HPLC-DAD-FLD determination of veterinary pharmaceuticals in wastewaters of pharmaceutical industry with and without precolumn derivatization by fluorescamine

Many veterinary pharmaceutical industries use the surface water as a recipient for wastewater effluent after treatment. Those wastewaters are usually loaded with very high concentration of pharmaceuticals, especially with veterinary antibiotics. Classical wastewater treatment plants are not suitable for removing pharmaceuticals from wastewaters of pharmaceutical industry, so, they can get into the receiving surface waters with the effluents. From these reasons it is necessary to monitor pharmaceuticals in wastewaters before and after treatment. In this purpose it was developed a robust, simple, realistic and practical analytical method capable of simultaneous determination of target compounds [sulfonamides(sulfaguanidine, sulfadiazine, and sulfadimidine), a sulfonamide synergist (trimethoprim), a tetracycline (oxytetracycline), fluoroquinolones (ciprofloxacin, enrofloksacin, norfloksacin) and local anesthetic procaine], by HPLC with diode array (DAD) and fluorescence (FLD) detectors. A liquid chromatographic method with fluorimetric detection is proposed for the determination of pharmaceuticals in wastewater samples because of highly fluorescent fluoroquinolones (FQ). Limit of detection for FQ is almost ten times lower with HPLC-FLD then with HPLC-DAD. Other investigated pharmaceuticals need to be derivatized with a fluorescamine solution in acetonitrile (5 mg/10 mL acetonitrile) and acetic buffer for determination on HPLC-FLD. The fluorescent compounds are detected at different λex/λem (nm/nm) values: sulfonamides, trimethoprim, oxytetracycline and procaine at 405/495 and fluoroquinolones at 277/445. The analytes are preconcentrated by solid phase extraction (SPE) using Strata-X cartridges and acetonitrile as eluent. The chromatographic separation is performed on a Synergy Fusion-RP18 80 A, 150 mm × 4.6 mm, particle size 4 microm column with a gradient elution program based on binary mixtures of solvents A (0.1% acetic acid in water) and B (0.1% acetic acid in acetonitrile). The flow rate was 1.0 mL/min. The performance characteristics of the SPE-HPLC-FLD method were established by validation procedure. Selectivity, linearity, limits of detection (LOD) and quantification (LOQ), precision and recovery were studied. The described method is applied to the determination of the investigated pharmaceuticals in pharmaceutical industry wastewater.