In order to establish an HPLC method for the content determination of palbociclib, HPLC is performed on the column of Agilent C₁₈ （250 mm × 4.6 mm，5 μm）with the mobile phase of acetonitrile-water (300∶700，0.1% TFA included) at the flow rate of 1.0 mL/min, with the sample size of 10 μL, the detection wavelength of 234 nm and the column temperature of 40 ℃. The result shows that the main drug and the related substances could be well separated; the calibration curve of palbociclib is in good linearity over the range of 50~150 μg/mL, and r=0.999 5; the RSD of instrument precision, intermediate precision and stability test of related substances and content are no more than 2.0%; the average recovery is 100.0% with RSD of 0.8% (n=9). The method can rapidly and accurately determine the PD content, effectively eliminate the effects of other external factors. It provides a new reference to the content determination of active pharmaceutical ingredients.