Rimantadine is an adamantane derivative, known for its antiviral activity against infections caused by influenza A viruses. The purpose of the present study was to synthesize 6 new rimantadine derivatives, containing short-chain (Gly, Ala, β-Ala) and bulky (Leu, Ile, Val) amino acids and to investigate their antimicrobial activity against the strains Bacillus subtilis NBIMCC 3562 and Escherichia coli NBIMCC 8785. All derivatives were successfully obtained in good yields by using the TBTU/TEA condensation system. Their antimicrobial properties were established by determining the minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC) against both test strains. The MIC was obtained via a microdilution method, whereas the MBC – via a spread plate method. Most derivatives showed antimicrobial activity, with stronger effects against the Gram-positive strain B. subtilis NBIMCC 3562. Among them, L-Ile-Rim was the most effective derivative against both bacterial strains.

Inflammation is part of the defense mechanism of the organism, which provokes different signaling pathways. It is also related to several inflammatory mediators, such as cytokines and enzymes from the phosphodiesterase group, especially phosphodiesterase 4 (PDE4). On the other hand, peptides play diverse physiological roles as neurotransmitters, neuromodulators, hormones, and enzyme modulators. Taking these facts into account, the anti-inflammatory activity of several C-terminal amide analogs of tetrapeptide Phe-Glu-Leu-Leu was investigated using the carrageenan-induced paw edema model, and their inhibitory activity against PDE4 was evaluated. It was found that Val and Ile substitutions at positions 3 and 4 improved anti-inflammatory activity in vivo, with compounds BB3 and BB4 producing statistically significant reductions in carrageenan-induced edema at the 3 h time point, although these effects were not sustained at later observations. However, the same substitutions did not consistently enhance PDE4 inhibition. The parent compounds BB1 (C-terminal amide) and BB11 (C-terminal free acid) demonstrated the most balanced overall profile, combining moderate PDE4 inhibitory activity (49% and 58% inhibition at a concentration of 5 mM, respectively) with favorable in vivo anti-inflammatory properties. These findings highlight the complex relationship between peptide structure, PDE4 inhibition, and anti-inflammatory efficacy.

Biologically active peptides are a promising alternative of the existing medical drugs used in the medicinal practice. Especially, antimicrobial peptides are such an alternative to conventional antibiotics considering the growing prevalence of antimicrobial resistance. Anoplin, a short decapeptide isolated from the venom of the solitary wasp Anoplius samariensis, has attracted attention due to the simple structure and membrane - targeting feature. This review summarizes current knowledge on the synthesis, structural characteristics, and biological activity of anoplin, with emphasis on structure - activity relationships and modification strategies. The activity of anoplin is closely related to its cationic and amphipathic nature. This allows interaction with bacterial membranes and leads to membrane disruption and cell death. Various modification approaches, including amino acid substitution, lipidation, and structural stabilization, have been applied to improve anoplin’s antimicrobial activity, selectivity, and stability. In addition, environmental factors such as ionic strength, pH, and proteolytic degradation significantly affect biological performance and bioavailability. Although anoplin shows relatively low susceptibility to resistance development, limitations related to stability and cytotoxicity is still a main disadvantage. Therefore, further optimization is required to enhance its therapeutic potential. Anoplin can serve as a useful model for the rational design of new antimicrobial peptides and peptide - based therapeutic agents.

Stable lipid systems with well-preserved physicochemical properties warrant particular attention as promising platforms for the development of drug delivery systems. In this work, two natural temporins A and F were investigated together with a series of analogues modified at position 7 with non-proteinogenic amino acids – citrulline, ornithine, 2,4-diaminobutanoic acid, or 2,3-diaminopropanoic acid – to examine how such substitutions influence peptide-membrane interactions and the overall membrane properties. Fast Fourier-transform electrochemical impedance spectroscopy and thermal shape fluctuation analysis were employed to probe changes in membrane capacitance and bending rigidity in the presence of the studied peptides. Model phosphatidylcholine bilayers were characterized using Fourier-transform infrared and Raman spectroscopy to identify specific interaction sites. Arginine-, citrulline-, and lysine-containing temporins increase membrane bending rigidity through distinct modes of membrane interfacial reorganization. A similar decrease in membrane capacitance was accompanied by peptide-specific FTIR spectroscopic signatures for the arginine-, diaminopropanoic acid-, and ornithine-containing analogues, suggesting different modes of membrane interfacial reorganization. The spectral changes are consistent with enhanced interfacial hydrogen-bonding and electrostatic interactions for the arginine analogue, whereas the remaining temporin analogues appear to perturb the membrane interface primarily through partial dehydration of the lipid carbonyl region. The reported findings advance the molecular-level understanding of the modulation of the physicochemical properties of lipid membranes by peptide sequence variations, with potential relevance to applications in biosensing and drug delivery.

Antimicrobial peptides (AMPs) have emerged as a promising therapeutic scaffold for combating antibiotic-resistant infections and cancer. Aurein 1.2, the most studied peptide in the aurein family, displays a broad spectrum of bi-functionality against bacterial and cancer cells, making it one of the smallest active amphibian peptides reported to date. The phenylalanine residues at positions 3 and 13 are critical for biological activity because they serve as essential anchors for membrane binding. In this study, a series of new analogues of aurein 1.2 containing fluorinated phenylalanine were synthesized by solid-phase peptide synthesis (SPPS), Fmoc-(9-fluorenylmethoxycarbonyl) chemistry, purified by High Performance Liquid Chromatography (HPLC), analysed by mass spectrometry (MS). The antiproliferative activity was determined in normal and tumour cell lines (MCF-12F, MCF-7, MDA-MB-231, U87 and MG63 cells) by MTT dye reduction assay. The target compounds exhibited different concentration-dependent antiproliferative effect against the tumour cell lines after 72 h treatment. The highest antiproliferative activity was observed with the peptide EH [Phe (4-F)]3,13[Lys]4 (IC50 = 9.79 ± 0.96 µM) in U87 cells and (IC50 = 9.96 ± 0.8 µM) in MG63 cells. Significant selectivity (SI > 2) was observed for the peptide analogue EH [Phe (4-F)]3 in MCF-7 cells (SI = 2.08). Antimicrobial activity was assessed against Gram-positive and Gram-negative bacteria. The aurein 1.2 and its analogues possessed better antibacterial activity against Gram-positive bacteria than Gram-negative bacteria. The obtained results reveal that replacement of Phe with Phe(4-F) and Asp with Lys can increase both the potency and activity. The best results were obtained with the peptide EH [Phe(4-F)]3,13[Lys]4 (MIC = 10 µg/ml), against S. aureus 3703. However, compared to the antibacterial data of the positive controls, the results showed weaker antibacterial activity.

Atopic dermatitis is a chronic inflammatory skin disease with significant health and social importance. Unfortunately, currently there is no well-defined effective treatment. Thus, the search for new therapeutic approaches is currently ongoing. This study presents the clinical impact on skin inflammation induced by 1-chloro-2,4-dinitrobenzene in a rat model, after local treatment with the peptide (KLAKLAK)2-NH2 (Si1) and a shortened analogue containing non-proteinogenic amino acid β-Ala conjugated with 1,8-naphthalimide (Npht) as second pharmacophore (NphtG-KLβAKLβAK-NH2 (Si2)). The changes in skin lesions were estimated along with ear thickness determination, and plasma IL4 and IL13 levels evaluation. The results showed attenuation of skin lesions by NphtG-KLβAKLβAK-NH2 (Si2), accompanied by respective changes in ear thickness, and IL plasma levels. The beneficial effect of the topically applied specifically designed bioconjugate could be attributed to attenuation of skin inflammation due to the pro-apoptotic effect of KLAKLAK moiety on M2 macrophages engaged in the pathological process.

Background/Objectives: With growing antimicrobial resistance, the overuse of antibiotics, and stagnation in the discovery of new antibiotics, a novel alternative is required to overcome hard-to-treat infections. Antimicrobial peptides (AMPs) show great potential as a possible alternative to standard chemotherapeutics. Temporins are a group of AMPs that have been under the spotlight in numerous studies. Herein, we report the design and synthesis of Temporin A modified in position 1, where the proteinogenic amino acid Phe is replaced by Tyr or fluorinated Phe. In addition, in other analogues, in position 10, the Ser residue is replaced by Tyr or Thr. The aim of all modifications in the primary structure of the native Temporin A is to study the influence of the changes made on the antibacterial properties, antiproliferative activity, and hydrolytic stability of the newly synthesized molecules. Methods: The Fmoc/OBut SPPS strategy was employed for the synthesis of the novel-designed analogues. The antibacterial activity was evaluated with both disk diffusion and broth microdilution methods. The BALB 3T3 NRU test and MTT dye reduction assay were used to determine safety and antiproliferative activity. Results: The investigated analogues have low toxicity and are photosafe. The greatest selectivity was shown by DTTyr10 towards MCF-7 cells. DT4F, containing fluorinated Phe in position 1, was the most effective antibacterial agent among the new compounds. The incorporation of Thr in position 10, in comparison with the natural Ser residue, led to an increase in the antiproliferative effect of the new peptide. Conclusions: The obtained structure–activity relationship data show that the most promising compound in the tested series is FLPLIGRVL-Y-GILNH2, where the Ser residue in position 10 is replaced by a more hydrophobic OH-containing Tyr residue. The analogue containing fluorinated Phe in position 1, DT4F, has the highest antiproliferative effect against both tested tumor cell lines, combined with good antibacterial properties at the lowest MIC (80 µg/mL), but it is more cyto- and phototoxic than the parent DTA molecule and is not stable at pH 9 for a 24 h period.

The high prevalence of pain affecting millions of people worldwide made it a major health problem. At the same time often, inflammation is closely associated to the pain. Thus, creation of molecules with a double effect is a good alternative to the currently existing in the medicinal practice non-steroidal medications. Herein, some modifications in the N- and C-terminus of the tetrapeptide FELL with proven anti-inflammatory properties are performed and the newly synthesized compounds are tested for both analgesic and anti-inflammatory potential. The analgesic and anti-inflammatory activity of the newly synthesized molecules was investigated using Paw-pressure and Carrageenan-Induced Paw Edema tests, respectively, on experimental animals. The results showed a certain “discrepancy” between the analgesic and the anti-inflammatory effects of the newly synthesized substances. Newly synthesized BB9 and BB15, L-Tyr containing C-terminal amide and free acid, respectively, exhibit a more significant analgesic activity compared to those of parent molecules BB1 and BB11, containing L-Phe in the N-terminus. Moreover, taking into account the obtained results for the compounds BB10 and BB16, the preferable substitution is L-Tyr, instead of D-Tyr. It could be concluded that the aromatic OH-function is probably important for the connection with the opioid and cannabinoid receptors. However, the designed structural modifications did not lead to improvement in anti-inflammatory effect compared to those of parent molecules. Moreover, in a context of the positive finding is that all modifications done save the hydrolytic stability of the molecules.

Oxidative stress and metal-driven redox processes are key contributors to the pathogenesis of chronic diseases and cancer, motivating the search for novel antioxidant molecules. In this study, the antioxidant potential of a series of synthetic peptides previously reported to possess antitumor and antibacterial properties was evaluated using two complementary electron-transfer assays: ferric reducing antioxidant power (FRAP) and cupric ion reducing antioxidant capacity (CUPRAC). Both assays were calibrated against caffeic acid, and results were expressed as caffeic acid equivalents (CAE). The FRAP assay revealed substantial differences in reducing activity, with Si8 exhibiting the highest value (0.558 ± 0.132), followed by Si12 (0.478 ± 0.0240), Si10 (0.293 ± 0.0220), and Si15 (0.250 ± 0.0200), whereas Si1 (0.00439 ± 0.00240) and Si11 (0.00260 ± 0.000500) showed negligible responses. A comparable pattern was observed in the CUPRAC assay, where Si8 again displayed the strongest reducing capacity (0.381 ± 0.0948), with Si12 (0.290 ± 0.0225), Si15 (0.262 ± 0.0223), and Si10 (0.224 ± 0.0290) also demonstrating appreciable activity, while Si1 (0.001800 ± 0.000400) and Si11 (0.0132 ± 0.000500) remained inactive. The combined application of FRAP and CUPRAC provided complementary and reproducible measures of peptide antioxidant capacity, establishing a framework for systematic characterization of redox-active peptides in relation to oxidative stress.

Extensive use of classical antibiotics has led to the growing emergence of many resistant strains of pathogenic bacteria. To combat this challenge, researchers have turned to the antimicrobial peptides (AMPs). Aurein 1.2 (GLFDIIKKIAESF-NH2) was demonstrated to have broad spectrum bi-functionality against bacterial and cancer cells. The Solid Phase Peptide Synthesis (Fmoc-strategy) was used for the synthesis of new analogs of aurein 1.2. The purity of all compounds was monitored by HPLC, and their structures were proven using mass spectrometry. Cytotoxicity and antiproliferative effects were studied using 3T3 NRU and MTT tests, respectively. The antibacterial activity was estimated against Gram-positive and Gram-negative bacteria using broth microdilution method in concentrations from 0 to 320 µg/mL to determine the minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC). The antiproliferative activity test shows that the peptide analog EH [Orn]8 has the highest activity (IC50 = 44 ± 38 μM) for the three cell lines studied (MCF-12F, MCF-7, and MDA-MB-231). The same compound exhibited good antimicrobial activity. The obtained results reveal that replacement of Lys with non-proteinogenic amino acids can increase both the potency and activity spectra of natural template peptides, making them suitable candidates for new drug development.

The adaptation of the body when exposed to a lower-than-usual temperature is a challenge that involves neuro-endocrine-immune mechanisms and affects the pharmacokinetics and/or pharmacodynamics of drugs taken before or after cold exposure. The experiments presented in this study clearly show differences in the analgesic effect of an exogenously introduced model substance (C-terminal fragment of calcium-binding protein, spermatid-specific 1) before and after cold exposure compared to its effect at an ambient temperature. The model substance used for the experiments is an octapeptide, TDIFELLK, which was synthesized via standard solid-phase peptide synthesis. Preliminary studies proved TDIFELLK's analgesic activity. The ANOVA analysis performed showed statistically significant differences in the pain thresholds, measured by a paw pressure test, in 109 rats distributed among 14 groups and subjected to cold exposure according to different set-ups. Cold exposure immediately after TDIFELLK administration appears to enhance its analgesic effect, while cold exposure before administration reduces the effect. In some of the set-ups, antagonists of the most significant for analgesia receptors, i.e., opioid, cannabinoid, and serotonergic, were also introduced. The results showed that cold exposure had a modulating influence on the effect of the exogenously administered substances. The modulating effect was manifested differently depending on whether the intake occurred before or after cold exposure. The results also showed that the interaction with individual mediator systems was also subjected to differences depending on intake occurring before and after cold exposure.

Aim: The aim of this study was to compare the lesion depth and level of penetration of CPP-ACFP, SAP P11-4 and 2 newly synthesized self-assembling peptides by measuring the linear depth of fluorescence.Materials and methods: A total of 30 enamel samples were prepared from orthodontically extracted permanent premolars. Three zones were provided: a sound enamel surface (SC), a demineralization zone (DC), and a remineralization zone (TA). The samples were randomly divided into five groups (n=6): group 1 – Gr V (MI Varnish™), group 2 – Gr CR (Curodont™ Repair), group 3 – IU1, group 4 – IU2, and group 5 – Gr NT (control group, no treatment). Lesion depth was assessed by measuring the linear penetration depth of the fluorescent dye by laser confocal microscopy.Results: A statistically significant reduction in the depth of the lesion was found in the experimental groups compared to the control group, which was not treated (p<0.001). The depth of the lesion reveals the degree of mineralization and the cariostatic effect in the experimental groups. When measuring the confocal images, the results showed that there was a similar degree of remineralization for CPP-ACFP and for SAP P11-4, IU1, and IU2 for the two time points DC and TA, but without statistical significance between them (p=1.00). The samples treated with IU2 showed the highest degree of remineralization, followed by CPP-ACFP, SAP P11-4 and IU1.Conclusion: The self-assembling peptides have a significant level of infiltration and potential for remineralization of artificial carious lesions in permanent teeth. The combination of SAP with fluoride has an additive effect.

Many microorganisms pose a threat to human health due to the ever-increasing bacterial resistance to conventional drugs. Nowadays, searching for new alternatives to conventional antibiotics to fight bacterial resistance is a main task. Thus, natural molecules such as amino acids and peptides arise as possible solutions to the problem. The antimicrobial activity of targeted compounds was studied by the agar-diffusion method, using the prepared working solutions of the targeted peptides with the corresponding concentrations. The results of the antimicrobial activity against different test pathogens show specificity, as antimicrobial activity against the used test microorganisms was not found in the investigated short-chain synthetic peptides Si6, Si3 and Si13. Antimicrobial activity against Bacillus cereus, Staphylococcus aureus, Staphylococcus epidermidis, Propionibacterium acnes, Escherichia coli, Pseudomonas aeruginosa, and the yeasts Malassezia furfur and Candida albicans was established for the long-chain synthetic peptides Si1, Si5 and Si16, except Si5 which does not show activity against pathogenic fungal strain C. albicans. The compound Si16 where natural Leu in (KLAKLAK)2-NH2 is replaced by unnatural Nle is the best candidate for medical drug due to the combined antibacterial and antiproliferative effect as well as long hydrolytic stability.

Atopic dermatitis (AD) is a chronic inflammatory skin condition of significant health and social importance, which justifies the search for new means of treatment. Since the endogenous cannabinoid system appears to be involved in the pathogenesis of AD, the proposed article summarizes the clinical impact on skin inflammation in a rat model of 1-chloro-2,4-dinitrobenzene-induced atopic dermatitis-like condition after exogenous systemic administration of the cannabinoid receptor type 1 (CB1r) agonist anandamide, as well as after local treatment with a newly synthesized pyrrole moiety containing bioconjugate of FELL tetrapeptide with CB1r-dependent analgesic activity. The changes in skin lesions and ear thickness were estimated along with the CB1r expression immunohistochemically determined on skin punch biopsies. The results showed attenuation of skin lesions by anandamide and lack of positive effect after introduction of CB1r antagonist, accompanied by a change in CB1r expression, suggesting the involvement of the cannabinoid system in the defensive functions of the skin. The topically applied newly synthesized bioconjugate also favorably affected skin manifestations of inflammation, but without a change in CB1r expression, suggesting the involvement of other mechanisms in the reported effects.

Background: Specifically designed peptide mimetics offer higher selectivity regarding their toxicity to mammalian cells. In addition to the α-helix conformation, the specific activity is related to the peptide’s ability to penetrate the cell membrane. The alterations in lipid membrane properties were addressed in the presence of the peptide KLAKLAK-NH2 and analogs containing β-alanine, strengthening the antibacterial activity and/or naphtalimide with proven anticancer properties. Methods: The molecular interactions of the peptide mimetics with POPC bilayers were studied using FTIR-ATR spectroscopy. The thermal shape fluctuation analysis of quasispherical unilamellar vesicles was applied to probe the membrane bending elasticity. The impedance characteristics of bilayer lipid membranes were measured using fast Fourier-transform electrochemical impedance spectroscopy. Results: A lateral peptide association with the membrane is reported for β-alanine-containing peptides. The most pronounced membrane softening is found for the NphtG-KLβAKLβAK-NH2 analog containing both active groups that corroborate with the indications for 1,8-naphthalimide penetration in the lipid hydrophobic area obtained from the FTIR-ATR spectra analysis. The β-alanine substitution induces strong membrane-rigidifying properties even at very low concentrations of both β-alanine-containing peptides. Conclusions: The reported results are expected to advance the progress in tailoring the pharmacokinetic properties of antimicrobial peptides with strengthened stability towards enzymatic degradation. The investigation of the nonspecific interactions of peptides with model lipid membranes is featured as a useful tool to assess the antitumor and antimicrobial potential of new peptide mimetics.