Abstract Objectives This study assessed the influence of short-term freeze-dried strawberry powder (FDSP) intake on gut microbial composition and select microbial-derived metabolites from feces, serum, and urine, and determine their relationship to microvascular function. Methods In this double-blind, controlled crossover study, postmenopausal overweight or obese women not taking any medications were randomly assigned to consume 39 g FDSP (equivalent to 3 servings fresh strawberries) or an isocaloric control powder mixed in water, daily for 4 weeks, separated by a 4-week washout period. Serum, urine, and fecal metabolomics were performed by proton Nuclear Magnetic Resonance (1H NMR) spectroscopy, microbiome analysis was performed by targeting the v4 region of the 16S rRNA gene. Microvascular function was evaluated as the Framingham reactive hyperemia index (fRHI). Additional measures included anthropometrics, fasting plasma lipids, glucose, and blood pressure. Results Overall, 19 women completed the protocol; with two additional women completing at least one arm of the study. After 4 weeks of FDSP intake, there was a trend toward improved fRHI compared to control (P = 0.069), and a decrease in fasting blood glucose when individuals were stratified according to cardiovascular disease risk (P = 0.049). Further, FDSP intake was associated with a significant increase in propane-1,2-diol, which was negatively correlated with fRHI, as well as the urinary metabolites 1-methylnicotinamide and hypoxanthine. Potential FDSP induced shifts in gut microbial composition will be presented. Conclusions This study confirms previous reports concerning the positive effects of FDSP intake on microvascular function and provides evidence that these changes may be driven in part by changes in metabolism. Funding Sources This work was funded by a grant awarded by the California Strawberry Commission and USDA-NIFA Hatch projects.