To reduce the adverse consequences of exertion-related and acute intentional dehydration research has focused on monitoring hydration status. This investigation: 1) compared sensitivity of urine specific gravity (U_sg), urine osmolality (U_osm) and a criterion measurement of hydration, plasma osmolality (P_osm), at progressive stages of acute hypertonic dehydration and 2) using a medical decision model, determined whether U_sg or U_osm accurately reflected hydra-tion status compared to P_osm among 51 subjects tested throughout the day. Incremental changes in P_osm were observed as subjects dehydrated by 5% of body weight and rehydrated while U_sg and U_osm showed delayed dehydration-related changes. Using the medical decision model, sensitivity and specificity were not significant at selected cut-offs for U_sg and U_osm. At the most accurate cut-off values, 1.015 and 1.020 for U_sg and 700 m_osm/kg and 800 m_osm/kg for U_osm, only 65% of the athletes were correctly classified using U_sg and 63% using U_osm. P_osm, U_sg, and U_osm appear sensitive to incremental changes in acute hypertonic dehydration, however, the misclassified outcomes for U_sg and U_osm raise concerns. Research focused on elucidating the factors affecting accurate assessment of hydration status appears warranted.