Antimicrobial Peptides in Frog Skin Secretions

Skin secretions from many species of anurans (frogs and toads) are a rich source of peptides with broad-spectrum antimicrobial activities that may be developed into agents with therapeutic potential, particularly for topical applications. This chapter describes the use of norepinephrine (injection or immersion) to stimulate peptide release from granular glands in the skin in procedures that do not appear to cause distress to the animals. The peptide components in the secretions are separated using reversed-phase HPLC on octadecylsilyl-silica (C₁₈ ) columns after partial purification on Sep-Pak C₁₈ cartridges. Peptides with antimicrobial activity are then identified by demonstration of their abilities to inhibit growth of Gram-negative (Escherichia coli ) and Gram-positive (Staphylococcus aureus) bacteria in liquid phase microtiter plate assays. Individual peptides with activity are purified to near homogeneity by further chromatography on butylsilyl-(C₄ ) and diphenylmethylsilyl-silica columns and characterized structurally by automated Edman degradation and mass spectrometry.