Endophytic fungi of different species have already proved their efficacy against a range of pathogenic bacteria. However, the separation of the compounds contained in these extracts that actually have an effective activity is not a constant, but it is essential for the scientific knowledge to formulate a new medication. Thus, this study aimed to separate compounds from two endophytic fungi extracts of Uncaria tomentosa with positive results against Gram-positive and Gram-negative bacteria. The separation was carried out by thin-layer chromatography and bioautography. For this, it was necessary to produce fungal extracts from the endophytes Colletotrichum (23916) and Fusarium (23952), which were separated from the liquid state by filtration. The remaining filtrate was divided into ethyl acetate and then concentrated in a rotary evaporator at 40°C. Antimicrobial activity was evaluated and the Minimum Inhibitory Concentration (MIC) was determined with the following test microorganisms: Escherichia coli, Enterecoccus faecalis, Klebsiella pneumoniae and Staphylococcus aureus. Subsequently, it was carried out the thin layer chromatography (TLC) of the extracts using hexane, ethyl acetate/hexane, ethyl acetate and acetate/methanol. The reading was performed in 312 nm UV light, and right after the chromatography, the bioautography was carried out. The following Rf values were obtained: 0.30 for K. pneumoniae and 0.35 for E. coli, in ethyl acetate, for the extract 23952, and 0.81 for K. pneumoniae and 0.41 for S. aureus, in styrene acetate, for the extract 23916, confirming different compounds by bioautography. Therefore, it was concluded that several compounds present in the extracts tested have antibacterial activity.