Chiral separation of enantiomers of amino acid derivatives by high-performance liquid chromatography on a norvancomycin-bonded chiral stationary phase

A novel norvancomycin-bonded chiral stationary phase (NVC-CSP) was synthesized by using the chiral selector of norvancomycin. The chiral separation of enantiomers of several dansyl-amino acids by high-performance liquid chromatography (HPLC) in the reversed-phase mode is described. The effects of some parameters, such as organic modifier concentration, column temperature, pH and flow rate of the mobile phase, on the retention and enantioselectivity were investigated. The study showed that ionic, as well as hydrophobic interactions were engaged between the analyte and macrocycle in this chromatographic system. Increasing pH of buffers usually improved the chiral resolution for dansyl-alpha-amino-n-butyric acid (Dns-But), dansyl-methionine (Dns-Met) and dansyl-threonine (Dns-Thr), but not for dansyl-glutamic acid (Dns-Glu) which contains two carboxylic groups in its molecular structure. The natural logarithms of selectivity factors (In alpha) of all the investigated compounds depended linearly on the reciprocal of temperature (1/T), most processes of enantioseparation were controlled enthalpically. Interestingly, the process of enantioseparation for dansyl-threonine was enthalpy-controlled at pH of 3.5, while at pH of 7.0, it was entropy-controlled according to thermodynamic parameters Delta(R,S)DeltaHdegrees and Delta(R,S)DeltaSdegrees afforded by Van't Hoff plots. In order to get baseline separation for all the solutes researched, norvancomycin was also used as a chiral mobile phase additive. In combination with the NVC-CSP remarkable increases in enanselectivity were observed for all the compounds, as the result of a "synergistic" effect. (C) 2003 Elsevier B.V. All rights reserved.

Modeling of protein adsorption in membrane affinity chromatography

For the design of affinity membranes, protein adsorption in membrane affinity chromatography (MAC) was studied by frontal analysis. According to fast mass transfer, small thickness of affinity membranes and high affinity between the protein and the ligand, an ideal adsorption (IA) model was proposed for MAC and was used together with equilibrium-dispersive (E-D) model to describe the adsorption of bovine serum albumin (BSA) onto cellulose diacetate/polyethyleneimine (CA/PEI) blend membranes with and without Cu2+ chelating. E-D model was found to better describe the initial region of experimental breakthrough curves. The influence of axial dispersion was revealed and it showed the importance of design of the module to homogenously distribute feed solution. IA model was found to be better for the whole experimental breakthrough curve. According to it, the capacity of affinity membranes and the specificity of the interaction are of equal importance for the design of affinity membranes. An optimum feed concentration was also found in the operation of MAC. The discrepancy between experimental optimum feed concentrations and predicted ones from IA model may be due to the ignorance of some experimental effects such as axial dispersion.

An empirical equation for the calculation of retention time with extrapolation of temperature in gas chromatography

An empirical equation is proposed to accurately correlate isothermal data over a wide range of temperature With the equation ln k = A* + B*/T-lambda the retention times of different solutes tested on OV-101, SE-54 and PEG 20M capillary columns have been achieved even when lambda is assigned a constant value of 1.7 Comparison with ln k = A + B/T and in k = c + d/T+ h/T-2, shows that the proposed equation is of higher accuracy and is applicable to extrapolation calculation, especially from data at high temperature to those at low temperature. Parameters A* and B* as well as A and B are also discussed. The linear correlation of A* and B* is weaker than that of A and B.

A new method for determination of adsorption and desorption coefficients of pesticides with soil column liquid chromatography

The adsorption and desorption coefficients of atrazine, methiocarb and simazine on a sandy loam soil were measured in this study with soil column liquid chromatographic (SCLC) technique. The adsorption and desorption data of all the three pesticides followed Freundlich isotherms revealing the existence of hysteresis. In comparing with other methods, SCLC method showed some characteristics such as rapidity, online and accuracy.

Human plasma proteome analysis by multidimensional chromatography prefractionation and linear ion trap mass spectrometry identification

A resurgence of interest in the human plasma proteome has occurred in recent years because it holds great promise of revolution in disease diagnosis and therapeutic monitoring. As one of the most powerful separation techniques, multidimensional liquid chromatography has attracted extensive attention, but most published works have focused on the fractionation of tryptic peptides. In this study, proteins from human plasma were prefractionated by online sequential strong cation exchange chromatography and reversed-phase chromatography. The resulting 30 samples were individually digested by trypsin, and analyzed by capillary reversed-phase liquid chromatography coupled with linear ion trap mass spectrometry. After meeting stringent criteria, a total of 1292 distinct proteins were successfully identified in our work, among which, some proteins known to be present in serum in < 10 ng/mL were detected. Compared with other works in published literatures, this analysis offered a more full-scale list of the plasma proteome. Considering our strategy allows high throughput of protein identification in serum, the prefractionation of proteins before MS analysis is a simple and effective method to facilitate human plasma proteome research.

On-line concentration of peptides and proteins with the hyphenation of polymer monolithic immobilized metal affinity chromatography and capillary electrophoresis

An iminodiacetic acid (IDA)-type adsorbent is prepared at the one end of a capillary by covalently bonding IDA to the monolithic rods of macroporous poly(glycidyl methacrylate-co-ethylene dimethacrylate). Cu(II) is later introduced to the support via the interaction with IDA. By this means, polymer monolithic immobilized metal affinity chromatography (IMAC) materials are prepared. With such a column, IMAC for on-line concentration and capillary electrophoresis (CE) for the subsequent analysis are hyphenated for the analysis of peptides and proteins. The reproducibility of such a column has been proved good with relative standard deviations (RSDs) of dead time of less than 5% for injection-to-injection and 12% for column-to-column (n = 3). Through application on the analysis of standard peptides and real protein samples, such a technique has shown promising in proteome study.

Analysis of phenols by high-performance liquid chromatography with pre-column derivatization by 2-(9-carbazole)-ethyl-chloroformate

2-(9-Carbazole)-ethyl-chloroformate (CEOC), a novel pre-column fluorescence labeling reagent, has been synthesized and applied for the derivatization of phenols. Taken phenol, p-chlorophenol, 2,5-dimethylphenol, 2,4-dichlorophenol and 1,4-dihydroxybenzene as testing standards, the effects of derivatization conditions, such as pH of borate buffer, reaction time and fluorescent tagging reagent concentration, have been systematically studied. Under the optimized conditions, CEOC reacts readily with the phenols to form stable derivatives with excitation and emission wavelengths, respectively, at 293 and 360 nm. The single step derivatization reaction could be finished within 20 min even at room temperature. Such a method has been successfully applied to the analysis of phenols in printing ink by high-performance liquid chromatography. (c) 2005 Elsevier B.V. All rights reserved.