Methicillin-resistant Staphylococcus aureus (MRSA) is a notorious pathogen with biofilm-forming and drug-resistant properties that make it difficult to eradicate. In this study, the inhibition of MRSA (ATCC 43300) by Starmerella bacillaris CC-PT4 (CGMCC No. 23573) was evaluated. The results showed that the inhibition of MRSA growth and biofilm was caused by S. bacillaris CC-PT4 cell-free supernatant (CFS). The CFS of S. bacillaris CC PT4 at different times can effectively inhibit the formation of MRSA biofilm, remove the preformed biofilm, and down-regulate the related genes that promote the formation of biofilm. Afterwards, untargeted metabolomics was performed to analyze the CFS of S. bacillaris CC-PT4. Several molecules with antibacterial and inhibitory biofilm effects from the CFS were found, one of which, 2-amino-1-phenylethanol (APE), has not been reported to have antiMRSA ability before. In this study, molecular docking analysis and in vitro experiments were used to verify the function of APE to inhibit MRSA. These results indicate that S. bacillaris CC-PT4 CFS can effectively inhibit MRSA which has potential application value in controlling MRSA.