Project Summary/Abstract: Estrogen deficiency increases the risk for many chronic diseases including cardiovascular diseases (CVDs). Recent evidence demonstrating gut microbial dysbiosis and gut integrity dysfunction that occur in estrogen deficiency provides new mechanistic targets to counter the cardiometabolic disturbances associated with menopause. The gut also plays an important role in the metabolism of estrogen through the action of microbial- derived estrogen deconjugating enzyme (i.e., β-glucuronidase) which alters the enterohepatic recirculation of estrogen making it more bioavailable. β-glucuronidase can also deconjugate phenolic compounds that can prolong their circulation and enhance their bioactivity. It is well established that dietary fibers such as resistant starch (RS) and fructooligosaccharides have prebiotic activity and could be used to counter the effects of estrogen deficiency on gut health and metabolic outcomes. However, polyphenols are also capable of acting as prebiotics by shifting the microbiome to favor the colonization of beneficial gut microbes. Studies examining the prebiotic activity of combination of fiber and polyphenols found in natural products and their mechanisms of action in the context of estrogen deficiency are limited. Pinto beans (PB) are rich in in these bioactive compounds with prebiotic activity and can confer health benefits in estrogen deficiency. Our recent findings demonstrated that PB prevented gut dysbiosis, improved markers of gut integrity, reduced inflammation, and improved insulin sensitivity in mice fed a Western-style diet. In this application, we proposed to build on that work by investigating the mechanism through which PB modulates cardiometabolic outcomes in a mouse model of estrogen deficiency. We hypothesize that PB, due to its many bioactive compounds particularly its RS content, will modulate gut microbial composition and alter β-glucuronidase activity as well as the production of short chain fatty acids (SCFAs) which will consequently improve gut health and metabolic outcomes in estrogen deficiency. This hypothesis will be tested by: Aim 1) determining the alterations in gut microbial composition and metabolic outcomes induced by PB and RS in estrogen deficiency; and Aim 2) investigating the role of the gut-derived β-glucuronidase in the metabolic response due to estrogen deficiency. The proposed study will provide an excellent training opportunity for students and our findings will advance our understanding of how prebiotics such as PB mediate their effects on metabolic outcomes in estrogen deficiency. Our findings will lay the groundwork for future clinical studies, which is in line with our long-term goal of developing economical, safe and effective strategies for cardiovascular disease prevention.