Optimization of enrofloxacin and ciprofloxacin in Nile tilapia muscle by high-performance liquid chromatography coupled with fluorescence detector using tetracycline as internal standard

The objective of this study was to improve the determination techniques of enrofloxacin (ENR) and ciprofloxacin (CIP) in tilapia muscle by high-performance liquid chromatography coupled with a fluorescence detector (HPLC-FLD). A fluorescence detector was used to simultaneously detect ENR, CIP, and TC at the corresponding wavelengths; exciting 274 nm and emission wavelength 428 nm. Tetracycline (TC) was used as an internal standard. The target compounds were successfully separated when using combined column C18 VertiSep™ AQS 4.6x250 mm, 5 µm. The column temperature was set at 30oC. The isocratic elution was performed as mobile phase A 0.1% (v/v) formic acid in Milli-Q water, and mobile phase B was 0.1 % (v/v) formic acid in acetonitrile (82:18, v/v). The injection volume and the flow rate were 20 μL and 1.0 mL/min. ENR, CIP, and TC have been extracted by vortex-assisted extraction with an extractant. Finally, the extracted samples were filtered with 0.20 μm of nylon filter and injected into HPLC-FLD. The determination coefficient (R2) was 0.9999 for the concentration ranging from 1.56˗100 μg/kg. The recovery of CIP and ENR at 6.25 and 25 μg/kg were 95.52˗105.28 % and 67.68˗103.36 %, respectively. Relative standard deviations (%RSD) of CIP and ENR were 3.24˗5.79 % and 0.33˗5.46 %, respectively. This study revealed a successful optimization and effective determination of ENR and CIP in fish samples by HPLC-FLD.