Egg Phosphatidylcholine Research Articles

Ethanolamine and Vinyl–Ether Moieties in Brain Phospholipids Modulate Behavior in Rats

Plasmalogens are brain-enriched phospholipids with a vinyl–ether bond at the sn-1 position between the glycerol backbone and the alkyl chain. Previous studies have suggested that plasmalogens modulate locomotor activity, anxiety-like behavior, and cognitive functions in rodents; however, the specific moieties contributing to behavioral regulation are unknown. In this study, we examined the behavioral modulation induced by specific phospholipid moieties. To confirm the permeability of phospholipids in injected liposomes, we measured the fluorescence intensity following intravenous injection of liposomes containing ATTO 740-labeled dioleoylphosphatidylethanolamine. Then, we compared the behavioral effects following injection of liposomes composed of egg phosphatidylcholine (PC) and 1-stearoyl-2-docosahexaenoyl-sn-glycero-3-phosphoethanolamine (PE 18:0/22:6), PC 18:0/22:6, 1-(1Z-octadecenyl)-2-docosahexaenoyl-sn-glycero-3-phosphoethanolamine (PE P-18:0/22:6), or PC P-18:0/22:6, into the tail vein of male rats. The time spent in the central region of the open field was significantly reduced after injection of PE 18:0/22:6, harboring an ester bond at sn-1 compared to controls. Furthermore, the discrimination ratio in the novel object recognition test was significantly higher in PC 18:0/22:6 compared to PE 18:0/22:6, suggesting that the substitution of ethanolamine with choline can enhance recognition memory. We demonstrate that the structures of the sn-1 bond and the hydrophilic moiety in the phospholipids can modulate exploratory behaviors and recognition memory in rodents.

A size-switchable microsphere loaded with salmon calcitonin as two-weekly dosing for osteoporosis therapy

Osteoporosis is a disease with an increased incidence of fractures due to decreased bone mass and destruction of the microstructure of bone tissue. Salmon calcitonin (sCT), as a peptide, possesses the ability to inhibit osteoclast activity and thus regulate bone metabolism in clinical. However, short half-life and unstable physicochemical properties leading to rapid degradation of sCT have severely limited its clinical application. In this study, a size-switchable microsphere was developed to solve the problem of frequent administration and poor stability of sCT. sCT was encapsulated into Egg PC to form anhydrous reverse micelles (ARM) and then ARM was encapsulated into microspheres (MS@ARM). The degradable composite microspheres were utilized to provide a drug reservoir for sustained release of ARM encapsulated with sCT to reduce the frequency of drug administration, while the released ARM encapsulated with sCT entered the blood circulation to further protect sCT. In vitro release experiments demonstrated that the microspheres could sustain the release of sCT for at least 16 days. The microspheres MS@ARM showed the advanced therapeutic effect on the mouse model of glucocorticoid-induced osteoporosis (GIOP) at a low dosing frequency. The size-switchable microsphere is expected to be a new strategy for delivering sCT for osteoporosis treatment.

The effect of the liposomal form of alteplase on the effectiveness of thrombolysis in coronary arteries in acute myocardial infarction

A liposomal (Lip) formulation of tissue plasminogen activator, alteplase (AlT), has been developed. The quantitative and qualitative composition of the liposomes, as well as their physicochemical properties and proteolytic activity, have been studied in relation to the thrombolytic liposomal form. It was determined that a formulation consisting of liposomes with a phosphatidylcholine/cholesterol ratio of 1.5 : 1, and lipids/alteplase ratio of 1 : 1, is optimal for treating acute myocardial infarction (AMI) in experimental models. At different component ratios, liposomes had a negative zeta potential value greater than 30 mV, indicating their aggregative stability, even after storage for two days at 20 degrees Celsius. Liposomes derived from soy phosphatidylcholine showed greater colloidal stability with a zeta potential of approximately –57 mV and a lower hydrodynamic diameter of approximately 140 nanometers, compared to liposomes derived from egg phosphatidylcholine, which had a zeta potential around –35.4 mV and a hydrodynamic diameter around 220 nanometers. The initial content of free AlT in the liposome supernatant from egg phosphatidylcholine (Lipeg) was 15.0 ± 4.0 %. During the incubation period of 4 days, the concentration of free AlT decreased to 9.0 ± 4.5 %. In contrast, in liposomes derived from soy phosphatidylcholine (LipS), the content of free AlT increased from 11.0 ± 4.5 % to 32.5 ± 6.0 % over the same incubation period. The value of the proteolytic activity of tissue plasminogen activator (tPA) in the compositions of Lipeg(AlT) and Lips (AlT) depends on the type of phosphatidylcholine. The initial tPA activity in Lipeg (AlT) was 36.0 %, and after 1 day, it increased to 45 %. In Lips (AlT), the initial activity was 61.0 % and increased to 66 % after 1 day. When using the liposomal form of alteplase for delivery into the coronary arteries of rats with acute myocardial infarction (AMI), a more complete fibrin lysis is noted compared to animals receiving the native form of the drug. The developed system of targeted delivery of alteplase using soy liposomes has been shown to significantly improve the degree of coronary artery lumen restoration by more than 15 %, compared to the use of a conventional drug (p < 0.05).

Chitosan hybrid nanomaterials: A study on interaction with biomimetic membranes

This study examined the influence of nanomaterials (NMs) on the organization of membrane lipids and the resulting morphological changes. The cell plasma membrane is heterogeneous, featuring specialized lipid domains in the liquid-ordered (Lo) phase surrounded by regions in the liquid-disordered (Ld) phase. We utilized model membranes composed of various lipids and lipid mixtures in different phase states to investigate the interactions between the NMs and membrane lipids. Specifically, we explored the interactions of pure chitosan (CS) and CS-modified nanocomposites (NCs) with ZnO, CuO, and SiO2 with four lipid mixtures: egg-phosphatidylcholine (EggPC), egg-sphingomyelin/cholesterol (EggSM/Chol), EggPC/Chol, and EggPC/EggSM/Chol, which represent the coexistence of Ld, Lo, and Ld/Lo, respectively. The data show that CS NMs increase the membrane lipid order at glycerol level probed by Laurdan spectroscopy. Additionally, the interaction of CS-based NMs with membranes leads to an increase in bending elasticity modulus, zeta potential, and vesicle size. The lipid order changes are most significant in the highly fluid Ld phase, followed by the Lo/Ld coexistence phase, and are less pronounced in the tightly packed Lo phase. CS NMs induced egg PC vesicle adhesion, fusion, and shrinking. In heterogeneous Lo/Ld membranes, inward invaginations and vesicle shrinking via the Ld phase were observed. These findings highlight mechanisms involved in CS NM-lipid interactions in membranes that mimic plasma membrane heterogeneity.

Liposomal delivery of organoselenium-cisplatin complex as a novel therapeutic approach for colon cancer therapy

Cisplatin is a widely-used chemotherapeutic agent for the treatment of various solid neoplasms including colon cancer. Cisplatin-induced DNA damage is restricted due to dose-related adverse reactions as well as primary resistance mechanisms. Therefore, it is imperative to utilize novel therapeutic approaches to circumvent cisplatin limitations and attenuate its normal tissues toxicity. In this study, we exploited a novel PEGylated liposomes with greater efficiency to treat colon cancer. For this, an organoselenium compound (diselanediylbis decanoic acid (DDA)) was synthesized, and liposomes composed of Egg PC or HSPC, as well as DOPE, mPEG2000-DSPE, cholesterol and DDA at varying molar ratios were prepared by using thin-film method. Cisplatin loading was performed through incubation with liposomes. Characterization of nanoliposomes indicated a favarable size range of 91–122 nm and negative zeta potential of −9 to −22 mv. The organoselenium compound significantly improved cisplatin loading efficiency within the liposomes (83.4 %). Results also revealed an efficient bioactivity of cisplatin liposome on C26 cells compared to the normal cells. Further, DDA bearing liposomes significantly improved drug residence time in circulation, reduced toxicity associated with the normal tissues, and enhanced drug accumulation within the oxidative tumor microenvironment. Collectively, results indicated that cisplatin encasement within liposomes by using this method could significantly improve the therapeutic efficacy in vivo, and merits further investigations.

Liposomes bearing poly(hydroxyethyl acrylate-co-ethyl vinyl sulfide) and indocyanine green and their temperature, oxidation, and near infrared-responsive release property

Liposomes are widely used for smart delivery due to their biodegradability and biocompatibility. Recently, liposomes have been developed to respond to dual and multi-stimuli. This development leads to the potential for superior in vitro and in vivo drug delivery in the future. In this study, egg PC liposomes responsive to temperature, oxidation, and NIR irradiation were prepared by modifying the liposomal membranes with poly(hydroxyethyl acrylate-co-ethyl vinyl sulfide) (P(HEA-co-EVS)) and including indocyanine green (ICG) in the liposomes.The copolymer was found to exhibit a lower critical solution temperature (LCST) and it was oxidizable and surface-active.On the TEM photos of the liposomes, multi-lamellar vesicles were found and the size was in accordance with the hydrodynamic diameter. The liposomes released their payload in response to H2O2 concentration possibly because the EVS was oxidized by the oxidizing agent. The liposome exhibited a temperature-responsive release possibly due to the thermal contraction of the copolymer chains.In addition, the release was triggered when a near-infrared(NIR) laser was irradiated to the liposome suspensions. Under NIR irradiation, the photosensitizer (i.e. ICG) can generate heat and reactive oxygen species, which, in turn, would induce the conformational and polarity change of the polymer chains, leading to a release.

Enhancing encapsulation of filarial antigen Brugia malayi thioredoxin in nano-liposomes: The role of lecithin composition.

Lymphatic filariasis is a debilitating infectious disease prevalent in endemic areas, necessitating the development of an effective vaccine for eradication. Although recombinant vaccine candidates have been deemed safe, they often fail to provide sufficient protection, which can be overcome by encapsulating them in nano-liposomes. In this study, we have optimised the liposomal composition for enhanced stability and encapsulation of filarial antigen Brugia malayi thioredoxin (Bm-TRX). Nano-liposomes were prepared using egg phosphatidylcholine (EPC) and cholesterol via thin-film hydration, followed by sonication and characterizing. Encapsulation efficiency was optimised using different weight ratios of EPC to cholesterol (8:2, 7:3, and 6:4) and total lipid (EPC+Cholesterol) concentration to antigen Bm-TRX (10:1, 10:2, and 10:3) followed by release kinetics study. Optimised parameters yielded spherical liposomes measuring 209 nm in diameter with narrow polydispersity. Our findings demonstrated the highest encapsulation efficiency of 70.685 % and stability of 10 hours for an EPC to cholesterol weight ratio of 7:3. The in silico study proved the antigenic nature of TRX. The liposomal formulations loaded with TRX, as optimized in this study, hold promise for improving antigen efficiency by enhancing stability, bioavailability, and prophylactic effects by acting as immune potentiators.

Preparation, physicochemical and emulsifying properties of chicken liver phosphatidylcholine by enzymatic extraction

In this study, chicken liver phosphatidylcholine (PCCL) was efficiently extracted by protamex proteinase. The physicochemical and emulsifying properties of PCCL was investigated and compared by those of bean phosphatidylcholine (PCB) and egg phosphatidylcholine (PCE). The results showed that the EAI of PCCL emulsion system was 0.817 m2/g, significantly higher than that of PCB (0.624 m2/g) and PCE (0.446 m2/g). Furthermore, the PCCL emulsion system had higher turbidity but was slightly less stable than the other two kinds of phosphatidylcholine. At the same temperature storage, as the pH increases, the reduced fat floatation rate delayed delamination, resulting in the lowest fat floatation rate of emulsions formed by PCB. The PCCL emulsifier was relatively stable in the different temperature at 25 and 50 °C. All of the emulsions exhibited shear thinning, with PCCL emulsions forming less viscous and more fluid emulsions. Interfacial protein adsorption and interfacial protein concentration were significantly higher in the PCCL emulsions. Therefore, the results of the study will facilitate further expansion of the high-value use of chicken liver, especially in poultry by-products.

The Use of Soy and Egg Phosphatidylcholines Modified with Caffeic Acid Enhances the Oxidative Stability of High-Fat (70%) Fish Oil-in-Water Emulsions

This study investigated the effect of the combined use of sodium caseinate (CAS), commercial phosphatidylcholine (PC), and modified PCs on the physical and oxidative stability of 70% fish oil-in-water emulsions. Caffeic acid was covalently attached to both modified PCs (PCs originated from soy and eggs) in order to increase the antioxidant activity of PCs and investigate the advantage of bringing the antioxidant activity to the close proximity of the oil-water interface. Results showed that oxidative stability was improved when part of the PC was substituted with modified soy PC or egg PC. Emulsions containing a low concentration of modified PCs (10 wt.% of total PC) resulted in a prooxidative effect on the formation of hydroperoxides compared to emulsions with free caffeic acid. On the other hand, a decrease in the formation of volatile oxidation products was observed for emulsions containing higher levels of modified PCs (60 wt.% of total PC) compared to the emulsions with free caffeic acid added at its equivalent concentration. Increased concentrations of modified PCs provided better oxidative stability in high-fat emulsions, independent of the modified PC type. Moreover, when oxidation was initiated by producing singlet oxygen near a single oil droplet using a focused laser, fluorescence imaging showed that the oxidation did not propagate from one oil droplet to another oil droplet.

Beta-Caryophyllene Induces Significant Changes in the Lipid Bilayer at Room and Physiological Temperatures: ATR-FTIR Spectroscopy Studies

Beta-caryophyllene (BCP) is a natural bicyclic sesquiterpene with high biological activity. Potentially, it can be used in the treatment of a wide range of neurological diseases. However, to date, there are practically no data on the interaction of BCP with biological membranes. In the present work, we studied for the first time the interaction of BCP with model membranes—liposomes based on egg yolk phosphatidylcholine (Egg PC) with a variable cholesterol content (from 0 to 25 w.%). Using ATR-FTIR spectroscopy, we have shown that the membrane rigidity and cholesterol content dramatically affect the nature of the interaction of BCP with the bilayer both at room temperature and at physiological temperatures. The incorporation of BCP into the thickness of the bilayer leads to changes in the subpolar region of the bilayer, and at a high cholesterol content, it can provoke the formation of defects in the membrane.

Protein Corona of Anionic Fluid-Phase Liposomes Compromises Their Integrity Rather than Uptake by Cells.

Despite the undisputable role of the protein corona in the biointeractions of liposome drug carriers, the field suffers from a lack of knowledge regarding the patterns of protein deposition on lipid surfaces with different compositions. Here, we investigated the protein coronas formed on liposomes of basic compositions containing combinations of egg phosphatidylcholine (PC), palmitoyloleoyl phosphatidylglycerol (POPG), and cholesterol. Liposome-protein complexes isolated by size-exclusion chromatography were delipidated and analyzed using label-free LC-MS/MS. The addition of the anionic lipid and cholesterol both affected the relative protein abundances (and not the total bound proteins) in the coronas. Highly anionic liposomes, namely those containing 40% POPG, carried corona enriched with cationic proteins (apolipoprotein C1, beta-2-glycoprotein 1, and cathelicidins) and were the least stable in the calcein release assay. Cholesterol improved the liposome stability in the plasma. However, the differences in the corona compositions had little effect on the liposome uptake by endothelial (EA.hy926) and phagocytic cells in the culture (U937) or ex vivo (blood-derived monocytes and neutrophils). The findings emphasize that the effect of protein corona on the performance of the liposomes as drug carriers occurs through compromising particle stability rather than interfering with cellular uptake.

Development of glycyrrhetinic acid ligand-functionalized liposomes for targeting hepatocellular carcinoma: Synthesis, preparation, characterization, and evaluation

In this study, we successfully synthesized a novel glycyrrhetinic acid (GA) derivate compound as a liposomal carrier. Glycyrrhetinic acid-diaminododecane-cholesterol (GA-DA-CH) was synthesized from GA, diaminododecane, and cholesteryl hemisuccinate by two-step amidation under catalytic conditions. The chemical structures of GA-DA (glycyrrhetinic acid-diaminododecane) and GA-DA-CH were confirmed by Mass Spectrum (MS), Nuclear Magnetic Resonance (NMR) spectrometer, and analyzed by Fourier Transform Infrared spectroscopy (FTIR), High-Performance Liquid Chromatography (HPLC). Glycyrrhetinic acid ligand-modified liposomes (GA-DA-CH-Lips) were prepared, consisting of egg phosphatidylcholine, cholesterol and GA-DA-CH. GA-DA-CH was integrated into the liposomal capsule to promote hepatocellular carcinoma targeting. The particle size of GA-DA-CH-Lips was calculated to be between 122.87 and 133.30 nm, with an approximate polydispersity index (PDI) was 0.20, and a zeta potential was in the range of –32.50 to –26.53 mV. GA-DA-CH-Lips were constructed with more than 90% encapsulation efficient and drug loading efficiency of no<6.40%. In addition, environmental stability assays show that GA-DA-CH-Lips are sensitive to temperature, and thus should be preserved at low temperatures. Furthermore, in vitro cellular uptake from qualitative and quantitative analysis indicates that GA-DA-CH integrating into liposomes possess a specific target for hepatocellular carcinoma. Intracellular inhibition showed that GA-DA-CH-Lips effectively inhibited cell proliferation in dose-dependent features. Based on these results, we have reason to believe that GA-DA-CH-Lips could be a potential hepatic target drug delivery to improve the therapeutic effect of hepatic diseases.

Nanoformulations with exopolysaccharides from cyanobacteria: enhancing the efficacy of bioactive molecules in the Mediterranean fruit fly control

The increasing demand for food has required intensive use of pesticides which are hazardous to the ecosystem. A valid alternative is represented by biopesticides; however, these molecules are often insoluble in water, and poorly bioavailable. Nanopesticides can be engineered to reach a selected target with controlled release of the active principle. In this work, capsaicin, an irritant alkaloid from hot chili peppers, and hydroxytyrosol, a phenolic compound obtained from extra-virgin olive oil by-products, were loaded into innovative nanocarriers. These were designed ad hoc combining exopolysaccharides from the cyanobacteria Neocyanospira capsulata, and a lipid component, i.e., egg phosphatidylcholine. The polysaccharide was chosen for chemical affinity with the chitin of insect exoskeleton, while the lipids were introduced to modulate the carrier rigidity. The newly formed nanosystems were characterized by physico-chemical techniques and tested for their possible use in pest control programs. The Mediterranean Fruit Fly Ceratitis capitata Wiedemann, 1824 (Diptera, Tephriditae), a pest of the Mediterranean Region causing high economic losses, was used as a model insect. We found that the nanoformulations nanocarriers prepared in this work, were able to increase the ovicidal effect of hydroxytyrosol. Moreover, the formulation encapsulating either hydroxytyrosol or capsaicin were able to reduce the number of females landing on treated apricots.Graphical

Fabrication of egg white protein/phosphatidylcholine-EGCG co-assembled nanoparticles with sustained release in simulated gastrointestinal digestion and their transcellular permeability in Caco-2 cultures

Epigallocatechin-3-gallate (EGCG) is a kind of polyphenol with various biological activities but poor stability and bioavailability. The encapsulation of EGCG in a nano-delivery system effectively improves its shortcomings. In this research, co-assembled nanoparticles of egg white protein, phosphatidylcholine (PC), and EGCG (EW/PE1:5 NPs) were prepared by antisolvent precipitation to improve their bioavailability. The constructed EW/PE1:5 NPs had an average particle size of 172 nm, a ζ-potential of −26.1 mV, and a polymerization dispersion index (PDI) of 0.33. The encapsulation efficiency (EE) of EGCG was 88.09% and the loading capacity (LC) was 20.10%. UV spectra, FTIR, XRD and intermolecular force results indicated that hydrophobic, electrostatic and hydrogen bonding interactions contributed to the formation of EW/PE1:5 NPs. EW/PE1:5 NPs exhibited Fick diffusion and sustained release characteristics in simulated gastrointestinal digestion. In addition, the transcellular permeability of EW/PE1:5 NPs was significantly increased compared with free EGCG. At the same time, EW/PE1:5 NPs increased EGCG absorption transport and inhibited MRP2 and P-gp-mediated efflux transport. These results suggest that EW/PE1:5 NPs play a key role in improving the bioavailability of EGCG. Therefore, EW/PE1:5 NPs are expected to provide a theoretical basis for the design and development of functional foods or dietary supplements that synergistically enhance the bioavailability of EW, PC and other flavonoids.

Abstract 1999: Repurposing an antibiotic for breast cancer treatment using liposomes as a delivery system

Abstract Purpose: Scientific community is always searching for alternatives for cancer treatment and promoting drug repurposing is one of the ways. Rifamycin is a well-known antibiotic, that inhibits RNA synthesis and probably can be explored as cytotoxic agent. Thus, our goal was to repurpose rifamycin for breast cancer treatment using liposomes as a delivery system. Methods: Liposomes were produced by thin-film evaporation method using egg phosphatidylcholine, cholesterol and DOPE-PEG5k (70:25:5, molar ratio). Non-encapsulated drug was removed using dialysis bags and encapsulated drug was directly determined by UV spectrophotometer. Liposomes were characterized regarding size and surface charge by dynamic light scattering. Liposomes were stored at 4°C and 25°C for stability evaluation, monitoring nanoparticles parameters and drug encapsulation. Formulation biocompatibility was evaluated by hemolysis assay. Finally, cytotoxicity of rifamycin SV and liposomes was evaluated in MCF-7 and MDA-MB-231 cell lines. Results: Blank liposomes (without drug) and rifamycin-loaded liposomes showed size smaller than 100 nm and polydispersity around 0.2, with slightly negative zeta potential. Rifamycin encapsulation in liposomes was 3mg of drug/10mg of lipid films. Both formulations were stable for at least 21 days when stored at 4°C, keeping above 60% of drug encapsulated. In the concentrations tested (0.5mg/mL of drug, 2mg/mL of lipids), liposomes formulation (blank and loaded) and free rifamycin were biocompatible and non-hemolytic, since hemolysis was below 6%. Blank liposomes were not cytotoxic in tested cell lines. Free rifamycin and rifamycin-loaded liposomes demonstrated the IC50 in the same range between 41 and 46 µg/mL in 24h and 48h for both MCF-7 and MDA-MB-231. Conclusions: Rifamycin was successfully encapsulated in liposomes, leading to a stable, biocompatible, and non-hemolytic formulation. The liposomal rifamycin demonstrated strong cytotoxicity towards cancer cells similar to that of the free drug. Citation Format: Janaina Artem Ataide, Nina Filipczak, Satya Siva Kishan Yalamarty, Priscila Gava Mazzola, Vladimir P. Torchilin. Repurposing an antibiotic for breast cancer treatment using liposomes as a delivery system [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1999.

Emetic Tartar-Loaded Liposomes as a New Strategy for Leishmaniasis Treatment.

Emetic tartar (ET), was used in the treatment of leishmaniasis but its use was discontinued due to its low therapeutic index. Liposomes have been shown to be a promising strategy for delivery of bioactive substances in the region of interest, in order to reduce and/or eliminate undesirable effects. In the present study, liposomes containing ET were prepared and characterized to evaluate acute toxicity as well as their leishmanicidal action using BALB/c mice with an inoculum of Leishmania (Leishmania) infantum. Liposomes were composed of egg phosphatidylcholine and 3ß-[N-(N',N'-dimethylaminoethane)-carbamoyl]cholesterol, with an average diameter of 200 nm, zeta potential of +18 mV, and ET encapsulated into liposomes at a concentration near 2 g/L. Healthy mice were treated with ET or liposome containing ET (Lip-ET) in a single dose of 16 mg/kg of Sb3+ intravenously and observed for 14 days. The death of two animals in the ET-treated group and no deaths in the Lip-ET-treated group was observed. Higher hepatic and cardiac toxicity were observed in animals treated with ET when compared to animals treated with Lip-ET, blank liposomes (Blank-Lip) and PBS. The study of antileishmanial efficacy was conducted by intraperitoneal administration of Lip-ET, for ten consecutive days. It was observed by limiting dilution that treatments with liposomal formulations containing ET, as well as Glucantime®, led to a significant reduction in parasitic load in spleen and liver (p < 0.05) when compared to the untreated control group.

Lipids in Mitochondrial Macroautophagy: Phase Behavior of Bilayers Containing Cardiolipin and Ceramide

Cardiolipin (CL) is a key lipid for damaged mitochondrial recognition by the LC3/GABARAP human autophagy proteins. The role of ceramide (Cer) in this process is unclear, but CL and Cer have been proposed to coexist in mitochondria under certain conditions. Varela et al. showed that in model membranes composed of egg sphingomyelin (eSM), dioleoyl phosphatidylethanolamine (DOPE), and CL, the addition of Cer enhanced the binding of LC3/GABARAP proteins to bilayers. Cer gave rise to lateral phase separation of Cer-rich rigid domains but protein binding took place mainly in the fluid continuous phase. In the present study, a biophysical analysis of bilayers composed of eSM, DOPE, CL, and/or Cer was attempted to understand the relevance of this lipid coexistence. Bilayers were studied by differential scanning calorimetry, confocal fluorescence microscopy, and atomic force microscopy. Upon the addition of CL and Cer, one continuous phase and two segregated ones were formed. In bilayers with egg phosphatidylcholine instead of eSM, in which the binding of LC3/GABARAP proteins hardly increased with Cer in the former study, a single segregated phase was formed. Assuming that phase separation at the nanoscale is ruled by the same principles acting at the micrometer scale, it is proposed that Cer-enriched rigid nanodomains, stabilized by eSM:Cer interactions formed within the DOPE- and CL-enriched fluid phase, result in structural defects at the rigid/fluid nanointerfaces, thus hypothetically facilitatingLC3/GABARAP protein interaction.

Administration of Phosphatidylcholine Increases Brain Acetylcholine Concentration and Improves Memory in Mice with Dementia

Studies on the effect of phosphatidylcholine administration on memory are limited. We administered egg phosphatidylcholine to mice with dementia and to normal mice and compared the differences in memory and serum choline concentration, and choline and acetylcholine concentrations and choline acetyltransferase activities of three forebrain regions (cortex, hippocampus and the remaining forebrain). Mice with dementia were produced by mating sibling mice who had impaired memory for >20 generations. These mice had poor memory and low brain acetylcholine concentration. We administered 100 mg of egg phosphatidylcholine (phosphatidylcholine group) or water (control group) by gavage to each mouse daily for about 45 d. Control mice with dementia had poorer memory in passive avoidance performance and lower brain choline (cortex and hippocampus) and acetylcholine (hippocampus and forebrain excluding cortex and hippocampus) concentrations and lower cortex choline acetyltransferase activity than the control normal mice (P < 0.05). The administration of phosphatidylcholine to mice with dementia improved memory and generally increased brain choline and acetylcholine concentrations to or above the levels of the control normal mice. In normal mice, phosphatidylcholine treatment did not affect memory or acetylcholine concentrations in spite of the great increase in choline concentrations in the three brain regions. Serum choline concentration in mice treated with phosphatidylcholine increased to a similar level in both strains of mice, indicating that the absorption of phosphatidylcholine was not impaired in mice with dementia. The results suggest that administration of egg phosphatidylcholine to mice with dementia increases brain acetylcholine concentration and improves memory.

Development of a Technique for Chromatographic Analysis of Excipients in a Liposomal Dosage Form of a Photosensitizer Tetra-3-phenylthio-phthalocyanine Hydroxyaluminium

Introduction. The Laboratory of Drug Formulation Development at the N. N. National Medical Research Center of Oncology developed a liposomal dosage form of FS tetra-3-phenylthio-pthalocyanine hydroxyluminide (TPHA) for which a simple, accurate and easily reproducible method of quantitative determination of the main active substance and excipients in the drug product was needed. High-performance thin-layer chromatography (HPTLC) was chosen as such a technique.Aim. The aim of this work was to develop chromatographic methods for the quantitative determination of excipients in a liposomal dosage form of a tetra-3-phenylthiophthalocyanine hydroxyaluminium (TPHA).Materials and methods. Determination of substances was carried out by the method of high-performance thin-layer chromatography (HPTLC) followed by TLC-densitometry. Substance TPHA 98 % (Federal State Unitary Enterprise "SSC "NIOPIK", Russia), placebo lyophilized liposomal form without TFHA in the composition (FSBI "National Medical Research Center of Oncology. N. N. Blokhin" of the Ministry of Health of Russia), egg phosphatidylcholine (96 %, CAS № 97281-44-2, EPC Lipoid, Germany), cholesterol (≥99 %, CAS № C8667, Sigma-Aldrich, Japan), sucrose analytical grade (CHIMMED, Russia) and various organic solvents.Results and discussion. The selected mobile phase for HPTLC analysis: isopropanol: ammonia 25 % aqueous 80 : 20, separation time was 20– 25 min (l = 75mm), t = 20 °С. To visualize the spots of sucrose, phosphatidylcholine, and cholesterol, a 5 % solution of phosphomolybdic acid was chosen. The stability of the standard samples was confirmed on a series of prevalidation studies. Validation of the methodology was carried out.Conclusion. For quantitative determination of sucrose, phosphatidylcholine and cholesterol in a new liposomal dosage form, the method of HPTLC with TLC-densitometry was chosen, which allows identifying all three components in the dosage form simultaneously. The method was validated.

Optimization using response surface methodology of phospholipase C production from Bacillus cereus suitable for soybean oil degumming

This work deals with the optimization of an extracellular phospholipase C production by Bacillus cereus (PLCBc) using Response Surface Methodology (RMS) and Box-Behnken design. In fact, after optimization, a maximum phospholipase activity (51 U/ml) was obtained after 6 h of cultivation on tryptone (10 g/L), yeast extract (10 g/L), NaCl (8.125 g/L), pH 7.5 with initial OD (0.15). The PLCBc activity, esteemed by the model (51 U) was very approximate to activity gutted experimentally (50 U). The PLCBc can be considered as thermoactive phospholipase since it showed a maximal activity of 50 U/mL at 60 °C using egg yolk or egg phosphatidylcholine (PC) as substrate. In addition, the enzyme was active at pH 7 and is stable after incubation at 55 °C for 30 min. The application of B. cereus phospholipase C in soybean oil degumming was investigated. Our results showed that when using enzymatic degumming, the residual phosphorus decrease more than with water degumming, indeed, it passes from 718 ppm in soybean crude oil to 100 ppm and 52 ppm by degumming using water and enzymatic process, respectively. The diacylgycerol (DAG) yield showed an increase of 1.2% with enzymatic degumming compared to soybean crude oil. This makes our enzyme a potential candidate for food industrial applications such as enzymatic degumming of vegetable oils.