Ciprofloxacin is one of the most potent fluoroquinolone antibiotics in medical treatment with a widely effective antibacterial activity. Aim of the presented paper was to use boron doped diamond electrodes for a sensitive, simple and reliable voltammetric determination of ciprofloxacin in human urine samples. Prior to the electrochemical analyses, an optimal boron doping level was determined in order to achieve the highest sensitivity. A set of boron doped diamond electrodes with the doping level in the range from 0 to 20 000 ppm B/C was used for this purpose. Electrochemical behavior of ciprofloxacin was investigated using cyclic voltammetry in an ammonium acetate buffer (pH 5), where ciprofloxacin provided a well‐defined irreversible oxidation peak at a potential of + 1.15 V. Under optimal experimental conditions, the calibration curve obtained by square‐wave voltammetry was linear in a concentration range from 0.15 to 2.11 μmol/L (R2=0.9974). A very low limit of detection (0.05 μmol/L) was obtained for the BDD electrode with the highest doping level. The developed square wave method was successfully applied to the determination of ciprofloxacin in human urine samples with a very good recovery (from 97 to 102%).