In this study, a new first derivative synchronous fluorescence spectrometry (SFS) for the accurate and sensitive determination of pefloxacin (PFX) and fleroxacin (FLX) in wastewater was established. PFX and FLX were complexed with Al3+ to significantly enhance their fluorescence intensity. Because the conventional fluorescence spectra of PFX-Al3+ and FLX-Al3+ overlap significantly, the wavelength difference (Δλ = 155 nm) was chosen for SFS scanning to narrow spectral band. And the narrowed synchronous fluorescence spectra was subjected to first derivative processing. Finally, the zero intersections of the first derivative synchronous fluorescence spectra at 281.6 nm and 273.6 nm were chosen for the determination of PFX and FLX, cleverly used of spectral parameters to eliminate the serious interference between PFX-Al3+ and FLX-Al3+ and the interference of wastewater background. The limits of detection of PFX and FLX were 0.0086 ng·mL−1 and 0.0068 ng·mL−1 with linear ranges from 0.1–40 ng·mL−1 and 0.2–80 ng·mL−1, respectively. Recoveries of PFX and FLX in wastewater were 88.60 %–98.60 % and 90.00 %–95.60 %. The first derivative SFS established in this study used spectral separation instead of chromatographic separation, so that sample pretreatment was simple and the detection speed was fast. Compared the conventional method of HPLC, the test results of above two methods were close. And the new method had been successfully used for the determination of PFX and FLX residues in wastewater with good results. Therefore, the new first derivative SFS was expected to become a routine detection technology for PFX and FLX residues in wastewater and widely used.
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