Isoflavones were the commonly polyphenols capable of forming inclusion complexes with starch to slow starch enzymatic digestion. However, the impact of isoflavone structures on the formation of starch-isoflavone complexes was not well understood. In this study, isoflavones with distinct structurally differences, including daidzein, genistein, biochanin A, genistin, and puerarin, were selected to examine the interaction between starch and these isoflavones utilizing both experimental and molecular dynamics analysis. The experimental findings showed that daidzein and genistein produced more V-type crystallites with starch, resulting in a greater decrease in starch digestibility compared to other isoflavones. Molecular dynamics simulations suggested that daidzein and genistein, which had smaller molecular size and less hydroxyl groups, formed fewer hydrogen bonds but more inclusion complexes with starch. It appeared that the number of hydroxyl groups and molecular size of isoflavones played a crucial role in the interaction between starch and isoflavones, ultimately influencing the formation of V-type starch crystallites.