Chain Length of Free Fatty Acids Influences the Phase Behavior of Stratum Corneum Model Membranes (original) (raw)
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Very Long Chain Lipids Favor the Formation of a Homogeneous Phase in Stratum Corneum Model Membranes
Langmuir, 2020
The stratum corneum (SC), the outermost layer of mammal epidermis, acts as a barrier dictating the rate of absorption of exogenous molecules through the skin, as well as to prevent excessive water loss from the body. The SC consists of protein-rich corneocytes embedded into a complex lipid mixture. The lipid fraction is mainly constituted of an equimolar mixture of ceramides (Cer), free fatty acids (FFA), and cholesterol (Chol), forming a solid phase in the intracellular space; this lipid phase is supposed to play a fundamental role in the SC barrier function. An unusual characteristic of this biological membrane is that its lipids generally bear very long acyl chains, with the 24-carbon long ones being the most abundant. In this work, we used Raman microspectroscopy and infrared spectroscopy to study the influence of the acyl chain length on the lipid mixing properties in SC model membranes. Our results revealed that the combination of ceramides and FFA bearing a very long chain is required for the formation of homogeneous lipid mixtures, while lipids with shorter chains (16-carbon and 20-carbon atom long) lead to domains with micrometer dimensions. It is proposed that the biological machinery necessary for acyl chain elongation occurring at the mammalian skin level is required to inhibit lipid phase separation, a critical feature in the proper barrier functioning.
Phase behavior of isolated skin lipids
The Journal of Lipid Research
Ceramides were isolated from the pig stratum corneum (SC) and mixed in varying molar ratios with either cholesterol or with cholesterol and free fatty acids. The phase behavior of the mixtures was studied by small-(SAXD) and wide-angle (WAXD) X-ray diffraction. Ceramides alone did not exhibit a long range ordering. Upon addition of cholesterol to ceramides, lamellar phases were formed and a hexagonal lateral packing was detected similar to that seen in intact SC. At a cholesterol/ceramide molar ratio of 0.1, only one reflection at 5.9 nm was observed. At a cholesterol/ceramide molar ratio of 0.2, three reflections corresponding to 12.3, 5.56, and 4.26 nm appeared. The reflections were based on two phases. Increasing the cholesterol/ceramide ratio to 0.4, the peak positions were slightly shifted. The diffraction pattern revealed the presence of two lamellar phases with periodicities of 12.2 and 5.2 nm, respectively. The positions of the peaks remained unchanged when the cholesterol/ceramide ratio was increased up to 1.0. At a cholesterol/ceramide molar ratio of 2.0, the intensity of various peaks based on the 12.2 nm phase decreased in intensity. The phase behavior of the cholesterol/ceramide mixtures in a ratio between 0.4 and 1.0 was very similar to that found in intact pig SC in which two lamellar phases with periodicities of 6.0 and 13.2 nm are present. Our data further indicate that the formation of the 5.2 nm lamellar phase requires a higher cholesterol content than the formation of the 12.2 nm lamellar phase. Furthermore, when the relative amount of cholesterol is very high, the 5.2 nm phase is the most pronounced one. Addition of free fatty acids increased the solubility of cholesterol, indicating the role free fatty acids may play for the skin barrier function. The phase behavior of cholesterol/ceramide/fatty acid mixtures was found to be dependent on the chain length of fatty acids used. Namely, addition of short-chain free fatty acids (C14-Cl8) did not change the periodicity of the 12.2 and 5.2 nm phases, but induced the formation of an additional 4.2 nm phase. In the presence of long-chain free fatty acids (C16-C26), the periodicity of the lamellar phases was slightly increased (to 13.0 and 5.3 nm, respectively) but no additional 4.2 nm phase was formed. I These results indicate that the lipid phase behavior of the cholesterol/ceramide/free fatty acid mixtures closely mimics that of the intact stratum corneum only in the presence of long-chain free fatty acids.-Bouwstra, J. A, G. S. Gooris, K. Cheng, A. Weerheim, W. Bras, and M. Ponec. Phase behavior of isolated skin lipids. J. Lipid Res. 1996. 37: 999-101 1.
Molecular dynamics simulations of stratum corneum lipid models: fatty acids and cholesterol
Biochimica et Biophysica Acta (BBA) - Biomembranes, 2001
We report the results of an investigation on stratum corneum lipids, which present the main barrier of the skin. Molecular dynamics simulations, thermal analysis and FTIR measurements were applied. The primary objective of this work was to study the effect of cholesterol on skin structure and dynamics. Two molecular models were constructed, a free fatty acid bilayer (stearic acid, palmitic acid) and a fatty acid/cholesterol mixture at a 1:1 molar ratio. Our simulations were performed at constant pressure and temperature on a nanosecond time scale. The resulting model structures were characterized by calculating surface areas per headgroup, conformational properties, atom densities and order parameters of the fatty acids. Analysis of the simulations indicates that the free fatty acid fraction of stratum corneum lipids stays in a highly ordered crystalline state at skin temperatures. The phase behavior is strongly influenced when cholesterol is added. Cholesterol smoothes the rigid phases of the fatty acids: the order of the hydrocarbon tails (mainly of the last eight bonds) is reduced, the area per molecule becomes larger, the fraction of trans dihedrals is lower and the hydrophobic thickness is reduced. The simulation results are in good agreement with our experimental data from FTIR analysis and NIR-FT Raman spectroscopy.
Molecular organization of the lipid matrix in intact Stratum corneum using ATR-FTIR spectroscopy
Biochimica et Biophysica Acta (BBA) - Biomembranes, 2008
ATR-FTIR spectroscopy is useful in investigating the lateral organization of Stratum corneum (SC) lipids in full-thickness skin. Based on studies of the thermotropic phase transitions in n-tricosane and in excised human skin, the temperature dependence of the CH 2 scissoring bandwidth emerged as a measure of the extent of orthorhombic and hexagonal phases. This dependence provides a simpler measure of the lateral order in lipid assemblies than the common spectroscopic approaches based on difference spectra, curve fitting of the CH 2 scissoring region, and the position of the CH 2 stretching vibrations. It has the advantages of ease of determination, relatively low variability, and high discriminative power for the type of lateral intermolecular chain packing. A comparison of the lateral organization of the lipids at the SC surface of mammalian skin using the scissoring bandwidth revealed considerable differences between human abdominal skin (containing mostly orthorhombic phases), porcine ear skin (containing mostly hexagonal phases), and reconstructed human epidermis (containing mostly disordered phases). This parameter also correctly described the different effects of propylene glycol (minimally disturbing) and oleic acid (formation of a highly disordered phase) on the SC lipids in excised human skin. The procedure described here is applicable to in vivo studies in the areas of dermatology, transdermal drug delivery, and skin biophysics.
Biochimica et Biophysica Acta (BBA) - Biomembranes, 2014
The stratum corneum (SC) plays a fundamental role in the barrier function of the skin. The SC consists of corneocytes embedded in a lipid matrix. The main lipid classes in the lipid matrix are ceramides (CERs), cholesterol (CHOL) and free fatty acids (FFAs). The aim of this study was to examine the effect of the chain length of FFAs on the thermotropic phase behavior and mixing properties of SC lipids. Fourier transform infrared spectroscopy and Raman imaging spectroscopy were used to study the mixing properties using either protonated or deuterated FFAs. We selected SC model lipid mixtures containing only a single CER, CHOL and either a single FFA or a mixture of FFAs mimicking the FFA SC composition. The single CER consists of a sphingoid base with 18 carbon atoms and an acyl chain with a chain length of 24 carbon atoms. When using lignoceric acid (24 carbon atoms) or a mixture of FFAs, the CER and FFAs participated in mixed crystals, but hydration of the mixtures induced a slight phase separation between CER and FFA. The mixed crystalline structures did not phase separate during storage even up to a time period of 3 months. When using palmitic acid (16 carbon atoms), a slight phase separation was observed between FFA and CER. This phase separation was clearly enhanced during hydration and storage. In conclusion, the thermotropic phase behavior and the mixing properties of the SC lipid mixtures were shown to strongly depend on the chain length and chain length distribution of FFAs, while hydration enhanced the phase separation.
Bicellar systems to modify the phase behaviour of skin stratum corneum lipids
Physical Chemistry Chemical Physics, 2012
Bicellar systems are a fascinating category of versatile lipid assemblies that comprise bilayered disk-shaped nanoaggregates formed in water by long and short alkyl chain phospholipids. Bicelles bridge the gap between micelles and lipid vesicles by combining the attractive properties of both systems. These structures have recently been proposed in dermatological, cosmetic and pharmaceutical applications. Two new binary bicellar systems composed of cholesterol sulphate (SCHOL) and long-chain phospholipids (dimyristoyl-phosphatidylcholine, DMPC, or dipalmitoylphosphatidylcholine, DPPC) are characterised herein by differential scanning calorimetry, fluorescence spectroscopy, X-ray scattering and microscopy. Additionally, a comparative study on skin treated with the new SCHOL systems (DMPC/SCHOL and DPPC/SCHOL) and classic DHPC systems (DMPC/DHPC and DPPC/DHPC) was performed. These studies were conducted to determinate how deeply bicelles penetrate into the skin and the extension of their effect on the phase behaviour of stratum corneum (SC) lipids using attenuated total reflectance-Fourier transform infrared spectroscopy and two-photon excitation fluorescence microscopy. Our results show that SCHOL modified the typical discoidal morphology and the phase behaviour of the systems, inducing coexistence of two phases, liquid-ordered and ripple phases. The effect of the systems on SC lipids depends on their composition and is related to the fluidity of the SC lipid alkyl chains. Thus, systems with DMPC induced more disorder in SC lipids than systems with DPPC, and SCHOL did not modify the lipid arrangement. Perdeuterated systems in the infrared spectroscopy technique supported a different distribution in the tissue for every system. DMPC systems were primarily at the first layers of the SC, whereas DPPC systems were more widely distributed. Systems with SCHOL had enhanced distribution and penetration of bicellar systems throughout the SC.
Characterization of Stratum Corneum Molecular Dynamics by Natural-Abundance 13C Solid-State NMR
PLoS ONE, 2013
Despite the enormous potential for pharmaceutical applications, there is still a lack of understanding of the molecular details that can contribute to increased permeability of the stratum corneum (SC). To investigate the influence of hydration and heating on the SC, we record the natural-abundance 13 C signal of SC using polarization transfer solid-state NMR methods. Resonance lines from all major SC components are assigned. Comparison of the signal intensities obtained with the INEPT and CP pulse sequences gives information on the molecular dynamics of SC components. The majority of the lipids are rigid at 32uC, and those lipids co-exist with a small pool of mobile lipids. The ratio between mobile and rigid lipids increases with hydration. An abrupt change of keratin filament dynamics occurs at RH = 80-85%, from completely rigid to a structure with rigid backbone and mobile protruding terminals. Heating has a strong effect on the lipid mobility, but only a weak influence on the keratin filaments. The results provide novel molecular insight into how the SC constituents are affected by hydration and heating, and improve the understanding of enhanced SC permeability, which is associated with elevated temperatures and SC hydration.
Biochimica et Biophysica Acta (BBA) - Biomembranes, 2012
Ethanol is used in a variety of topical products. It is known to enhance the permeability of the skin by altering the ability of the stratum corneum (SC) intercellular membranes to form an effective barrier. In addition, ethanol and other alcohols are key components of antiseptic gels currently used for hand wash. Using infrared and deuterium NMR spectroscopy as well as calorimetry, we have investigated the effect of ethanol on a model membrane composed of lipids representing the three classes of SC lipids, an equimolar mixture of N-palmitoylsphingosine (ceramide), palmitic acid and cholesterol. Ethanol is found to influence the membrane in a dose dependent manner, disrupting packing and increasing lipid motion at low concentrations and selectively extracting lipids at moderate concentrations.