A rapid and sensitive method based on transient ITP and field enhancement in CE with electrochemical detection at copper disk electrode was developed for the simultaneous separation and determination of three estrogens: estrone, 17β-estradiol, and estriol. The effects of several important factors that influence the separation and detection were investigated. Under the optimum conditions, the estrogens could be separated in 0.06 mol/L sodium hydroxide solution within 14 min. With transient ITP by addition of 0.5% NaCl, a good linear response was obtained for three estrogens from 0.2 to 10 μmol/L, with correlation coefficients higher than 0.9993. The detection limits were 8.9 × 10−8, 6.7 × 10−8, and 1.1 × 10−7 mol/L (S/N = 3) for estriol, 17β-estradiol, and estrone, respectively. This method was successfully employed to analyze different water samples from waterworks, tap water, fishpond, and river samples with recoveries in the range of 90.8–108.9%, and RSDs < 4.69%. The satisfied results demonstrated that this method was of convenient preparation, high sensitivity, and good repeatability, which could be applied to the rapid determination of environmental water samples.