Background: Proper hydration is vital for both exercise and general health. Although various methods for hydration assessment exist, many are not valid for either use or never tested. Introduction: The purpose of this study was to determine whether the uChek© smart phone application can be used to diagnose underhydration based on elevated urine specific gravity (USG) assessed by refractometry. Methods: One hundred forty-seven (n = 147) fresh human urine samples from young and middle-age adults were analyzed for USG with a refractometer and the uChek© application by reading the Siemens Multistix 10G urine reagent strip. Results: Bland-Altman analysis showed agreement of the two methods of assessment. Overall diagnostic ability of the uChek© to identify underhydration was fair (area under the curve 79%). However, the sensitivity to correctly identify underhydration was poor (60%) as well as the specificity of correctly identifying euhydration (53%). Conclusion: The uChek© application does not accurately detect underhydration.