Enantiomeric Separation of Ofloxacin by Nano-Liquid Chromatography Using a Sulfated-beta-Cyclodextrin as a Chiral Selector in the Mobile Phase

The S-(-)-enantiomer of ofloxacin, a chiral fluoroquinolone antibiotic, exhibits an antibacterial activity 8-128 times higher than that of the R-(+)-isomer. A new stereospecific nano-LC method for the enantioseparation of the ofloxacin racemic mixture was developed using an achiral reversed phase column (Pinnacle II Phenyl 3 ¼m, 15 cm x 100 ¼m ID) and heptakis-(2,3-diacetyl-6-sulfo)-beta-cyclodextrin (HDAS-beta-CD) as a mobile phase chiral additive. Best enantioresolution was achieved using a mobile phase containing acetonitrile, 50 mM sodium acetate buffer pH 3 (30:70, v/v) and 5 mM of HDAS-beta-CD. The flow rate thorough the capillary column was estimated in 620 nL/min and the wavelength for the UV detection was set at 290 nm. Good separation of the enantiomers was obtained is less than 10 min with a resolution and selectivity factor of 1.66 and 1.88, respectively. The effect of several experimental parameters on the chiral separation was also evaluated and the method was successfully validated in terms of linearity, accuracy and precision, showing the enantiorecognition capability of the employed sulfated beta-CD toward the ofloxacin chiral drug.