In order to achieve rapid and highly sensitive detection of copper in natural water by laser-induced breakdown spectroscopy (LIBS), the water samples were analyzed by LIBS after copper therein was enriched by montmorillonite substrate. According to the emission spectral characteristics of copper enriched on the solid substrate, the characteristic spectral line of copper at 324.7 nm was chosen as the analytical line of copper. Measurements of the intensity and signal to noise ratio of the analytical line in different parameter occasions show that the optimal laser spot size is 100 μm, the optimal pulse laser energy was 45.9 mJ, and the optimal delay time is 3 μs. Under the optimized experimental conditions, the calibration curve of copper was established. The linear correlation coefficient with 0.996 indicates a good linear relationship between the copper concentration in water and the intensity of the analytical line. The detection limit for copper in water reached 0.03 mg/L. Furthermore, the method was used to determine the copper concentration in water samples collected from different sites. The measurement results are consistent with those measured by the inductively coupled plasma-atomic emission spectrometry, indicating that the method can be applied in the measurement of trace copper in natural water.