Clostridioides difficile infection (CDI) is one of the most common nosocomial infections which the prevalence has increased continuously. C. difficile is a spore-forming anaerobic bacteria which able to cause antibiotic-associated diarrhea. Although many antibiotics are used to treat CDI, approximately 15-35% of CDI patients recur within two months. Therefore, discovering new alternative CDI treatments is attractive. Recently, some research found that Enterococcus faecalis probiotics, can be used to treat CDI, but the effect of their healthy metabolic products, postbiotics, has not been investigated until now. Thus, this study aimed to confirm the activity of E. faecalis's postbiotics against toxigenic C. difficile to find a new alternative treatment for CDI. Firstly, cell-free supernatant of two E. faecalis strains, PK 1301 and PK 1801, were lyophilized. The lyophilized supernatant (LCFS) was evaluated Minimum inhibitory concentration (MIC) and observed morphology change of C. difficile after being treated with postbiotics by SEM. Subsequently, we used the postbiotics to evaluate anti-sporulation and anti-spore germination activities on C. difficile. LCFS showed ability in antimicrobial activity against C. difficile 630. MIC of LCFS 1301 was 250 mg/mL. whereas the MIC of LCFS 1801 was 41.67 ± 10.42 mg/mL. The morphology of C. difficile after being treated with LCFSs was shorter than the beginning and observed the damage on C. difficile cells. Interestingly, both postbiotics revealed anti-sporulation and anti-spore germination activities of toxigenic C. difficile dramatically compared to control. In summary, LCFS from E. faecalis significantly inhibit C. difficile growth. Able to inhibit sporulation and spore germination processes of C. difficile. Therefore, both LCFSs might be used as potential postbiotics to prevent and treat CDI.