OBJECTIVE: To establish a method for simultaneous determination of vitamin B1, dihydralazine sulfate, vitamin B6 and vitamin B5 in Compound Apocynum venetum tabletsⅠ. METHODS: HPLC method was used for content determination of 8 batches of samples from 2 enterprises. The determination was performed on Waters XBridge C18 column with mobile phase consisted of acetonitrile-0.2% phosphoric acid (gradient elution) at the flow rate of 0.5 mL/min. The detection wavelengths were set at 285 nm (vitamin B1, dihydralazine sulfate, vitamin B6) and 210 nm (vitamin B5). The column temperature was 30 ℃, and sample size was 10 μL. RESULTS: The linear ranges of vitamin B1, dihydralazine sulfate, vitamin B6 and vitamin B5 were 41.66-208.3 μg/mL (r=0.999 9), 127.0-635.0 μg/mL (r=0.999 9), 40.74-203.7 μg/mL (r=0.999 9) and 22.72-113.6 μg/mL (r=0.999 9), respectively. The limits of quantitation were 0.267, 0.274, 0.062, 0.124 μg/mL, respectively. The limits of detection were 0.095, 0.088, 0.021, 0.039 μg/mL, respectively. RSDs of precision, stability and reproducibility tests were all lower than 2.0% (n=6). The average recoveries were 98.70% (RSD=0.69%, n=9), 99.74% (RSD=1.2%, n=9), 99.89% (RSD=1.0%, n=9), 99.25% (RSD=1.2%, n=9), respectively. Among 8 batches of samples, the contents of vitamin B1, dihydralazine sulfate, vitamin B6 and vitamin B5 were 91.44%-98.74%, 36.68%-95.23%, 92.42%-97.90%, 91.36%-99.48% of labelled amount, respectively. The contents of dihydralazine sulfate were significantly different and differed by 2.6 times between the lowest and the highest threshold value. CONCLUSIONS: The method is accurate, reliable, simple and can be used for simultaneous determination of the above 4 components in Compound A. venetum tabletsⅠ.