Abstract: In order to ensure the accuracy and reliability of high resolution mass spectrometry for the determination of sildenafil derivatives in aphrodisiac oral liquid, it is necessary to evaluate the uncertainty of the determination results. In this study, the content of 6 sildenafil derivatives in oral liquid health food was determined by ultra-high performance liquid chromatography with quadrupole/electrostatic field Orbitrap high resolution mass spectrometry (UPLC-Q/Orbitrap HRMS). The mathematical model was established and the measurement uncertainty was evaluated according to JJF 1059.1-2012 “Evaluation and Expression of Measurement Uncertainty”. When the contents of N-demethyl sildenafil, dimethyl thiosildenafil, dimethyl sildenafil, hydroxythiocarmo sildenafil, thiocarmo sildenafil and thiosildenafil in oral solution were 0.90, 1.04, 1.22, 2.01, 1.51 and 1.14 mg/kg, the extended uncertainties were 0.07, 0.09, 0.10, 0.16, 0.13 and 0.09 mg/kg (k=2), respectively. The results show that the uncertainty mainly comes from UPLC-Q/Orbitrap HRMS, standard solution preparation, recovery and random effect.