Chemically unique environment of endophytes makes them to have various adaptive mechanisms for survival. One of such mechanisms involves the production of pharmacologically significant plant-specific metabolites. In the present study, 26 endophytic fungi were isolated from stem of Bacopa monnieri (L.) Wettst. plants. All the isolates were screened for bacopaside production property by HPLC. Among these, the fungal isolate BmF 16 which was identified as Aspergillus sp. was confirmed for bacopaside N1 production (m/z 796) by LC-MS/MS analysis. As the extract of BMF16 used in the study was prepared from the fifth generation of culture, the obtained result can be confirmed as due to fungal production of bacopaside. In addition, this property was identified only for one among the 26 fungi screened. As bacopaside N1 production in fungi has not yet been reported, the results of the study are novel.