A high-performance liquid chromatography (HPLC) method was developed for the determination of 5-fluorouracil (5-FU) in 5-FU injection and human serum. A Nova-park C18 column (3.9 x 150 mm, 4.6 μm) and mobile phase consisting of 5 mmol/L KH2PO4 (pH 6.0) - methanol (96:4, v/v) at a flow rate of 1.0 mL/min were used for separation. The detection wavelength was set at 254 nm. The human serum was treated with AgNO3 (20%) and NaCl (20%). The method showed a good linear relationship in the range of 0.1 - 100 μg/mL (R2 = 0.9999) with limits of quantification (LOQ) at 0.10 μg/mL and 0.85 μg/mL (S/N = 10) in 5-FU injection and human serum, respectively. The average recoveries of different concentrations in 5-FU injection and human serum were 95.79 - 105.1% and 90.2 - 108.8%, respectively. The relative standard deviation (RSD) of precision was 0.4%, and those of stability and repeatability were less than 1.3% and 2.1% (n = 5) in both samples, respectively. Impurities in the human serum did not interfere with the determination of 5-FU. The method is simple, rapid, sensitive and can accurately determine 5-FU in 5-FU injection and human serum.

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