Evaluation and comparison of a methylated teicoplanin aglycone to teicoplanin aglycone and natural teicoplanin chiral stationary phases

HPLC enantiomeric separations of a wide variety of racemic analytes was evaluated using chiral stationary phases (CSPs) based on the macrocyclic glycopeptides teicoplanin (T), teicoplanin aglycone (TAG), and methylated teicoplanin aglycone (Me-TAG) in two different mobile phase modes, i.e., the RP mode and the polar organic (PO) mode. Comparison of the enantiomeric separations using Chirobiotic T, Chirobiotic TAG, and the methylated form of TAG were conducted in order to gain a better understanding of the roles of the polar functional groups on the CSP. Substantial effects due to the cleavage of saccharides and/or methylation on chiral separations were observed in both separation modes. Improved separation efficiencies for many acidic analytes were obtained by methylating the H-bonding groups of TAG. These groups were believed to be a contributing factor to band broadening on TAG due to their negative effect on mass transfer between the stationary phase and mobile phase. Ionic/dipolar interactions between the carboxylate group of the analytes and the amine groups on T, TAG, or Me-TAG are important for chiral discrimination. Therefore, analytes possessing a carboxyl group are good candidates for successful separations on these CSPs. Hydrophobic interactions are important for enantiomeric separations in the RP mode where the H-bonding interactions between analytes and the chiral selectors are relatively weak. Me-TAG offers higher hydrophobicity, which can accentuate the interactions of analytes with hydrophobic moieties, but these interactions are not necessarily stereoselective. In the PO mobile phase, electrostatic/dipolar interactions between polar functional groups are the dominating interactions in chiral recognition. Another important factor is steric fit, which could be changed with every modification of the T structure. Therefore, substantial changes of enantioseparations were obtained within this studied group of CSPs. The PO mode was shown to be the most powerful mobile phase mode for enantiomeric separations on T-based stationary phases, mainly due to the improved efficiency. Methylation of the TAG proved to be a very useful tool for investigating the chiral recognition mechanism for this group of chiral selectors.     

Enantioseparation of dipeptides by capillary electrochromatography on a teicoplanin aglycone chiral stationary phase

This work deals with investigations on the enantioseparation of glycyl-dipeptides by capillary electrochromatography (CEC) on a capillary packed with teicoplanin aglycone immobilized on 3.5 μm silica gel. The results were compared to those obtained with micro-HPLC using the same chiral stationary phase. Polar organic and reversed-phase mode were checked, whereby the latter showed better results. Out of 12 glycyldipetides investigated, all compounds showed baseline separation with R_s values up to 20. Plate numbers were in the range of 10 000–300 000/m. The choice of organic modifier was found to be crucial. While methanol increased retention time, acetonitrile reduced it. A ternary mixture of ethanol–acetonitrile–aqueous triethylamine acetate solution pH 4.1 was found to be a useful compromise, providing excellent resolution with retention times less than 25 min. Efficiency and resolution were generally found to be higher in CEC than with micro-HPLC.     

Thermodynamic origin of the chiral recognition of tryptophan on teicoplanin and teicoplanin aglycone stationary phases

The D-, L-tryptophan binding and the chiral recognition properties of the teicoplanin and teicoplanin aglycone (TAG) chiral stationary phase (CSPs) were compared at various column temperatures. The solute adsorption isotherms (bi-Langmuir model) were determined for both the two CSPs using the perturbation method. It was demonstrated that the sugar units were involved in the reduction of the apparent enantioselectivity through two phenomena: (i) the inhibition of some enantioselective contacts with low-affinity binding regions of the aglycone and (ii) a decrease in the stereoselective properties of the aglycone high-affinity binding pocket. The phenomenon (ii) was governed by both a decrease in the ratio of the enantiomer adsorption constant and a strong reduction of the site accessibility for D- and L-tryptophan. In addition, a temperature effect study was performed to investigate the chiral recognition mechanism at the aglycone high-affinity pocket. An enthalpy-entropy compensation analysis derived from the Grunwald model as well as the comparison with the literature data demonstrated that the enantioselective binding mode was dependent on an interface dehydration process. The change in the enantioselective process observed between the TAG and teicoplanin CSP was characterized by a difference of ca. 2-3 ordered water molecules released from the species interface.     

替考拉宁高效液相色谱手性固定相的制备研究

替考拉宁属于大环抗生素,具有半篮状结构和多个手性中心,是常见的手性识别材料,广泛应用于对映体的色谱手性分离分析.本文研究了以替考拉宁为手性识别剂,采用键合的方法制备得到9种高效液相色谱手性固定相,用于苯甘氨酸和对羟基苯甘氨酸的拆分研究,并且考察了重现性和稳定性及进样量对拆分结果的影响.实验结果表明,9种手性固定相均具有拆分苯甘氨酸及对羟基苯甘氨酸的能力.     

Chiral resolution of diastereomeric di- and tripeptides on a teicoplanin aglycone phase by capillary electrochromatography

The chiral separation ability of a capillary packed with teicoplanin aglycone as a chiral stationary phase was investigated. This stationary phase was used successfully for chiral resolution of both diastereomeric dipeptides and tripeptides possessing one or two chiral centers. The composition of the mobile phase was shown to be crucial for separation. The use of reversed-phase mode was clearly superior to the polar-organic mode. The nature of the organic modifier was found to have a marked influence on separation. After optimizing conditions, all diasteromeric dipeptides and tripeptides investigated were baseline-resolved, however, it was not possible to find a uniform mobile phase showing optimal results for all peptides investigated.     

Enantioselective reversed-phase and non-aqueous capillary electrochromatography using a teicoplanin chiral stationary phase

Enantiomeric separation of chiral pharmaceuticals is carried out in aqueous and non-aqueous packed capillary electrochromatography (CEC) using a teicoplanin chiral stationary phase (CSP). Capillaries were slurry packed with 5 microm 100-A porous silica particles modified with teicoplanin and initially evaluated using a non-aqueous polar organic mode system suitability test for the separation of metoprolol enantiomers (Rs = 2.3 and 53000 plates m(-1)). A number of pharmaceutical drugs were subsequently screened with enantioselectivity obtained for 25 racemic solutes including examples of neutral, acidic and basic molecules such as coumachlor (Rs = 3.0 and 86000 plates m(-1)) and alprenolol (Rs = 3.3 and 135000 plates m(-1)) in reversed-phase and polar organic mode, respectively. A statistical experimental design was used to investigate the effects of non-aqueous polar organic mobile phase parameters on the CEC electroosmotic flow, resolution and peak efficiency for two model solutes. Results primarily indicated that higher efficiency and resolution values could be attained at higher methanol contents which is similar to findings obtained on this phase in liquid chromatography.     

Validation of a chiral capillary electrochromatographic method for metoprolol on a teicoplanin stationary phase

Summary A chiral capillary electrochromatographic (CEC) method for determination of the enantiomeric purity of either enantiomer of metoprolol has been validated. High resolution and efficiency separations (R _s =2.5 and 80000 plates m⁻¹, respectively) were achieved by use of a teicoplanin chiral stationary phase in the polar organic mode. Method validation showed that detection linearity, robustness, accuracy, and repeatability were adequate. The method was also shown to be sufficiently sensitive for the determination of a minor enantiomer; the limit of quantitation (LOQ) was determined to be 0.09% of the peak area of the enantiomer under investigation. A similar commercial column was subsequently evaluated by use of the validated method and found to yield results for metoprolol comparable with those obtained on the homepacked columns. Acceptable separations on this commercial column were also obtained for other β-blocking drugs; those for alprenolol were particularly noteworthy (R _S =3.8 and 265000 plates m⁻¹).     

Chromatographic characterization of a bonded liquid crystal stationary phase for HPLC

Summary A chromatographic and thermodynamic study of the compound [4-(allyloxy)benzoyl]-4-methoxyphenyl (ABMP) as a model of a chemically bonded liquid crystal stationary phase for HPLC was undertaken. A number of polycyclic aromatic hydrocarbons (PAHs) and two small solutes, carvone and pulegone, were studied under varying solvent and temperature conditions. Plots of log k vs. % organic in the mobile phase were not completely linear in all cases. The van't Hoff plots revealed at least one phase transition. The enthalpies of solute transfer from the mobile phase to the ABMP phase were determined for several PAHs. All tests indicate that ABMP possess liquid crystal properties when bonded to particulate silica.     

High-performance liquid chromatography of metal ions on a bonded stationary phase

Summary High performance liquid chromatography was used to separate Cu²⁺, Ni²⁺, and Zn²⁺ ions. A column packed with a -diketone bonded phase on silica gel and a mobile phase composed of trifluoroacetylacetone in acetone was used for the separation. After post-column reaction with a color-forming reagent, the metals were detected by absorption in the visible region.Currently assigned to 1155th Technical Operations Squadron, McClellan Air Force Base, California, 95652, USA     

Temperature stability of reversed phase and normal phase stationary phases under aqueous conditions

In this study the temperature stability of several normal phase and RP columns was investigated using a water-only mobile phase. The temperature was adjusted to 120 degrees C for the bare silica stationary phases and to 185 degrees C for the metal oxide and carbon stationary phases. It could be shown that metal oxide stationary phases exhibited excellent thermal stability over the duration of the test period and are therefore suitable for high temperature LC applications.     

Synthesis of a propyldimethylphenylsilane stationary phase using the allyl bonded phase as an intermediate

Summary A propyldimethylphenylsilane stationary phase was prepared by a hydrosilylation reaction on the double bond of an allyl bonded phase intermediate. The carbon load on the silica was between 8.8–9.0%. Evaluation of the material by FTIR indicated high conversion of the double bond by the hydrosilylation reaction. The bonded material displayed reversedphase properties as determined by the retention behavior of alkylarylketones but it was less hydrophobic than either C-8 or C-18. The stability at both low and high pH was excellent. Separations of pharmaceutical compounds, a mixture of anilines, and a mixture of dopamine and epinephrine were satisfactory.     

Adsorption isotherms on nicotinamide-imprinted polymer stationary phase

The molecular-imprinted technique is applied for the preparation of a polymer selector by using methacrylic acid as functional monomer, ethyleneglycol dimethacrylate as the cross-linker, 2,2'-azobisisobutyronitrile as the initiator, and nicotinamide as the template. The adsorption isotherms of nicotinamide and nicotinic acid and the competitive adsorption isotherms of nicotinamide and nicotinic acid on the imprinted stationary phase are determined using rectangular pulse frontal analysis and static method. Aqueous solution is used as the mobile phase in frontal analysis. It is found that the adsorption data fit well to both Langmuir and Freundlich isotherm models.     

Adsorption behavior of hexafluorophosphate on selected bonded phases

The adsorption behavior of ammonium hexafluorophosphate was studied on four HPLC columns packed with adsorbents of different ability for dispersive interactions using frontal chromatography with LC/MS detection in negative ESI mode. Hexafluorophosphate (PF(6)(-)) adsorption isotherms were measured from acetonitrile/water and methanol/water mixtures. Increased PF(6)(-) adsorption with increased acetonitrile content was found between 0 and 15% of acetonitrile in the eluent. Further increase of the acetonitrile concentration leads to an exponential decrease of PF(6)(-) adsorption. Methanol, on the other hand, causes a steady decrease of PF(6)(-) adsorption with increased organic concentration in the mobile phase.     

Capillary electrochromatographic separation of enantiomers under aqueous mobile phases on a covalently bonded cellulose derivative chiral stationary phase

A chemically bonded cellulose tris(3,5-dimethylphenylcarbamate) chiral stationary phase (CSP) was prepared by a radical polymerization reaction. The prepared CSP was packed into fused-silica capillaries with inner diameter of 75 microm to perform enantiomer separations in CEC. The electrochromatographic behavior of the CSP was investigated. On the prepared CSP, high EOF could be generated under acidic mobile phases, which represented an advantage for the separation of acidic enantiomers. Several neutral, acidic, and basic enantiomers were resolved on the prepared CSP under aqueous mobile phases. The column efficiencies were between 20,000 and 100,000 plates/m, which were much higher than those of HPLC. In addition, it was observed that the separation of some enantiomers benefited from the adoption of THF as mobile phase modifier.     

叔丁基异氰酸酯全衍生化β-环糊精SBA-15硅胶手性固定相的研究

合成和表征了一种叔丁基异氰酸酯全衍生化β-环糊精SBA-15硅胶手性固定相.在RP-HPLC条件下,采用5-cm短柱,分别实现了对包含β-受体阻滞剂普萘洛尔(propranolol)在内的8种含氮原子手性药物对映体的拆分,初步考察了流动相的组成、pH及流速对手性色谱分离的影响.上述药物对映体均在短时间(t ＜6 min...     

Enantioselective ion-exclusion chromatography on teicoplanin aglycone and (+)-(18-crown-6)-2,3,11,12-tetracarboxylic acid stationary phases

Ion-exclusion chromatography is a well established technique for the analysis of achiral ionic species, but it has rarely been applied to chiral analytes. In this paper enantioselective ion-exclusion separations were developed on two commercially available HPLC phases: Chirobiotic TAG, based on teicoplanin aglycone, and Opticrown RCA (+), based on (+)-(18-crown-6)-2,3,11,12-tetracarboxylic acid. Chirobiotic TAG columns have a carboxylic acid group on the chiral ligand, which can be partially ionized to exclude anionic analytes by ionic repulsion. Under acidic conditions Opticrown columns have a cationic sublayer generated from the aminopropyl base silica that excludes cationic analytes. Both columns demonstrate a large dependence of efficiency on flow-rate, with the highest efficiencies at 0.1 ml/min on a 4.6 mm inner diameter column.     

Dansyl amino acid enantiomer separation on a teicoplanin chiral stationary phase: effect of eluent pH

A sensitive method for the simultaneous determination of loratadine and its major active metabolite descarboethoxyloratadine (DCL) in plasma was developed, using high-performance liquid chromatographic separation with tandem mass spectrometric detection. The samples were extracted from plasma with toluene followed by back-extraction into formic acid (2%) for DCL after which the toluene containing the loratadine was evaporated, the analyte reconstituted and combined with the DCL back-extract. Chromatography was performed on a Phenomenex Luna C18 (2) 5-microm, 150x2.1-mm column with a mobile phase consisting of acetonitrile-0.1% formic acid using gradient elution (10 to 90% acetonitrile in 2 min) at a flow-rate of 0.3 ml/min. Detection was achieved by a Perkin-Elmer API 2000 mass spectrometer (LC-MS-MS) set at unit resolution in the multiple reaction monitoring mode. TurbolonSpray ionisation was used for ion production. The mean recovery for loratadine and descarboethoxyloratadine was 61 and 100%, respectively, with a lower limit of quantification at 0.10 ng/ml for both the analyte and its metabolite. This is the first assay method described for the simultaneous determination of loratadine and descarboethoxyloratadine in plasma using one chromatographic run. The method is sensitive and reproducible enough to be used in pharmacokinetic studies.