Permeabilization Properties Research Articles

LL37-Derived Fragments Improve the Antibacterial Potential of Penicillin G and Ampicillin against Methicillin-Resistant Staphylococcus aureus.

Methicillin-resistant Staphylococcus aureus (MRSA) infections are a severe threat to public health. Antimicrobial peptides (AMPs) are novel and potential antimicrobials with specific antibacterial mechanisms. Our aim was to study the potential of LL37, FK16, and FK13 to enhance the anti-MRSA activity of antibiotics in vitro, particularly penicillin G and ampicillin. Our results showed that FK16 and FK13 have more synergistic inhibitory effects to MRSA strains when combined with penicillin G and ampicillin. In addition, AMPs exhibited strong membrane permeabilizing properties, and membrane permeabilizing effects can provide a possible explanation for the improved antibacterial effects of antibiotics, since permeabilizing AMPs have the potential to increase the access of antibiotics. To further study the electrostatic interactions among cationic AMPs with negatively charged bacteria, we measured the zeta potentials of three MRSA strains and also neutralized three MRSA strains with the addition of cationic AMPs. Further, we demonstrated the connection between membrane permeabilization and zeta potential neutralization. Finally, we treated MRSA strains with AMPs and characterized the MICs of penicillin G and ampicillin. FK16 was the most promising AMP among the three AMPs, since exposure to FK16 decreased the MICs of both penicillin G and ampicillin for all MRSA strains and also demonstrated more synergistic combinations when combined with antibiotics. AMP exposure and subsequent membrane permeabilization provide a possible pathway to re-sensitize drug-resistant bacteria to traditional antibiotics. Re-sensitization may help preserve the effectiveness of traditional antibiotics, thus providing a potential new strategy for fighting MRSA infections.

Acute Toxicity of the Dinoflagellate Amphidinium carterae on Early Life Stages of Zebrafish (Danio rerio).

Dinoflagellates of the genus Amphidinium can produce a variety of polyketides, such as amphidinols (AMs), amphidinoketides, and amphidinin, that have hemolytic, cytotoxic, and fish mortality properties. AMs pose a significant threat to ecological function due to their membrane-disrupting and permeabilizing properties, as well as their hydrophobicity. Our research aims to investigate the disparate distribution of AMs between intracellular and extracellular environments, as well as the threat that AMs pose to aquatic organisms. As a result, AMs containing sulphate groups such as AM19 with lower bioactivity comprised the majority of A. carterae strain GY-H35, while AMs without sulphate groups such as AM18 with higher bioactivity displayed a higher proportion and hemolytic activity in the extracellular environment, suggesting that AMs may serve as allelochemicals. When the concentration of extracellular crude extracts of AMs reached 0.81 µg/mL in the solution, significant differences in zebrafish embryonic mortality and malformation were observed. Over 96 hpf, 0.25 μL/mL of AMs could cause significant pericardial edema, heart rate decrease, pectoral fin deformation, and spinal deformation in zebrafish larvae. Our findings emphasized the necessity of conducting systematic research on the differences between the intracellular and extracellular distribution of toxins to gain a more accurate understanding of their effects on humans and the environment.

Guanidinylated Polymyxins as Outer Membrane Permeabilizers Capable of Potentiating Rifampicin, Erythromycin, Ceftazidime and Aztreonam against Gram-Negative Bacteria.

Polymyxins are considered a last-line treatment against infections caused by multidrug-resistant (MDR) Gram-negative bacteria. In addition to their use as a potent antibiotic, polymyxins have also been utilized as outer membrane (OM) permeabilizers, capable of augmenting the activity of a partner antibiotic. Several polymyxin derivatives have been developed accordingly, with the objective of mitigating associated nephrotoxicity. The conversion of polymyxins to guanidinylated derivatives, whereby the L-γ-diaminobutyric acid (Dab) amines are substituted with guanidines, are described herein. The resulting guanidinylated colistin and polymyxin B (PMB) exhibited reduced antibacterial activity but preserved OM permeabilizing properties that allowed potentiation of several antibiotic classes. Rifampicin, erythromycin, ceftazidime and aztreonam were particularly potentiated against clinically relevant MDR Gram-negative bacteria. The potentiating effects of guanidinylated polymyxins with ceftazidime or aztreonam were further enhanced by adding the β-lactamase inhibitor avibactam.

Cationic polymer contributes to broaden the spectrum of vancomycin activity achieving eradication of Pseudomonas aeruginosa.

Vancomycin (VAN) is unable to penetrate the outer membrane of Gram-negative bacteria and reach the target site. One approach to overcome this limitation is to associate it with compounds with permeabilizing or antimicrobial properties. EudragitE100® (Eu) is a cationic polymer insufficiently characterized for its potential antimicrobial action. Eu-VAN combinations were characterized, the antimicrobial efficacy against Pseudomonas aeruginosa was evaluated and previous studies on the effects of Eu on bacterial envelopes were extended. Time-kill assays showed eradication of P. aeruginosa within 3-6h exposure to Eu-VAN, whilst VAN was ineffective. Eu showed regrowth in 24h and delayed colony pigmentation. Although permeabilization of bacterial envelopes or morphological alterations observed by TEM and flow cytometry after exposure to Eu were insufficient to cause bacterial death, they allowed access of VAN to the target site, since Eu-VAN/Van-FL-treated cultures showed fluorescent staining in all bacterial cells, indicating Van-FL internalization. Consequently, Eu potentiated the activity of an otherwise inactive antibiotic against P. aeruginosa. Moreover, Eu-VAN combinations exhibited improved physicochemical properties and could be used in the development of therapeutic alternatives in the treatment of bacterial keratitis.

In Vitro Anti-Candida Activity and Action Mode of Benzoxazole Derivatives.

A newly synthetized series of N-phenacyl derivatives of 2-mercaptobenzoxazole, including analogues of 5-bromo- and 5,7-dibromobenzoxazole, were screened against Candida strains and the action mechanism was evaluated. 2-(1,3-benzoxazol-2-ylsulfanyl)-1-(4-bromophenyl)ethanone (5d), 2-(1,3-benzoxazol-2-ylsulfanyl)-1-(2,3,4-trichloro-phenyl)ethanone (5i), 2-(1,3-benzoxazol-2-ylsulfanyl)-1-(2,4,6-trichlorophenyl)ethanone (5k) and 2-[(5-bromo-1,3-benzoxazol-2-yl)sulfanyl]-1-phenylethanone (6a) showed anti-C. albicans SC5314 activity, where 5d displayed MICT = 16 µg/mL (%R = 100) and a weak anti-proliferative activity against the clinical strains: C. albicans resistant to azoles (Itr and Flu) and C. glabrata. Derivatives 5k and 6a displayed MICP = 16 µg/mL and %R = 64.2 ± 10.6, %R = 88.0 ± 9.7, respectively, against the C. albicans isolate. Derivative 5i was the most active against C. glabrata (%R = 53.0 ± 3.5 at 16 µg/mL). Benzoxazoles displayed no MIC against C. glabrata. Benzoxazoles showed a pleiotropic action mode: (1) the total sterols content was perturbed; (2) 2-(1,3-benzoxazol-2-ylsulfanyl)-1-(3,4-dichlorophenyl)ethanol and 2-(1,3-benzoxazol-2-ylsulfanyl)-1-(2,3,4-trichlorophenyl)ethanol (8h–i) at the lowest fungistatic conc. inhibited the efflux of the Rho123 tracker during the membrane transport process; (3) mitochondrial respiration was affected/inhibited by the benzoxazoles: 2-(1,3-benzoxazol-2-ylsulfanyl)-1-(4-chlorophenyl)ethanol and 2-(1,3-benzoxazol-2-ylsulfanyl)-1-(4-bromophenyl)ethanol 8c–d and 8i. Benzoxazoles showed comparable activity to commercially available azoles due to (1) the interaction with exogenous ergosterol, (2) endogenous ergosterol synthesis blocking as well as (3) membrane permeabilizing properties typical of AmB. Benzoxazoles display a broad spectrum of anti-Candida activity and action mode towards the membrane without cross-resistance with AmB; furthermore, they are safe to mammals.

Effect of Disulfide Cyclization of Ultrashort Cationic Lipopeptides on Antimicrobial Activity and Cytotoxicity.

Ultrashort cationic lipopeptides (USCLs) are considered to be a promising class of antimicrobials with high activity against a broad-spectrum of microorganisms. However, the majority of these compounds are characterized by significant toxicity toward human cells, which hinders their potential application. To overcome those limitations, several approaches have been advanced. One of these is disulfide cyclization that has been shown to improve drug-like characteristics of peptides. In this article the effect of disulfide cyclization of the polar head of N-palmitoylated USCLs on in vitro biological activity has been studied. Lipopeptides used in this study consisted of three or four basic amino acids (lysine and arginine) and cystine in a cyclic peptide. In general, disulfide cyclization of the lipopeptides resulted in peptides with reduced cytotoxicity. Disulfide-cyclized USCLs exhibited improved selectivity between Candida sp., Gram-positive strains and normal cells in contrast to their linear counterparts. Interactions between selected USCLs and membranes were studied by molecular dynamics simulations using a coarse-grained force field. Moreover, membrane permeabilization properties and kinetics were examined. Fluorescence and transmission electron microscopy revealed damage to Candida cell membrane and organelles. Concluding, USCLs are strong membrane disruptors and disulfide cyclization of polar head can have a beneficial effect on its in vitro selectivity between Candida sp. and normal human cells.

Comparative assessment of purified saponins as permeabilization agents during respirometry

Saponins are a diverse group of secondary plant metabolites, some of which display hemolytic toxicity due to plasma membrane permeabilization. This feature is employed in biological applications for transferring hydrophilic molecules through cell membranes. Widely used commercial saponins include digitonin and saponins from soap tree bark, both of which constitute complex mixtures of little definition. We assessed the permeabilization power of pure saponins towards cellular membranes in an effort to detect novel properties and to improve existing applications.In a respirometric assay, we characterized half-maximal permeabilization of the plasma membrane for different metabolites, of the mitochondrial outer membrane for cytochrome C and the full solubilization of mitochondrial inner membrane protein complexes.Beyond the complete list as repository for the field, we highlight several findings with direct applicability. First, we identified and validated α-chaconine as alternative permeabilization agent in respirometric assays of cultured cells and isolated synaptosomes, superior to digitonin in its tolerability for mitochondria. Second, we identified glycyrrhizic acid to form exceptionally small pores impermeable for adenosine diphosphate. Third, in a concentration dependent manner, tomatine proved to be able to selectively permeabilize the mitochondrial outer, but not inner membrane, allowing for novel states in which to determine cytochrome C oxidase activity.In summary, we provide a list of the permeabilization properties of 18 pure saponins. The identification of two saponins, namely tomatine and chaconine, with direct usability in improved or novel cell biological applications within this small subgroup demonstrates the tremendous potential for further functional screening of pure saponins.

C1-CBP-vancomycin: Impact of a Vancomycin C-Terminus Trimethylammonium Cation on Pharmacological Properties and Insights into Its Newly Introduced Mechanism of Action.

C1-CBP-vancomycin (3) was examined alongside CBP-vancomycin for susceptibility to acquired resistance upon serial exposure against two vancomycin-resistant enterococci strains where its activity proved more durable and remarkably better than many current therapies. Combined with earlier studies, this observation confirmed an added mechanism of action was introduced by incorporation of the trimethylammonium cation and that C1-CBP-vancomycin exhibits activity against vancomycin-resistant organisms through two synergistic mechanisms of action, both independent of d-Ala-d-Ala/d-Lac binding. New insights into this added mechanism of action, induced cell membrane permeabilization, can be inferred from studies that show added exogenous lipoteichoic acid reduces antimicrobial activity, rescues bacteria cell growth inhibition, and blocks induced cell permeabilization properties of C1-CBP-vancomycin, suggesting a direct binding interaction with embedded teichoic acid is responsible for the added mechanism of action and enhanced antimicrobial activity. Further studies indicate that the trimethylammonium cation does not introduce new liabilities in common pharmacological properties of the analogue and established that 3 is well tolerated in mice, displays substantial PK improvements over both vancomycin and CBP-vancomycin, and exhibits in vivo efficacy against a challenging multidrug-resistant and vancomycin-resistant S. aureus strain that is representative of the resistant pathogens all fear will emerge in the general population.

Magainin-H2 effects on the permeabilization and mechanical properties of giant unilamellar vesicles

Among the potential novel therapeutics to treat bacterial infections, antimicrobial peptides (AMPs) are a very promising substitute due to their broad-spectrum activity and rapid bactericidal action. AMPs strongly interact with the bacterial membrane, and the need to have a correct understanding of the interaction between AMPs and lipid bilayers at a molecular level prompted a wealth of experimental and theoretical studies exploiting a variety of AMPs. Here, we studied the effects of magainin H2 (Mag H2), an analog of the well-known magainin 2 (wt Mag 2) AMP endowed with a higher degree of hydrophobicity, on giant unilamellar vesicles (GUVs) concentrating on its permeabilization activity and the effect on the lipid bilayer mechanical properties. We demonstrated that the increased hydrophobicity of Mag H2 affects its selectivity conferring a strong permeabilization activity also on zwitterionic lipid bilayers. Moreover, when lipid mixtures including PG lipids are considered, PG has a protective effect, at variance from wt Mag 2, suggesting that for Mag H2 the monolayer curvature could prevail over the peptide-membrane electrostatic interaction. We then mechanically characterized GUVs by measuring the effect of Mag H2 on the bending constant of lipid bilayers by flickering spectroscopy and, by using micropipette aspiration technique, we followed the steps leading to vesicle permeabilization. We found that Mag H2, notwithstanding its enhanced hydrophobicity, has a pore formation mechanism compatible with the toroidal pore model similar to that of wt Mag 2.

Structure and Function in Antimicrobial Piscidins: Histidine Position, Directionality of Membrane Insertion, and pH-Dependent Permeabilization.

Piscidins are histidine-enriched antimicrobial peptides that interact with lipid bilayers as amphipathic α-helices. Their activity at acidic and basic pH in vivo makes them promising templates for biomedical applications. This study focuses on p1 and p3, both 22-residue-long piscidins with 68% sequence identity. They share three histidines (H3, H4, and H11), but p1, which is significantly more permeabilizing, has a fourth histidine (H17). This study investigates how variations in amphipathic character associated with histidines affect the permeabilization properties of p1 and p3. First, we show that the permeabilization ability of p3, but not p1, is strongly inhibited at pH 6.0 when the conserved histidines are partially charged and H17 is predominantly neutral. Second, our neutron diffraction measurements performed at low water content and neutral pH indicate that the average conformation of p1 is highly tilted, with its C-terminus extending into the opposite leaflet. In contrast, p3 is surface bound with its N-terminal end tilted toward the bilayer interior. The deeper membrane insertion of p1 correlates with its behavior at full hydration: an enhanced ability to tilt, bury its histidines and C-terminus, induce membrane thinning and defects, and alter membrane conductance and viscoelastic properties. Furthermore, its pH-resiliency relates to the neutral state favored by H17. Overall, these results provide mechanistic insights into how differences in the histidine content and amphipathicity of peptides can elicit different directionality of membrane insertion and pH-dependent permeabilization. This work features complementary methods, including dye leakage assays, NMR-monitored titrations, X-ray and neutron diffraction, oriented CD, molecular dynamics, electrochemical impedance spectroscopy, surface plasmon resonance, and quartz crystal microbalance with dissipation.

The mechanism of action of pepR, a viral-derived peptide, against Staphylococcus aureus biofilms.

ObjectivesTo investigate the mechanism of action at the molecular level of pepR, a multifunctional peptide derived from the Dengue virus capsid protein, against Staphylococcus aureus biofilms.MethodsBiofilm mass, metabolic activity and viability were quantified using conventional microbiology techniques, while fluorescence imaging methods, including a real-time calcein release assay, were employed to investigate the kinetics of pepR activity at different biofilm depths.ResultsUsing flow cytometry-based assays, we showed that pepR is able to prevent staphylococcal biofilm formation due to a fast killing of planktonic bacteria, which in turn resulted from a peptide-induced increase in the permeability of the bacterial membranes. The activity of pepR against pre-formed biofilms was evaluated through the application of a quantitative live/dead confocal laser scanning microscopy (CLSM) assay. The results show that the bactericidal activity of pepR on pre-formed biofilms is dose and depth dependent. A CLSM-based assay of calcein release from biofilm-embedded bacteria was further developed to indirectly assess the diffusion and membrane permeabilization properties of pepR throughout the biofilm. A slower diffusion and delayed activity of the peptide at deeper layers of the biofilm were quantified.ConclusionsOverall, our results show that the activity of pepR on pre-formed biofilms is controlled by its diffusion along the biofilm layers, an effect that can be counteracted by an additional administration of peptide. Our study sheds new light on the antibiofilm mechanism of action of antimicrobial peptides, particularly the importance of their diffusion properties through the biofilm matrix on their activity.

Synergistic antifungal effect of chitosan-stabilized selenium nanoparticles synthesized by pulsed laser ablation in liquids against Candida albicans biofilms.

BackgroundCandida albicans is a major opportunistic fungal pathogen. One of the most important virulence factors that contribute to the pathogenesis of candidiasis is its ability to form biofilms. A key characteristic of Candida biofilms is their resistance to antifungal agents. Due to significant morbidity and mortality rates related to biofilm-associated drug resistance, there is an urgency to develop novel nanotechnology-based approaches preventing biofilm-related infections.MethodsIn this study, we report, for the first time, the synthesis of selenium nanoparticles by irradiating selenium pellets by nanosecond pulsed laser ablation in liquid chitosan as a capping agent. Synergy of the fungicidal effect of selenium nanoparticles and chitosan was quantified by the combination index theorem of Chou–Talalay.ResultsThis drug combination resulted in a potent fungicidal effect against a preformed C. albicans biofilm in a dose–response manner. By advanced electron microscopy techniques, we documented the adhesive and permeabilizing properties of chitosan, therefore allowing selenium nanoparticles to enter as the cell wall of the yeast became disrupted and distorted. Most importantly, we demonstrated a potent quantitative synergistic effect when compounds such as selenium and chitosan are combined.ConclusionThese chitosan-stabilized selenium nanoparticles could be used for ex vivo applications such as sterilizers for surfaces and biomedical devices.

Ultrafast Delivery of Aggregation-Induced Emission Nanoparticles and Pure Organic Phosphorescent Nanocrystals by Saponin Encapsulation.

Saponins are a class of naturally occurring bioactive and biocompatible amphiphilic glycosides produced by plants. Some saponins, such as α-hederin, exhibit unique cell membrane interactions. At concentrations above their critical micelle concentration, they will interact and aggregate with membrane cholesterol to form transient pores in the cell membrane. In this project, we utilized the unique permeabilization and amphiphilic properties of saponins for the intracellular delivery of deep-red-emitting aggregation-induced emission nanoparticles (AIE NPs) and pure organic room-temperature phosphorescent nanocrystals (NCs). We found this method to be biocompatible, inexpensive, ultrafast, and applicable to deliver a wide variety of AIE NPs and NCs into cancer cells.

Studying the Mechanism of Membrane Permeabilization Induced by Antimicrobial Peptides Using Patch-Clamp Techniques.

Many short peptides selectively permeabilize the bacteria plasma membrane, leading to their lyses and death: they are therefore a source of antibacterial molecules and inspiration for novel and more selective drugs, which may have wider application in many other fields, as selective anticancer drugs. In this chapter, it is presented a new method to investigate the permeabilization properties of antimicrobial peptides under strict physiological conditions, employing the patch-clamp technique coupled to a fast perfusion system.

Walking dead: Permeabilization of heat-treated Geobacillus stearothermophilus ATCC 12980 spores under growth-preventing conditions

Although heat treatment is probably the oldest and the most common method used to inactivate spores in food processes, the specific mechanism of heat killing of spores is still not fully understood. The purpose of this study is to investigate the evolution of the permeabilization and the viability of heat-treated spores during storage under growth-preventing conditions. Geobacillus stearothermophilus spores were heat-treated under various conditions of temperature and pH, and then stored under conditions of temperature and pH that prevent growth. Spore survival was evaluated by count plating immediately after heat treatment, and then during storage over a period of months. Flow cytometry analyses were performed to investigate the Syto 9 permeability of heat-treated spores. Sub-lethally heat-treated spores of G. stearothermophilus were physically committed to permeabilization after heat treatment. However, prolonged heat treatment may abolish the spore permeabilization and block heat-treated spores in the refractive state. However, viability loss and permeabilization during heat treatment seem to be two different mechanisms that occur independently, and the loss of permeabilization properties takes place at a much slower rate than spore killing. Under growth-preventing conditions, viable heat-treated spores presumably lose their viability due to the permeabilization phenomena, which makes them more susceptible to the action of adverse conditions precluding growth.

Membrane Interactions of Natural Cyclic Lipodepsipeptides of the Viscosin Group

Many Pseudomonas spp. produce cyclic lipodepsipeptides (CLPs), which, besides their role in biological functions such as motility, biofilm formation and interspecies interactions, are antimicrobial. It has been established that interaction with the cellular membrane is central to the mode of action of CLPs. In this work, we focus on the CLPs of the so-called viscosin group, aiming to assess the impact of the main structural variations observed within this group on both the antimicrobial activity and the interaction with model membranes. The antimicrobial activity of viscosin, viscosinamide A, WLIP and pseudodesmin A were all tested on a broad panel of mainly Gram-positive bacteria. Their capacity to permeabilize or fuse PG/PE/cardiolipin model membrane vesicles is assessed using fluorescent probes. We find that the Glu2/Gln2 structural variation within the viscosin group is the main factor that influences both the membrane permeabilization properties and the minimum inhibitory concentration of bacterial growth, while the configuration of the Leu5 residue has no apparent effect. The CLP-membrane interactions were further evaluated using CD and FT-IR spectroscopy on model membranes consisting of PG/PE/cardiolipin or POPC with or without cholesterol. In contrast to previous studies, we observe no conformational change upon membrane insertion. The CLPs interact both with the polar heads and aliphatic tails of model membrane systems, altering bilayer fluidity, while cholesterol reduces CLP insertion depth.

Incorporation of β-Silicon-β3-Amino Acids in the Antimicrobial Peptide Alamethicin Provides a 20-Fold Increase in Membrane Permeabilization.

Incorporation of silicon-containing amino acids in peptides is known to endow the peptide with desirable properties such as improved proteolytic stability and increased lipophilicity. In the presented study, we demonstrate that incorporation of β-silicon-β3-amino acids into the antimicrobial peptide alamethicin provides the peptide with improved membrane permeabilizing properties. A robust synthetic procedure for the construction of β-silicon-β3-amino acids was developed and the amino acid analogues were incorporated into alamethicin at different positions of the hydrophobic face of the amphipathic helix by using SPPS. The incorporation was shown to provide up to 20-fold increase in calcein release as compared with wild-type alamethicin.

Comparative functional properties of engineered cationic antimicrobial peptides consisting exclusively of tryptophan and either lysine or arginine.

We previously reported a series of de novo engineered cationic antibiotic peptides (eCAPs) consisting exclusively of arginine and tryptophan (WR) that display potent activity against diverse multidrug-resistant (MDR) bacterial strains. In this study, we sought to examine the influence of arginine compared to lysine on antibacterial properties by direct comparison of the WR peptides (8-18 residues) with a parallel series of engineered peptides containing only lysine and tryptophan. WR and WK series were compared for antibacterial activity by bacterial killing and growth inhibition assays and for mechanism of peptide-bacteria interactions by surface plasmon resonance and flow cytometry. Mammalian cytotoxicity was also assessed by flow cytometry, haemolytic and tetrazolium-based assays. The shortest arginine-containing peptides (8 and 10 mers) displayed a statistically significant increase in activity compared to the analogous lysine-containing peptides. The WR and WK peptides achieved maximum antibacterial activity at the 12-mer peptide (WK12 or WR12). Further examination of antibacterial mechanisms of the optimally active 12-mer peptides using surface plasmon resonance and flow cytometry demonstrates stronger interactions with Pseudomonasaeruginosa, greater membrane permeabilizing activity, and lower inhibitory effects of divalent cations on activity and membrane permeabilization properties of WR12 compared to WK12 (P < 0.05). Importantly, WK12 and WR12 displayed similar negligible haemolytic and cytotoxic effects at peptide concentrations up to ten times the MIC or 20 times the minimum bactericidal concentration. Thus, arginine, compared to lysine, can indeed yield enhanced antibacterial activity to minimize the required length to achieve functional antimicrobial peptides.

Synthesis and evaluation of membrane permeabilizing properties of cationic amphiphiles derived from the disaccharide trehalose.

Cationic amphiphiles derived from trehalose have been synthesized; trehalose analogues substituted with n-pentyl or n-hexyl ethers exhibited membrane disrupting activities against clinically important Gram positive and Gram negative bacteria and fungi. Our results demonstrate that trehalose is a useful disaccharide scaffold for the development of broad-spectrum antimicrobial and antifungal agents.

Ceramides with a pentadecasphingosine chain and short acyls have strong permeabilization effects on skin and model lipid membranes

The composition and organization of stratum corneum lipids play an essential role in skin barrier function. Ceramides represent essential components of this lipid matrix; however, the importance of the individual structural features in ceramides is not fully understood. To probe the structure–permeability relationships in ceramides, we prepared analogs of N-lignoceroylsphingosine with shortened sphingosine (15 and 12 carbons) and acyl chains (2, 4 and 6 carbons) and studied their behavior in skin and in model lipid membranes. Ceramide analogs with pentadecasphingosine (15C) chains were more barrier-perturbing than 12C- and 18C-sphingosine ceramides; the greatest effects were found with 4 to 6C acyls (up to 15 times higher skin permeability compared to an untreated control and up to 79 times higher permeability of model stratum corneum lipid membranes compared to native very long-chain ceramides). Infrared spectroscopy using deuterated lipids and X-ray powder diffraction showed surprisingly similar behavior of the short ceramide membranes in terms of lipid chain order and packing, phase transitions and domain formation. The high- and low-permeability membranes differed in their amide I band shape and lamellar organization. These skin and membrane permeabilization properties of some short ceramides may be explored, for example, for the rational design of permeation enhancers for transdermal drug delivery.