Alkali-solution and acid-isolation method (ASAI) and aqueous two-phase system separation method (ATSS) have been reported to extract natural colourant carthamin from Carthamus tinctorius. In this study, these two methods were compared based on the optimisation results of extraction conditions by response surface methodology. In ASAI, the maximum extraction yield reached 0.779% at pH value of potassium carbonate solution of 11.16, ratio of potassium carbonate solution to raw material of fifteen and extraction time of 18 min, while 2.652% was achieved with concentration of acetone of 58%, ratio of acetone solution to raw material of twenty-three and extraction time of 41 min in ATSS. From the point of view of extraction yield, ATSS had more superiority than ASAI. However, more pure carthamin was provided in ASAI according to HPLC assay. The obtained results in our experiments could be utilised for further researches of carthamin.