A comparative application of spectrophotometric and spectrofluorimetric methods to estimate levofloxacin-DNA and...

Ofloxacin (OFL) and its (S)-enantiomer, levofloxacin (LEV), are among members of the fluoroquinolone antibiotic class, renowned for their broad-spectrum efficacy against both gram-negative and gram-positive bacteria. These potent drugs have been widely used in both human and veterinary medicine, working as bactericidal agents by binding to DNA gyrase, an essential enzyme for bacterial DNA replication. Understanding the binding constants of these drugs to DNA is vital for elucidating their interaction mechanisms and enhancing our grasp of gene expression regulation. The interactions of LEV and OFL with calf thymus DNA under a physiological medium (0.02 M tris-HCl buffer, pH 7.4) using UV spectrophotometry and spectrofluorimetry were investigated. The assay results obtained by applying two spectroscopic approaches confirmed the presence of the interaction of LEV and OFL antibiotics with DNA. In the LEV-DNA and OFL-DNA interactions, hyperchromic effect and fluorescence quenching were observed for UV spectrophotometric and spectrofluorometric measurements, respectively. In the spectrophotometric analysis, the binding constants for the LEV-DNA and OFL-DNA complexes at 298 K were determined as (1.24 ± 0.047) x 10³ and (1.39 ± 0.040) x 10³ M^- 1, respectively. In the spectrofluorimetric analysis of the interaction of LEV and OFL with DNA, the thermodynamic properties were examined at three distinct temperatures. Based on the fluorescence signal changes the binding constants at 293, 298, and 310 K were calculated as (8.91 ± 0.161) x 10³, (7.62 ± 0.098) x 10³, and (6.08 ± 0.041) x 10³ M^- 1 for LEV-DNA and, (3.14 ± 0.053) x 10³, (3.04 ± 0.031) x 10³, and (2.78 ± 0.023) x 10³ M^- 1 for OFL-DNA, respectively. In these assays, the Gibbs free energy (ΔG⁰), entropy (ΔS⁰), and enthalpy (ΔH⁰) were determined using the Van't Hoff equation. The negative ΔG⁰ values indicate that both LEV-DNA and OFL-DNA interactions are spontaneous. Furthermore, the positive ΔS⁰ and negative ΔH⁰ values revealed that electrostatic forces played a significant role in the binding LEV and OFL to DNA.