Chiral discrimination of phenoxypropionic acid herbicides on teicoplanin phase: Effect of mobile phase modifier

Summary The chiral recgonition mechanism for a series of phenoxypropionic acid herbicides was investigated in reversed phase liquid chromatography (RPLC) on a teicoplanin stationary phase over a wide range of column temperatures. Thermodynamic constants, of the transfer of an enantiomer from the mobile to the teicoplanin stationary phases were determined. The van't Hoff plots for all solutes had a break at a critical temperature T^* showing a variation in the enantiomer retention mechanism due to a change in the conformational state of the teicoplanin, structure. Additionally, enthalpy-entropy compensation confirmed both the change in enantiomer interaction mechanism observed for regions T<T^* and T>T^* and the independence of this mechanism from (i) herbicide molecular structure,s i.e. the position of the chloro group on the phenol ring and (ii) the carbon absolute configuration. Moreover, the increasing enantioselectivity with increasing methanol fraction in the mobile phase was attributed to restriction of the solute association in the teicoplain, stationary phase, leading to favorable stereoselective interactions. This behavior was used to optimize chromatographic conditions for separation of herbicide enantiomers on teicoplanin.     

Chiral separation of beta-adrenergic antagonists, profen non-steroidal anti-inflammatory drugs and chlorophenoxypropionic acid herbicides using teicoplanin as the chiral selector in capillary liquid chromatography

Three groups of structurally diverse chiral compounds were used to study the interaction mechanism responsible for stereoselective recognition with teicoplanin as chiral selector in capillary liquid chromatography. Teicoplanin-based chiral stationary phase (CSP) was used. The effect of the variation of mobile phase composition on retention and enantioselective separation was studied. The mobile phase composition suitable for enantioresolution of the various chiral compounds differed according to the interaction forces needed for chiral recognition. Mobile phases with high buffer portion (70-90 vol.%) were preferred for separation of enantiomers of profen non-steroidal anti-inflammatory drugs and chlorophenoxypropionic acid herbicides that require hydrophobic interactions, inclusion and pi-pi interactions for stereoselective recognition with teicoplanin. Higher concentration triethylamine in the buffer (0.5-1.0%) increased resolution of these acids. On the other hand, H-bonding and electrostatic interactions are important in stereoselective interaction mechanism of beta-adrenergic antagonists with teicoplanin. These interaction types predominate in the reversed phase separation mode with high organic modifier content (95% methanol) and in polar organic mobile phases. For this reason beta-adrenergic antagonists were best enantioresolved in the polar organic mode. The mobile phase composed of methanol/acetic acid/triethylamine, 100/0.01/0.01 (v/v/v), provided enantioresolution values of all the studied beta-adrenergic antagonists in the range 1.1-1.9. Addition of teicoplanin to the mobile phase, which was suitable for enantioseparation of certain compounds on the CSP, was also investigated. This system was used to dispose of nonstereoselective interactions of analytes with silica gel support that often participate in the interaction with CSPs. Very low concentration of teicoplanin in the mobile phase (0.1 mM) resulted in enantioselective separation of 2,2- and 2,4-chlorophenoxypropionic acids.     

Contribution to the study of the alteration of lipase activity ofCandida rugosa by ions and buffers

A semipurifiedC. rugosa lipase (LS) has been prepared from commercial lipase (LC) using an economical procedure. The presence of sugars and glycopeptides has been detected in LS and LC. Pure lipase only has covalently bonded sugars. The hydrolysis of olive oil catalyzed by LS and commercial lipase (LC) is sensitive to the presence of cations Na(I), Mg(II), Ca(II), and Ba(II) and to the nature of buffer. Highest enzyme activity is obtained with 0.1M Tris/HCl buffers and the combination of NaCl 0.11M and CaCl₂ 0.11M. Fluorescence spetroscopy analysis of LC, LS, and both pure isoenzymes lipases A and B, was used to analyze the interaction of the lipase with these effecttors. Inorganic cations Na or Ca do not interact with pure enzyme LA but do interact with LC and LS and do so slightly with LB. The organic cations (morfolinium ortris) interact with pure lipases. We postulate that the increase in the lipase activity produced by Na(I) or Ca(II) is related with interfacial phenomena, but the increase might be more specific in the hydrolysis of olive oil in the presence of Tris-HCl or morfoline-HCl buffer, owing to enzyme-buffer interaction.     

Chiral discrimination of dansyl-amino-acid enantiomers on teicoplanin phase: sucrose-perchlorate anion dependence

The chiral recognition mechanism for a series of d,l-dansyl-amino-acids (test solutes) on a teicoplanin stationary phase was investigated in reversed phase liquid chromatography (RPLC). The effect of both a surface tension modifier (sucrose) and a chaotropic agent (perchlorate anion) on the enantiomeric separation was studied by varying their concentration, c, in the mobile phase. The thermodynamic data supported the fact that the sucrose molecule acted only on the hydrophobic part of the interaction teicoplanin/dansyl-amino-acid and not on the specific chiral part. It was demonstrated that the enhancement of the separation factor observed as the perchlorate salt concentration increased in the mobile phase was enthalpically controlled owing to stereoselective bonding interactions. Such behavior was used to optimize the chromatographic conditions for separation of dansyl-amino-acids on teicoplanin.     

Interfacing microbore and capillary liquid chromatography to continuous-flow fast atom bombardment mass spectrometry for the analysis of glycopeptides

This work describes a system to interface either microbore or packed capillary gradient liquid chromatography (LC) to fast atom bombardment mass spectrometry (FAB-MS). The interface incorporates on-line ultraviolet detection and post-column matrix addition to enable independent optimization of both LC and FAB-MS. The glycopeptide antibiotic teicoplanin was chosen as a model system because this group of compounds places severe demands on the chromatographic separation and is difficult to analyze by FAB-MS. For both microbore and capillary LC, high-quality mass spectra of the major components in teicoplanin were obtained; however, the increased sensitivity of the capillary system allowed spectra to be obtained at low picomole concentrations. The sensitivity and ease of use make capillary LC the preferred system for use in LC-FAB-MS.     

Chiral discrimination of phenoxypropionic acid herbicide enantiomers on teicoplanin phase: methanol dependence and eluent pH consideration

The chiral recognition mechanism for a series of d,l phenoxypropionic acid herbicides (PPAs) on a teicoplanin stationary phase was investigated in reversed phase liquid chromatography (RPLC) over a wide range of mobile phase pH and column temperature. The effect of methanol on the enantiomeric separation was studied by varying its fraction (v/v) in the mobile phase. The thermodynamic data indicated that the chiral recognition was controlled by the interaction between the anionic form of the solute and the teicoplanin phase while those with the neutral form played a minor role. In addition, it was demonstrated that the enhancement of the separation factor observed as the methanol fraction increased in the mobile phase was enthalpically controlled owing to stereoselective binding interactions. Such behavior was used to optimize the chromatographic conditions for separation of PPAs herbicides on teicoplanin.     

New Insights into Glycopeptide Antibiotic Binding to Cell Wall Precursors using SPR and NMR Spectroscopy

Glycopeptide antibiotics, such as vancomycin and teicoplanin, are used to treat life‐threatening infections caused by multidrug‐resistant Gram‐positive pathogens. They inhibit bacterial cell wall biosynthesis by binding to the D ‐Ala‐D ‐Ala C‐terminus of peptidoglycan precursors. Vancomycin‐resistant bacteria replace the dipeptide with the D ‐Ala‐D ‐Lac depsipeptide, thus reducing the binding affinity of the antibiotics with their molecular targets. Herein, studies of the interaction of teicoplanin, teicoplanin‐like A40926, and of their semisynthetic derivatives (mideplanin, MDL63,246, dalbavancin) with peptide analogues of cell‐wall precursors by NMR spectroscopy and surface plasmon resonance (SPR) are reported. NMR spectroscopy revealed the existence of two different complexes in solution, when the different glycopeptides interact with Ac2Kd AlaD AlaOH. Despite the NMR experimental conditions, which are different from those employed for the SPR measurements, the NMR spectroscopy results parallel those deduced in the chip with respect to the drastic binding difference existing between the D ‐Ala and the D ‐Lac terminating analogues, confirming that all these antibiotics share the same primary molecular mechanism of action and resistance. Kinetic analysis of the interaction between the glycopeptide antibiotics and immobilized AcKd AlaD AlaOH by SPR suggest a dimerization process that was not observed by NMR spectroscopy in DMSO solution. Moreover, in SPR, all glycopeptides with a hydrophobic acyl chain present stronger binding with a hydrophobic surface than vancomycin, indicating that additional interactions through the employed surface are involved. In conclusion, SPR provides a tool to differentiate between vancomycin and other glycopeptides, and the calculated binding affinities at the surface seem to be more relevant to in vitro antimicrobial activity than the estimations from NMR spectroscopy analysis.     

Comparative study of three teicoplanin-based chiral stationary phases using the linear free energy relationship model

Teicoplanin (T) is a macrocyclic glycopeptide that is highly effective as a chiral selector for enantiomeric separations. In this study, we used three teicoplanin-based chiral stationary phases (CSPs) - native teicoplanin, teicoplanin aglycon (TAG) and recently synthesized methylated teicoplanin aglycon (MTAG). In order to examine the importance of various interaction types in the chiral recognition mechanism the three related CSPs were evaluated and compared using a linear free energy relationship (LFER). The capacity factors of 19 widely different solutes, with known solvation parameters, were determined on each of the columns under the same mobile phase conditions used for the chiral separations. The regression coefficients obtained revealed the magnitude of the contribution of individual interaction types to the retention on the compared columns under those specific experimental conditions. Statistically derived standardized regression coefficients were used to evaluate the contribution of individual molecular interactions within one stationary phase. It has been concluded that intermolecular interactions of the hydrophobic type significantly contribute to retention on all the CSPs studied here. Other retention increasing factors are n- and pi-electron interactions and dipole-dipole or dipole-induced dipole ones, while hydrogen donating or accepting interactions are more predominant with the mobile phase than with the stationary phases. However, these types of interactions are not equally significant for all the CSPs studied.     

糖肽大环抗生素作选择试剂的高效液相色谱和毛细管电泳的手性识别机理

万古霉素(vancomycin)、替考拉宁(teicoplanin)和瑞斯西丁素A(ristocetin A)等大环抗生素是在HPLC和CE中成功使用的新手性选择试剂。这3种选择试剂的对映识别机理与其分子结构密切相关。本文报道了替考拉宁中糖部分在氨基酸和非氨基酸分离中的作用,万古霉素和替考拉宁中糖苷配基上的最基本结合位点的确定,以及pH值、温度等实验条件和有机改性剂的性质等对保留值和识别机理的影响的研究成果,综述了它们在对映体分离中的应用。     

A chromatographic approach to analyze dansyl amino acid-HP-beta-CD association using macrocyclic antibiotic as the stationary phase

The retention mechanism for a series of D,L-dansyl amino acids in high-performance liquid chromatography is investigated using a teicoplanin stationary phase and hydroxypropyl-beta-cyclodextrin (HP-beta-CD) as the mobile phase additive (0-16mM). A theoretical treatment is developed to determine the HP-beta-CD influence on the equilibrium between the teicoplanin phase and the aqueous medium, respectively. From the experimental data, the association constants of the D,L-dansyl amino acids-HP-beta-CD inclusion complexes are determined and discussed in relation to the enantiomer structure. A thermodynamic study confirms that both the retention and complexation mechanisms are independent of the dansyl amino acid molecular structure and its absolute carbon configuration.     

On-Line HPLC-UV-mass spectrometry and tandem mass spectrometry for the rapid delineation and characterization of differences in complex mixtures: a case study using toxic oil variants

An integrated differential approach to the characterization of complex mixtures is presented which includes the targeting of liquid chromatography (LC) peaks for identification using characteristic UV adsorption of the LC peak, subsequent molecular weight and formula determination using accurate mass LC mass spectrometry (MS), and structure characterization using accurate mass LC-tandem mass spectrometry. The use of differential UV adsorption aids in narrowing the scope of the study to only specific peaks of interest. Accurate mass measurement of the molecular ion species provides molecular weight information as well as atomic composition information. The tandem MS (MS/MS) spectra provide fragmentation information which allows for structural characterization of each component. Accurate mass assignment of each of the fragment ions in the MS/MS spectrum provides atomic composition for each of the fragment ions and thus further aids in the structural characterization. These experiments are facilitated through the use of on-line LC-MS and LC-MS/MS with in-line UV detection. A synthetic toxic oil (STO) related to Toxic Oil Syndrome is studied with a focus on possible contaminants resulting from the interaction of aniline, used as a denaturant, with the normal components of the oil. A differential analysis between the STO and a control oil is performed. LC peaks were targeted using UV absorbance to indicate the possible presence of the aniline moiety. Further differential analysis was performed through the determination of the MS signals associated with each component separated on the LC. Finally, the MS/MS data was also used to determine if the fragmentation of the targeted components indicated the presence of aniline. The MS/MS and accurate mass data were used to assign the structures for the targeted components.     

Orientational processes in the yarns of high-molecular-mass cellulose acetate in vapors of nitromethane and its mixture with water

The phenomenon of spontaneous changes in the linear dimensions (elongation-contraction) is described for commercial and laboratory samples of acetate yarns processed by wet spinning from high- molecular -mass cellulose diacetate in vapors of a mesophas ogenic solvent (which is able to form a lyotropic LC phase with the polymer), nitromethane, and its mixtures with water. The thermodeformational, elastic- plastic, physicomechanical, and surface characteristics of the yarns at different stages of their spontaneous deformation are analyzed. When samples are treated with probe vapors, orientational processes in the polymer matrix are activated, and this observation makes it possible to prepare yarns with improved physicomechanical characteristics and with low linear density. A new phenomenon (to our knowledge) of cyclic three- stage deformation of acetate monofilaments in nitromethane vapors is discovered. This evidence is interpreted from the standpoint of phase (amorphous glassy-liquid crystalline state) and conformational transitions as well as by the mechanism of interaction between polymers and solvent vapors and related development of the LC phase.     

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.     

Alignment of liquid crystals on ultrathin organized molecular films

Liquid crystal (LC) alignment techniques based on various kinds of ultrathin organized molecular films are reviewed. The mechanisms of LC alignment on the organized films are discussed. For the homeotropic alignment of LCs the main anchoring mechanism is due to the dipole–dipole interaction between polar groups of an aligning agent and LC molecules while the homogeneous alignment is mainly attributed to the orientation of polymer chains or polymer aggregates. An experimental system for an anchoring transition induced by a conformation change of aligning molecules is introduced. Finally the AFM experimental observations on the rubbed polymer films and its mechanisms are summarized.     

On the interaction of the morphological structure and the LC behaviour of LC side group block copolymers

The interaction between morphological structure and phase behaviour of a LC side group block copolymer has been investigated using DSC, TEM and small angle X-ray diffraction. All samples of Polystyrene-block-2-(3-cholesteryloxycarbonyloxy)ethyl methacrylate (PS-b-PChEMA) show a phase separation between the two blocks. It was found that in the case of those samples where the liquid crystalline sub-phase is not continuous (spheres), only a nematic phase is seen, whereas in all samples in which there is a continuous liquid crystalline sub-phase, the smectic A phase of the homopolymer is formed. On the other hand, the block copolymer seems to stabilize the LC phase; no dependency of the clearing temperatures on the molecular weight of the LC blocks has been observed.     

Determination of the molecular orientation of very thin films on solid substrates: surface liquid crystal layers and rubbed polyimide films

The molecular orientation of very thin films on solid substrates can be determined quantitatively by measuring the polarized infrared (IR) absorption spectra of samples as a function of angle of incidence. The quantitative molecular orientation is derived by fitting the incident angle dependence and the dichroic ratio with theoretical calculations. We applied this method to a technologically important system: liquid crystal (LC)/rubbed polyimide film. To understand the alignment mechanism of LC molecules in contact with rubbed polyimide films, we have quantitatively determined the molecular orientation of rubbed polyimide films and a surface LC layer in contact with a rubbed polyimide film. In this paper two relations are discussed: (1) correlation between the inclination angle of polyimide backbone structures in rubbed films and the pretilt angle of bulk LC in contact with them, and (2) relation among the molecular orientation of a rubbed polyimide film and those of surface and bulk LC layers in contact with it.     

Side‐chain functionalized liquid crystalline polymers and blends, 2. Phase behavior of comb‐shaped liquid crystalline ionomers containing ions of alkaline metals

An effective way to synthesize liquid crystalline ionomers by an exchange reaction between the acid groups of functionalized LC polymers and a metal acetate was examined. A number of LC ionomers containing ions of the alkaline metals Na and Rb (1.2–18.3 mol‐%) were obtained. The introduction of small amounts (1.2–5 mol‐%) of metal ions into a functionalized LC polymer matrix leads to significant changes in the phase state of copolymers – the nematic phase is replaced by a SmA phase; an increase of the content of alkaline metal ions causes linear growth of clearing temperatures. The influence of the nature of the metal ions on the phase behavior of LC ionomers was determined. To describe the peculiarities of the LC ionomers' phase behavior a model based on the assumption that an ion multiplet is formed was proposed. Here, the rigidity of the functionalized LC copolymer chains used as a matrix for the preparation of LC ionomers was taken into consideration.     

NaX型沸石分子筛中吸附质分子结构与吸附热的关系

近年来,沸石分子筛结构与性能关系的理论计算研究工作愈来愈受到人们的重视[‘－0．人工合成的N     

Moving pieces in a proteomic puzzle: mass fingerprinting of toxic fractions from the venom of Tityus serrulatus (Scorpiones, Buthidae).

Scorpion venoms are very complex mixtures of molecules, most of which are peptides that display different kinds of biological activity. These venoms have been studied in the light of their pharmacological targets and their constituents are able to bind specifically to a variety of ionic channels located in prey tissues, resulting in neurotoxic effects. Toxins that modulate Na(+), K(+), Ca(++) and Cl(-) currents have been described in scorpion venoms. Mass spectrometry was employed to analyze toxic fractions from the venom of the Brazilian scorpion Tityus serrulatus in order to shed light on the molecular composition of this venom and to facilitate the search for novel pharmacologically active compounds. T. serrulatus venom was first subjected to gel filtration to separate its constituents according to their molecular size. The resultant fractions II and III, which account for 90 and 10% respectively of the whole venom toxic effect, were further analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS), on-line liquid chromatography/electrospray mass spectrometry (LC/ESMS) and off-line LC/MALDI-TOFMS in order to establish their mass fingerprints. The molecular masses in fraction II were predominantly between 6500 and 7500 Da. This corresponds to long-chain toxins that mainly act on voltage-gated Na(+) channels. Fraction III is more complex and predominantly contained molecules with masses between 2500 and 5000 Da. This corresponds to the short-chain toxin family, most of which act on K(+) channels, and other unknown peptides. Finally, we were able to measure the molecular masses of 380 different compounds present in the two fractions investigated. To our knowledge, this is the largest number of components ever detected in the venom of a single animal species. Some of the toxins described previously from T. serrulatus venom could be detected by virtue of their molecular masses. The interpretation of this large set of data has provided us with useful proteomic information on the venom, and the implications of these findings are discussed.     

Linear free energy relationship as a tool for characterization of three teicoplanin-based chiral stationary phases under various mobile phase compositions

Teicoplanin, teicoplanin aglycon, and methylated teicoplanin aglycon chiral stationary phases (CSPs) have been compared on the basis of the regression coefficients calculated from the linear free energy relationship (LFER) equation. The parameters have been obtained from the measurements of a set of 34 structurally diverse solutes. Influence of mobile phase composition - variation of methanol (MeOH) content - on the participation of different interactions types in the retention mechanism has been evaluated. Retention of the various interaction forces in analytes differs with both the CSP and the mobile phase composition. Hydrophobic interactions play a major role in mobile phases for high buffer contents. The more hydrophobic the CSP, the more important are they in the retention mechanism. With increase of MeOH contents in the mobile phase the major role in the interaction mechanism is shifted to more polar forces in which basicity and dipolarity/polarizability dominate. Although the LFER model does not address chiral aspects, we have attempted to explore the importance of the individual interactions in chiral discrimination of amino acids and their N-tert-butyloxycarbonyl derivatives.