The main causes of uncertainty in determination of sexivalent chromium in groundwater by diphenylcarbazide spectrophotometric method is analyzed, calculated and synthesized, using the calculating model of standard curve of double error fitting uncertainty in the course of standard curve fitting. The result of determination of sexivalent chromium in groundwater suggests that the main causes of uncertainty that affect the determination results are preparation of sexivalent chromium standard series solution and repeated measurement of the sample. In addition, the uncertainty in the determination of sexivalent chromium in groundwater is evaluated.