A simple procedure is described for the simultaneous extraction and determination of betulin and betulinic acid in white birch bark. The extraction was checked using different solvents: dichloromethane, ethyl acetate, acetone, chloroform, methanol and 95% ethanol (aqueous solution, v/v). It was found 95% ethanol was a good extraction solvent that allowed extraction of triterpenoid with a highest content. Separation was achieved on a reversed phase C18 column with acetonitrile–water 86:14 (v/v). Detection was accomplished with UV detection at λ = 210 nm. Using this method, the bioactive triterpenoid in white birch bark were simultaneously determined. Significant variations in the content of betulin and betulinic acid in white birch bark growing in different locations of China were also observed.