To establish an HPLC method for the simultaneous determination of rhein, emodin, chrysophanol and physcion in Niuhuangqingwei pills. Methods: Diamonsil C18 (250 mm×4.6 mm, 5 μm) at 30 ℃ was used with methanol and 0.1% phosphoric acid (75: 25) as the mobile phase, the flow rate was 1.0 ml·min-1 and the detection wavelength was 254 nm. Results: The linear ranges of rhein, emodin, chrysophanol and physcion were at 22.68-113.40 μg·mL-1(r=0.9991), 6.25～31.24 μg·mL-1(r=0.9996), 14.74～73.68 μg·mL-1(r=0.9990) and 5.22～26.12 μg·mL-1(r=0.9999) respectively, the average recoveries (n=6) were 99.9%（RSD=1.2%）, 99.9%（RSD=1.9%）, 100.0%（RSD=1.2%） and 99.8%（RSD=1.8%).