As an important limiting factor, lignin hinders the utilization rate of maize straw in ruminants. CaO treatment increases the feed digestibility of maize straw by disrupting the ester bonds between hemicellulose, cellulose, and lignin in maize straw. Our previous research found that CaO treatment of corn straw may increase its feed digestibility by altering the rumen microbes' abundance. This study further investigated the molecular mechanism of CaO treatment to enhance feed utilization and also examined its ongoing effects on rumen metabolites. Rumen fluid was collected to analyze microbial metabolites using liquid chromatography-mass spectrometry (LC-MS) non-targeted metabolomics. Maize straw (moisture content of 60%) treated with four levels of CaO (0%, 3%, 5%, and 7%) was used as the fermentation substrate for a 6 h