Abstract: Objective: To optimize the preparation conditions of trans-crocetin from crocin by alkaline hydrolysis for obtaining trans-crocetin with high purity. Methods: A single factor experiment was used to preliminarily investigate the influence of the hydrolysis reaction time, temperature, and alkali concentration on the yield of trans-crocetin. Then the Box-Behnken design response surface experiment was used to optimize the parameters. Results: The best hydrolysis conditions were determined as follows: the reaction time, temperature and alkali concentration were 60.0 min, 80.0 ℃ and 1.70 mg/mL respectively. Under this condition, crude trans-crocetin with purity of 86.0% was obtained, the yield was 80.09%. After recrystallization, the purity of the crude product was increased to 99.3%. The purified product was determined to be all-trans-crocetin by carbon nuclear magnetic resonance (¹³C NMR) and hydrogen spectrum (¹H NMR) analysis. Conclusion: It is an effective method of alkaline hydrolysis from crocin and recrystallization to prepare trans-crocetin with high purity.