The invention discloses a method for accurately preparing an acetic acid standard curve solution and detecting protected amino acid acetic acid, and the method comprises the following steps: S1, preparing an acetic acid stock solution, precisely weighing a designed amount of anhydrous potassium acetate, and adding ultrapure water to a constant volume according to a detection concentration demand to obtain the acetic acid stock solution with a designed concentration; S2, preparing an acetic acid standard curve solution, adding ultrapure water to dilute the stock solution to obtain an acetic acid standard curve solution with a designed concentration, and adding trifluoroethanol or methanol with a designed amount during preparation. By improving the formula of the standard curve, the preparation process of the standard curve is simpler, more convenient and more accurate, the use of an organic solvent is avoided, the volatilization of the solvent is reduced, and the detection cost is reduced; meanwhile, the interference peak and the target peak are effectively separated by adopting ion chromatography for detection and designing reasonable gradient conditions, and finally, the target peak is obvious in performance and easy to judge and calculate, so the problems of misjudgment and misjudgment in the prior art are solved.