Identifying plant water sources is fundamental for elucidating ecohydrological processes and improving water resource management in arid zones under climate change. Stable hydrogen and oxygen isotopes are commonly used to trace plant water uptake; however, cryogenic vacuum extraction (CVE), the standard method for extracting plant xylem water, may induce deuterium depletion, thereby biasing source attribution. To systematically assess the effects of CVE-induced deuterium depletion across species, size classes, and habitats, we excavated five representative soil profiles along the mainstream of the Tarim River in northwestern China, in mid-July 2022. A total of 29 individuals, comprising both Populus euphratica and Tamarix ramosissima, were sampled. We divided P. euphratica individuals into four groups based on diameter at breast height (<50, 50-100, 100-250, and >250 cm), while categorized T. ramosissima individuals into four groups according to plant height (<1.0, 1.0-2.0, 2.0-4.0, and >4.0 m). Plant xylem water was extracted using CVE, and five deuterium depletion scenarios (-5.00%o, -7.00%o, -9.00%o, -11.00%o, and-13.00%o) were simulated. The Bayesian Mixing Model for Stable Isotope Analysis in R (MixSIAR) was applied under six input modes to quantify the proportional contributions of potential water sources and associated prediction errors. Model evaluation revealed that P. euphratica achieved the highest accuracy with a-9.00%o correction of depletion, whereas a-11.00%o correction was optimal for T. ramosissima, reducing relative prediction errors by 68.65% and 67.73%, respectively, compared with uncorrected scenario. Small-sized P. euphratica individuals exhibited less deuterium depletion, whereas no clear size-dependent pattern was observed for T. ramosissima. Spatially, plant individuals located farther from the river exhibited reduced deuterium depletion in xylem water. Despite differences in species traits and habitat conditions, both species predominantly relied on deep soil water and groundwater, which together contributed, on average, 61.45% and 59.95% for P. euphratica and T. ramosissima, respectively. These findings highlight the necessity of accounting for CVE-induced deuterium depletion when identifying plant water-use strategies and provide methodological guidance for isotope-based ecohydrological studies in arid environments.