The study aimed to evaluate the endophytic fungi isolated from Cocos nucifera (L.) cotyledon for their possible antimicrobial activity. A total of twelve endophytic fungal species were successfully isolated, from which five were selected and presumptively identified, including Aspergillus niger, Raffaelea sp., Aspergillus flavus, Cladosporium sp. and Pseudallescheria spp. based on their macro and microscopic characters. The agar blocks of actively growing pure culture were assessed for antibacterial activity against five human pathogenic bacterial and fungal strains: Escherichia coli (BIOTECH 1634), Bacillus subtilis (BIOTECH 1697), Staphylococcus aureus (BIOTECH 1592), Serratia marcescens (BIOTECH 1748), and Candida tropicalis (BIOTECH 2085). Among the selected five endophytic used for antimicrobial activity screening, the Raffaelea sp. (CN-EF 2) has shown a high zone of inhibition of 11 mm against Bacillus subtilis, 18 mm against Escherichia coli, 15 mm against Staphylococcus aureus, and 11 mm against Candida tropicalis. Correspondingly, Aspergillus niger (CN-EF 1) has shown a zone of inhibition of 16 mm against C. tropicalis. The endophytic fungus Raffaelea sp. was effective against all test organisms except for Serratia marcescens, while Aspergillus niger was only effective for pathogenic fungi C. albicans. Hence, further in-depth studies of the isolated endophytic fungi are needed to determine the mechanisms behind the endophytes' inhibitory effects.