Peptaibols are linear α-aminoisobutyric acid-containing peptide antibiotics originating from soil fungi mainly of the genus Trichoderma and biosynthesized in complex mixtures of closely related analogues by a polyenzymatic pathway. Addition of amino acids such as α-aminoisobutyric acid (Aib), glutamic acid or arginine, to the fermentation medium of two Trichoderma strains, T. harzianum and T. longibrachiatum, has been shown to result in the simplification of the natural peptaibol mixtures, leading in each case to the almost exclusive biosynthesis of a single peptide. Surprisingly, the obtained peptides are Aib-enriched, whether the added amino acid is Aib, Glu or Arg. By adding Aib to the fermentation medium of T. harzianum, two new Aib-rich peptaibols were isolated. Moreover, adding glutamic acid to the culture medium of T. longibrachiatum, which produces both neutral and acidic 20-residue peptaibols with either glutamine or glutamic acid at position 18, increases the production of the acidic peptides. However, arginine which is a positively charged amino acid generally absent from peptaibol sequences, is not incorporated in trichorzins when added to the fermentation medium of T. harzianum.