Teicoplanin Aglycone Research Articles

Liquid Chromatographic Enantioseparation of Newly Synthesized Fluorinated Tryptophan Analogs Applying Macrocyclic Glycopeptides-Based Chiral Stationary Phases Utilizing Core-Shell Particles.

Due to the favorable features obtained through the incorporation of fluorine atom(s), fluorinated drugs are a group with emerging pharmaceutical importance. As their commercial availability is still very limited, to expand the range of possible candidates, new fluorinated tryptophan analogs were synthesized. Control of enantiopurity during the synthesis procedure requires that highly efficient enantioseparation methods be available. In this work, the enantioseparation of seven fluorinated tryptophans and tryptophan was studied and compared systematically to (i) develop analytical methods for enantioselective separations and (ii) explore the chromatographic features of the fluorotrytophans. For enantioresolution, macrocyclic glycopeptide-based selectors linked to core-shell particles were utilized, applying liquid chromatography-based methods. Application of the polar-ionic mode resulted in asymmetric and broadened peaks, while reversed-phase conditions, together with mobile-phase additives, resulted in baseline separation for all studied fluorinated tryptophans. The marked differences observed between the methanol and acetonitrile-containing eluent systems can be explained by the different solvation abilities of the bulk solvents of the applied mobile phases. Among the studied chiral selectors, teicoplanin and teicoplanin aglycone were found to work effectively. Under optimized conditions, baseline separations were achieved within 6 min. Ionic interactions were semi-quantitatively characterized and found to not influence enantiorecognition. Interestingly, fluorination of the analytes does not lead to marked changes in the chromatographic characteristics of the methanol-containing eluents, while larger differences were noticed when the polar but aprotic acetonitrile was applied. Experiments conducted on the influence of the separation temperature indicated that the separations are enthalpically driven, with only one exception. Enantiomeric elution order was found to be constant on both teicoplanin and teicoplanin aglycone-based chiral stationary phases (L < D) under all applied chromatographic conditions.

Enantioseparation of a-substituted proline analogs with macrocyclic glycopeptide-based chiral stationary phases immobilized on superficially porous particles of silica applying liquid chromatography with ultraviolet and mass spectrometric detection

In this study, the liquid chromatography-based direct enantioseparation of the stereoisomers of α-substituted proline analogs has been investigated utilizing chiral stationary phases with UV and/or mass spectrometric (MS) detection. Macrocyclic antibiotics, such as vancomycin, teicoplanin, modified teicoplanin, and teicoplanin aglycone, all covalently immobilized to 2.7 μm superficially porous silica particles have been applied as stationary phases. Mobile phases utilizing mixtures of methanol and acetonitrile with different additives (polar-ionic mode) were optimized during method development. Best separations were achieved with mobile phases of 100% MeOH containing either 20mM acetic acid or 20mM triethylammonium acetate. Special attention was given to the applicability of MS-compatible mobile phases. Acetic acid was found to be advantageous as a mobile phase additive for MS detection. Enantioselective chromatographic behaviors are interpreted based on the explored correlations between the analytes' structural features and those of the applied chiral stationary phases. For the thermodynamic characterization, separations were studied in the temperature range of 5-50°C. Generally, retention and selectivity decreased with increasing temperature, and in most cases, enthalpy-driven enantiorecognition was observed, but entropic contributions also were present. Unexpectedly, unusual shapes for the van Deemter curves were registered in the kinetic evaluations. General trends could be observed in the enantiomeric elution orders: S < R on VancoShell and NicoShell, and opposite R < S on TeicoShell and TagShell columns.

Teicoplanin aglycone media and carboxypeptidase Y: Tools for finding low-abundance D-amino acidsandepimericpeptides.

D-amino acids and epimeric peptides/proteins can play crucial biological roles and adversely affect protein folding and oligopeptide aggregation in age-related pathologies in humans. This has ignited interest in free D-amino acids as well as those incorporated in peptides/proteins and their effects in humans. However, such stereoisomeric analytes are often elusive and in low abundance with few existing methodologies capable of scouting for and identifying them. In this work, we examine the feasibility of using teicoplanin aglycone, a macrocyclic antibiotic, which has been reported to strongly retain D-amino acids and peptides with a D-amino acid on the C-terminus, for use as asolidphaseextraction(SPE)medium. The HPLC retention factors of L-/D-amino acids and C-terminus modified D-amino acid-containing peptides and their L-amino acid exclusive counterparts on teicoplanin aglycone are presented. Retention curve differences between amino acids and peptides highlight regions of solvent composition that can be utilized for their separation. This approach is particularly useful when coupled with enzymatic hydrolysis via carboxypeptidase Y to eliminate all L-amino acid exclusive peptides. The remaining peptides with carboxy-terminal D-amino acids are then more easily concentrated and identified.

Macrocyclic glycopeptides- and derivatized cyclofructan-based chiral stationary phases for the enantioseparation of fluorinated ß-phenylalanine analogs

The enantioseparation of five fluorinated β-phenylalanine analogs together with the nonfluorinated α- and β-phenylalanines has been investigated utilizing chiral stationary phases. The employed chiral selectors include macrocyclic antibiotics, such as vancomycin, teicoplanin, and teicoplanin aglycone, isopropyl carbamate functionalized cyclofructan-6, and Cinchona alkaloid-based tert.-butyl carbamate quinine, all covalently bonded to 2.7 µm superficially porous silica particles. The applied conditions included reversed-phase and polar-ionic modes where the vancomycin-, and the cyclofructan-6-based core-shell particles proved to offer suitable efficiency. Under reversed-phase conditions typical hydrophobic chromatographic behavior was observed, especially in the H2O/MeOH system. The improved selectivity with increasing MeOH content observed in polar ionic mode suggests that H-bonding may not play a major role in the chiral recognition. The stoichiometric displacement model was probed to gather information on the ionic interactions. The ion-exchange process was found to affect retention, but it has no essential contribution to chiral recognition. Without paying special attention to the optimization of the system volume of the UHPLC instrument plate heights varying in the range of 10–50 µm were obtained. In all cases, retention and selectivity decreased with increasing temperature, and enthalpy-driven enantiorecognition was observed. Elution sequences were determined in all cases.

Enantiomeric Separation and Molecular Modelling of Bioactive 4-Aryl-3,4-dihydropyrimidin-2(1H)-one Ester Derivatives on Teicoplanin-Based Chiral Stationary Phase

The enantiomeric separation of 15 racemic 4-aryl-3,4-dihydropyrimidin-2(1H)-one (DHP) alkoxycarbonyl esters, some of which proved to be highly active as A2B adenosine receptor antagonists, was carried out by HPLC on ChirobioticTM TAG, a chiral stationary phase (CSP) bearing teicoplanin aglycone (TAG) as the chiral selector. The racemic compounds were separated under polar organic (PO) conditions. Preliminarily, the same selectands were investigated on three different Pirkle-type CSPs in normal-phase (NP) conditions. A baseline separation was successfully obtained on TAG-based CSPs for the majority of compounds, some of which achieved high enantioselectivity ratios (α &gt; 2) in contrast with the smaller α values (1–1.5) and the lack of baseline resolution observed with the Pirkle-type CSPs. In particular, the racemic tetrazole-fused DHP ester derivatives, namely compounds 8 and 9, were separated on TAG-based HPLC columns with noteworthy α values (8.8 and 6.0, respectively), demonstrating the potential of the method for preparative purposes. A competition experiment, carried out with a racemic analyte (6) by adding N-acetyl-d-alanine (NADA) to the mobile phase, suggested that H-bonding interactions involved in the recognition of the natural dipeptide ligand d-Ala-d-Ala into the TAG cleft should be critical for enantioselective recognition of 4-aryl DHPs by TAG. The X-ray crystal structure of TAG was elucidated at a 0.77 Å resolution, whereas the calculation of molecular descriptors of size, polar, and H-bond interactions, were complemented with molecular docking and molecular dynamics calculations, shedding light on repulsive (steric effects) and attractive (H-bond—polar and apolar) interactions between 4-aryl DHP selectands and TAG chiral selectors.

Enantioseparation of ß2-amino acids by liquid chromatography using core-shell chiral stationary phases based on teicoplanin and teicoplanin aglycone

Enantioseparation of nineteen ß2-amino acids has been performed by liquid chromatography on chiral stationary phases based on native teicoplanin and teicoplanin aglycone covalently bonded to 2.7 µm superficially porous silica particles. Separations were carried out in unbuffered (water/methanol), buffered [aqueous triethylammonium acetate (TEAA)/methanol] reversed-phase (RP) mode, and in polar-ionic (TEAA containing acetonitrile/methanol) mobile phases. Effects of pH in the RP mode, acid and salt additives, as well as counter-ion concentrations on chromatographic parameters have been studied. The structure of selectands (ß2-amino acids possessing aliphatic or aromatic side chains) and selectors (native teicoplanin or teicoplanin aglycone) was found to have a considerable influence on separation performance. Analysis of van Deemter plots and determination of thermodynamic parameters were performed to further explore details of the separation performance.

Development and validation of a fast HPLC method for methyldopa enantiomers using superficially porous particle based macrocyclic glycopeptide stationary phase

Methyldopa is a centrally acting antihypertensive agent, whose pharmacological activity is attributed to the S-α-methyldopa enantiomer. The assessment of enantiomeric purity is an essential quality control test for chiral drugs, as the presence of certain impurities in a drug can interfere with the desired pharmacological effect. The present study was undertaken to evaluate different chiral stationary phases composed of macrocyclic glycopeptides (vancomycin, teicoplanin and aglycone teicoplanin) and the use of superficially porous particles for the efficient separation of methyldopa enantiomers. The reversed-phase and polar ionic elution modes were evaluated, with different proportions of solvents and additives. The influence of the flow rate and temperature on the analysis were also evaluated. Different stationary phases in different elution modes exhibited suitable selectivity and fast analysis. Total separation of the enantiomers were obtained with the teicoplanin and teicoplanin aglycone stationary phases. The teicoplanin aglycone column produced excellent resolution values and very fast separation analyzes. The use of the teicoplanin aglycone chiral stationary phase for the analysis of MDopa enantiomers is innovative and has not yet been explored in the literature. The best method was obtained and validated with teicoplanin aglycone in the reversed-phase mode using 20 mM ammonium acetate buffer pH 4.0/MeOH (20:80), column temperature of 45 °C and flow rate of 1.0 mL/min, which presented a total run time less than 5 min, a resolution of 5.05 and good peak symmetry. The method can be used in pharmaceutical quality control laboratories and support the stereospecific synthesis of the drug.

Molecular docking and density functional theory calculations of vinpocetine and teicoplanin aglycone chiral selector

Chiral separation has been an essential issue as stereochemistry significantly influences the biological activity. In some cases, one stereoisomer may show effectiveness in pharmacology activities while the other may appear toxic. Thus, devising technology with knowledge at the fundamental level becomes crucial in the field of chiral separation. This work presented molecular docking simulation and quantum chemical calculation of vinpocetine (VP) drug stereoisomers complexation with teicoplanin aglycone (TAG) chiral selector. Docking simulation was used to find the best conformation of VP stereoisomers with TAG chiral selector followed by density functional theory (DFT) calculation at B3LYP-D3(BJ)/6–31 g(d) level of theory representing the second phase of this work. The stability of VP–TAG inclusion complexes based on binding energy differences obtained from the quantum-chemical calculation was in the order of (3R16S)VP–TAG > (3R16R)VP–TAG > (3S16S)VP–TAG > (3R16S)VP–TAG. The significant differences in the binding energy suggested that the chiral separation of VP stereoisomers can be achieved using TAG.

Effects of high concentrations of mobile phase additives on retention and separation mechanisms on a teicoplanin aglycone stationary phase in supercritical fluid chromatography

The objective of this study is to understand the behavior of a peptide in a medium containing supercritical carbon dioxide mixed with an alcohol (methanol) and acidic or basic additives in uncommonly high concentrations. Chirobiotic TAG is a chromatographic column made of silica bonded with a macrocyclic peptide, teicoplanin aglycone. With this stationary phase, two additives (trifluoroacetic acid and isopropylamine) were tested under extreme concentration conditions to observe the behavior of this peptide. Indeed, concentrations exceeding 1 M in the methanol co-solvent (>0.1 M overall concentration in the CO2-methanol mixture) were used whereas usual additive concentrations employed in supercritical fluid chromatography (SFC) rarely exceed 50 mM in the co-solvent. One purpose was to modify the apparent pH of the fluid, which is normally slightly acidic (around 5) and consequently possibly changing the ionization state of the peptide. Firstly, the effect of acidic and basic additives on the polarity and the apparent pH were evaluated with the help of color indicators. This served to assess the ionization state of the peptide under the selected operating conditions. Secondly, 54 achiral and 24 chiral molecules were injected in the chromatographic column at different levels of additives. The achiral species served at establishing retention models based on linear solvation energy relationships (LSER), while the chiral species were examined for their enantioresolution. From the LSER equations and observation of chromatograms, it appeared that specific interactions between the peptide-based stationary phase and the analytes evolved when increasing the concentration of additives, particularly hydrogen bonds and ionic interactions. A bare silica stationary phase (Acquity BEH) served as reference to deconvolute the contributions of silica support and bonded peptide. This study, with these extreme conditions of mobile phase, could be useful to understand the behavior of such peptides in SFC mobile phases and also improve the knowledge of the effects of additives in SFC, which should be helpful in the future prospect of analyzing large biomolecules in SFC.

Macrocyclic Antibiotics as Chiral Selectors in High-Performance Liquid Chromatography and Capillary Electrophoresis

Data on the use of macrocyclic antibiotics (vancomycin, teicoplanin, teicoplanin aglycone, and eremomycin) for the enantioseparation of amino acids, various amino acid derivatives, α-phenylcarboxylic acids, β-blockers, and some pharmaceutical preparations in reversed-phase and polar-organic HPLC modes are summarized. It is shown that mixed chiral selectors (eremomycin–vancomycin, eremomycin–bovine serum albumin) combine the properties of two selectors. Eremomycin and macrolides (azithromycin, erythromycin, and clarithromycin) are successfully used as chiral selectors in capillary electrophoresis. Aqueous and aqueous–organic supporting electrolytes (SEs) with the addition of eremomycin or nonaqueous supporting electrolytes with the addition of a macrolide are used for enantioseparation. The use of nonaqueous supporting electrolytes decreases the adsorption of the selectors on the quartz capillary surface and enables the separation of enantiomers at a low concentration of a chiral selector. Additions of boric acid into the supporting electrolyte improve the selectivity of separation.

Hydrophobic Derivatives of Glycopeptide Antibiotics as Inhibitors of Protein Kinases.

As key regulators of cell signaling, protein kinases (PKs) are attractive targets for therapeutic intervention in a variety of diseases. Herein, we report for the first time the inhibitory activity of polycyclic peptides, particularly, derivatives of glycopeptide antibiotics teicoplanin and eremomycin, against a panel of 12 recombinant human protein kinases and two protein kinases (CK1 and CK2) isolated from rat liver. Several of the investigated compounds inhibited various PKs with IC50 values below 10 μM and caused >90% suppression of the enzyme activity at 10 μM concentration. Kinetic analysis of the protein kinase CK2α inhibition by the teicoplanin aglycon analogue (7) demonstrated the non-competitive mechanism of inhibition (with regard to ATP). Interestingly, the inhibitory activity of some investigated compounds correlated with the earlier described antiviral activity against HIV, HCV, and other corona- and flaviviruses.

Simultaneous Preconcentration, Identification, and Quantitation of Selenoamino Acids in Oils by Enantioselective High Performance Liquid Chromatography and Mass Spectrometry.

Selenium is an essential micronutrient for humans. In food, selenium can be present in both inorganic and organic forms, the latter mainly being selenomethionine, Se-methyl-selenocysteine, and selenocystine. Selenoamino acid speciation rarely involves the chirality of selenoamino acids. In this work, a 5 cm long CHIROBIOTIC TAG chromatographic column was used for enantioresolution of selenoamino acids (d- and l-selenomethionine, Se-methyl-l-selenocysteine, d-, l- and meso-selenocystine); in the optimized conditions, the complete resolution of the analytes was achieved within 15 min by using a very polar aqueous mobile phase (gradient elution by methanol/acetonitrile/H2O, 45:45:10 ( v/ v/ v) with 10 mmol L-1 of ammonium formate and 0.5% formic acid as the mobile phase A and acetonitrile/H2O, 20:80 ( v/ v) with 20 mmol L-1 of ammonium formate at apparent pH 4 as the mobile phase B). The affinity of the teicoplanin aglycone was further exploited to devise a preconcentration method for selenoamino acids in oils. In particular, the CHIROBIOTIC TAG precolumn was used to directly concentrate the selenoamino acids after simple dilution of oil samples with dichloromethane. An optimized procedure for selenoamino acid trapping and preconcentration under normal phase conditions was developed. The enrichment procedure also ensured band focusing during the subsequent separation. The target analytes were finally identified and quantified by triple quadrupole selected reaction monitoring. The method allowed obtainment of recovery values up to 73%, with limits of detection between 280 and 750 ng and limits of quantification between 375 and 960 ng for the different selenoamino acids. The method was applied to commercial oil samples, and only l-selenomethionine was detected.

Determination of the enantiomeric purity of albuterol on sorbents modified by macrocyclic antibiotics

The separation of albuterol enantiomers on sorbents with macrocyclic glycopeptide antibiotics immobilized on the silica surface was investigated. Commercial columns—Nautilus-E (BioKhimMak, Russia) with eremomycin as a chiral selector and ChirobioticTAG (Astec, United States) with teicoplanin aglycone as a chiral selector—were used for enantioseparation. Levalbuterol is the (R)-enantiomer of albuterol. We managed to separate albuterol enantiomers on both columns in a polar organic mode, but selectivity was higher on the ChirobioticTAG column (R s = 1.7). The maximum resolution of enantiomer peaks (1.7) was observed in methanol–acetonitrile–triethylamine–acetic acid (90: 10: 0.05: 0.05) as the mobile phase. The detection limit of the compound calculated by a signal–background ratio of 3: 1 was 0.00002 mg/mL, which corresponds to 0.1% of (S)-enantiomer with respect to the total amount. The results made it possible to determine the enantiomeric purity of the active pharmaceutical substances of levalbuterol.

Thermodynamic study of HPLC enantioseparations of some sulfur-containing amino acids on teicoplanin columns in ion-pairing reversed-phase mode

ABSTRACTHomocysteine is an essential amino acid produced in the metabolism of methionine. It can be regulated through two metabolic routes either to cysteine or back to methionine. The imbalance in the methionine or homocysteine metabolism is connected with many disorders. In this work, HPLC enantioseparations of cysteine, homocysteine, and methionine were performed. Enantiomers were separated on natural teicoplanin and teicoplanin aglycone chiral stationary phases when buffered mobile phase with the presence of an ion-interaction reagent was used. The column temperature effect on the enantiomeric retention, selectivity, and resolution was studied in the range of 5–45°C at 5°C increment. Thermal stability of homocysteine standard was investigated by X-ray powder-diffraction measurements at elevated temperature in the range of 25–100°C at 5°C increment in air. Moreover, the thermodynamic analysis was used to quantify contributions of entropic and enthalpic terms on separations of investigated amino acid enantiomers. The elution order of enantiomers was determined in all cases.

HPLC Enantioseparation of Phenylcarbamic Acid Derivatives by Using Macrocyclic Chiral Stationary Phases

Abstract The HPLC by using chiral stationary phases based on macrocyclic antibiotics, dimethylphenyl carbamate cyklofructan 7 and β-cyclodextrin in terms of polar-organic separation mode (mobile phase methanol/acetonitrile/acetic acid/triethylamine) were used for enantioseparation of alkoxy derivatives of phenylcarbamic acid. The effect of the analyte structures on the efficiency of enantioseparation was investigated. The most suitable stationary phase was teicoplanin aglycone, where the separations of the enantiomers were obtained (the resolution value from 0.65 to 2.90, depending on the structure of the analyte). Significant effect on the resolution of the enantiomers has position of alkoxy substituent in the hydrophobic part of the molecule. The enantiorecognition was achieved for 3-alkoxysubstituted derivatives.

Regulation of the P450 Oxygenation Cascade Involved in Glycopeptide Antibiotic Biosynthesis.

Glycopeptide antibiotics (GPAs) are nonribosomal peptides rich in modifications introduced by external enzymes. These enzymes act on the free peptide aglycone or intermediates bound to the nonribosomal peptide synthetase (NRPS) assembly line. In this process the terminal module of the NRPS plays a crucial role as it contains a unique recruitment platform (X-domain) interacting with three to four modifying Cytochrome P450 (P450) enzymes that are responsible for cyclizing bound peptides. However, whether these enzymes share the same binding site on the X-domain and how the order of the cyclization steps is orchestrated has remained elusive. In this study we investigate the first two reactions in teicoplanin aglycone maturation catalyzed by the enzymes OxyBtei and OxyAtei. We demonstrate that both enzymes interact with the X-domain via the identical interaction site with similar affinities, irrespective of the peptide modification stage, while their catalytic activity is restricted to the correctly cross-linked peptide. On the basis of steady state kinetics of the OxyBtei-catalyzed reaction, we propose a model for P450 recruitment and peptide modification that involves continuous association/dissociation of the P450 enzymes with the NRPS, followed by specific recognition of the peptide cyclization state by the P450 (scanning). This leads to an induced conformational change that enhances the affinity of the enzyme/substrate complex and initiates catalysis; product release then occurs, with the product itself becoming the substrate for the second enzyme in the pathway. This model rationalizes our experimental findings for this complex enzyme cascade and provides insights into the orchestration of the sequential peptide tailoring reactions on the terminal NRPS module in GPA biosynthesis.

Determination of the enantiomeric purity of Pemetrexed on the macrocyclic glycopeptides bonded phases

The enantioseparation of Pemetrexed on chiral sorbents with macrocyclic glycopeptide antibiotics as chiral selectors was investigated. The pharmacological group of Pemetrexed is antimetabolites with antitumor activity. Pemetrexed is a structural analog of folic acid. IUPAC name of this drug is disodium (2 S )-2-[[4-[2-(2-amino-4-oxo-4,7-dihydro-1 H -pyrrolo[2,3- d ]pyrimidin-5-yl)ethyl]benzoyl]amino]pentanedioate heptahydrate. The mechanism of Pemetrexed antitumor action is based on the inhibition of the synthesis of purine and pyrimidine. Chiral stationary phases with antibiotics of different structures were used. Columns with silica modified antibiotic eremomycin (commercial column Nautilus-E, Bio-Chem-Mac, Russia) and silica with teicoplanin aglycone (ChirobioticTAG, Astec, USA) were applied. The separation of the enantiomers was carried out by the reversed-phase and polar-ion chromatography modes. The mobile phase in the reversed-phase chromatography mode was a mixture of ammonium dihydrogenphosphate solution and organic solvents (methanol, acetonitrile), and in polar-ion mode – organic solvents (methanol, acetonitrile) with additives of acids (acetic acid, formic acid) and bases (triethylamine, diethylamine). The influence of the components’ nature and concentration in the mobile phase, their composition and pH on the separation of Pemetrexed enantiomers was investigated using the silica modified eremomycin and teicoplanin aglycone. The resolution of Pemetrexed enantiomers was 2.8 - 3.3 in the reverse phase mode on the column with eremomycin. In the polar-ion mode chromatography, Pemetrexed eluted with dead volume (capacity factor k ` = 0.1 ÷ 0.3) independently of the additives concentrations in the mobile phase. The enantiomers of Pemetrexed were not separated on the column with teicoplanin aglycone. Teicoplanin aglycone has one amino-group only, whereas eremomycin has three amino-groups. Electrostatic interactions between teicoplanin aglycone and Pemetrexed were less than interactions between eremomycin and Pemetrexed. Eremomycin has more chiral centers than teicoplanin aglycone. In addition, hydrophobic interactions, hydrogen bonds and dipolar interactions with amide and hydroxyl groups of eremomycin provide high enantioselectivity of sorbent with eremomycin. The determination of enantiomeric purity of Pemetrexed drug on the column with eremomycin was conducted. Mobile phase was (55:15:30) MeOH:ACN:NH 4 H 2 PO 4 (50mM, pH = 2.5). The time of analysis was less than 10 minutes. D-isomer was not detected in the drug. The limit of detection of Pemetrexed D-enantiomer was 0.0003 mg/ml (assumed 3:1 signal to noise ratio) which constituted 0.12 % from the total amount drug. Keywords : macrocyclic glucopeptides, chiral selector, enantiomers, enantiomeric purity of the pharmaceutical substances (Russian) DOI: h ttp://dx.doi.org/10.15826/analitika.2016.20.2.002 E.N. Shapovalova, I.A. Fedorova, A.A. Priporova, I.A. Ananieva, O.A. Shpigun M.V.Lomonosov Moscow State University, 1-3 Leninskie gory, Moscow, , 119991, Russian Federation

High efficiency, narrow particle size distribution, sub-2 μm based macrocyclic glycopeptide chiral stationary phases in HPLC and SFC

State of the art chiral chromatography still employs 3–5 μm bonded or immobilized chiral selectors in 10–25 cm columns. With the availability of 1.9 μm narrow particle size distribution (NPSD) silica, it is now possible to make ever shorter, high efficiency columns practical for sub-minute chiral separations. Three macrocyclic glycopeptides (teicoplanin, teicoplanin aglycone, and vancomycin) were bonded onto 1.9 μm NPSD particles. Such packed columns had ∼80% lower backpressure as compared to polydisperse (PD) 1.7 μm silica materials when using the same mobile phase. The decreased backpressure allowed for diminution of frictional heating and allowed for the use of the 1.9 μm NPSD particle based columns at high flow rates. The 1.9 μm NPSD particle based columns showed up to 190,000 plates m−1 for chiral molecules and 210,000 plates m−1 for achiral probes. Representative enantiomeric separations are shown for wide classes of compounds, including different types of amino acids, β-blockers, and pharmaceutically important heterocyclic compounds such as oxazolidinones. Applications in three liquid chromatography modes, namely, reversed phase, polar organic mode and normal phase chiral separations were shown with resolution values ranging from 1.5 to 5.7. Additionally, the same columns were used with supercritical fluid chromatography (SFC) for ultrafast separations.

Three-dimensional structure of cyclic antibiotic teicoplanin aglycone using NMR distance and dihedral angle restraints in a DMSO solvation model.

The three-dimensional solution conformation of teicoplanin aglycone was determined using NMR spectroscopy. A combination of NOE and dihedral angle restraints in a DMSO solvation model was used to calculate an ensemble of structures having a root mean square deviation of 0.17 Å. The structures were generated using systematic searches of conformational space for optimal satisfaction of distance and dihedral angle restraints. Comparison of the NMR-derived structure of teicoplanin aglycone with the X-ray structure of a teicoplanin aglycone analog revealed a common backbone conformation with deviation of two aromatic side chain substituents. Experimentally determined backbone (13)C chemical shifts showed good agreement with those computed at the density functional level of theory, providing a cross validation of the backbone conformation. The flexible portion of the molecule was consistent with the region that changes conformation to accommodate protein binding. The results showed that a hydrogen-bonded DMSO molecule in combination with NMR-derived restraints together enabled calculation of structures that satisfied experimental data.

Separation Of Methionine Enantiomers By Using Teicoplanin And Cyclofructan Columns

Abstract Methionine is a naturally occurring amino acid. Its enantiomeric separation by using high performance liquid chromatography on various types of chiral stationary phases was studied. The effect of mobile phase composition on enantioselectivity and retention was considered. The separation of the enantiomers was attained in different separation modes – reversed phase mode for the macrocyclic antibiotic chiral stationary phases (teicoplanin, teicoplanin aglycone), normal phase and polar organic phase modes for the isopropyl carbamate cyclofructan 6 chiral stationary phase. It was shown that the hydrogen bonding, dipole interactions, steric effects between methionine molecules and stationary phases play an important role in the separation of enantiomers.