Abstract: OBJECTIVE To establish an analytical method for simultaneous determination of sacubitril, valsartan and its main metabolite LBQ-657 in human plasmas by high performance liquid chromatography-fluorescence detection(HPLC-FD). METHODS Precipitation method was used to treat plasma samples via acetonitrile and SN-38 applied as internal standard. Agilent Eclipse SB-C₁₈(250 mm×4.6 mm, 5 μm) was used. The chromatographic separations were accomplished using multi-wavelength with a mobile phase composed of methanol and sodium dihydrogen phosphate buffer(pH 3.0) at ratio of 67:33. The column temperature was maintained at 25℃ with the flow rate of 1.0 mL·min^-1.The injection volume was 20 μL. The multi-wavelength conditions were 0-6 min, λ_EX=380 nm, λ_EM=430 nm, for SN-38; 6-9.4 min, λ_EX=255 nm, λ_EM=314 nm, for sacubitril; 9-12 min, λ_EX=255 nm, λ_EM=374 nm, for valsartan; 12-18 min, λ_EX=255 nm, λ_EM=314 nm, for LBQ-657. Sacubitril and valsartan sodium tablets(dose 100 mg) was administered to 5 patients and sampling after 1.5, 3 h. HPLC-FD was used to detect concentration of sacubitril, valsartan and its main metabolite LBQ-657 in human plasma. RESULTS The calibration curves were linear over the range of 0.1-20 μg·mL^-1(r>0.999). The extraction recovery of sacubitril, valsartan and LBQ-657 were found from 80.30% to 96.45%. According to the intra- and inter-day precisions, accuracy and stability, demonstrating that the method was confirmed to be accurate, precise and stable. CONCLUSION The developed method is successfully applied for determination of sacubitril, valsartan and its main metabolite LBQ-657 in human plasma.