(1) Background: Antimicrobial resistance is growing at an extreme pace and has proven to be an urgent topic, for research into alternative treatments. Such a prospective possibility is hidden in antimicrobial peptides because of their low to no toxicity, effectiveness at low concentrations, and most importantly their ability to be used for multiple treatments. This work was focused on the study of the effect of the modification in position 7 of Temporin A on its biological activity; (2) Methods: The targeted peptides were synthesized using Fmoc/Ot-Bu SPPS. The antibacterial activity of the analogs was determined using the broth microdilution method and disk-diffusion method. In vitro tests were performed to determine the cytotoxicity, phototoxicity, and antiproliferative activity of the peptide analogs on a panel of tumor and normal cell lines; (3) Results: All analogs except DTCit showed good antibacterial activity, with DTDab having the best activity according to the disk-diffusion method. However, DTCit had an acceptable cytotoxicity, combined with good selectivity against the test MCF-7 cell line; (4) Conclusions: The obtained results revealed the importance of the basicity and length of the side chain at position 7 in the Temporin A sequence for both tested activities.

In view of an appreciable increase in resistance of Staphylococcus aureus to the conventional antibiotics, it is desired to develop new effective drugs. Antimicrobial peptides (AMPs) seem to be attractive candidates. In general, AMPs samples used for in vitro studies consist of a peptide, counter-ion, and water. The presence of the counter-ion could be significant as it affects peptide secondary structure and biological activity. The purpose of this study was to estimate the impact of counter-ion on antistaphylococcal activity of selected AMPs (CAMEL, citropin 1.1, LL-37, pexiganan, temporin A). To do this, three kinds of salts were prepared, namely, acetates, hydrochlorides, and trifluoroacetates. In addition, the hemolytic activity against human red blood cells (hRBCs) and cytotoxicity (HaCaT) were determined. The results indicate that there is a substantial difference between different salts, but the pattern is not consistent for the peptides. In general, the antistaphylococcal activity decreased in the order: CAMEL > temporin A > pexiganan > citropin 1.1 ≫ LL-37. The highest selectivity indexes were determined for CAMEL hydrochloride, pexiganan acetate, and temporin A trifluoroacetate. This study shows how important is to take into account the kind of counter-ions when designing novel peptide-based antimicrobials.