Tartary buckwheat has established itself as a functional food source because of its basic nutrition and phenolic compound contents, such as dietary fiber (DF) and rutin (RU). However, little information has been obtained concerning the comparative effects of DF and RU on the in vitro and in vivo glucose responses of tartary buckwheat flour. Moreover, the relationship between the flour’s in vitro starch digestibility and its components’ blood glucose response is not well-known. This study found that DF and RU reduced rapidly digestible starch (RDS) by 37.32→33.88% and 41.71→30.28%, whereas they increased resistant starch (RS) by 30.47→31.46% and 28.41→36.78%, respectively. Furthermore, RU had a lower glycemic index (GI) compared to DF. The regression equation for the in vitro and in vivo data from RU exhibited positive correlation (R 2 = 0.99); however, DF did not display positive correlation, which indicates that the in vitro and in vivo GI mechanisms by DF and RU are different.