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Nonribosomal lipopeptides (NRLPs) are complex natural products of bacterial origin that not only fulfill important ecological functions but also serve as lead structures for the development of new pharmaceuticals. In order to carry out detailed structure–activity relationship studies and to decipher the biological activities of NRLPs, the primary structure, including stereochemical assignment, of every new member of this natural product family has to be established first. In this review, we want to focus on analytical techniques and tools that can be employed to elucidate the structure of bacterial NRLPs.

Graphical abstract: Structure elucidation of bacterial nonribosomal lipopeptides

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