A high-performance liquid chromatography (HPLC) method for determining furfural and 5-hydroxymethylfurfural (HMF) in biomass hydrolysate was developed. The extraction procedure used n-butanol as the solvent to collect furfural and HMF from biomass hydrolysate. The C18 column and ultraviolet visible detector were used as the separation and detection units for determination of the furfural and HMF, respectively. The new method performed well with wide linear ranges of 0.01-0.5 mg/mL (R-2 > 0.9994), low limits of detection and quantitation (less than 0.003 and 0.009 mg/mL), and good recoveries (98.34-100.21% for furfural and 98.78-101.45% for HMF). In comparison to the widely accepted National Renewable Energy Laboratory (NREL) method, it did not differ significantly in determining furfural and HMF in biomass hydrolysate but reduced the running time to less 28 min (from the pretreatment to the finish of analysis), almost half of the 55 min running time for the NREL method. Moreover, if large amounts of samples need to be analyzed, the time-saving advantage of the current method is more obvious as a result of having the capacity of batch extraction.