Fourier transform infrared (FTIR) spectrometry can be used to measure broadband infrared spectra, and it can analyze various gases in the air simultaneously. A new method of measuring stable isotopes in atmospheric water vapor by open-path FTIR spectrometry is described. An open-path FTIR system is utilized to continuously measure the stable isotopes of water vapor in ambient air in a field experiment, which is based on the analysis of collected mid-infrared spectrum. The deuterium isotope ratios δD are also obtained. The measurement error of this system is about 0.25% and 1.60% for H216O and HD16O respectively, and the measurement precision is about 1.32‰ for δD. A detailed analysis of five-day data is presented to explore the variation of H216O, HD16O and δD with time. Furthermore, Keeling plot method is employed to study the deuterium isotopic signature of evapotranspiration. The results of the field experiment show the ability of this proposed measuring method combined with the open-path FTIR system for long-term measurement of the stable isotopes in the air.