New chiral crown ether stationary phase for the liquid chromatographic resolution of alpha-amino acid enantiomers

A new chiral stationary phase (CSP) for the liquid chromatographic separation of enantiomers was prepared by bonding a novel enantiopure (diphenyl-substituted 1,1'-binaphthyl) crown ether to 5 microm silica gel. The resulting CSP was applied to the separation of the enantiomers of various natural and unnatural alpha-amino acids. All alpha-amino acids tested were resolved very well on the new CSP, with the exception of proline, which does not contain a primary amino group. The resolution of alpha-amino acid enantiomers on this new CSP was found to be dependent on the type and amounts of organic and acidic modifiers, and on column temperature.     

Isocratic high-performance liquid chromatographic resolution of glutethimide enantiomers and their 4-hydroxyglutethimide metabolites using cellulose tribenzoate chiral stationary phase

Glutethimide (2-ethyl-2-phenylglutarimide) enantiomers and their corresponding 4-hydroxyglutethimide metabolites (RS and RR) are separated using newly developed commercially available cellulose tris(4-methylphenyl benzoate) ester (Chiralcel OJ) chiral stationary phase and hexane-ethanol or hexane-2-propanol as the mobile phase. The effects of ethanol or 2-propanol concentration in the mobile phase and of column temperature on retention and enantioselectivity of glutethimide enantiomers are also demonstrated. Maximum resolutions of 14.23 and 7.09 are obtained for glutethimide and their 4-hydroxyglutethimide metabolites, respectively, with hexane-ethanol (60:40) at 23 degrees C and a flow rate of 1 mL/min.     

Stereochemical resolution of enantiomeric 2-aryl propionic acid non-steroidal anti-inflammatory drugs on a human serum albumin based high-performance liquid chromatographic chiral stationary phase

Summary The native enantioselectivity in binding of human serum albumin (HSA) towards 2-aryl propionic acid non-steroidal anti-inflammatory drugs (2-APA-NSAIDs, the profens) was found to be preserved when the protein was immobilized within a commercially available diol high-performance liquid chromatographic column. High capacity factors were obtained, reflecting the previously observed extensive binding of the 2-APA-NSAIDs to free HSA. The capacity factors were modified by the addition of octanoic acid to the mobile phase. Chiral resolution of the enantiomers of all nine 2-APA-NSAIDs studied was achieved. Preliminary studies show that in addition to being a useful chiral analytical tool for this therapeutically important series of compounds, the HSA chiral stationary phase may provide useful information on the affinity and binding mechanism of small molecules to HSA.     

Synthesis and chromatographic properties of a chiral stationary phase derived from bovine serum albumin immobilized on magnesia-zirconia using phosphonate spacers

A novel bovine serum albumin (BSA)-modified magnesia-zirconia stationary phase was prepared using the sodium salt of cis-(3-methyloxiranyl)phosphonic acid (fosfomycin) as spacer and glutaraldehyde as coupler. Baseline separation of six derivatized amino acids (DNB-Leu, Dansyl-Val, etc.) was achieved on this column using ammonium acetate buffer-isopropanol mobile phase at a flow rate of 1.0 mL/min. The effects of mobile phase composition, eluent pH value, column temperature, and flow rate on the retention and separation of chiral compounds were also investigated. The BSA chiral stationary phase (BSA-CSP) was relatively stable under experimental conditions. The coupling reaction in this method was mild, reliable, and reproducible; thus it was also suitable for the immobilization of various biopolymers with amino groups in the preparation of chromatography stationary phases.     

Preparation and application of a new ligand exchange chiral stationary phase for the liquid chromatographic resolution of alpha-amino acid enantiomers

A new liquid chromatographic ligand exchange CSP has been prepared by covalently bonding (S)-N,N-carboxymethyl undecyl leucinol monosodium salt onto silica gel and employed in resolving various alpha-amino acids. The new CSP was quite good in resolving various a-amino acids and the resolution results were dependent on the type and content of organic modifier in the mobile phase. From these results, a chiral recognition model using a lipophilic interaction between the tethering alkyl group of the CSP and the substituent at the chiral center of alpha-amino acids was proposed. The liquid chromatographic resolution of alpha-amino acids on the new CSP was also found to be dependent on the Cu(II) concentration in the mobile phase and the column temperature.     

Direct chromatographic resolution of carnitine and O-acylcarnitine enantiomers on a teicoplanin-bonded chiral stationary phase.

R-(-)-Carnitine (vitamin B(T)) plays an important role in human energy metabolism, by facilitating the transport of long-chained fatty acids across the mitochondrial membranes. Its (S)-enantiomer acts as a competitive inhibitor of carnitine acetyltransferase, causing depletion of the body R-(-)-carnitine stock. Consequently, the separation of carnitine enantiomers is very important both to study their biological activities and to control the enantiomeric purity of pharmaceutical formulations. In the present paper we describe an easy, fast and convenient procedure for the separation of the enantiomers of carnitine and O-acylcarnitines by enantioselective HPLC on a laboratory-made chiral column containing covalently bonded teicoplanin as selector. High enantioselectivity factors (alpha values ranging from 1.31 to 3.02) and short-time analyses characterize the analytical procedure; in addition, analytes are easily detected by evaporative light scattering with no need for preliminary derivatization. The effects of pH and ionic strength of the mobile phase and of the nature of the organic modifier on the enantioselective separations were also investigated.     

Direct high-performance liquid chromatographic resolution of dihydropyridine enantiomers

A direct high-performance liquid chromatographic enantioseparation on a chiral stationary phase using a protein as chiral selector was achieved for some dihydropyridines with different structural groups (amine, amide, acid and hydroxyl groups). The protein involved was alpha 1-acid glycoprotein and the work was mainly carried out with an EnantioPac column. An improved version of this chiral column (Chiral-AGP) was compared with the previous one and gave interesting results.     

Polysaccharide-based chiral stationary phases for high-performance liquid chromatographic enantioseparation

Recent developments of polysaccharide-based chiral stationary phases (CSPs) for the direct separation of enantiomers in high-performance liquid chromatography (HPLC) are mainly reviewed together with the results on mechanistic studies by means of chromatography, NMR and mass spectroscopies, and computational methods. Miscellaneous applications of polysaccharide derivatives to the newly developed, chiral dynamic high-performance liquid chromatography (DHPLC) for obtaining a nonracemic compound are also described.     

Synthesis and chromatographic properties of an HPLC chiral stationary phase based upon human serum albumin

Summary A new HPLC stationary phase was synthesized by thein situ covalent immobilization of human serum albumin (HSA). The protein was immobilized on a commerically available diol column which had been activated with 1,1-carbonyldiimidazole. Initial chromatographic studies show that this phase can be used for chiral separations of enantiomeric solutes and that these separations may reflectin vitro binding to the HSA. The effects of mobile phase composition and temperature on the stereochemical resolutions are reported.     

New chiral crown ether stationary phase for the liquid chromatographic resolution of α-amino acid enantiomers

A new chiral stationary phase (CSP) for the liquid chromatographic separation of enantiomers was prepared by bonding a novel enantiopure (diphenyl-substituted 1,1′-binaphthyl) crown ether to 5 μm silica gel. The resulting CSP was applied to the separation of the enantiomers of various natural and unnatural α-amino acids. All α-amino acids tested were resolved very well on the new CSP, with the exception of proline, which does not contain a primary amino group. The resolution of α-amino acid enantiomers on this new CSP was found to be dependent on the type and amounts of organic and acidic modifiers, and on column temperature.     

Preparation of chiral stationary phase via activated carbamate intermediate for liquid chromatographic optical resolution

Summary Chiral stationary phases (CSPs) for liquid chromatography were prepared by the way of an activated carbamate intermediate. The amino group of aminopropylsilyl silica gel was first activated by carbamylation with disuccinimido carbonate (DSC). The obtained activated carbamate silica gel (ACsil) proved useful as an intermediate for the preparation of urea-type CSPs. The reaction of ACsil with (S)- of (R)-1-(α-naphthyl)-ethylamine gave naphthylethylurea type CSPs. These CSPs were also obtained directly from aminopropylsilyl silica gel by its reaction with optically active (S)- or (R)-succinimido 1-(α-naphthyl)ethyl carbamate (SINEC). Several phenylthiohydantoin amino acid enantiomers and p-bromophenylcarbamyl amino acid enantiomers were resolved on the CSPs by elution with aqueous mobile phase.     

Chiral separations of pesticide enantiomers by high-performance liquid chromatography using cellulose triphenylcarbamate chiral stationary phase

The direct chiral separations of pesticide enantiomers by high-performance liquid chromatography by applying self-prepared cellulose triphenylcarbamate chiral stationary phase are performed. The mobile phase is n-hexane modified by isopropanol as a polar modifier. Nine chiral pesticides (benalaxyl, vinclozolin, diclofop-methyl, tebuconazole, quizalofop-ethyl, hexaconazole, lactofen, isocarbophos, and paclobutrazol) show enantioselectivity on the chiral stationary phase. An online circular dichrorism detector is used for identifying the pesticide enantiomers. The influences of the volume content of isopropanol and column temperature on the separations are investigated. The thermodynamic parameters related to the chiral distinguish mechanisms are also calculated.     

High performance liquid chromatographic separation of clausenamide enantiomers with chiral –AGP stationary phase

<正>A method of high performance liquid chromatographic separation of clausenamide enantiomers with chiral-AGP(α_1-acid glycoprotein) stationary phases has been established.The absolute configurations of(-)clausenamide and(+)clausenamide are 3S, 4R,5R,6S and 3R,4S,5S,6R,respectively.The present method has been used to analyze the(-)clausenamide and(+)clausenamide and its analogues such as the major metabolite and synthetic derivatives of clausenamide.     

Stereoselective high-performance liquid chromatographic assay for propranolol enantiomers in serum

A simple and sensitive stereoselective high-performance liquid chromatographic assay for the quantitation of propranolol enantiomers in serum is described. The method involves conversion of the propranolol enantiomers to diastereomeric urea derivatives by reaction with the clinical reagent (+)-phenylethylisocyanate, followed by chromatographic separation of the diastereomeric products. Conditions of the derivatization reaction were optimized to achieve rapid and quantitative yield with either of the enantiomers. Baseline resolution of the diastereomers was achieved on a reversed phase C8 column with an isocratic mobile phase. Fluorescence detection afforded an absolute on-column detection limit of 100 pg. The assay has been applied to pharmacokinetic studies in humans and small laboratory animals.     

Steric and electronic effects in the resolution of enantiomeric amides on a commercially available Pirkle-type high-performance liquid chromatographic chiral stationary phase

The steric and electronic effects in the resolution of enantiomeric amides on a commercially available (R)-N-(3,5-dinitrobenzoyl)phenylglycine chiral stationary phase (CSP) have been investigated. Several homologous series of enantiomeric amides were synthesized from alkyl and aromatic amines and from alkyl and aromatic acids. The results of the study indicate that chiral recognition is based on the formation of diastereomeric solute-CSP complexes that are due to attractive interactions located on a single bond in both the solute and CSP and on steric interactions within the complexes. The magnitude of the chiral resolution appears to depend on the steric bulk at the chiral center. In addition, when the amides synthesized from chiral amines were chromatographed, the (R)-enantiomers eluted first, whereas the opposite elution order was found for the amides synthesized from enantiomeric carboxylic acids. Thus, the amide moiety not only provides the sites of attractive interaction between the solute and CSP, but also influences the spatial orientation of the two molecules, thereby affecting the relative stabilities of the two diastereomeric complexes and determining the enantiomeric elution order.     

High performance liquid chromatographic separation of dipeptide and tripeptide enantiomers using a chiral crown ether stationary phase

A chiral stationary phase (CSP) based on (-)-(18-crown-6)-2,3,11,12-tetracarboxylic acid was evaluated for the direct resolution of the enantiomers of dipeptides and tripeptides. The type and concentration of the acid and the methanol content were optimized with regard to retention time and resolution using Ala-Phe as model peptide. A mobile phase consisting of 10 mM sulfuric acid in 70% aqueous methanol was applied to the separation of a set of 16 structurally diverse dipeptides and tripeptides. Generally, the configuration of the amino acid at the N-terminus determined the enantiomer elution order. With a few exceptions the LL- and LD-enantiomers interacted stronger with the CSP compared to the corresponding DD- or DL-enantiomers. The experimental conditions also allowed the simultaneous separation of all four stereoisomers of Ala-Phe. Addition of ammonium sulfate generally reduced retention times and enantiomer resolution. Addition of triethylamine as modifier led to an overall increase of the retention times while the resolution did not show a general trend, increasing in the case of Ala-Ala but decreasing in the case of Ala-Phe.     

Direct high-performance liquid chromatographic enantioseparation of apolar beta-amino acids on a quinine-derived chiral anion-exchanger stationary phase

A quinine-derived chiral anion-exchanger stationary phase was applied for the direct high-performance liquid chromatographic separation of the enantiomers of N-protected unusual beta-amino acids, i.e. 3-aminobutanoic acid, 3-aminopentanoic acid, 3-amino-4-methylpentanoic acid, 3-amino-4,4-dimethylpentanoic acid, 3-amino-4-methylhexanoic acid, 3-amino-4-ethylhexanoic acid, 3-amino-3-cyclohexylpropanoic acid, 3-amino-3-(3-cyclohexen-1-yl)propanoic acid and 3-amino-3-phenylpropanoic acid. The readily prepared N-2,4-dinitrophenyl derivatives were well separable, with good efficiency and high resolution. The chromatographic conditions (eluent composition, pH and buffer concentration) were varied to achieve optimal separation. In some cases, the elution sequences of the enantiomers of the derivatives were determined.     

Liquid chromatographic separation of enantiomers of beta-amino acids using a chiral stationary phase

The enantiomers of both alpha-substituted beta-alanines and beta-substituted beta-alanines may be chromatographically separated using silica-bonded chiral stationary phases derived from N-acetylated alpha-arylalkylamines. The amino acids are chromatographed as alkyl esters of N-3,5-dinitrobenzoyl derivatives; separability factors range from 1.11 to 1.65 for nine alpha-substituted beta-alanines and from 1.08 to 1.20 for nine beta-substituted beta-alanines. The enantiomers of beta-aminoisobutyrate and beta-leucine, chiral beta-amino acids occurring in animal tissues and physiological fluids, are among those resolved. The enantiomers of R,S-beta-aminoisobutyrate and several related alpha-alkyl-beta-alanines were prepared by chromatographic resolution of diastereomeric dipeptides.