The control of phytopathogenic fungi in agricultural crops requires the use of synthetic chemical fungicides, which have damaged the environment for decades. Biocontrol with microorganisms is one option to reduce their use, with the fungi of the Trichoderma genus standing out for their ability to interact with soil pathogens through different control mechanisms through antibiosis or production of substances harmful to other microorganisms. The objective of this work was to evaluate the biological control mechanism using Trichoderma asperellum antibiotics on the growth of Fusarium oxysporum and F. equiseti. Antibiosis bioassays were performed using the cellophane test (diffusible metabolite assay), the reverse plate technique (volatility assay), and poisoned foods (T. asperellum mycelium extracts and extracellular metabolite assays). The diffusible metabolites of T. asperellum presented the greatest inhibition of growth. The highest percentage of inhibition was observed on F. oxysporum in plates where T. asperellum developed for 72 h (>25 %), while F. equiseti inhibition was more effective in plates with 48 h (>40 %). In both species, no significant inhibitory effect was observed in volatility tests (>10 %), while extracellular metabolites showed no inhibition. In contrast, metabolites extracted from T. asperellum mycelium with ethyl acetate inhibited Fusarium between 18 and 40 %; with hexane, between 9 and 20 %; and with methanol, no inhibition was observed. The direct analysis in real-time mass spectrometry (DART-MS) analysis showed the presence of pyrones, fatty acids, alcohols, and carbohydrates in extracts and liquid culture of T. asperellum, which suggests that the control mechanism through antibiotics on F. oxysporum and F. equiseti is fungistatic.