Trimethylamine N-oxide (TMAO), a compound generated by the gut microbiota, may be associated with acute pancreatitis (AP). Nonetheless, the exact crosstalk mechanism remains unidentified. This study used Mendelian randomization, bioinformatics methods, and in vitro analyses to assess the causal relationship among eating behavior, TMAO, obesity, and AP. Our findings indicate that high-calorie meals elevate circulating TMAO levels, which is significantly associated with obesity risk. An in-depth examination of the GSE24883 and GSE194331 datasets indicated that TMAO-induced macrophage inflammation might be crucial in the obesity–AP relationship. In vitro studies indicate that TMAO may induce macrophage pyroptosis. Our results suggest that high-calorie meals significantly elevate circulating TMAO levels and that TMAO-induced macrophage pyroptosis affects the causal relationship between obesity and AP.