Abstract:The content of resistant starch affects the physiological functions of food and directly determines the processing characteristics of raw materials rich in resistant starch. Accurately measuring the content of resis‐tant starch in food is a prerequisite for its application. The key factors affecting the determination of resistant starch content are the removal of non‐resistant starch from the sample and the hydrolysis of resistant starch in the sample. The experiment selected high amylose corn resistant starch（7012），red bean，and sweet potato samples as research objects，and optimized the optimal conditions for removing non‐resistant starch and hydro‐lyzing resistant starch through single‐factor experiments. The optimal conditions for removing non‐resistant starch from the sample were determined by adding 30 U/mL α‐amylase and 3 U/mL starch glucosidase and shaking in a water bath at 37 ℃for 16 h of hydrolysis，followed by washing twice with 50% ethanol solution.The optimal conditions for hydrolyzing resistant starch involved adding 0.10 mL of 3 260 U/mL starch glucosi‐dase and shaking in a water bath at 50 ℃for 30 min. The method was validated for precision，accuracy，detec‐tion limit，and quantification limit. The results showed that the relative standard deviation of 10 measurements of resistant starch content in the sample was less than 10%，indicating good precision of the method. The error between the measured value and the labeled value of the quality control sample with known resistant starch content was 2.62%，indicating high accuracy of the method. In conclusion，the method established in this ar‐ticle is suiTablefor the determination of resistant starch content in food.