Chia, amaranth, quinoa and buckwheat grains have been widely used in food formulations because of their high and balanced nutritional properties. Since all grains are not equally nutritious, there is a requirement for comparing the health-related effects and processing performance of a variety of whole grains. The expected glycemic index (eGI) flours of chia was determined to be quite low, and flours except quinoa can be classified as low GI foods. The highest resistant starch (RS) content (4.76 g/100 g) was found in amaranth flour, and it was followed by buckwheat (1.27 g/100 g). The amaranth had the highest stimulation effect on the growth of probiotics and increased the count of L. acidophilus and B. bifidum as 4.57 and 2.26 log CFU/ml, respectively. Moreover, chia flour showed a positive effect on the growth of L. acidophilus whereas it negatively affected B. bifidum compared to the control. A significant correlation was detected between rapidly available glucose content and eGI. On the other hand, a significant relationship between RS and the growth rate of probiotics was reported.