Repeated cooking and freezing of whole wheat flour increases resistant starch with beneficial impacts on in vitro fecal fermentation properties

• Increased resistant starch in whole wheat flour may improve prebiotic properties.
• Cooking–freezing cycles of whole wheat flour change starch digestion fractions.
• Minor increase in resistant starch modulated metabolic activity of gut microbiota.
• Total propionate increased as increased cooking–freezing cycles.

Resistant starch (RS) has shown benefits to gastrointestinal health, but it is present in only small amounts in most grain-based foods. The purpose of this study was to increase RS in whole wheat flour to improve its potential health benefits. Zero to 7 cycles of cooking (20 min, boiling water) and freezing (−18 °C, 23 h) of whole wheat flour in water (1:15 %w/v) were performed. Increasing cooking–freezing cycles increased RS from 1.03 to 8.07% during in vitro starch digestion. During in vitro fecal fermentation, increasing cooking–freezing cycles increased short chain fatty acids, mainly propionate. Increases in butyrate were also noted during the first 8 h of fermentation. All flours resulted in significant increases in Bifidobacterium of >0.5 log during fermentation compared to baseline. Thus, even modest increases in the RS content of whole wheat flour modulated the metabolic activity of gut microbiota to increase production of beneficial metabolites.