)-(S)-zopiclone and (−)-(S)-ofloxacin, respectively. The mobile phase enabling successful resolution of the drugs was ethanol–acetonitrile–glacial acetic acid–diethylamine–distilled water containing 0.5% HP-β-CD (4:2:3:1:1, by volume), pH 4, for zopiclone and ethanol–acetonitrile–glacial glacial acetic acid–diethylamine–distilled water containing 0.3% HP-β-CD (4:4:3:2:1 by volume), pH 4.5, for ofloxacin at 25 ± 2°C. All variables affecting the resolution, such as concentration of different chiral selectors, temperature, and pH, were investigated, and the conditions were optimized. Furthermore, some thermodynamic parameters were calculated. The procedure provided a linear response over the concentration range of 1–4 and 2–7 μg spot−1 for determination of pure active isomers, (+)-(S)-zopiclone and (−)-(S)-ofloxacin, respectively, with acceptable precision (relative standard deviation [% RSD] <2.0). The developed method was validated and proved to be robust. The proposed method was found to be selective and accurate for the identification and quantitative determination of enantiomeric purity of the two active isomers in their drug substances and drug products.

Enantioseparation of mandelic acid derivatives by high performance liquid chromatography with substituted β-cyclodextrin as chiral mobile phase additive and evaluation of inclusion complex formation.

The enantioseparation of ten mandelic acid derivatives was performed by reverse phase high performance liquid chromatography with hydroxypropyl-β-cyclodextrin (HP-β-CD) or sulfobutyl ether-β-cyclodextrin (SBE-β-CD) as chiral mobile phase additives, in which inclusion complex formations between cyclodextrins and enantiomers were evaluated. The effects of various factors such as the composition of mobile phase, concentration of cyclodextrins and column temperature on retention and enantioselectivity were studied. The peak resolutions and retention time of the enantiomers were strongly affected by the pH, the organic modifier and the type of β-cyclodextrin in the mobile phase, while the concentration of buffer solution and temperature had a relatively low effect on resolutions. Enantioseparations were successfully achieved on a Shimpack CLC-ODS column (150×4.6mm i.d., 5μm). The mobile phase was a mixture of acetonitrile and 0.10molL(-1) of phosphate buffer at pH 2.68 containing 20mmolL(-1) of HP-β-CD or SBE-β-CD. Semi-preparative enantioseparation of about 10mg of α-cyclohexylmandelic acid and α-cyclopentylmandelic acid were established individually. Cyclodextrin-enantiomer complex stoichiometries as well as binding constants were investigated. Results showed that stoichiometries for all the inclusion complex of cyclodextrin-enantiomers were 1:1.

A simple and rapid method for chiral separation of amlodipine using dual chiral mobile phase additives

A simple, rapid and sensitive method for chiral separation of the amlodipine racemate was developed. (R)- and (S)-amlodipine enantiomers were separated by achiral HPLC, utilising dual chiral mobile phase additives, within 16 min, which was much faster than the previous similar studies. The method was established on an achiral WondaSil C18 column, using methanol–water (45:55 (v/v), pH 2.5) containing 7.5 mmol L−1 sulfobutylether-β-cyclodextrin (SBE-β-CD) and 0.3 mmol L−1 polyethylene glycol-20000 (PEG-20M) as the mobile phase. Effects of additives and concentration, content of methanol and pH on the enantioseparation were investigated. The method was validated for linearity, repeatability, limits of detection (LOD) and limits of quantification (LOQ). The calibration curve (R(S)2 = 0.9998) was constructed in the range of 10–500 μg mL−1 for (S)-amlodipine. Repeatability (n = 6) showed less than 2% relative standard deviation (R.S.D.). LOD and LOQ were found to be 0.032 and 0.106 μg mL−1 for (S)-amlodipine. This method was flexible, simple and economically advantageous over the use of a chiral stationary phase, and was successfully applied to the determination of the amlodipine enantiomers in a commercial pharmaceutical tablets study.

Enantiomeric separations of four basic drugs containing N-alkyl groups by a RP-HPLC system using SBE-β-CD as chiral mobile phase additive

Abstract The chiral separations of four pharmaceutical racemates which contain N -alkyl groups were satisfactorily resolved using SBE- β -CD as a chiral mobile phase additive (CMPA) in a RP-HPLC system (the resolution is 2.701 for ondansetron hydrochloride, 1.996 for sulpiride, 1.293 for clenbuterol hydrochloride and 0.816 for omeprazole). In addition, the effects of different parameters such as CD type and CD concentration were investigated. The separation mechanism arises through the combination of several potential interactions, including electrostatic interactions as well as hydrogen bonding interactions and hydrophobic inclusion interactions, which allow for the SBE- β -CD–drug complexation with strong stereoselectivity and stability. The resolution also relates to the number and location of N atoms in the enantiomers. This method will be applicable to the isolation of various types of biologically important enantiomers containing N -alkyl groups.

The Influence of Temperature, Ultrasonication and Chiral Mobile Phase Additives on Chiral Separation: Predominant Influence of β-Cyclodextrin Chiral Mobile Phase Additive Under Ultrasonic Irradiation

This paper introduces a technique for resolving amino acids that combines the advantages of the conventional CSP (chiral stationary phase) method with the CMPA (chiral mobile phase additive) method. A commercially available chiral crown ether column, CROWNPAK CR(+), was used as the CSP and three cyclodextrins (-CD, -CD, HP--CD) were used as the mobile phase additives. Chromatographic resolution was performed at and with or without sonication. A comparison of the chromatographic results under ultrasonic conditions with those under non-ultrasonic conditions showed that ultrasound decreased the elution time and enantioselectivity at all temperatures. In the case of the -CD mobile phase additive, the elution time and enantioselectivity under ultrasonic condition were significantly higher than under non-sonic condition at all temperatures. Commercially available Chiralpak AD, Whelk-O2 and Pirkle 1-J columns were used as CSPs to examine more meticulously the effects of ultrasonication and temperature on the optical resolution. The optical resolution of some chiral samples analyzed at and with or without sonication was compared. As in the previous case, the enantioselectivity was lower at but similar enantioselectivity was observed at .

P.4.006 Ketamine infusion in bipolar disorder resistant to treatment with antidepressants

Racemate separation can be achieved according to two modes. The first mode is direct separation method, and the second is indirect method. The first mode of separation includes the application of a chiral stationary phase (cellulose and its derivatives, CLEC-system, or silica), a stationary phase impregnated with a chiral selector (enantiomers of various chiral compounds, for example acids, alkaloids, amino acids, and their derivatives) or chiral mobile phase additives (e.g., various cyclodextrins and their derivatives, alkaloids). The second mode of separation principle consists of a reaction between a racemate and a chiral derivatizing reagent, which yields separate diastereomers in an achiral chromatographic system. The examples of both types of chiral resolution of a racemate using thin-layer chromatography will be presented.

Cyclodextrins as a chiral mobile phase additive in nano-liquid chromatography: comparison of reversed-phase silica monolithic and particulate capillary columns

Enantioseparations of racemic nonsteroidal anti-inflammatory drugs (naproxen, ibuprofen, ketoprofen, flurbiprofen, suprofen, indoprofen, cicloprofen, and carprofen) were performed by nano-liquid chromatography, employing achiral capillary columns and heptakis(2,3,6-tri-O-methyl)-β-cyclodextrin (TM-β-CD) or hydroxylpropyl-β-cyclodextrin (HP-β-CD) as a chiral mobile phase additive (CMPA). Working under the same experimental conditions (in terms of mobile phase and linear velocity), the performance of a RP-C18 monolithic column was compared with that of a RP-C18 packed column of the same dimensions (100 μm i.d. × 10 cm). Utilizing a mobile phase composed of 30% ACN (v/v) buffered with 50 mM sodium acetate at pH 3, and containing 30 mM TM-β-CD, the monolithic column provided faster analysis but lower resolution than the packed column. This behavior was ascribed to the high permeability of the monolithic column, as well as to its minor selectivity. HP-β-CD was chosen as an alternative to TM-β-CD. Employing the monolithic column, the effects of different parameters such as HP-β-CD concentration, mobile phase composition, and pH on the retention factor and the chiral resolution of the analytes were studied. For the most of the analytes, enantioresolution (which ranged from R(s) = 1.80 for naproxen to R(s) = 0.86 for flurbiprofen) was obtained with a mobile phase consisting of sodium acetate buffer (25 mM, pH 3), 10% MeOH, and 15 mM HP-β-CD. When the same experimental conditions were used with the packed column, no compound eluted within 1 h. Upon increasing the percentage of organic modifier to favor analyte elution, only suprofen eluted within 30 min, with an R(s) value of 1.14 (20% MeOH). Replacing MeOH with ACN resulted in a loss of enantioresolution, except for naproxen (R(s) = 0.89).

Direct separation of clopidogrel enantiomers by reverse-phase planar chromatography method using β-cyclodextrin as a chiral mobile phase additive

Summary A simple and rapid TLC method using β-cyclodextrin as a chiral mobile phase additive (CMPA) was developed for direct separation of S-clopidogrel and its impurity R-clopidogrel. The influence of different factors (stationary phases, organic modifiers, chiral selectors and their concentrations in the mobile phase, and optimal saturation time of the chamber) on enantioseparation was studied. The best resolution of clopidogrel enantiomers was achieved on Polygram®cel 300 Ac-10% plates using isopropanol-0.5 mM β-cyclodextrin (6:4, υ/υ) as mobile phase in TLC chamber previously equilibrated with the mobile phase for 20 min. The spots were detected under UV light and using iodine vapours. The method enables rapid separation of clopidogrel enantiomers and can be successfully used in control of stereoselective synthesis of clopidogrel and in control of its purity. Finally, the molecular modelling of the inclusion complexes between the analytes and α-, β-, and γ-cyclodextrin was performed to investigate the mechanism of the enantiorecognition.

Recent Developments of Enantioseparations for Fluoroquinolones Drugs Using Liquid Chromatography and Capillary Electrophoresis

Fluoroquinolones antibiotics are more effective pathogens resistant to other antibacterial, thus being regularly prescribed drugs in human and veterinary medicine. Many fluoroquinolones posses one or two stereogenic centers giving rise to enantiomers whose pharmacological properties and toxicity are often different. At the successive stages of drug discovery, the enantiomers of any chiral molecule must be isolated and analyzed and their enantiomeric purity determined. The present review summarizes recent developments on chiral separation of fluoroquinolones using chromatographic and electrophoretic techniques. Keywords: Fluoroquinolones, Enantioseparation, Liquid chromatography, Capillary electrophoresis, Enantioseparations, Fluoroquinolones Drugs, enantiomers, chiral separation, Gram-negative, Gram-positive, nalidixic acid, racemates, ofloxacin, gemifloxacin, antofloxacin, levofloxacin, diastereoisomers, sparafloxacin, High-Performance Liquid Chromatography, counter current chromatography, chiral stationary phases, chiral mobile phase additives, microchip techniques, cyclodextrins, macrocyclic antibiotics, antofloxacin hydrochloride, N-n-dodecyl-Lproline, R-ofloxacin enantiomer, ODS column, pazufloxacin mesilate, CLC-ODS column, acid glycoproteincoated chiral stationary phase, Highly-sulfated-gamma-cyclodextrin, Bovine serum albumin, Crown Ethers