Resolution Of Pharmaceutical Intermediate Enantiomers By HPLC With Chiral Stational Phase

OBJECTIVE:Enantiomers differ in their pharmacodynamics, pharmacokinetics and toxicology significantly. A single enantiomer is greater therapeutic benefits and low toxicity . Therefore, these properties of the enantiomers have created an interest to search the enantiomerically pure compounds, as well as to establish enantiomeric separation and determination technology of high property, high resolution and high sensitivity . At present the separation methods employed mainly included gas chromatography, liquid chromatography and capillary electrophoresis , high performance liquid chromatography (HPLC) employing a chiral stationary phase (CSP) is used most widely, which is a simple, rapid, reliable, accurate, selective method for the separation of chiral drugs.In this assay, the separation behavior of peptidase-inhibitor intermediate (3-(n-boc-amino)-1-oxirane-4-phenylbutane) and darifenacin intermediate (3-(cyanodiphenylmethyl)pyrrolidine-1-tert-butylcarboxylate) enantiomers on polysaccharide chiral columns (Chiralcel OD column , Chiralcel OJ column , Chiralpak AD column and Chiralpak AS column), as well as the resolution influence of mobile phase for the two intermediate enantiomers was respectively investigated for the establishment of methods for the resolution of the two intermediate enantiomers.METHODS:Chromatography conditions: column: Chiralcel OD column (250 mm×4.6 mm, 10μm), Chiralcel OJ column (250 mm×4.6 mm, 10μm), Chiralpak AD column (250 mm×4.6 mm, 10μm), Chiralpak AS column (250 mm×4.6 mm , 10μm). Mobile phase: hexane- alcohol (methanol, ethanol, n-propanol, isopropanol). The flow rate of mobile phase was 1.0 mL/min. The absorbance wavelength was carried out at 220 nm. The column temperature was carried out at 25℃and the injection volume was 20μL.RESULTS:Chiralcel OD column was the most suitable for resolution of 3-(n-boc-amino)-1-oxirane-4-phenylbutane enantiomers, on which the effective separation of the four isomers was achieved simultaneously with the mobile phase of hexane-ethanol-isopropanol (99:0.5:0.5, v/v/v); The effective separation of 3-(cyanodiphenylmethyl)pyrrolidine-1-tert-butylcarboxylate enantiomers was obtained on Chiralpak AD or Chiralcel OJ column , and its better resolution was achieved on Chiralpak AD column using hexane-ethanol (95:5, v/v) as the mobile phase(Rs=7.50).CONCLUSION:The established normal-phase HPLC separation of 3-(n-boc-amino)-1-oxirane-4-phenylbutane can be used for the analytical research of optical purity of (2S,3S)-isomer or (2R,3S)-isomer , and the established method of 3-(cyanodiphenylmethyl)pyrrolidine-1-tert-butylcarboxylate enantiomers can be used for the analytical research of optical purity of (S)-isomer.