Abstract: OBJECTIVE To evaluate the extraction process by establishing the UPLC fingerprint of Indigo Naturalis. METHODS The study was designed as a single factor experiment, the number of the common peak, total area of the characteristic fingerprint peak and active ingredient content of the UPLC fingerprint were taken as evaluation indexes to optimize factors including extraction solvent, extraction temperature, extraction time and reactant ratio that might affect heating-circumfluence extraction, ultrasonic extraction and microwave extraction, so as to screen and evaluate the extraction process. RESULTS The optimum extraction solvent of the 3 extraction methods was N,N-dimethylformamide, and the optimum extraction technique was heating-circumfluence extraction (reactant ratio was 0.25 g︰30 mL, extraction temperature was 20 ℃, and extraction time was 60 min), ultrasonic extraction (reactant ratio was 0.25 g︰30 mL, extraction temperature was 50 ℃, and extraction time was 60 min) and microwave extraction (reactant ratio was 0.25 g︰30 mL, extraction temperature was 50 ℃, and extraction time was 8 min), respectively. With the application of the optimum extraction technique, the characteristic peak number of microwave extraction, ultrasonic extraction and heating-circumfluence extraction was 34, 30 and 11, respectively; and corresponding total peak area was 3 495.6, 2 904.9 and 420.9, respectively. In addition, the content of indirubin extracted by microwave extraction was 1.63-fold and 22.57-fold of that by ultrasonic extraction and heating-circumfluence extraction; meanwhile, the content of indigo obtained by ultrasonic extraction was 2.52-fold and 11.40-fold of that by microwave extraction and heating-circumfluence extraction. CONCLUSION Microwave extraction may be superior than ultrasonic extraction and heating-circumfluence extraction with the number of the common peak and total area of the characteristic fingerprint peak in the UPLC fingerprint, and the content of indirubin as evaluation indexes; besides, ultrasonic extraction may be better than microwave extraction and heating-circumfluence extraction when using the number of the common peak and total area of the characteristic fingerprint peak in the UPLC fingerprint, and the content of indigo as the evaluating indicators.