The aim of this study was to evaluate potential drug-drug interaction between lithium and levofloxacin. The study was conducted on New Zealand white rabbits with three groups having two subgroups each (n = 12). The first group compared the pharmacokinetic (Pk) parameters of lithium when lithium was given as a single dose (56 mg/kg) and when lithium was co-administered with levofloxacin (35 mg/kg). The second group compared the Pk parameters of lithium when lithium was given for 6 days alone and when levofloxacin was given on the sixth day after lithium steady-state levels were achieved. The third group compared the Pk parameters of lithium when lithium was given alone for 8 days and levofloxacin was given on days 6, 7, and 8 along with lithium. Apart from this, creatinine levels were also measured to detect nephrotoxicity effects because of this co-administration. It was found that there was increase in lithium levels in all three groups. The increase was significant when a single dose of levofloxacin was administered with steady-state level of lithium (C(max) of lithium: 2.54 ± 0.15 vs 2.79 ± 0.12 mm, P < 0.001 and AUC(0-α) of lithium: 24.36 ± 3.68 vs 31.88 ± 4.83 mmol/mL h, P < 0.001). The increase in lithium levels was also significant when levofloxacin was coprescribed for 3 days after lithium steady-state levels were achieved (C(max) increased from 2.72 ± 0.29 to 3.96 ± 0.29 mm, P < 0.001 and AUC(0-α) increased from 27.1 ± 4.96 to 42.64 ± 4.94 mmol/mL h). Levofloxacin increases lithium levels when they are co-administered, and this interaction might be clinically significant as they may be coprescribed.