Rice origin authenticity is important for food safety and consumer confidence. The stable isotope composition of rice is believed to be closely related to its water source, which affects its origin characteristics. However, the influence of water availability on the distribution of rice stable isotopes (δ 2 H and δ 18 O) is not clear. In this study, three irrigation waters with different isotopic values were used to investigate isotopic water use effects of Indica and Japonica rice, using pot experiments. Under three different water isotope treatments, the δ 2 H values of Indica polished rice showed significant differences (−65.0 ± 2.3, −60.5 ± 0.8 and −55.8 ± 1.7‰, respectively, p < 0.05) compared to δ 13 C and δ 15 N, as did Japonica polished rice. The values of δ 2 H and δ 18 O of rice became more positive when applying more enriched (in 2 H and 18 O) water, and the enrichment effect was higher in rice than in the corresponding plant tissue. In addition, the δ 2 H and δ 18 O values of Indica rice leaves decreased at the heading stage, increased at the filling stage, and then decreased at the harvest stage. Japonica rice showed a similar trend. δ 2 H changes from stem to leaf were more negative, but δ 18 O changes were more positive, and δ 2 H and δ 18 O values from leaf to rice were more positive for both brown and polished rice. The results from this study will clarify different water isotopic composition effects on rice and provide useful information to improve rice origin authenticity using stable isotope-based methods.