Abstract: In order to provide insights into developing low-glycemic index (GI) flour products, this study focus on the effect of processing accuracy on the in vitro digestion characteristics of starch in wheat flour. A series of wheat flours with different processing accuracy (flour yield of 55% to 95%, with 55% to 75% considered as high processing accuracy, and 75% to 95% as low processing accuracy) were prepared based on dietary fiber content and cumulative flour yield using different system powders, middling, and bran as raw materials. The starch digestion characteristics were determined using the in vitro Englyst method. Combined with the determination of wet gluten content, swelling power, pasting properties, thermal properties, and correlation analysis of those with the starch digestion characteristics, the mechanism of processing accuracy affecting starch digestibility was elucidated. The results showed that as the processing accuracy decreased, the in vitro hydrolysis rate of starch and the content of rapidly digestible starch decreased, while the content of slowly digestible starch and resistant starch (RS) increased. The decrease of maximum degree of starch hydrolysis of wheat flour with high processing accuracy was higher than those with low processing accuracy. For all wheat flours with different processing accuracy, RS content showed a significant negative correlation with peak viscosity and setback value (P < 0.01). At low processing accuracy, the correlation between dietary fiber content and wet gluten, as well as free sulfhydryl content, was stronger. The starch in wheat flour with low processing accuracy is more resistant to digestion, which is significantly correlated with the deterioration of gluten network exacerbated by the higher dietary fiber content. For wheat flour with high processing accuracy, dietary fiber and gluten synergistically competed with starch for water absorption, which had a certain physical barrier effect on starch granules, limited starch swelling, inhibited amylose leaching, and limited the movement of starch chains, thereby improving the resistance to enzymatic hydrolysis. For wheat flour with low processing accuracy, dietary fiber deteriorated the gluten network, resulting in a decrease in the barrier effect of gluten on starch granules decreased, and the starch digestibility decreased slightly.