Abstract: Spatial and temporal variability of hydrogen and oxygen isotope compositions of water provides a very effective means for studying on the water cycle,inversion of the paleoclimate and paleo-environment, identification of water source, etc. This paper presents a method for online simultaneous determination of δD and δ¹⁸O in micro-liter water samples by thermal conversion/elemental analysis-isotope ratio mass spectrometry (TC/EA-IRMS). The sample size is reduced to only 0.2 μL of water and the determination precision are 0.81‰ and 0.06‰ for δD and δ¹⁸O respectively. Sample classification (classification based on δD difference between samples and standards <50‰) can eliminate memory effect and achieve accurate calibration for the determination results without carefully calibrating reference gases and measuring H⁺₃-factor.