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LC Enantioseparation of 30-Component Diastereomeric Mixture of Amino Acids and Detection of d-Isomers Using New Reagents with Amines as Chiral Auxiliaries in Cyanuric Chloride

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Abstract

Two enantiomerically pure amines, viz., (R)-(+)-naphthylethyl amine and (S)-(+)-1-benzyl-3-aminopyrrolidine, were used as chiral auxiliaries for nucleophilic substitution of chlorine atoms in cyanuric chloride or its 6-butoxy derivative. The chiral derivatizing reagents so obtained were characterized and their chiral purity was ascertained. Diastereomers of 15 dl-proteinogenic amino acids were synthesized under microwave irradiation using these reagents. Separation of diastereomeric pairs along with separation of a mixture of 30 diastereomers in a single chromatographic run was carried out on a reversed-phase C18 column. Mixtures of acetonitrile with aqueous trifluoroacetic acid were used as mobile phase. The detection was made at 230 nm using photo diode array detector. The separation behavior in terms of retention times and resolutions was compared on the basis of effect of chiral auxiliaries (i.e. amines) and achiral substituents (i.e. chlorine or butoxy group) in the chiral derivatizing reagents and the hydrophobic side chains of amino acids. The separation method was validated in terms of accuracy, precision, linearity, recovery, limit of detection and limit of quantitation. The method was successful for determination of d-amino acids in the absence of pure d-enantiomers and for separation of 19 diastereomers from a mixture of 30.

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Acknowledgments

The authors are thankful to University Grants Commission (UGC) New Delhi, India, for awarding a senior research fellowship (SRF) (to M.L.). Thanks are due to AvH-Stiftung, Bonn, Germany for donating Knauer HPLC equipment (to R.B.).

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  1. Department of Chemistry, Indian Institute of Technology Roorkee, Roorkee, 247667, India

    Ravi Bhushan & Manohar Lal

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  1. Ravi Bhushan
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  2. Manohar Lal
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Correspondence to Ravi Bhushan.

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Bhushan, R., Lal, M. LC Enantioseparation of 30-Component Diastereomeric Mixture of Amino Acids and Detection of d-Isomers Using New Reagents with Amines as Chiral Auxiliaries in Cyanuric Chloride. Chromatographia 76, 1087–1096 (2013). https://doi.org/10.1007/s10337-013-2515-6

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  • DOI: https://doi.org/10.1007/s10337-013-2515-6

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