Synthesis and characteristics of the human serum albumin-triazine chiral stationary phase

Human serum albumin (HSA) was successfully bonded to silica with s-triazine as activator. The coupling reaction by this method was rapid and effective. The triazine-activated silica is relatively stable and can be installed for at least 1 month without obvious loss of reactivity when stored below 30 degreesC, pH below 7. It was observed that the amount of bound HSA reached 120 mg/g silica calculated from the UV absorbance difference of the HSA solution. d,l-tryptophan was selected as the probe solute to characterize the properties of HSA bonded s-triazine chiral stationary phase, and separation factor of 9.4 was obtained for d,l-tryptophan. Furthermore, the amount of effective HSA on silica was measured by high-performance frontal analysis, and only 16.8 mg/g silica was responsible for the resolution of d,l-tryptophan. These results indicate that the amount of both the bound and effective HSA on silica with triazine as activator was much higher than those by the Schiff base coupling method. Different kinds of enantiomers were resolved successfully on the aminopropylsilica-bonded HSA s-triazine chiral stationary phase. (C) 2000 Wiley-Liss, Inc.

Synthesis of a silica-bonded bovine serum albumin s-triazine chiral stationary phase for high-performance liquid chromatographic resolution of enantiomers

A novel method of synthesizing protein chiral stationary phase (protein-CSP) is proposed with 2,4,6-trichloro-1,3,5-triazine as the activator. The bovine serum albumin (BSA) based chiral columns (150x4.6 mm I.D.) were prepared successfully within 8 h. With tryptophan as the probe solute, it was observed that the BSA immobilized by this method had a better ability to distinguish enantiomers than that activated by glutaric dialdehyde. This may be due to the well-maintained BSA conformation and the larger amount of BSA immobilized on the silica gel. The BSA-CSP prepared by this method was relatively stable under experimental conditions, and the resolution of 13 chiral compounds was achieved. The coupling reaction in this method is mild, reliable and reproducible; it is also suitable for the immobilization of various biopolymers in the preparation of bioreactor, biosensor and affinity chromatography columns. (C) 2000 Elsevier Science B.V. All rights reserved.

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.

Multimilligram enantioresolution of sulfoxide proton pump inhibitors by liquid chromatography on polysaccharide-based chiral stationary phase

The enantiomers of sulfoxide proton pump inhibitors - omeprazole, lansoprazole, rabeprazole and Ro 18-5364 - were enantiomerically separated by liquid chromatography at multimilligram scale on a poly saccharide-based chiral stationary phase using normal and polar organic conditions as mobile phase. The values of the recovery and production rate were significant for each enantiomer; better results were achieved using a solid-phase injection system. However, this system was applied just for the enantionteric separation of omeprazole to demonstrate the applicability of this injection mode at milligram scale. The chiroptical characterization of the compounds was performed using a polarimeter and a circular dichroism detector. The higher enantiomeric purity obtained for the isolated enantiomers suggests that the methods here described should be considered as a simple and rapid way to obtain enantiomeric pure standards for analytical purpose. (C) 2007 Elsevier B.V. All rights reserved.

Analysis of carvedilol enantiomers in human plasma using chiral stationary phase column and liquid chromatography with tandem mass spectrometry

Carvedilol is an antihypertensive drug available as a racemic mixture. (-)-(S)-carvedilol is responsible for the nonselective beta-blocker activity but both enantiomers present similar activity on a1-adrenergic receptor. To our knowledge, this is the first study of carvedilol enantiomers in human plasma using a chiral stationary phase column and liquid chromatography with tandem mass spectrometry. The method involves plasma extraction with diisopropyl ether using metoprolol as internal standard and direct separation of the carvedilol enantiomers on a Chirobiotic T (R) (Teicoplanin) column. Protonated ions [M + H]+ and their respective ion products were monitored at transitions of 407 > 100 for the carvedilol enantiomers and 268 > 116 for the internal standard. The quantification limit was 0.2 ng ml-1 for both enantiomers in plasma. The method was applied to study enantioselectivity in the pharmacokinetics of carvedilol administered as a single dose of 25 mg to a hypertensive patient. The results showed a higher plasma concentration of (+)-(R)-carvedilol (AUC08 205.52 vs. 82.61 (ng h) ml-1), with an enantiomer ratio of 2.48. Chirality, 2012. (C) 2012 Wiley Periodicals, Inc.

Separation of enantiomers by nanoliquid chromatography and capillary electrochromatography using a bonded cellulose trisphenylcarbamate stationary phase

A cellulose trisphenylcarbamate-bonded chiral stationary phase was applied to nano-liquid chromatography (nano-LC) and capillary electrochromatography (CEC) with nonaqueous and aqueous solutions as the mobile phases. Several chiral compounds were successfully resolved on the prepared phase by nano-LC. The applicability of nonaqueous CEC on a cellulose derivative stationary phase was investigated with the organic solvents methanol, hexane, 2-propanol, and tetrahydrofuran (THF) containing acetic acid, as well as triethylamine as the mobile phases. Enantiomers of warfarin and praziquantel were baseline-resolved with plate numbers of 82 300 and 38 800 plates/m, respectively, for the first eluting enantiomer. The influence of applied voltage, concentration of nonpolar solvent, apparent pH, and buffer concentration in the mobile phase on the electroosmotic flow (EOF) and the mobility of the enantiomers was evaluated. Enantioseparations of traps-stilbene oxide and praziquantel were also achieved in aqueous CEC with plate numbers of 111 100 and 107 400 plates/m, respectively, for the first eluting enantiomer. A comparison between nonaqueous CEC and aqueous CEC based on a cellulose trisphenylcarbamate stationary phase was discussed. Pressure-assisted CEC was examined for the chiral separation of praziquantel and faster analysis with high enantioselectivity was acquired with the proper pressurization of the inlet vial.

Capillary electrochromatographic separation of enantiomers on chemically bonded type of cellulose derivative chiral stationary phases with a positively charged spacer

Positively charged chiral stationary phases (CSPs) were prepared for capillary electrochromatography (CEC) separation of enantiomers by chemically immobilizing cellulose derivatives onto diethylenetriaminopropylated silica (DEAPS) with tolylene-2,4-diisocyanate (TDI) as a spacer reagent. Anodic electroosmotic mobility was observed in both nonaqueous and aqueous mobile phases due to the positively charged amines on the surface of the prepared CSPs. For comparison, the traditionally used 3-aminopropyl silica (APS) was also adopted as the base material instead of DEAPS to prepare CSP. It was observed that the EOF on the DEAPS-based CSP was 18%-60% higher than that on the APS-based CSP under nonaqueous mobile phase conditions. Separation of enantiomers in CEC was performed on the positively charged CSPs with the nonaqueous mobile phases of pure ethanol or mixture of hexane-alcohol and the aqueous phases of acetonitrile-water or 95% ethanol. Fast separation of enantiomers was achieved on the newly prepared CSPs.

Synthesis and characteristics of composite chiral stationary phases based on cellulose derivatives

Composite chiral stationary phases (CSPs) were prepared on the basis of cellulose derivatives coated or bonded onto silica. "Molecular exterior" type CSPs were prepared by mixing together two different cellulose tris-derivatives before or after being coated or bonded onto silica, and the "molecular interior" type was obtained by synthesizing non-regioselectively heterosubstituted cellulose derivatives coated or bonded onto silica. For the sake of comparison, the individual phases were also prepared with corresponding cellulose derivatives by coating or bonding approaches, respectively. All of the prepared CSPs were characterized and their chiral recognition properties were evaluated by HPLC with several test racemates. The experimental results demonstrated that the "molecular exterior" CSPs generally exhibit chiral recognition capacities intermediate between those of the two individual phases. However, in the separation of some racemates higher enantioselectivity may be achieved on the "molecular interior" phases than on individual phases, thus broadening the application range of a single cellulose-based CSP.

Improved procedure for preparation of covalently bonded cellulose tris-phenylcarbamate chiral stationary phases

The classical method for preparation of covalently boned cellulose derivative chiral stationary phases (CSP) with diisocyanate as spacer was improved. Diisocyanate was firstly allowed to react with 3-aminopropyltriethoxysilane, and the resulting product was then applied as the spacer reagent to immobilize cellulose derivatives onto silica gel. Influences of the amount and the length of the spacer on the optical resolution ability of the CSP were investigated. Comparing improved procedure to classical diisocyanate method, the cross-linking between the glucose units of the cellulose derivatives was avoided to the most extent. With the improved procedure, regio-nonselective ways could be adopted to prepare covalently bonded CSP, which showed an advantage for the rapid preparation.