In this paper， the preparation technology， analysis and stability of hypouricemic peptide from Katsuwonus pelamis were discussed. Taking the inhibitory activity of xanthine oxidase (XO) and the hydrolysis degree as indicators， enzymatic hydrolysis process of hypouricemic peptide was optimized， and the molecular weight， amino acid composition and stability of the hydrolysate were also analyzed. The results showed that the neutral protease was the most suitable protease for preparing the hypouricemic peptide of Katsuwonus pelamis， through single factor and response surface optimization tests， the optimal process conditions were determined of 40 ℃ temperature， pH 7.1， 10 400 U/g enzyme addition， enzymolysis time 4 h and material to liquid ratio 1∶5(g/mL). Under these conditions， the XO inhibition rate and hydrolysis degree of the prepared enzymatic hydrolysate were (83.91±0.16)% and (38.67±0.39)% respectively， and its yield was (18.24±1.02)%. Molecular weight distribution and amino acid analysis showed that the prepared hypouricemic hydrolysate was mainly composed of small molecular peptides， whose molecular weight less than 1 ku and hydrophobic amino acids accounted for 91.9% and 49.61% respectively. The prepared hypouricemic hydrolysate had good stability to heat， acid and alkali as well as metal ions such as Ca2+， Mg2+， AI3+， K+， Na+ and NH+. Adding Zn2+ and Cu2+ could significantly increase the XO inhibitory activity of hydrolysate， whereas the presence of Fe2+ and Fe3+ significantly reduced the XO inhibitory activity. In addition， the enzymatic hydrolysate was relatively stable in the gastrointestinal simulated digestion environment， and its XO inhibitory activity was significantly enhanced after in vitro intestinal simulated digestion. This study provides a theoretical basis for the deep and high value-added utilization of Katsuwonus pelamis resources.