In Vitro Evaluation of Cytotoxicity and Permeation Study on Lysine- and Arginine-Based Lipopeptides with Proven Antimicrobial Activity

Owing to their excellent antimicrobial activities with a relatively low cost of production, lipopeptides are being intensively investigated as potential alternatives to popular antimicrobials. However, a critical obstacle for their application is a relatively high toxicity, hence a lot of attention has been paid to designing new molecules with optimal properties. In this study we synthesized the following lipopeptides: C(16)-KK-NH(2), C(16)-KεK-NH(2), C(16)-KKK-NH(2), C(16)-KRK-NH(2), C(16)-RR-NH(2), C(16)-RRR-NH(2), (C(10))(2)-KKKK-NH(2) and (C(12))(2)-KKKK-NH(2). Their antimicrobial activity against representative strains of Gram-positive bacteria, Gram-negative bacteria and fungi has been confirmed. The compounds have been evaluated with regard to the safety of their application in dermatology. The cytotoxicity was determined in HaCaT keratinocytes using MTT assay, whereas Strat M membranes placed in Franz diffusion cells were used to assess their ability to skin permeation. The compounds containing one hexadecanoic acid chain turned out to be very toxic towards human keratinocytes, while lipopeptides containing two fatty acid chains (decanoic and dodecanoic) demonstrated much lower cytotoxicity. For the most promising lipopeptide, (C(10))(2)-KKKK-NH(2), the measured IC(50) on HaCaT keratinocytes was few times higher as compared to MICs obtained for the tested bacteria. Both groups of lipopeptides did not permeate the model membranes and therefore lack of permeation through human skin could be expected. The results of this work encourage further research on the potential application of lipopeptides with two fatty acids as novel antimicrobials.