Hydrogen and oxygen stable isotopes in terrestrial ecosystems can provide unique trace information for studies on water exchange between land and the atmosphere and the hydrological cycle in terrestrial ecosystems. We observed stable isotopic composition and environmental factors in a Canadian deciduous forest during the 2009 growing season. The spatial pattern and temporal variability in isotopic composition for various ecosystem water pools and atmospheric water vapor were analyzed. Further, the variability of the isotopic composition of ecosystem evapotranspiration, and soil evaporation, and their isoflux were analyzed. The main controlling environmental factors were discussed. The results indicated that the isotopic composition varied in different water pools. The isotopic composition of leaf water was more enriched and more variable with comparison to twig and soil water. The H₂¹⁸O and HDO composition of atmospheric water vapor (δ¹⁸O_v and δD_v) decreased with increasing height, and the opposite occurred for deuterium-excess. The diurnal pattern of water vapor isotopes showed a "W" pattern, decreased during the morning, rose and fell slightly at noon, and increased in the afternoon. The isotopic composition of water vapor was significantly correlated with atmospheric moisture, and the mid-day means of the deuterium excess of atmospheric water vapor were both negatively correlated with surface relative humidity and the water vapor mixing ratio, with correlation coefficients of -0.61 (P < 0.01) and -0.57 (P < 0.01), respectively. The H₂¹⁸O composition of evapotranspiration was higher at noon and in the evening and lower in the afternoon because of the influence of evapotranspiration rate and leaf-water enrichment. The isoflux indicated that evapotranspiration could promote the isotopic enrichment of atmospheric water vapor, with a maximum isoflux of 147.5 mmol m^-2 s^-1 ‰. Our results could provide data support and theory reference to the development of isotopic hydrological models.