A new analytical method using Back-Propagation(BP) artificial neural networks and spectrophotometry for simultaneous determination of calcium and magnesium in tap water, the Yellow River water and seawater is established. By condition experiment, the optimum analytical conditions for calcium, magnesium and Arsenazo(Ⅲ) color reactions are obtained. Levenberg-Marquart(L-M) algorithm is used for calculation in BP neural network. The topological structure of three-layer BP ANN network architecture is chosen as 11-10-2(nodes). The initial value of gradient coefficient μ is fixed at 0. 001 and the increase factor and reduction factor of μ take the default values of the system. The data are processed by computers with our own programs written in MATLAB 7. 0. The relative standard deviations of the calculated results for calcium and magnesium are 2. 31% and 2. 14%, respectively. The results of standard addition method show that the recoveries of calcium and magnesium are 103. 6% and 100. 8% in the tap water, 103. 2% and 96. 6% in the Yellow River water(Lijin district of Shandong Province), and 98. 8%-103. 3% and 98. 43%-103. 4% in seawater from Jiaozhou Bay of Qingdao. It is found that 14 common cations and anions do not interfere with the determination of calcium and magnesium under the optimum experimental conditions. The comparative experiments do not show any obvious difference between the results obtained by this new method and those obtained by the classical complexometric titration method in seawater medium. This method exhibits good reproducibility and high accuracy in the determination of calcium and magnesium and can be used for the simultaneous determination of Ca2+ and Mg2+ in tap water and natural water.