Models of the Three-Component Bilayer of Stratum Corneum: A Molecular Simulation Study.

The construction of the stratum corneum (SC) is crucial to the problems of transdermal drug delivery. SC consists of the keratinocyte layers and the lipid matrix surrounding it. Among them, the lipid matrix is the barrier for many exogenous molecules, mainly composed of ceramides (CERs), free fatty acids (FFA), and cholesterol (CHOL). In this work, we developed single-component (CERs, CER-NS, and CER-EOS) and six three-component models, and each model was simulated by using the GROMOS-54A7 force field. Short-period phase (SPP) and long-period phase (LPP) systems were established separately, and area per lipid (APL), thickness, order of carbon chain (SCD), and density distribution were analyzed. The transition of CER-NS and CER-EOS in LPP was observed. The results of hydrogen bonds in the lipid systems indicated that a strong hydrogen-bond network was formed between the skin-lipid bilayers. Umbrella sampling method simulations were performed to calculate the free energy change of ethanol moving into the skin-lipid bilayer. The results revealed that ethanol molecules pulled some water molecules into the membrane when they passed through SPP-1. Our findings provided some insights and models of the stratum corneum that could be used for the subsequent mechanism of macromolecule permeation through membranes in drugs, cosmetics, and so on.

Small-angle x-ray scattering investigation of the integration of free fatty acids in polysorbate 20 micelles.

A critical quality attribute for liquid formulations is the absence of visible particles. Such particles may form upon polysorbate hydrolysis resulting in release of free fatty acids into solution followed by precipitation. Strategies to avoid this effect are of major interest for the pharmaceutical industry. In this context, we investigated the structural organization of polysorbate micelles alone and upon addition of the fatty acid myristic acid (MA) by small-angle x-ray scattering. Two complementary approaches using a model of polydisperse core-shell ellipsoidal micelles and an ensemble of quasiatomistic micelle structures gave consistent results well describing the experimental data. The small-angle x-ray scattering data reveal polydisperse mixtures of ellipsoidal micelles containing about 22-35 molecules per micelle. The addition of MA at concentrations up to 100 µg/mL reveals only marginal effects on the scattering data. At the same time, addition of high amounts of MA (>500 µg/mL) increases the average sizes of the micelles indicating that MA penetrates into the surfactant micelles. These results together with molecular modeling shed light on the polysorbate contribution to fatty acid solubilization preventing or delaying fatty acid particle formation.

Comparative analysis of intercellular lipid organization and composition between psoriatic and healthy stratum corneum.

The lipid composition and organization of the stratum corneum (SC) in patients with psoriasis and healthy subjects were compared using X-ray diffraction, Fourier-transform infrared spectroscopy (FT-IR), and ultraperformance liquid chromatography, combined with time-of-flight mass spectrometry (UPLC-TOFMS). In healthy SC (HSC), SC lipids formed two lamellar phases (long and short periodicity phases). Hexagonal and orthorhombic hydrocarbon-chain packing were observed in the lateral lipid organization at 30 °C via X-ray diffraction. In HSC, the lamellar phases and the hydrocarbon-chain packing organizations changed with elevated temperatures and finally disappeared. In these behaviors, the high-temperature hexagonal hydrocarbon-chain packing organization, which appeared above the orthorhombic hydrocarbon-chain packing organization, transformed to the liquid phase at about 90 °C in HSC. In psoriatic SC (PSC), hexagonal hydrocarbon-chain packing organization disappeared at about 65 °C with elevated temperatures. No high-temperature hexagonal hydrocarbon-chain packing organization were observed in PSC during heating process. Disorder of the hydrocarbon-chain packing of SC lipids was observed in PSC via FT-IR. In UPLC-TOFMS, free fatty acid (FFA) and ceramide (CER) compositions differed between patients with PSC and HSC. Specifically, the levels of ultra-long chain fatty acids containing CER and phytosphingosine-containing CER were decreased, while those of sphingosine and dihydrosphingosine-containing CER and unsaturated FFA were increased in PSC. Furthermore, FFA and CER carbon chain lengths decreased in patients with PSC. These results suggest that the alteration of SC lipid composition and the reduction of carbon chain lengths in PSC lowered the structural transformation temperature, thereby reducing barrier function.

Microphase transitions of Langmuir-Blodgett lipid-assembled monolayers with new types of ceramides, ultra-long-chain ceramide and 1-O-acylceramide.

HYPOTHESIS: Intercellular lipid lamellae, consisting of ceramide, cholesterol, and free fatty acids, are the primary pathways for substances in the stratum corneum (SC). The microphase transition of lipid-assembled monolayers (LAMs), mimicking an initial layer of the SC, would be affected by new types of ceramides: ceramide with ultra-long chain (CULC) and 1-O-acylceramide (CENP) with three chains in different direction. EXPERIMENTS: The LAMs were fabricated with varying the mixing ratio of CULC (or CENP) against base ceramide via Langmuir-Blodgett assembly. Surface pressure-area isotherms and elastic modulus-surface pressure plots were obtained to characterize π-dependent microphase transitions. The surface morphology of LAMs was observed by atomic force microscopy. FINDINGS: The CULCs favored lateral lipid packing, and the CENPs hindered the lateral lipid packing by lying alignment, which was due to their different molecular structures and conformations. The sporadic clusters and empty spaces in the LAMs with CULC were presumably due to the short-range interactions and self-entanglements of ultra-long alkyl chains following the freely jointed chain model, respectively, which was not noticeably observed in the neat LAM films and the LAM films with CENP. The addition of surfactants disrupted the lateral packing of lipids, thus weakening the LAM elasticity. These findings allowed us to understand the role of CULC and CENP in the lipid assemblies and microphase transition behaviors in an initial layer of SC.

Colour and pigment in raw ground meat incorporated crushed garlic during the refrigerated storage: Their relationship to lipolytic and volatilomic changes.

The effects of freshly crushed garlic incorporated in raw ground meat at different rates 0 % to 2 % on colour, pigment forms, TBARS, peroxide, free fatty acids and volatilomic were investigated during 96 h storage at 4 °C. With advancing storage time and increasing garlic rate from 0 % to 2 %, the redness (a*), colour stability, oxymyoglobin and deoxymyoglobin decreased, but metmyoglobin, TBARS, peroxide, free fatty acids (C6, C15-C17), and aldehydes and alcohols, especially hexanal, hexanol, benzaldehyde, increased. Principal component analysis based on pigment, colour, lipolytic and volatilomic changes classified successfully the meat samples. Metmyoglobin was positively correlated with lipid oxidation products (TBARS, hexanal), but the other pigment forms and colour parameters (a* and b* values) were negatively correlated. Increased metmyoglobin proportion and decreased redness and colour stability may be used as reliable indicators of lipid oxidation. Also, the incorporation of fresh garlic into ground meat was not promising to increase oxidative stability.

Biodiesel Production from Chlorella homosphaera by Two-Step Catalytic Conversion Using Waste Radish Leaves as a Source for Heterogeneous Catalyst.

The economic viability of algal biodiesel can be improved by enhancing the microalgal lipid accumulation and using agricultural waste as a cheap and sustainable source of catalysts. In the current study, the effect of various nitrogen concentrations on the growth and lipid of Chlorella homosphaera were investigated. Furthermore, two-step catalytic conversion was applied to convert the oil of C. homosphaera with high free fatty acids (FFA) to biodiesel using waste radish leaves as a source of a heterogeneous base catalyst. The result revealed that the maximum lipid productivity of 25.0 mg L-1 day-1 and lipid content of 30.83% were obtained under nitrogen-depleted and limited nitrogen conditions, respectively. The FFA was reduced from 18.79 to 0.76%, and the acid value was decreased from 37.4 to 1.52 mg KOH g-1 using a 15:1 methanol to oil molar ratio (MTOR), 1.5 wt.% H2SO4, at 60 °C for 150 min. Under the optimized conditions, i.e., MTOR of 10:1, 3 wt.% of catalyst ratio for 120 min at 60 °C, the highest oil conversion of 96.61% was obtained. The physicochemical properties of the produced biodiesel were in the range of the standard specification norms for biodiesel. Hence, the proposed two-step catalytic conversion using calcined radish leaves as a heterogeneous catalyst has thus exhibited good potential for biodiesel production using algal oil with high FFA.

Estradiol production of granulosa cells is unaffected by the physiological mix of nonesterified fatty acids in follicular fluid.

Ovarian cycle is controlled by circulating levels of the steroid hormone 17-ß-estradiol, which is predominantly synthesized by the granulosa cells (GCs) of ovarian follicles. Our earlier studies showed that unsaturated fatty acids (USFs) downregulate and saturated fatty acids (SFAs) upregulate estradiol production in GCs. However, it was unclear whether pituitary gonadotropins induce accumulation of free fatty acids (FFAs) in the follicular fluid since follicle-stimulating hormone induces and luteinizing hormone inhibits estradiol production in the mammalian ovary. Interestingly, we show here the gas chromatography analysis of follicular fluid revealed no differential accumulation of FFAs between pre- and post-luteinizing hormone surge follicles. We therefore wondered how estradiol production is regulated in the physiological context, as USFs and SFAs are mutually present in the follicular fluid. We thus performed in vitro primary GC cultures with palmitate, palmitoleate, stearate, oleate, linoleate, and alpha-linolenate, representing >80% of the FFA fraction in the follicular fluid, and analyzed 62 different cell culture conditions to understand the regulation of estradiol biosynthesis under diverse FFA combinations. Our analyses showed co-supplementation of SFAs with USFs rescued estradiol production by restoring gonadotropin receptors and aromatase, antagonizing the inhibitory effects of USFs. Furthermore, transcriptome data of oleic acid-treated GCs indicated USFs induce the ERK and Akt signaling pathways. We show SFAs inhibit USF-induced ERK1/2 and Akt activation, wherein ERK1/2 acts as a negative regulator of estradiol synthesis. We propose SFAs are vital components of the follicular fluid, without which gonadotropin signaling and the ovarian cycle would probably be shattered by USFs.

Using molecular simulation to understand the skin barrier.

Skin's effectiveness as a barrier to permeation of water and other chemicals rests almost entirely in the outermost layer of the epidermis, the stratum corneum (SC), which consists of layers of corneocytes surrounded by highly organized lipid lamellae. As the only continuous path through the SC, transdermal permeation necessarily involves diffusion through these lipid layers. The role of the SC as a protective barrier is supported by its exceptional lipid composition consisting of ceramides (CERs), cholesterol (CHOL), and free fatty acids (FFAs) and the complete absence of phospholipids, which are present in most biological membranes. Molecular simulation, which provides molecular level detail of lipid configurations that can be connected with barrier function, has become a popular tool for studying SC lipid systems. We review this ever-increasing body of literature with the goals of (1) enabling the experimental skin community to understand, interpret and use the information generated from the simulations, (2) providing simulation experts with a solid background in the chemistry of SC lipids including the composition, structure and organization, and barrier function, and (3) presenting a state of the art picture of the field of SC lipid simulations, highlighting the difficulties and best practices for studying these systems, to encourage the generation of robust reproducible studies in the future. This review describes molecular simulation methodology and then critically examines results derived from simulations using atomistic and then coarse-grained models.

Phytosphingosine ceramide mainly localizes in the central layer of the unique lamellar phase of skin lipid model systems.

Understanding the lipid arrangement within the skin's outermost layer, the stratum corneum (SC), is important for advancing knowledge on the skin barrier function. The SC lipid matrix consists of ceramides (CERs), cholesterol, and free fatty acids, which form unique crystalline lamellar phases, referred to as the long periodicity phase (LPP) and short periodicity phases. As the SC lipid composition is complex, lipid model systems that mimic the properties of native SC are used to study the SC lipid organization and molecular arrangement. In previous studies, such lipid models were used to determine the molecular organization in the trilayer structure of the LPP unit cell. The aim of this study was to examine the location of CER N-(tetracosanoyl)-phytosphingosine (CER NP) in the unit cell of this lamellar phase and compare its position with CER N-(tetracosanoyl)-sphingosine (CER NS). We selected CER NP as it is the most prevalent CER subclass in the human SC, and its location in the LPP is not known. Our neutron diffraction results demonstrate that the acyl chain of CER NP was positioned in the central part of the trilayer structure, with a fraction also present in the outer layers, the same location as determined for the acyl chain of CER NS. In addition, our Fourier transformed infrared spectroscopy results are in agreement with this molecular arrangement, suggesting a linear arrangement for the CER NS and CER NP. These findings provide more detailed insight into the lipid organization in the SC lipid matrix.

The importance of ceramide headgroup for lipid localisation in skin lipid models.

The stratum corneum's lipid matrix is a critical for the skin's barrier function and is primarily composed of ceramides (CERs), cholesterol (CHOL) and free fatty acids (FFAs). The lipids form a long periodicity phase (LPP), a unique trilayer unit cell structure. An enzyme driven pathway is implemented to synthesize these key lipids. If these enzymes are down- or upregulated as in inflammatory diseases, the final lipid composition is affected often altering the barrier function. In this study, we mimicked down regulation of enzymes involved in the synthesis of the sphingosine and CER amide bond. In a LPP lipid model, we substituted CER N-(tetracosanoyl)-sphingosine (CER NS) with either i) FFA C24 and free sphingosine, to simulate the loss of the CER amide bond, or ii) with FFA C24 and C18 to simulate the loss of the sphingosine headgroup. Our study shows the lipids in the LPP would not phase separate until at least 25% of the CER NS is substituted keeping the lateral packing and conformational ordering unaltered. Neutron diffraction studies showed that free sphingosine chains localized at the outer layers of the unit cell, while the remaining CER NS head group was concentrated in the inner headgroup layers. However, when FFA C18 was inserted, CER NS was dispersed throughout the LPP, resulting in an even distribution between the inner and outer water layers. The presented results highlight the importance of the CER NS headgroup structure and its interaction in combination with the carbon chain invariability for optimal lipid arrangement.

Monitoring polysorbate hydrolysis in therapeutic proteins using an ultrasensitive extraction-free fatty acid quantitation method.

Polysorbates (PSs) are surfactants commonly added to therapeutic protein drug product formulations to protect proteins from denaturation and aggregation during storage, transportation, and delivery. However, enzymatic hydrolysis of PSs has been recognized as the primary route of PS degradation in monoclonal antibody formulations, resulting in the release of free fatty acids that drive undesired particulate formation. Here, we present a rapid lipase activity assay with optimized incubation conditions for accurate quantitation of free fatty acids without a fatty acid extraction step. This assay can detect low levels of PS degradation (0.000024% PS20 degradation) within 1 day with minimal sample preparation. The levels of released free fatty acids were found to strongly correlate with the degree of PS20 degradation. The case study described herein suggests that this approach can detect low levels of PS20 degradation caused by sub-ppm lipase levels within 1 day, compared with the duration of 14 days needed for PS degradation assays based on two-dimensional liquid chromatography-charge aerosol detection.

Antiadipogenic Effects of Different Molecular Forms of Conjugated Linoleic Acid on 3T3-L1 Cells: Comparison between Free Fatty Acid and Phosphatidylcholine Forms.

The antiadipogenic activity of conjugated linoleic acids (CLA) in the form of phosphatidylcholine-bound (CLA-PC) or free fatty acids (FFA; CLA-FFA) was evaluated using 3T3-L1 adipocytes. Phosphatidylcholine from soya (soy-PC) was used as the comparison of PC form. Both the lipid accumulation and activity of glycerol-3-phosphate dehydrogenase were measured to determine lipogenesis, whereas the glycerol content was measured to evaluate lipolysis. The CLA uptake also measured to find out the utilization of CLA by the cells. As a results, lipid accumulation in 3T3-L1 adipocytes was inhibited in a dose-dependent manner following treatment with CLA-PC (50-400 µM). Both CLA-PC and soy-PC significantly suppressed lipid accumulation compared with CLA-FFA, even though the amount of CLA in CLA-PC was a half than CLA-FFA. The CLA uptake of PC form was superior to FFA form, however, no difference was noted between CLA-PC and soy-PC. These forms exerted their antiadipogenic activity via the suppression of lipogenesis, and not by increasing lipolysis. Short-term treatment, especially in the middle stage of differentiation, was more effective than long-term treatment; especially for CLA-FFA. The antiadipogenic effect of CLA-PC was partially attributed to the chemical structure of the PC molecule. These results provide important information for the utilization of physiologically functional fatty acids and particularly CLA in the food and medical fields.

Evaluating a Modified High Purity Polysorbate 20 Designed to Reduce the Risk of Free Fatty Acid Particle Formation.

PURPOSE: To evaluate a modified high purity polysorbate 20 (RO HP PS20)-with lower levels of stearate, palmitate and myristate esters than the non-modified HP PS20-as a surfactant in biopharmaceutical drug products (DP). RO HP PS20 was designed to provide functional equivalence as a surfactant while delaying the onset of free fatty acid (FFA) particle formation upon hydrolytic degradation relative to HP PS20. METHODS: Analytical characterization of RO HP PS20 raw material included fatty acid ester (FAE) distribution, higher order ester (HOE) fraction, FFA levels and trace metals. Functional assessments included 1) vial and intravenous bag agitation; 2) oxidation via a placebo and methionine surrogate study; and 3) hydrolytic PS20 degradation studies to evaluate FFA particle formation with and without metal nucleation. RESULTS: Interfacial protection and oxidation propensity were comparable between the two polysorbates. Upon hydrolytic degradation, FFA particle onset was delayed in RO HP PS20. The delay was more pronounced when HOEs of PS20 were preferentially degraded. Furthermore, the hydrolytic degradants of RO HP PS20 formed fewer particles in the presence of spiked aluminum. CONCLUSION: This work highlights the criticality of having tighter control on long chain FAE levels of PS20 to reduce the occurrence of FFA particle formation upon hydrolytic degradation and lower the variability in its onset. By simultaneously meeting compendial PS20 specifications while narrowing the allowable range for each FAE and shifting its composition towards the shorter carbon chain species, RO HP PS20 provides a promising alternative to HP PS20 for biopharmaceutical DPs.

Hierarchical Zeolites as Catalysts for Biodiesel Production from Waste Frying Oils to Overcome Mass Transfer Limitations.

Hierarchical crystals with short diffusion path, conventional microcrystals and nanocrystals of ZSM-5 zeolites were used for biodiesel production from waste frying oils and were assessed for their catalytic activity in regard to their pore structure and acidic properties. Produced zeolites were characterized using XRD, nitrogen adsorption-desorption, SEM, TEM, X-ray fluorescence, and FTIR. Pore size effect on molecular diffusion limitation was assessed by Thiele modulus calculations and turnover frequencies (TOF) were used to discuss the correlation between acidic character and catalytic performance of the zeolites. Owing to the enhanced accessibility and mass transfer of triglycerides and free fatty acids to the elemental active zeolitic structure, the catalytic performance of nanosponge and nanosheet hierarchical zeolites was the highest. A maximum yield of 48.29% was reached for the transesterification of waste frying oils (WFOs) using HZSM-5 nanosheets at 12:1 methanol to WFOs molar ratio, 180 °C, 10 wt % catalyst loading, and 4 h reaction time. Although HZSM-5 nanosponges achieved high conversions, these more hydrophilic zeolites did not function according to their entire acidic strength in comparison to HZSM-5 nanosheets. NSh-HZSM5 catalytic performance was still high after 4 consecutive cycles as a result of the zeolite regeneration.

A simulated strategy for analysis of Short- to Long- chain fatty acids in mouse serum beyond chemical standards.

Broadening coverage in fatty acid (FA) analysis benefits the understanding of metabolic regulation in biological system. However, the limited access of chemical standards makes it challenging. In this work, we introduced a simulation assisted strategy to analyze short-, medium-, long- and very-long-chain fatty acids beyond the use of chemical standards. This targeted analysis in selected reaction monitoring (SRM) mode incorporated 3-nitrophenylhydrazine derivatization and mathematical simulation of ion transitions, collision energies, RF values and retention times to identify and quantify the fatty acids without chemical standards. Serum analysis using high resolution mass spectrometry coupled with paired labeling was employed to refine the computational retention times. Based on the simulation, 116 free fatty acids from C1 to C24 were covered in a single analysis on use of 34 standard chemicals. Background interference is commonly observed in fatty acid analysis. For certain fatty acids, e.g. acetic acid or palmitic acid, reliable quantitation is largely restricted by contamination level instead of detection limit. Therefore, the background interference and quantifiable serum volume required for each fatty acid were also evaluated. At least 20 µL serum was suggested to cover most molecules. Using this approach, a total of 66 free fatty acids with various chain lengths and saturations were detected in NTCP knockout mice serum, of which 34 FAs were confirmed by chemical standards and 32 FAs were potentially assigned based on the simulation. Gender dependent fatty acid regulation was observed by NTCP knockout. This work provides a unique strategy that enables to broaden the fatty acid coverage with the absence of chemical standards and is applicable to other derivatizations.

Changes of lipids in noodle dough and dried noodles during industrial processing.

This study investigated the changes of lipids during the industrial preparation of noodle dough and dried noodles, including the hydration, sheeting, and drying processes. The results showed that industrial processing markedly influenced the stability of lipids during the preparation of dried noodles. The contents of total free fatty acids, polyunsaturated fatty acids, and free lipids were reduced, while peroxide values increased during the hydration and sheeting processes, showing the instability of lipids. The increase in lipid oxidation may have been due to the activation of lipoxygenase. Although its activity declined by 45.7% in the hydration process compared to that of the native wheat flour (198.5 ± 20.4 U/g/min), the residue activity should have been high enough to oxidize lipids. Interestingly, lipase activity remained relatively stable. In addition, an obvious increase of carbon-centered free radicals was observed during the entire processing. In conclusion, the industrial processing, especially the hydration process, markedly changed the lipid profile and promoted lipid oxidation during the preparation of dried noodles. PRACTICAL APPLICATION: The present study showed the positive relationship between endogenous lipid degrading enzymes and the degradation of lipids and elucidated the role of industrial processing on lipid stability in noodle dough and dried noodles. The results of the present study will also help us to understand more about the sensory quality of dried noodles during preparation, as well as to develop high quality of wheat-based food products.

Development of a liquid chromatography-electrospray ionization tandem mass spectrometric method for the simultaneous analysis of free fatty acids.

Fatty acids (FAs) play important roles in several physiological and pathophysiological processes, functioning as both nonesterified free FAs (FFAs) and components of other lipid classes. Although many lipid classes are readily measured using liquid chromatography-electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS), the measurement of FFAs by this method is not straightforward because of inconsistent fragmentation behaviours. In this study, we describe a strategy to measure FFAs using conventional reverse-phase LC-ESI-MS/MS, without derivatization. The strategy combines three key methods: (i) an isocratic LC separation with a high organic solvent ratio, (ii) postcolumn base addition, and (iii) pseudo-multiple reaction monitoring. The method facilitates the measurement of ultra-long-chain FAs, the accumulation of which is a common biochemical abnormality in peroxisomal disorders. This study delivers a broad strategy that measures a wide spectrum of FFA species in complex biological samples.

The Importance of Free Fatty Chain Length on the Lipid Organization in the Long Periodicity Phase.

The skin's barrier ability is an essential function for terrestrial survival, which is controlled by intercellular lipids within the stratum corneum (SC) layer. In this barrier, free fatty acids (FFAs) are an important lipid class. As seen in inflammatory skin diseases, when the lipid chain length is reduced, a reduction in the barrier's performance is observed. In this study, we have investigated the contributing effects of various FFA chain lengths on the lamellar phase, lateral packing. The repeat distance of the lamellar phase increased with FFA chain length (C20-C28), while shorter FFAs (C16 to C18) had the opposite behaviour. While the lateral packing was affected, the orthorhombic to hexagonal to fluid phase transitions were not affected by the FFA chain length. Porcine SC lipid composition mimicking model was then used to investigate the proportional effect of shorter FFA C16, up to 50% content of the total FFA mixture. At this level, no difference in the overall lamellar phases and lateral packing was observed, while a significant increase in the water permeability was detected. Our results demonstrate a FFA C16 threshold that must be exceeded before the structure and barrier function of the long periodicity phase (LPP) is affected. These results are important to understand the lipid behaviour in this unique LPP structure as well as for the understanding, treatment, and development of inflammatory skin conditions.

Two-stage continuous production process for fatty acid methyl ester from high FFA crude palm oil using rotor-stator hydrocavitation.

Two-stage continuous production process for fatty acid methyl ester (FAME) from crude palm oil was performed using the rotor-stator hydrocavitation reactor. The novel ABS filament printed rotor having spherical holes on the surface of the rotor which is an efficient, fast and cost-effective procedure, was installed in the stainless steel stator of hydrosonic reactor. The 3D printed hydrosonic reactor was used to treat the FFA-rich in MCPO by esterification and followed by transesterification to produce the methyl ester. The optimum conditions of both esterification and transesterification processes were determined using the response surface methodology (RSM). For the 1st step esterification, the conditions of methanol 17.7 vol%, sulfuric acid 2.9 vol%, 3000 rpm rotor speed, hole's diameter and depth 4 and 6 mm, and 25 L/h MCPO, were used for decreasing the FFA from 11.456 to 1.028 wt%. For the 2nd step, transesterification was employed with the optimal condition of 28.6 vol% methanol, 6.2 g/L of potassium hydroxide, 3000 rpm rotor speed, the dimension of the spherical holes on the rotor's surface having diameter of 6.4 mm and 6.2 mm in depth, and esterified oil flow rate 25 L/h, for producing the methyl ester to over 99.163 wt%. Moreover, the purified biodiesel yields and the average energy consumption for the entire two-stage continuous process between hydrosonic and ultrasonic clamp reactors were compared. The results of purified methyl ester clearly indicate that the methyl esters of 99.163 wt% and 97.814 wt% were achieved from hydrosonic and ultrasonic clamp reactors, respectively, under the same optimized conditions. The maximum yields of purified biodiesel were 97.51 vol% and 88.69 vol% using hydrosonic and ultrasonic clamp reactors, respectively. The average energy consumption for the entire continuous two-stage process for both hydrosonic and ultrasonic clamp reactors were 0.049 and 0.056 kW h/L, respectively. For practical industrial processes, stainless steel rotors inside the stator was manufactured by CNC machine, which was also verified under the optimum conditions. The results showed that 1.07 wt% FFA and 99.221 wt% methyl ester of were obtained from first step and second step, respectively.

Acidic pH Is Required for the Multilamellar Assembly of Skin Barrier Lipids In Vitro.

Lipid membrane remodeling belongs to the most fundamental processes in the body. The skin barrier lipids, which are ceramide dominant and highly rigid, must attain an unusual multilamellar nanostructure with long periodicity to restrict water loss and prevent the entry of potentially harmful environmental factors. Our data suggest that the skin acid mantle, apart from regulating enzyme activities and keeping away pathogens, may also be a prerequisite for the multilamellar assembly of the skin barrier lipids. Atomic force microscopy on monolayers composed of synthetic or human stratum corneum lipids showed multilayer formation (approximately 10-nm step height) in an acidic but not in a neutral environment. X-ray diffraction, Fourier transform infrared spectroscopy, and permeability studies showed markedly altered lipid nanostructure and increased water loss at neutral pH compared with that at acidic pH. These findings are consistent with the data on the altered organization of skin lipids and increased transepidermal water loss under conditions such as inadequate skin acidification, for example, in neonates, the elderly, and patients with atopic dermatitis.