Abstract: A growing number of strains of the filamentous soil fungus Trichoderma arecommercially available as plant strengtheners, plant growth promoters or as biocontrol agents(BCA) of phytopathogens. When interacting with the pathogenic fungus and the plant,Trichoderma spp. produce secondary metabolites, such as peptaibiotics, which are believed toplay a key role in the crosstalk between the respective organisms. Peptaibiotics are linear nonribosomalpeptides with antibiotic properties, which contain the non-proteinogenic amino acid a-aminoisobutyric acid (Aib). In this study, we screened Trichoderma strains present in commercialBCA products from five suppliers for their ability to produce peptaibiotics, when grown in liquidculture medium. Using an analytical workflow consisting of liquid chromatography–massspectrometry (LC-MS) and subsequent LC–tandem MS (LC-MS/MS) experiments, we were ableto detect 12 peptaibols of the group of trichotoxins, two members of the trichorzin HA group and11 novel peptaibols in cultures of three of the investigated BCA strains.