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Resolution and isolation of enantiomers of ( ± )‐isoxsuprine using thin silica gel layers impregnated with l ‐glutamic acid, comparison of separation of its diastereomers prepared with chiral derivatizing reagents having l ‐amino acids as chiral auxiliaries
Author(s) -
Bhushan Ravi,
Nagar Hariom
Publication year - 2015
Publication title -
biomedical chromatography
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.4
H-Index - 65
eISSN - 1099-0801
pISSN - 0269-3879
DOI - 10.1002/bmc.3284
Subject(s) - diastereomer , chemistry , chromatography , enantiomer , detection limit , elution , silica gel , high performance liquid chromatography , trifluoroacetic acid , derivatization , chiral derivatizing agent , reagent , chiral column chromatography , organic chemistry
Thin silica gel layers impregnated with optically pure l ‐glutamic acid were used for direct resolution of enantiomers of ( ± )‐isoxsuprine in their native form. Three chiral derivatizing reagents, based on DFDNB moiety, were synthesized having l ‐alanine, l ‐valine and S ‐benzyl‐ l ‐cysteine as chiral auxiliaries. These were used to prepare diastereomers under microwave irradiation and conventional heating. The diastereomers were separated by reversed‐phase high‐performance liquid chromatography on a C 18 column with detection at 340 nm using gradient elution with mobile phase containing aqueous trifluoroacetic acid and acetonitrile in different compositions and by thin‐layer chromatography (TLC) on reversed phase (RP) C 18 plates. Diastereomers prepared with enantiomerically pure (+)‐isoxsuprine were used as standards for the determination of the elution order of diastereomers of ( ± )‐isoxsuprine. The elution order in the experimental study of RP‐TLC and RP‐HPLC supported the developed optimized structures of diastereomers based on density functional theory. The limit of detection was 0.1–0.09 µg/mL in TLC while it was in the range of 22–23 pg/mL in HPLC and 11–13 ng/mL in RP‐TLC for each enantiomer. The conditions of derivatization and chromatographic separation were optimized. The method was validated for accuracy, precision, limit of detection and limit of quantification. Copyright © 2014 John Wiley & Sons, Ltd.