A new, cost-effective, rapid, and precise spectrophotometric method was developed for the quantitative determination of ciprofloxacin in its pure form and some pharmaceutical formulations. This method utilizes the second derivative of the zero-order spectrum. The study demonstrated that this technique enables the quantitative determination of ciprofloxacin within a concentration range of 5–100 µg/mL. The peak height above the baseline at the wavelength of 316 nm was utilized for measurements. The results indicated that the method is accurate and exhibits good reproducibility, with a recovery percentage ranging from 99.5% to 102.5% and a relative standard deviation (RSD%) of 2.5–2.807 for the respective wavelengths. The method was successfully applied to several pharmaceutical formulations.