Dorothee Wistuba

Enantiomer separation by pressure-supported electrochromatography using capillaries packed with a permethyl-β-cyclodextrin stationary phase

Abstract Efficient enantiomer separation by pressure-assisted, micro-packed capillary electrochromatography (CEC) has been carried out using a permethyl-β-cyclodextrin-modified silica support (PM–β-CD–silica). When comparing this method with micro-packed-high-performance liquid chromatography in the single-column-mode, CEC displays higher column efficiencies (about three times higher theoretical plate numbers at comparable elution times). The pressure support (about 10 bar), applied to avoid bubble formation, has a negligible influence on elution times in CEC. The influence of the type and composition of organic modifiers is described.

Recent progress in enantiomer separation by capillary electrochromatography

Enantiomer separation by electrochromatography (CEC) can be performed in three modes: (i) open‐tubular capillary electrochromatography (o‐CEC), in which the chiral selector is physically adsorbed coated, and thermally immobilized or covalently attached to the internal capillary wall; (ii) packed capillary electrochromatography (p‐CEC), in which the capillary is either filled with chiral modified silica particles or with an achiral packing material, and a chiral selector is added to the mobile phase; and (iii) monolithic (rod)‐capillary electrochromatography (rod‐CEC) in which the chiral stationary phase (CSP) consists of a single piece of porous solid. We present an overview on methods and new trends in the field of electrochromatographic enantiomer separation such as CEC with either nonaqueous mobile phases or stationary phases with incorporated permanent charges, or with packing beds consisting of nonporous silica particles or particles with very small internal diameters .

Biological activation of 1,3-butadiene to vinyl oxirane by rat liver microsomes and expiration of the reactive metabolite by exposed rats

SummaryWhen 1,3-butadiene is incubated with rat liver microsomes and NADPH both enantiomers of vinyl oxirane are formed, the amount of epoxide being dependent on incubation time, microsomal protein, and substrate concentration. Inhibition by SKF 525 A or dithiocarb as well as induction by pretreatment with phenobarbital or 20-methylcholanthrene suggest participation of cytochrome P-450 in this reaction. The amount of epoxide is enhanced by addition of 1,1,1-trichloropropene oxide and reduced by glutathione, especially in the presence of hepatic cytosol. When rats are exposed to 1,3-butadiene in a closed chamber (conditions of maximal metabolism) vinyl oxirane is exhaled and can be quantitatively determined from the gas phase. The concurrent exhalation of acetone is consistent with the idea of biologic action of a reactive metabolite.

A silica monolithic column prepared by the sol-gel process for enantiomeric separation by capillary electrochromatography

A method for the preparation of a silica monolithic capillary electrochromatography (CEC) column for the separation of enantiomers has been developed. The porous silica monolith was fabricated inside a fused‐silica capillary column by using the sol‐gel process. After gelation for 24 h, hydrothermal treatment at 100°C for 24 h was performed to prevent the sol‐gel matrix from cracking. The prepared monolith was then coated with Chirasil‐β‐Dex which represents a chiral polymer prepared by grafting permethyl‐β‐cyclodextrin to polymethylsiloxane with an octamethylene spacer. Immobilization of Chirasil‐β‐Dex was performed by heat treatment at 120°C for 48 h to give a nonextractable coating. The column performance was evaluated by using racemic hexobarbital as a model compound. The efficiency of 9.2×104 theoretical plates/m for the first eluted enantiomer of hexobarbital was obtained at an optimal flow rate of the mobile phase. The effect of mobile phase composition on enantiomeric separation of hexobarbital was also investigated. The column proved to be stable for more than one hundreds of runs during a two‐months period. The enantiomers of several neutral and negatively charged chiral compounds were baseline separated on this column.

Recent innovations in enantiomer separation by electrochromatography utilizing modified cyclodextrins as stationary phases

Enantiomer separation by electrochromatography employing modified cyclodextrins as stationary phases is performed in two ways. (i) Polysiloxane‒linked permethylated β‒cyclodextrin (Chirasil‒Dex 1) or related selectors are coated and immobilized onto the inner surface of a capillary column. Enantiomer separation is performed in the open tube and the method is referred to as open‒tubular capillary electrochromatography (o‒CEC). (ii) Silica‒linked native β‒cyclodextrin, permethylated β‒cyclodextrin (Chira‒Dex 2) or hydroxypropyl‒β‒cyclodextrin are filled into a capillary column and the bed is secured by two frits. Enantiomer separation is performed in a packed column and the method is referred to as packed capillary electrochromatography (p‒CEC). In a unified instrumental approach, method (i) as well as method (ii) can be operated both in the electro‒ and pressure‒driven modes (o‒CEC vs. open‒tubular liquid chromatography (o‒LC) and p‒CEC vs. p‒LC). It is demonstrated that the electro‒driven variant affords higher efficiencies at comparable elution times. Employing a single open‒tubular column coated with Chirasil‒Dex 1, a unified enantioselective approach can be realized in which the same selectand is separated using all existing chromatographic modes for enantiomers, i.e., gas chromatography (GC), supercritical fluid chromatography (SFC), o‒LC and o‒CEC. As the chiral selector is utilized as a stationary phase, an additional chiral selector may be added to the mobile phase. In the resulting dual chiral recognition systems, enhancement of enantioselectivity (matched case) or compensation of enantioselectivity (mismatched case) may be observed. The overall enantioselectivity is dependent on the sense of enantioselectivity of the selectors chosen and their influence on the electrophoretic and electroosmotic migration of the enantiomers of a selectand.

Stereospecific formation of 1,2-epoxypropane, 1,2-epoxybutane and 1-chloro-2,3-epoxypropane by alkene-utilizing bacteria

Abstract Resting cells of ethene grown Mycobacterium 2W produced 1,2-epoxypropane stereospecifically from propene as revealed by optical rotation, 1H n.m.r. using a chiral shift reagent, and also by complexation gas chromatography involving a glass capillary column coated with an optically active metal chelate. The gas-liquid chromatography method allowed the rapid screening of 11 strains with regard to stereospecific formation of 1,2-epoxypropane, 1,2-epoxybutane and 1-chloro-2,3-epoxypropane. Bacteria grown on either ethene, propene or butadiene all predominantly produced the R form of 1,2-epoxypropane from propene and 1,2-epoxybutane from 1-butene while the strains tested for 1-chloro-2,3-epoxypropane production from 3-chloro-1-propene predominantly accumulated the S enantiomer.

Enantiomer separation by capillary electrochromatography on a cyclodextrin‐modified monolith

A chiral monolithic stationary phase was prepared by packing a capillary with bare porous silica and sintering the silica bed at high temperature. The resulting silica monolith was polymer‐coated with Chirasil‐Dex, a permethylated β‐cyclodextrin covalently linked via an octamethylene spacer to dimethylpolysiloxane. Subsequently, Chirasil‐Dex was thermally immobilized on the silica support and a chiral monolith of very high stability (30 kV, more than 400 bar pressure) was obtained. The enantiomer separation of various chiral compounds by monolithic (rod) capillary electrochromatography (rod‐CEC) was feasible. This method was compared with capillary liquid chromatography (LC) in a single‐column mode using unified equipment. About two to three times higher efficiency was found in the rod‐CEC mode as compared to rod‐LC. The influence of pressure‐driven flow support on efficiency, resolution, elution time and baseline stability was investigated. The amount and nature of organic modifier strongly influences efficiency and resolution.

Enantiomer separation by pressure-supported electrochromatography using capillaries packed with Chirasil-Dex polymer-coated silica.

Pressure‒supported electrochromatography using capillaries packed with permethyl‒cyclodextrin covalently linked via an octamethylene spacer to dimethylpolysiloxane and immobilized on silica (Chirasil‒Dex silica) has been employed as an efficient and rapid method for the enantiomer separation of various racemic compounds. By comparing this method with micropacked liquid chromatography (LC), employing the same column in a unified instrumental setup, micropacked capillary electrochromatography (CEC) shows higher column efficiencies and hence better resolution factors. The influence of type and concentration of buffer, amount and nature of organic modifier, and pressure support is investigated.

Enantiomer separation by nonaqueous and aqueous capillary electrochromatography on cyclodextrin stationary phases

Native β‐ and γ‐cyclodextrin bound to silica (Chira Dex‐beta and Chira Dex‐gamma) were packed into capillaries and used for enantiomer separation by capillary electrochromatography (CEC) under aqueous and nonaqueous conditions. Negatively charged analytes (dansyl‐amino acids) were resolved into their enantiomers by nonaqueous CEC (NA‐CEC). The addition of a small amount of water to the nonaqueous mobile phase enhanced the enantioselectivity but increased the elution time. The choice of the background electrolyte (BGE) determined the direction of the electroosmotic flow (EOF). With 2‐(N‐morpholino) ethanesulfonic acid (MES) or triethylammonium acetate (TEAA) as BGE an inverse EOF (anodic EOF) was observed while with phosphate a cathodic EOF was found. The apparent pH (pH*), the concentration of the BGE, and the nature of the mobile phase strongly influenced the elution time, the theoretical plate number and the chiral separation factor of racemic analytes.

Analytical enantiomer separation of aliphatic diols as boronates and acetals by complexation gas chromatography

Abstract Cyclic boronates and acetals of mono- and dialkylsubstituted 1,2-, 2,3- 1,3- & 1,4- diols have been quantitatively separated into enantiomers by complexation gas chromatography utilizing optically active metal chelates. An efficient, precise & sensitive method for determining enantiomeric purities for volatile glycols is thus available.