Fluorescence spectra, fluorescence quantum yield and dissociation constant of sarafloxacin

In this study, the fluorescence spectra of sarafloxacin (SAR) under different pH conditions were investigated to determine the structural changes due to protonation that result from change in pH. At pH < 1.02, SAR exists in the H₃ L²⁺ form for which the maximum fluorescence emission wavelength was about 455 nm. At pH 1.87-4.94, SAR exists in the H₂ L⁺ form in which H₃ L²⁺ loses one proton in the nitrogen molecule at the 1-position in the quinoline ring. Fluorescence intensity was strong and steady and the maximum emission wavelength was 458 nm. At pH 7.14-9.30, the maximum emission wavelengths were gradually blue shifted to 430 nm with increase in pH, here SAR exists in the form of a bipolar ion HL in which H₂ L⁺ loses a carboxyl group proton. At pH > 11.6, HL transforms into anionic L^- in which HL loses one proton from the piperazine ring, leading to a decrease in fluorescence intensity, and the maximum emission wavelength was red shifted to approximately 466 nm. The two-step dissociation constant pKa for SAR was calculated, pK_a1 was 6.06 ± 0.37 and pK_a2 for SAR was 10.53 ± 0.19. In a pH 3.62 buffer solution with quinine sulfate as the reference, the fluorescence quantum yield of SAR at the maximum excitation wavelength of 276 nm was 0.09.