Critical points in the analysis of ciprofloxacin by high-performance liquid chromatography

Chromatographic analysis of the antibiotic compound ciprofloxacin on a C18 column proved to show some critical points. On a C18 column, ciprofloxacin analysis resulted in two peaks in the chromatogram. Effects of stock solution composition, mobile phase composition and type of stationary phase were investigated to understand the occurrence of both peaks. Esterification of the carboxylic group of ciprofloxacin was shown when methanol was used as stock solution solvent in acidic conditions. Secondly, a ciprofloxacin pKa value of 3.64 in water at position N-1 was found which could not fully explain the observation of the two peaks: mobile phase composition and type of stationary phase were also found to be important parameters with respect to the peak area ratio of both peaks.