Cellulose Chiral Stationary Phases Research Articles

Enantiomeric Determination of Carvedilol by a Newly Developed and Validated Chiral HPLC Method

A new simple, selective, linear and accurate chiral HPLC method for assay of carvedilol enantiomers was developed with immobilized cellulose chiral stationary phases under normal-phase mode using hexane: isopropyl alcohol: diethyl amine: acetic acid (40: 60: 0.7: 0.3, v/v) as the mobile phase at a flow rate of 1.0 mL/min and detection at 220 nm. The method was validated for linearity, precision, accuracy, ruggedness, robustness and stability as per the guidelines of USP and ICH. The regression coefficients (r2) of the linearity were found to be 0.999 for both S- and R- carvedilol, detection limit (LOD) were 1.67 and 1.78 μg/mL and quantitation limit were also found to be 5.06 and 5.41 μg/mL for S- and R- carvedilol, respectively. The average percentage of recovery was found to be 99.59% to 100.83% for S- and 98.42% to 100.18% for R- carvedilol, respectively. The new method can be used for enantiomeric separation and estimation of carvedilol in pharmaceutical dosage forms without any interference.
 Dhaka Univ. J. Pharm. Sci. 18(1): 61-68, 2019 (June)

Graphene quantum dots functionalized β-cyclodextrin and cellulose chiral stationary phases with enhanced enantioseparation performance

Graphene quantum dots (GQD) functionalized β-cyclodextrin (β-CD) and cellulose silica composites were first prepared and applied in HPLC as chiral stationary phases (CSP) to investigate the effect of GQDs on chiral separation. Through comparing the enantioseparation performance of GQDs functionalized β-CD or cellulose CSPs and unmodified β-CD or cellulose CSPs, we found GQDs enhanced the enantioseparation performance of nature β-CD, β-CD-3,5-dimethylphenylcarbamate derivative and cellulose-3,5-dimethylphenylcarbamate derivative. Molecular modeling was applied to understand and theoretically study the enhancement mechanism of GQDs for enantioseparation. According to molecular simulation results, GQDs provide extra interactions such as hydrophobic, hydrogen bond and π-π interaction when chiral selector interacts with enantiomers, which enhances the chiral recognition ability indirectly. The molecular simulation results showed a good agreement with the experimental results. Our work reveals the enhancement performance of GQDs for chiral separation, it can be expected that GQDs-based chiral composites and chiral GQDs have great prospect in chiral separation and other research fields such as asymmetric synthesis, chiral catalysis, chiral recognition and drug delivery.

Nitro-imidazoles in ferrocenyl alkylation reaction. Synthesis, enantiomeric resolution and in vitro and in vivo bioeffects

Ferrocenylalkyl nitro-imidazoles (4a-h, 5a-h) were prepared via the regiospecific reaction of the α-(hydroxy)alkyl ferrocenes, FcCHR (OH) (1a–h; Fc = ferrocenyl; R = H, Me, Et, Pr, i-Pr, Ph, ortho-Cl-Ph, ortho-I-Ph), with nitro-imidazoles in aqueous organic medium (H2O-CH2Cl2) at room temperature in the presence of HBF4, within several minutes in good yields. X-ray structural data for racemic (R,S)-1-N-(benzyl ferrocenyl)-2-methyl-4-nitroimidazole (5f) were determined. The resulting enantiomers were resolved into enantiomers by analytical HPLC on modified amylose or cellulose chiral stationary phases. The viabilities of 4b, 4d, 5b, 5cin vitro, and in experiments in vivo antitumor effects of 1-N-ferrocenylethyl-4-nitroimidazole (4b) against murine solid tumor system Ca755 carcinoma were evaluated.

Enantioselective separation of (±)-β-hydroxy-1,2,3-triazoles by supercritical fluid chromatography and high-performance liquid chromatography.

This paper reports the enantioseparation of β-hydroxy-1,2,3-triazole derivatives, which present a broad range of biological properties, by supercritical fluid chromatography (SFC) and high-performance liquid chromatography techniques (HPLC). Polysaccharide-based chiral columns (cellulose and amylose) were used to evaluate the separation in SFC and HPLC. Time of analyses, consumption of solvent, and parameter optimization were reduced using SFC technique. The columns based on cellulose chiral stationary phase using 2-propanol and ethanol as modifiers showed the best results for the enantioresolution of the (±)-β-hydroxy-1,2,3-triazoles by SFC analyses. These techniques were applied to evaluate the selectivity of biocatalytic reduction of β-keto-1,2,3-triazoles by marine-derived fungus Penicillium citrinum CBMAI 1186 to obtain the (±)-β-hydroxy-1,2,3-triazoles.

Solid-phase extraction combined with dispersive liquid-liquid microextraction and chiral liquid chromatography-tandem mass spectrometry for the simultaneous enantioselective determination of representative proton-pump inhibitors in water samples.

This report describes, for the first time, the simultaneous enantioselective determination of proton-pump inhibitors (PPIs-omeprazole, lansoprazole, pantoprazole, and rabeprazole) in environmental water matrices based on solid-phase extraction combined with dispersive liquid-liquid microextraction (SPE-DLLME) and chiral liquid chromatography-tandem mass spectrometry. The optimized results of SPE-DLLME were obtained with PEP-2 column using methanol-acetonitrile (1/1, v/v) as elution solvent, dichloroethane, and acetonitrile as extractant and disperser solvent, respectively. The separation and determination were performed using reversed-phase chromatography on a cellulose chiral stationary phase, a Chiralpak IC (250mm × 4.6mm, 5μm) column, under isocratic conditions at 0.6mLmin(-1) flow rate. The analytes were detected in multiple reaction monitoring (MRM) mode by triple quadrupole mass spectrometry. Isotopically labeled internal standards were used to compensate matrix interferences. The method provided enrichment factors of around 500. Under optimal conditions, the mean recoveries for all eight enantiomers from the water samples were 89.3-107.3% with 0.9-10.3% intra-day RSD and 2.3-8.1% inter-day RSD at 20 and 100ngL(-1) levels. Correlation coefficients (r (2)) ≥ 0.999 were achieved for all enantiomers within the range of 2-500μgL(-1). The method detection and quantification limits were at very low levels, within the range of 0.67-2.29ngL(-1) and 2.54-8.68ngL(-1), respectively. This method was successfully applied to the determination of the concentrations and enantiomeric fractions of the targeted analytes in wastewater and river water, making it applicable to the assessment of the enantiomeric fate of PPIs in the environment. Graphical Abstract Simultaneous enantioselective determination of representative proton-pump inhibitors in water samples.

Simultaneous enantioselective determination of phenylpyrazole insecticide flufiprole and its chiral metabolite in paddy field ecosystem by ultra-high performance liquid chromatography/tandem mass spectrometry

A novel and sensitive ultra-high performance liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) method was developed and validated for simultaneous enantioselective determination of flufiprole and its hydrolysis metabolite in paddy field ecosystem. The separation and determination were performed using reversed-phase chromatography on a novel cellulose chiral stationary phase, a Lux Cellulose-4 (150mm×2.0mm) column, under isocratic conditions at 0.25mL/min flow rate. The effects of other four different polysaccharide-based chiral stationary phases (CSPs) on the separation and simultaneous enantioseparation of the two target compounds were also evaluated. The elution orders of the eluting enantiomers were identified by an optical rotation detector. Modified QuEChERS (acronym for Quick, Easy, Cheap, Effective, Rugged and Safe) method and solid-phase extraction (SPE) were used for the enrichment and cleanup of paddy water, rice straw, brown rice and paddy soil samples, respectively. Parameters including the matrix effect, linearity, precision, accuracy and stability were evaluated. Under the optimal conditions, the mean recoveries for all enantiomers from the above four sample matrix were ranged from 83.6% to 107%, with relative standard deviations (RSD) in the range of 1.0–5.8%. Coefficients of determination R2≥0.998 were achieved for each enantiomer in paddy water, rice straw, brown rice and paddy soil matrix calibration curves within the range of 5–500μg/kg. The limits of quantification (LOQ) for all stereoisomers in the above four matrices were all below 2.0μg/kg. The methodology was successfully applied for simultaneously enantioselective analysis of flufiprole enantiomers and their chiral metabolite in the real samples, indicating its efficacy in investigating the environmental stereochemistry of flufiprole in paddy field ecosystem.

Enantioseparation of Six Antihistamines with Immobilized Cellulose Chiral Stationary Phase by HPLC.

A stereoselective high performance liquid chromatography method has been developed for the chiral separation of the enantiomers of six antihistamines, doxylamine, carbinoxamine, dioxopromethazine, oxomemazine, cetirizine and hydroxyzine. The effects of mobile phase additive, column temperature and flow rate on the retention time and resolution were studied. Enantiomeric separation of cetirizine, doxylamine and hydroxyzine were achieved on cellulose tris-(3,5-dichlorophenylcarbamate) immobilized on silica gel chiral stationary phase known as Chiralpak IC (RS = 3.74, RS = 1.85 and RS = 1.74, respectively).

Enantioseparation of the Six Important Intermediates of Homoharringtonine With Immobilized Cellulose Chiral Stationary Phase.

A new liquid chromatographic method has been developed for the chiral separation of the enantiomers of intermediates in the preparation of the ester side-chain of homoharringtonine. The enantiomers were separated by a Chiralpak IC (250 × 4.6 mm, 5 µm) in normal phase high-performance liquid chromatography (HPLC). Four compounds were baseline resolved. By comparing the chromatographs of racemates and single enantiomers of the six intermediates, the enantiomeric excess values of the single enantiomers were evaluated, and the elution orders of the enantiomers were established.

Enantioseparation of Three Important Intermediates of Tanikolide with Immobilized Cellulose Chiral Stationary Phase.

Three stereoselective HPLC methods have been developed for the chiral separation of the enantiomers of three intermediates in the preparation of (+)-tanikolide. The enantiomers were separated on Chiralpak IC (250 × 4.6 mm, 5 μm) in normal phase HPLC. Three intermediates were all baseline separated (RS = 2.84, 2.58 and 3.58, respectively). By comparing the chromatograms of racemates and single enantiomers of the three intermediates, the e.e. values of the single enantiomers were determined and calculated.

Enantiomeric Separations of N–H/N–Me Aziridines Utilizing GC and HPLC

High-performance liquid chromatographic (HPLC) and gas chromatographic (GC) methods were developed for the separation of enantiomers of 12 novel aziridines. The HPLC separations were performed on cyclodextrin, cyclofructan, amylose, cellulose and macrocyclic glycopeptide-based chiral stationary phases, whereas the GC separations were performed only on cyclodextrin-based chiral stationary phases. The amylose-based HPLC chiral stationary phases showed good selectivity toward the aziridines, while having low retention factors. It was shown that the HPLC generally provided better separations for most of the compounds as compared to GC. Effective enantiomeric separations for ten aziridines were developed.

Synthesis and enantioseparation behaviors of novel immobilized 3,5-dimethylphenylcarbamoylated polysaccharide chiral stationary phases

Two new polysaccharide-derived chiral selectors, namely, 6-azido-6-deoxy-3,5-dimethylphenylcarbamoylated amylose and 6-azido-6-deoxy-3,5-dimethylphenyl carbamoylated cellulose, were synthesized under homogeneous conditions and immobilized onto aminized silica gel by the Staudinger reaction, resulting in two new immobilized polysaccharide chiral stationary phases (CSPs). Their enantioseparation performances were investigated under normal-phase mode by HPLC. Among 17 analytes, baseline separations of 12 pairs of enantiomers are achieved on the immobilized cellulose CSP, which demonstrates that this new cellulose material exhibits almost the same enantioseparation performance as the coated cellulose CSP. In addition, the amylose-derived CSP presents limited enantiorecognition ability but certain complementarity with the immobilized and coated cellulose-based materials. Neither metolachlor nor paclitaxel side chain acids are separated on two cellulose-derived CSPs, but effective separations are obtained on the immobilized amylose column.

Preparation of a new immobilized cellulose-based chiral stationary phase and its enantioseparation behaviors

The immobilized polysaccharide-based chiral stationary phase has attracted considerable attention over the past decades due to its high chemical stability, good solvent resistance, great enantioseparation ability, etc. In this study, a new immobilized cellulose chiral stationary phase (denoted as ImCel) was prepared through the Staudinger reaction of 6-azido-6-deoxy-cellulose-3,5-dichlorophenylcarbamate and aminopropyl silica gel. The enantioseparation performance of the ImCel for 20 pairs of chiral analytes and the effect of non-standard solvents have been investigated by high performance liquid chromatography. Baseline separations of 17 pairs of enantiomers were achieved on the ImCel. The separation ability of the ImCel in the normal mode was much better than in reversed mode. In addition, the ImCel showed good chemical stability in the non-standard mobile phase due to the covalent bonds between the cellulose chiral selectors and silica support. Moreover, it exhibited complementarity with another immobilized-cellulose chiral stationary phase containing 3, 5-dimethylphenylcarbamate groups for the separation of a series of 9-fluorenylmethyloxycarbonyl (fmoc) -derived amino acids. The reversal of enantiomer elution order induced by the difference of the substituents in chiral stationary phase was observed under the same chromatographic conditions. In brief, a new immobilized cellulose chiral stationary phase with high stability and good separation performance was developed in this work.

Separation of mandelic acid and its derivatives with new immobilized cellulose chiral stationary phase

A new liquid chromatographic method has been developed for the chiral separation of the enantiomers of mandelic acid and their derivatives 2-chloromandelic acid, 4-hydroxymandelic acid, 4-methoxymandelic acid, and 3,4,5-trismethoxymandelic acid. The enantiomers were separated by a CHIRALPAK(®) IC (250 mm×4.6 mm, 5 μm). Mandelic acid, 4-methoxymandelic acid, and 3,4,5-trismethoxymandelic acid were baseline resolved (resolution factor (RS)=2.21, RS=2.14, and RS=3.70, respectively). In contrast, the enantioselectivities between CHIRALPAK(®) IC and 2-chloromandelic acid and 4-hydroxymandelic acid investigated were low. By comparing the chromatographs of mandelic acid enantiomers and mandelic acid spiked with (R)-mandelic acid, it was determined that the first effluent was (R)-mandelic acid.

Cellulose 2,3-Di(p-chlorophenylcarbamate) Bonded to Silica Gel for Resolution of Enantiomers

In this study, 6-azido-2,3-di(p-chlorophenylcarbamoylated) cellulose was synthesized and bonded onto aminized silica gel to obtain a new chiral stationary phase. Enantioselectivity of the chiral stationary phase and Chiralcel OF suggested promising chiral separation ability of the new cellulose chiral stationary phase. In addition, the effect of trifluoroacetic acid, diethylamine on enantioselectivity and retention factors on the chiral stationary phase in high performance liquid chromatography was investigated. Experimental results revealed that resolution increased as the trifluoroacetic acid concentration increased to 0.3% while resolution declined as the diethylamine concentration increased. Therefore, the optimal concentrations of trifluoroacetic acid and diethylamine were determined to be 0.3 and 0.1%, respectively. In most cases, trifluoroacetic acid shortened the retention of the first eluted enantiomer while it increased the retention of the other. For acidic compounds, with the existence of diethylamine in the mobile phase, the retention of both enantiomers decreased. But for basic compounds, the retention of both enantiomers increased.

Analytical and Preparative Chiral Separation of β-Carboline Derivatives, LDL Oxidation Inhibitors, Using HPLC and CE Methodologies: Determination of Enantiomeric Purity

In the present study, baseline separation of the enantiomers of 16 β-carboline derivatives was successfully achieved using both capillary electrophoresis (CE) and high-performance liquid chromatography (HPLC) techniques in short run times (<15 min) and thus permit the determination of enantiomeric excess. In HPLC methodology, cellulose chiral stationary phase (Chiralcel OD-H) was used with a binary mobile phase constituted of n-hexane/ethanol 85/15 leading to a resolution factor of 12.6 in 15 min. Preparative HPLC allowed to obtain pure enantiomers of two compounds. In CE, chiral selectivity was developed with an in-capillary stacking strategy using anionic (highly sulfated-γ) cyclodextrins 5% (w/v) as chiral selectors and a 60 mM phosphate buffer (pH 2.5) resulting in a resolution of 10.26 in 14 min of analysis. The analytical characteristics of the two developed methods were studied in terms of repeatability, limits of detection and limits of quantification showing their suitability to be extended to all the other molecules.

Simultaneous enantioselective determination of triazole fungicides in soil and water by chiral liquid chromatography/tandem mass spectrometry

The manuscript concerns the development and validation of a novel and sensitive multi-residue method for simultaneous enantiomeric analysis of 8 triazole fungicides (tetraconazole, fenbuconazole, epoxiconazole, diniconazole, hexaconazole, triadimefon, paclobutrazol, and myclobutanil) in soil and water using chiral liquid chromatography coupled with tandem mass spectrometry. The separation and determination were performed using reversed-phase chromatography on a cellulose chiral stationary phase, a Chiralcel OD-RH (150 mm × 4.6 mm) column, under isocratic conditions using a mixture of ACN-2 mM ammonium acetate in water (55/45, v/v) as the mobile phase at 0.45 mL/min flow rate. The effects of three cellulose-based columns and three amylose-based columns on the separation were also investigated. The QuEChERS (acronym for quick, easy, cheap, effective, rugged and safe) method and solid-phase extraction (SPE) were used for the extraction and clean-up of the soil and water samples, respectively. Parameters including the matrix effect, linearity, precision, accuracy and stability were undertaken. Under optimal conditions, the mean recoveries for all sixteen enantiomers from the soil samples were 76.4–108.1% with 2.6–12.0% intra-day relative standard deviations (RSD) and 4.2–14.1% inter-day RSD at 5, 25 and 50 μg/kg levels; the mean enantiomer recoveries from the water samples were 81.2–106.5% with 2.1–11.5% intra-day RSD and 3.4–13.6% inter-day RSD at 0.25, 0.5 and 2.5 μg/L levels. Coefficients of determination R 2 ≥ 0.9989 were achieved for all studied analytes in the soil and water matrix calibration curves within the range of 1.0–125 μg/L. The limits of detection (LOD) (S/N = 3) for all enantiomers in the soil and water were less than 1.0 μg/kg or μg/L, whereas the limit of quantification (LOQ) (S/N = 10) did not exceed 3.0 μg/kg or μg/L. The results of the method validation confirm that this proposed method is convenient and reliable for the enantioselective determination of the enantiomers of triazole fungicides in soil and water.

Simultaneous enantioselective determination of fenbuconazole and its main metabolites in soil and water by chiral liquid chromatography/tandem mass spectrometry

A novel and sensitive method was developed for the simultaneous determination of fenbuconazole and its main metabolites enantioselectively using chiral liquid chromatography coupled with tandem mass spectrometry. The separation and determination were performed using reversed-phase chromatography on a cellulose chiral stationary phase, a Chiralcel OD-RH (150 mm × 4.6 mm) column, under isocratic conditions at 0.5 mL/min flow rate. The effects of three cellulose-based columns and three amylose-based columns on the separation were also investigated. The elution orders of the eluting enantiomers were identified by an optical rotation detector. The QuEChERS (acronym for Quick, Easy, Cheap, Effective, Rugged and Safe) method and solid-phase extraction (SPE) were used for the extraction and clean-up of the soil and water samples, respectively. Parameters including the matrix effect, linearity, precision, accuracy and stability were evaluated. Under optimal conditions, the mean recoveries for all enantiomers from the soil samples were 82.5–104.1% with 2.7–9.5% intra-day relative standard deviations (RSD) and 5.7–11.2% inter-day RSD at 5, 25 and 50 μg/kg levels; the mean enantiomer recoveries from the water samples were 81.8–104.6% with 2.6–11.4% intra-day RSD and 5.3–10.4% inter-day RSD at 0.25, 0.5 and 2.5 μg/L levels. Coefficients of determination R 2 ≥ 0.9991 were achieved for each enantiomer in the soil and water matrix calibration curves within the range of 1.0–125 μg/L. The limits of detection (LOD) for all enantiomers in the soil and water were less than 0.8 μg/kg, whereas the limit of quantification (LOQ) did not exceed 2.5 μg/kg. The results of the method validation confirm that this proposed method is convenient and reliable for the enantioselective determination of the enantiomers of fenbuconazole and its main metabolites in soil and water.

Characterization of an unusual adsorption behavior of racemic methyl-mandelate on a tris-(3,5-dimethylphenyl) carbamoyl cellulose chiral stationary phase

An interesting adsorption behavior of racemic methyl mandelate on a tris-(3,5-dimethylphenyl)carbamoyl cellulose chiral stationary phase was theoretically and experimentally investigated. The overloaded band of the more retained enantiomer had a peculiar shape indicating a type V adsorption isotherm whereas the overloaded band of the less retained enantiomer had a normal shape indicating a type I adsorption behavior. For a closer characterization of this separation, adsorption isotherms were determined and analyzed using an approach were Scatchard plots and adsorption energy distribution (AED) calculations are combined for a deeper analysis. It was found that the less retained enantiomer was best described by a Tóth adsorption isotherm while the second one was best described with a bi-Moreau adsorption isotherm. The latter model comprises non-ideal adsorbate–adsorbate interactions, providing an explanation to the non-ideal adsorption of the more retained enantiomer. Furthermore, the possibility of using the Moreau model as a local model for adsorption in AED calculations was evaluated using synthetically generated raw adsorption slope data. It was found that the AED accurately could predict the number of adsorption sites for the generated data. The adsorption behavior of both enantiomers was also studied at several different temperatures and found to be exothermic; i.e. the adsorbate–adsorbate interaction strength decreases with increasing temperature. Stochastic analysis of the adsorption process revealed that the average amount of adsorption/desorption events increases and the sojourn time decreases with increasing temperature.

Insights into chiral recognition mechanisms in supercritical fluid chromatography. II. Factors contributing to enantiomer separation on tris-(3,5-dimethylphenylcarbamate) of amylose and cellulose stationary phases

In this second part of our work on enantioselective supercritical fluid chromatography (SFC), we investigate the factors participating in the chiral recognition process on tris-(3,5-dimethylphenylcarbamate) of amylose and cellulose chiral stationary phases (CSPs). 135 racemates with diverse structures were analysed under identical SFC conditions on both stationary phases. The possibility of identifying the differential interactions of an enantiomer pair within the chromatographic system is assessed using a modified version of the solvation parameter model and factorial discriminant analysis. It is illustrated that one relationship of intermolecular interactions is insufficient to express the enantioseparation of different groups of racemates. An innovative approach is used in unravelling the interactions taking part in the enantiorecognition process. Different intermolecular interactions participating in the enantiomeric separation are demonstrated between the two stationary phases.

Study of Elution Order in Chiral Separation of Organophosphonate Esters Using Tris-(3,5-dimethylphenyl Carbamate) Cellulose Chiral Stationary Phase by HPLC

It is known that the separation of several organic phosphonate esters using a tris(3,5-dimethylphenyl carbamate) cellulose (Chiralcel OD) column are better than the separation obtained using the Pirkle type chiral stationary phase (CSP), namely the N-(3,5-dinitrobenzoyl) leucine column. However, the elution order of the organic phosphonate esters enantiomers separated on tris(3,5-dimethylphenyl carbamate) cellulose chiral stationary phase is not known. The tandem use of these two CSPs together showed that the R-enantiomer had shorter retention time than S-enantiomer when tris(3,5-dimethylphenyl carbamate) cellulose chiral stationary phase was used.