Enniatins are promising antimicrobial and anticancer agents based on their ionophoric, enzyme inhibiting and mitochondriotoxic properties. Using the combination of SEIRA spectroscopy and tBLMs assisted by DFT-based computational spectroscopy, we have characterized the cation-dependent behaviour of EBn:M z+ -complexes modulating the incorporation and interaction with the lipid membrane, in which they can act as ionophores. Overall, we find (as summarized in Figure ) -that the monovalent K + and Na + promote EB-membrane incorporation with distinctly different kinetics. In presence of K + , a direct incorporation is observed, which we associate with the a dominating 2:1 EB2:K + sandwich structure. -Instead in presence of Na + , a sigmoidal shaped kinetics of incorporation occurs. This can be rationalized in terms of an positively cooperative/autocatalytic process that is mediated via a mixture of distorted 2:1 EB2:Na + sandwich structures and 1:1 open-faced EB:Na + complexes.