R-Besifloxacin hydrochloride is a novel chiral broad-spectrum fluoroquinolone developed for treatment of bacterial conjunctivitis and used in clinics due to its higher antibacterial activity compared to S-enantiomer. UHPLC separation of R- and S-enantiomers of Besifloxacin hydrochloride on a new Chiralpak IE-3 column consisting of an immobilized amylose tris(3,5-dichlorophenylcarbamate) chiral stationary phase on 3 µm silica was studied. The selection and effects of mobile phase solvents, additives, organic modifiers, and column temperature on retention, separation, and resolution were investigated and optimized. The separation was accomplished under normal-phase conditions using a mobile phase composed of dichloromethane:isopropyl alcohol:trifluoroacetic acid:diethylamine (90:10:0.3:0.05 v/v/v/v) under isocratic mode of elution at 15 °C. The interactions between the analytes, additives, and the stationary phase were discussed and the most probable separation mechanism was proposed. The developed method was validated according to ICH guidelines and found to be suitable for determining the enantiomeric purity of R-Besifloxacin in bulk drugs and formulations.
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The authors thank the Director, IICT, for encouragement and permission to communicate the results for publication. A. M. Swamy has received Senior Research (SRF) Fellowship from Council of Scientific and Industrial Research (CSIR) [Grant No. 31/14(2540/2013-EMR-I)] and N. N. Varma has received Senior Research (SRF) Fellowship from University Grants Commission (UGC) (Grant No. 2013/14-10454-94).
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All the authors declare that they have no conflict of interest.
This article does not contain any studies with human participants or animals performed by any of the authors.
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Ramisetti, N.R., Arnipalli, M.S., Nimmu, N.V. et al. UHPLC Determination of Besifloxacin Enantiomers on Immobilized Amylose Tris(3,5-dichlorophenylcarbamate) Chiral Stationary Phase. Chromatographia 80, 1509–1515 (2017). https://doi.org/10.1007/s10337-017-3346-7