Cardiovascular disease (CVD) is strongly associated with the gut microbiota and its metabolites, including trimethylamine-N-oxide (TMAO) formed from ʟ-carnitine. Raw garlic juice, with allicin as its primary compound, has been shown to powerfully impact the gut microbiota. This study validated the benefits of raw garlic juice against CVD risk via modulation of the gut microbiota and its metabolites. Allicin supplementation significantly decreased serum TMAO in ʟ-carnitine-fed C57BL/6J mice. It also reduced aortic lesions and altered the fecal microbiota in carnitine-induced, atherosclerosis-prone, apolipoprotein E-deficient mice. In human subjects exhibiting high TMAO production, raw garlic juice intake for a week reduced TMAO formation, improved gut microbial diversity, and increased the relative abundances of beneficial bacteria. In in vitro study, raw garlic juice inhibited γ-butyrobetaine (γBB) and trimethylamine (TMA) production by the gut microbiota. Thus, raw garlic juice can potentially prevent cardiovascular disease by decreasing TMAO production through gut microbiota modulation.