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September 2009
2.4. Preparative Applications 2.4.1. HPLC
One of themost appealing features of Cinchona carbamate-type stationary phases, in addition to their broad chiral recognition capabilities for chiral acids, is their inherently high loading capacity.
Recently, the preparative potential of a
commercially available quinidine tert- butylcarbamate CSP was evaluated using Fmoc-allylglycine enantiomers asmodel analyte andmethanol–glacial acetic acid– ammoniumacetate as eluent [37]
. In this
particular study, the properties of themobile phase were incorporated into the adsorption model and the inversemethod was used to measure the competitive adsorption isotherms of both the solute enantiomers as well as of the invisible adsorbing acetic acid additive. The adsorption of the Fmoc- allylglycine enantiomers could be sufficiently well described by a non-heterogeneous adsorptionmodel, indicating a close-to- homogenous interaction in this chiral preparative phase system. The saturation capacities under these conditions were exceptionally high, amounting to 185mg/mL for themore strongly retained (R)-enantiomer and 110mg/mL of the less strongly (S)- enantiomer. The effective loading capacity, corresponding to a touching band chromatogram, was estimated to be in the range of 20.0mg/g CSP for amethanol– acetic acid–ammoniumacetate 99:1:0.25 (v/v/w)mobile phase. This figure ranks amongst the highest values reported so far for chiral stationary phases. Considering that the studied analytes show a relativelymodest
enantioselectivity value (α = 2.0) higher loading capacitiesmay be well in reach with cinchona carbamate-type CSPs.
2.4.2. Liquid-liquid Extraction Based Enantiomer Separation
The exceptional high enantioselectivity values and binding affinities achievable with cinchona carbamate-type selectorsmakes thesemolecular recognition elements attractive candidates for the development of preparative liquid-liquid extraction based enantiomer separation technologies.
Specifically for these applications a special class of chiral extractants has been developed by enhancing the lipophilicity of the parent Cinchona carbamate-type selectors by attachment of highly lipophilic alkyl chains. This type ofmodification ensures high solubility of the extractants in apolar organic solvents and effective retention in organic solvents even upon protonatation. Several of these chiral extractants have been employed to study the feasibility of various liquid-liquid extraction enantiomer separation formats, including supported liquid
Figure 9. Enantiomer separation of 3,5-dinitrobenzoyl chloride (DND-Leu) by multistage countercurrent liquid-liquid extraction using coupled centrifugal contactors and a quinine carbamate-type extractant. a) Chemical structures of Dichlorprop and the employed quinine-based chiral extractant. b) Schematic representation of the pilot-scale process configuration employed. Reproduced from [41] with permission.
membrane [38] , centrifugal partition chromatography [39, 40] liquid-liquid extraction techniques [41]
and countercurrent .
Appealing results were achieved when these enantioselective extractants were utilized as chiral selectors for preparative separation of chiral acids in centrifugal partition chromatography (CPC) applications. CPC is a carrier-free chromatographic technique employing immiscible liquids as stationary as well asmobile phase. Separation is carried out with a rotor-type column comprising a systemof several hundred interconnected extraction compartments. In operational state, one of the liquid phases is kept immobilized within the column compartments
by a centrifugal field generated by rotation, while the other functioning asmobile phase is forced through the stationary phase by pumping. Duringmigration of themobile through the rotating column separation is achieved viamultiple sequential liquid-liquid extractions steps.
In studies performed with a lab-scale CPC instrument,O-9-(1-adamantylcarbamoyl)- 10,11-dihydro-11-octadecylsulfinylquinine employed as chiral extractant in acetone/isobutylmethylketone (1:2, v/v) as stationary phase and 100mMammonium phosphate buffer pH 8.0 asmobile phase systemcould separate up to 300mg of racemic 3,5-dinitrobenzoyl leucine [40]
Figure 8. Enantiomer separation of Dichlorprop by centrifugal partition chromatography (CPC) using Cinchona-type extractant as a chiral stationary phase additives. a) Chemical structures of Dichlorprop and the employed quinine-based chiral extractant. b) Preparative CPC enantiomer separation of 366 mg dichlorprop achieved under optimized operation conditions. Conditions: flow rate of 3 mL/min, mobile phase: aqueous sodium phosphate buffer (100 mM, pH 8.0) as mobile phase; stationary phase: 10 mM (DHQD)2PHAL-type CSPA inMTBE; rotor speed: 1100 rpm; T: 25 °C. r refers to the molar ratio of the loaded amount of racemic dichlorprop and the total amount of extractant employed. Reproduced from [39] with permission.
. Even
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