Egg Phosphatidylcholine Liposomes Research Articles

Chimeric liposomes decorated with P407: an alternative biomaterial for producing stealth nano-therapeutics

The aim of the present study is the development and evaluation of the physicochemical properties of chimeric hydrogenated soya phosphatidylcholine (HSPC) and egg phosphatidylcholine (EggPC) liposomes with incorporated triblock copolymer Poloxamer P407 (P407). The physicochemical assay was held in water HPLC-grade and Foetal Bovine Serum (FBS), in order to determine whether these systems can be used as drug or antigen delivery nanosystems. Dynamic and electrophoretic light scattering (DLS/ELS) techniques were used for the measurement of the hydrodynamic diameter, the polydispersity index, and the ζ-potential of the prepared nanosystems. The incorporation of the P407 resulted in a size reduction of all systems. A decrease in the hydrodynamic diameter and polydispersity index were also found as a result of increasing the storage temperature from 4 °C to 25 °C, attributed to P407. The experiments that were carried out in FBS, showed that the addition of P407 improved systems stealth properties. Concluding, we propose P407 as a promising alternative to PEG in the development of lipid nanoparticles with optimized bio- and shelf-stability.

Modeling the saturation of detergent association in mixed liposome systems

Cells tune the lipid types present in their membranes to adjust for thermal and chemical stability, as well as to promote association and dissociation of small molecules and bound proteins. Understanding the influence of lipid type on molecule association would open doors for targeted cell therapies, in particular when molecular association is observed in the presence of competing membranes. For this reason, we modeled and experimentally observed the association of a small molecule with two membrane types present by measuring the association of the detergent Triton X-100 with two types of liposomes, egg phosphatidylcholine (ePC) liposomes and egg phosphatidic acid (ePA) liposomes, at varying ratios. We called this mixed liposomes, as each liposome population was formed from a different lipid type. Absorbance spectrometry was used to observe the stages of detergent association with mixed liposomes and to determine the detergent concentration at which the liposomes were fully saturated. A saturation model was also derived that predicts the detergent associated with each liposome type when the lipid bilayers are fully saturated with detergent. The techinical input parameters for the model are the detergent to lipid ratio and the relative absorbance intensity for each of the pure liposome species at saturation. With that, the association of detergent with any mixture of those liposome types at saturation can be determined.

Interactions of natural deep eutectic solvents (NADES) with artificial and natural membranes

Natural deep eutectic solvents (NADES) are a third type of liquids, apart from water and lipids. NADES show promise as drug carriers in dermal formulations or formulations for local administration in the oral cavity due to their low toxicity, tunability and biodegradability as well as their solubilizing and stabilizing properties. However, a thorough investigation of their effects in a biological system is needed. The present study aimed to investigate the physicochemical properties of six selected NADES and dilutions thereof. Further, the suitability of NADES as vehicles in topical preparations was evaluated through the application of liposome model membranes and ex vivo pig skin models. The effects of NADES on the membrane stability of egg-phosphatidylcholine (Egg-PC) liposomes and 1,2-dipalmitoyl-sn-glycero-3-phosphocholine 16:0 PC (DPPC) liposomes were investigated by physical stability studies, membrane permeability studies, calorimetric analysis and visual representation by transmission electron microscopy (TEM). The liposomes appeared in most cases physically stable for 24 h in NADES, although there were changes in size and morphology most likely related to increased osmolality. Some NADES were also able to disaggregate formed DPPC liposome aggregates. The ex vivo permeation studies indicated a slightly reduced permeability of chloramphenicol through the pig skin after NADES application, which most likely was caused by hydrogen bonding between the NADES and proteins in the skin based on FT-IR-results. Altogether, our findings suggest that NADES are promising drug carriers in new drug delivery systems for topical applications.

P.725 ACC neurometabolites in Bipolar Disorder are influenced by mood state and medication: a systematic review and meta-analysis of 1H-MRS studies

Local anesthetics promote analgesia by interacting with excitable membranes. Articaine (ATC) has a unique composition among local anesthetics as it possesses a thiophene instead of the typical phenyl ring. Aiming to characterize the interaction of neutral articaine (nATC) with phospholipid membranes, we have employed a synergistic approach of experimental and computational techniques. Fluorescence measurements supported nATC partitioning into the membranes, since its intrinsic fluorescence anisotropy increased from 0.03 in water to 0.29 in the presence of egg phosphatidylcholine (EPC) liposomes, and the fluorescence of AHBA, a probe that monitors the water-membrane interface, was quenched by nATC. 1H NMR experiments revealed changes in the chemical shifts of articaine and EPC hydrogens after partitioning, and shorter T1 values of nATC hydrogens when inserted into the EPC vesicles. Contacts of nATC and the phospholipid polar head group were inferred from 2D-NOE. Taken together, these results indicate a superficial insertion of the nATC molecules inside EPC bilayers. This conclusion was confirmed by molecular dynamics simulations, which allowed the identification of the key interactions underlying the preferential location of nATC in the bilayer. Contrary to what is often stated (that articaine is a high lipophilic local anesthetic agent) our results place ATC among the hydrophilic ones, such as lidocaine, prilocaine, and mepivacaine, for which the water/membrane interface is the preferred location.

A Phospholipase-A Activity in Soluble Leishmania Antigens Causes Instability of Liposomes.

This study aimed to investigate the existence of phospholipase-A (PLA) activity in Soluble L. major Antigens (SLA) because of no reports for it so far. Liposomes were used as sensors to evaluate PLA activity. Liposomal SLA consisting of Egg Phosphatidylcholine (EPC) or Sphingomyelin (SM) were prepared by two different methods in different pH or temperatures and characterized by Dynamic Light Scattering (DLS) and Thin Layer Chromatography (TLC). Lipid hydrolysis led to the disruption of EPC liposomal SLA in both methods but the Film Method (FM) produced more stable liposomes than the Detergent Removal Method (DRM). The preparation of EPC liposomal SLA at pH 6 via FM protected liposomes from hydrolysis to some extent for a short time. EPC liposomes but not SM liposomes were disrupted in the presence of SLA. Therefore, a phospholipid without ester bond such as SM should be utilized in liposome formulations containing PLA as an encapsulating protein.

Articaine interaction with phospholipid bilayers

Local anesthetics promote analgesia by interacting with excitable membranes. Articaine (ATC) has a unique composition among local anesthetics as it possesses a thiophene instead of the typical phenyl ring. Aiming to characterize the interaction of neutral articaine (nATC) with phospholipid membranes, we have employed a synergistic approach of experimental and computational techniques. Fluorescence measurements supported nATC partitioning into the membranes, since its intrinsic fluorescence anisotropy increased from 0.03 in water to 0.29 in the presence of egg phosphatidylcholine (EPC) liposomes, and the fluorescence of AHBA, a probe that monitors the water-membrane interface, was quenched by nATC. 1H NMR experiments revealed changes in the chemical shifts of articaine and EPC hydrogens after partitioning, and shorter T1 values of nATC hydrogens when inserted into the EPC vesicles. Contacts of nATC and the phospholipid polar head group were inferred from 2D-NOE. Taken together, these results indicate a superficial insertion of the nATC molecules inside EPC bilayers. This conclusion was confirmed by molecular dynamics simulations, which allowed the identification of the key interactions underlying the preferential location of nATC in the bilayer. Contrary to what is often stated (that articaine is a high lipophilic local anesthetic agent) our results place ATC among the hydrophilic ones, such as lidocaine, prilocaine, and mepivacaine, for which the water/membrane interface is the preferred location.

Liposomal Formulation of a Melphalan Lipophilic Prodrug: Studies of Acute Toxicity, Tolerability, and Antitumor Efficacy.

Recently we developed a scalable scheme of synthesis of melphalan ester conjugate with 1,2-dioleoyl-sn-glycerol (MlphDG) and a protocol for the fabrication of its lyophilized liposomal formulation. Herein we compared this new convenient in use formulation of MlphDG with parent drug Alkeran® in rats concerning several toxicological parameters and evaluated its antitumor efficacy in the model of breast cancer in mice. Liposomes of approximately 100 nm in diameter, consisting of egg phosphatidylcholine, soybean phosphatidylinositol, and MlphDG, or placebo liposomes without the drug were produced by extrusion and lyophilized. Alkeran® or liposomes recovered by the addition of water were injected into the tail vein of animals. Clinical examination of rats consisted of detailed inspection of the behavior, general status, and hematological parameters. Mice with transplanted breast cancer WNT-1 were subjected to multiple treatments with the drugs; tumor growth inhibition was assessed, together with cellular immunity parameters. Liposomes showed approximately two times lower acute toxicity and better tolerability than Alkeran® in terms of behavioral criteria. The toxic effects of liposomes on hemopoiesis were manifested at higher doses than in the case of Alkeran®, proportionally to the difference in LD50 values. The formulation inhibited tumor growth significantly more effectively than Alkeran®, delaying the start of the exponential growth phase and exhibiting no additional toxic effects toward bone marrow. Lower toxicity of the liposomal formulation of MlphDG promises improved quality of life for cancer patients in need of treatment with melphalan. Presumably, the list of indications for melphalan therapy could be extended.

Lipophilic Prodrug of Methotrexate in the Membrane of Liposomes Promotes Their Uptake by Human Blood Phagocytes.

Previously, we showed that incorporation of methotrexate (MTX) in the form of a lipophilic prodrug (MTXDG) in 100-nm lipid bilayer liposomes of egg phosphatidylcholine can allow one to reduce toxicity and improve the antitumor efficiency of MTX in a mouse model of T-cell leukemic lymphoma. However, in our hemocompatibility tests in vitro, MTX liposomes caused complement (C) activation, obviously due to binding on the liposome surface and fragmentation of the C3 complement factor. In this work, we studied the interactions of MTX liposomes carrying stabilizing molecules phosphatidylinositol (PI), ganglioside GM1, or a lipid conjugate of N-carboxymethylated oligoglycine (CMG) in the bilayer with subpopulations of human blood leukocytes. Liposomes labeled with BODIPY-phosphatidylcholine were incubated with whole blood (30 min and 1 h, 37°C), blood cells were lysed with a hypotonic buffer, and the fluorescence of the liposomes bound but not internalized by the leukocytes was quenched by crystal violet. Cell suspensions were analyzed by flow cytometry. Incorporation of MTXDG dramatically enhanced the phagocytosis of liposomes of any composition by monocytes. Neutrophils consumed much less of the liposomes. Lymphocytes did not accumulate liposomes. The introduction of PI into MTX liposomes practically did not affect the specific consumption of liposomes by monocytes, while CMG was likely to increase the consumption rate regardless of the presence of MTXDG. The GM1 ganglioside presumably shielded MTX liposomes from phagocytosis by one of the monocyte populations and increased the efficiency of monocyte uptake by another population, probably one expressing C3b-binding receptors (C3b was detected on liposomes after incubation with blood plasma). MTX liposomes were shown to have different effects on TNF-α production by activated leukocytes, depending on the structure of the stabilizing molecule.

Anaesthetic benefits of a ternary drug delivery system (Ropivacaine-in-Cyclodextrin-in-Liposomes): in-vitro and in-vivo evaluation.

To evaluate whether a ternary system composed of hydroxypropyl-β-cyclodextrin (HP-βCD) further encapsulated into egg phosphatidylcholine liposomes (LUV) could prolong the action and reduce the toxicity of ropivacaine (RVC). Dynamic light scattering and NMR were used to characterize the inclusion complex (RVC:HP-βCD), liposomal (RVC:LUV) and ternary (LUV:RVC:HP-βCD) systems containing 0.25% RVC. Their encapsulation efficiency, release kinetics, in-vitro cytotoxicity and in-vivo anaesthetic effect (paw-withdraw tests in mice) were also evaluated. 1:1 RVC:HP-βCD inclusion complex was encapsulated in liposomes (220.2±20.3nm size, polydispersity <0.25, zeta potentials=-31.7±1.4mV). NMR (diffusion-ordered spectroscopy (DOSY)) revealed stronger anaesthetic binding to LUV:RVC:HP-βCD (Ka =342m-1 ) than to RVC:HP-βCD (Ka =128m-1 ) or liposomal formulation (Ka =22m-1 ). The formulations promoted in-vitro sustained drug release and partially reverted the cytotoxicity of RVC against 3T3 fibroblasts in the profile: LUV:RVC:HP-βCD≥RVC:HP-βCD>RVC:LUV. Accordingly, in-vivo sensory block of free RVC (180min) was prolonged ca. 1.7 times with the ternary system and RVC:HP-βCD (300min) and 1.3 times with RVC:LUV (240min). These results confirm the suitability of this double-carrier system in clinical practice, to decrease the toxicity and prolong the anaesthesia time evoked by RVC.

Liposomes incorporating cinnamoyl gelatin and cinnnamoyl alginate and their pH and UV-responsive release property

pH and UV-responsive liposome was prepared by decorating the surface of egg phosphatidylcholine (EPC) liposome using cinnamoyl gelatin (CinGel) and cinnamoyl alginate (CinAlg). CinGel and CinAlg were prepared through condensation reaction between cinnamoyl chloride and each polymer. Liposomes bearing CinGel/CinAlg were prepared by a film hydration method while CinGel/CinAlg mass ratio was kept to be 7:3. The fluorescence quenching of carboxyfluorescein (CF) encapsulated in the liposome and the diameter of the liposomes decreased as the EPC to cinnamoyl polymer mass ratio increased. Liposome without cinnamoyl polymers hardly released its payload for 120 sec at all the pH values tested (pH 5.0, pH 6.0, pH 7.4, and pH 9.0). Meanwhile, liposome with CinGel/CinAlg released its payload markedly at pH 5.0 and pH 6.0. The complex coacervation was thought to cause the release of dye out of the liposomes. Liposome scarcely released its payload at pH 7.4 for 120 min without UV irradiation regardless of whether it included the cinnamoyl polymers. Meanwhile, liposome with cinnamoyl polymers showed marked release under UV irradiation.

Phospholipid multilamellar vesicles entrapping phenothiazine photosensitizers. Preparation, characterization and evaluation of their photodynamic properties

The aim of the present paper was to optimize the properties of phenothiazine photosensitizers (PSs), Azure B and its monobrominated derivative. They were entrapped in multilamellar liposomes of egg phosphatidylcholine (EPC) and dipalmitoylphosphatidylcholine (dpPC). The higher partition coefficient of both PSs in the EPC/water system compared with that of both PSs in the dpPC/water system allowed the selection of EPC to be used for the development of third generation photosensitizers. The optimal phospholipid/phenothiazine ratio and the location of these dyes in the lipid bilayer systems were established. These dyes were found to be located within the polar head group region of the lipid bilayer, which would enhance the effectiveness of the irradiation procedure. In addition, the encapsulation of both photosensitizers in liposomes led to a decrease in the aggregation of these compounds, and consequently to an increase in their singlet oxygen quantum yield (ϕΔ). The encapsulation of the PSs in lipid vesicles significantly increased their photochemical reactivity, doubling the ϕΔ value of AzBBr and increasing that of AzB by 60%. The results obtained demonstrate that the vehiculization of these PSs in liposomes allowed the development of third generation photosensitizers with better photochemical properties to be used in potential therapeutic applications.

Orientation assignment of LH2 and LH1-RC complexes from Thermochromatium tepidum reconstituted in PC liposome and their ultrafast excitation dynamics comparison between in artificial and in natural chromatophores

In this study, the artificial chromatophore was constructed by reconstitution of light-harvesting complex 2 (LH2) and core-complex (LH1-RC) from Thermochromatium tepidum in egg phosphatidylcholine (PC) liposome. By calcium ion dependent absorption changes, the orientations of protein complexes in liposomes solely were assigned. Ultrafast EET dynamics were studied by fs time-resolved absorption spectroscopy on both the artificial and natural chromatophores at their RC ‘open’ and ‘close’ states, respectively. The results indicated that these two membrane systems have similar EET dynamic profiles, and the orientations of LH2 and LH1-RC complexes in artificial chromatophores were opposite to those in natural chromatophores.

Revealing Dynamics of Accumulation of Systemically Injected Liposomes in the Skin by Intravital Microscopy.

Accumulation of intravenously injected cytotoxic liposomes in the skin induces serious toxicity. We used single time point and longitudinal intravital microscopy to understand skin accumulation dynamics of non-PEGylated and PEGylated liposomes after systemic injection into mice. Non-PEGylated egg phosphatidylcholine (PC) liposomes showed short circulation half-life (1.3 h) and immediate aggregation in the blood, with some aggregates lodging in skin microvasculature soon after the injection. At 24 h, and more prominently at 48 h postinjection, liposomes appeared in dermal and subdermal cells. PEGylated egg PC liposomes showed long circulation half-life (22 h) and no aggregation in the blood. PEGylated liposomes started to accumulate in the skin microvasculature as soon as 5 min after the injection. Within 3 h postinjection, PEGylated liposomes accumulated in extravascular cells in the dermis and subdermis. Liposomes were present in the skin for at least 7 days postinjection. A regulatory approved PEGylated liposomal doxorubicin (LipoDox) and empty liposomes of the same composition as LipoDox showed similar skin distribution as PEGylated egg PC liposomes, suggesting that this phenomenon is relevant to liposomes of different lipid composition. Decorating liposomes with shorter PEGs (350 or 700) in addition to PEG 2000 did not decrease the deposition. Outside the capillaries, liposomes partially colocalized with CD45-, F4/80+ cells. The accumulation of liposomes was not due to prior neutrophil/platelet binding and transport across endothelium. Moreover, our studies have excluded a role of complement in the skin accumulation of liposomes. Further understanding of mechanisms of this important phenomenon can improve the safety of liposomal nanocarriers.

Bioactivity In Vitro of Quercetin Glycoside Obtained in Beauveria bassiana Culture and Its Interaction with Liposome Membranes.

Quercetin (Q) was used as substrate for regioselective glycosylation at the C-7 position catalyzed by Beauveria bassiana AM278 strain. As a result the glycoside quercetin 7-O-β-d-(4″-O-methyl)glucopyranoside (Q 7-MeGlu) was formed. The goal of the studies was to determine the anti-oxidative (liposome membrane protection against free radicals IC50Q 7-MeGlu = 5.47 and IC50Q = 4.49 µM) and anti-inflammatory (COX-1 and COX-2 enzymes activity inhibition) properties of Q 7-MeGlu as compared to Q. Every attempt was made to clarify the antioxidant activity of these molecules, which are able to interact with egg phosphatidylcholine liposomes, using a fluorometric method (by applying the probes MC540, TMA-DPH and DPH). The results indicated that Q 7-MeGlu and Q are responsible for increasing the packing order, mainly in the hydrophilic but also in hydrophobic regions of the membrane (Q > Q 7-MeGlu). These observations, confirmed by a 1H-NMR method, are key to understanding their antioxidant activity which is probably caused by the stabilizing effect on the lipid membranes. The results showed that Q 7-MeGlu and Q have ability to quench the human serum albumin (HSA) intrinsic fluorescence through a static quenching mechanism. The results of thermodynamic parameters indicated that the process of formation complexes between studied molecules and HSA was spontaneous and caused through Van der Waals interactions and hydrogen bonding.

Preparation of liposome bearing disulfide proteinoid and its reduction-responsive release property

Egg phosphatidylcholine (Egg PC) liposome bearing a disulfide proteinoid exhibited a reduction-responsive release property. Proteinoid composed of Asp, Leu, and cystamine (Prot(ALC)) and Asp and Leu (Prot(AL)) were synthesized by thermal condensation, confirmed by Raman, FT-IR, 1H NMR and 13C NMR spectroscopy. Egg PC liposome bearing the proteinoid was prepared by a film hydration and sonication method. The fluorescence quenching of dye (i.e. calcein) loaded in liposome bearing the proteinoid was 68.1–78.1%. The mean hydrodynamic diameter of liposome bearing the proteinoid was less than 200 nm and it decreased with increasing amount of the proteinoid. On the TEM photo, multi-lamellar vesicles were observed and the vesicle diameter was 100–300 nm. At all the phospholipid to proteinod ratios tested (i.e. 1:0.01, 1:0.02, and 1:0.05 (w/w)), the release degree in 12 h of dye loaded in liposome bearing Prot(AL) was less than 2%, and it was almost the same regardless of dithiothreitol (DTT, a reducing agent) concentration (i.e. 0, 10, and 20 mM). Whereas the release degree of dye loaded in liposome bearing Prot(ALC) was 1.5–8.4% and it was significantly dependent on DTT concentration.

Effects of flavonol glycosides on liposome stability during freezing and drying

Flavonoids are a large and diverse group of plant secondary metabolites that are mainly present as glycosides. They are often accumulated in response to abiotic stresses such as UV radiation, drought, cold and freezing. The most extensively studied function of flavonoids is their antioxidant activity although their importance as antioxidants in plants has been questioned. We therefore aim to study effects of flavonols on cellular stress tolerance that are independent of their antioxidant function. Here we investigate the effects of the glycosylated flavonols kaempferol-3-O-glucoside, kaempferol-7-O-glucoside, quercetin-3-O-glucoside and quercetin-3-O-rhamnoside on liposome stability after freezing and drying. Insertion of flavonols in lipid bilayers destabilized egg phosphatidylcholine (EPC) liposomes and to a lesser extent vesicles made from equal proportions of EPC and egg phosphatidylethanolamine (EPE) during a freeze-thaw cycle, while liposomes containing the unsaturated non-bilayer lipid 18:2 PE were either unaffected or slightly stabilized. In general, the kaempferol derivatives were more destabilizing for liposomes during freezing than the quercetin derivatives. Fourier-transform infrared spectroscopy revealed that all flavonols were localized in the interfacial region of the lipid bilayers, forming H-bonds with the lipid phosphate and carbonyl groups. The phase transition temperature of dry 16:0/18:1 PC (POPC) and POPC/EPE liposomes was decreased by 75°C and 55°C, respectively. Changes in the vibration bands attributed to the phenolic ring structures of the flavonols in the presence of liposomes provided further evidence of interactions of these molecules in particular with the interfacial region of the bilayers.

In vitro anti-inflammatory efficacies of liposomal suspensions of acetylsalicylic acid

Egg phosphatidylcholine (EPC) liposome containing acetylsalicylic acid (ASA) (EPC/ASA liposome) was prepared by a film hydration and sonication method, and the effect of liposome on the in vitro anti-inflammatory efficacy and the in vitro skin permeation of ASA was investigated. The mean diameter of EPC liposome and EPC/ASA liposome was about 271 and 175 nm, respectively. Both of liposomes were multi-lamellar vesicles on transmission electron microscopy photos. The in vitro viability of cell (Raw264.7) treated with EPC/ASA liposome suspension was significantly lower than the viability of cell treated with ASA solution. The amount of nitrite and prostaglandin E2 produced by cell treated with EPC/ASA liposome suspension was significantly lower than the amount produced by cells treated with ASA solution, indicating EPC liposome boosted the anti-inflammatory efficacy of ASA. The amount of ASA permeated through hairless mouse skin was inappreciable for 24 h when ASA solution was applied, whereas the permeation amount markedly increased to about 185 μg/cm2 for 24 h when EPC/ASA liposome suspension was applied. EPC liposome also enhanced the permeation into stratum corneum and epidermis/dermis.

Reduction-responsive release property of egg phosphatidylcholine liposomes incorporating benzyl disulfide

Reduction-responsive liposome was prepared by incorporating benzyl disulfide in egg phosphatidylcholine (egg PC) liposomal bilayer. The fluorescence quenching degree of calcein enveloped in egg PC liposome bearing benzyl disulfide decreased from 62.7% to 49.2% and the mean hydrodynamic diameter of the liposome decreased from 276nm to 206nm when the egg PC to benzyl disulfide weight ratio increased from 20:0 to 20:10. The liposomes bearing benzyl disulfide were multi-lamellar vesicles on TEM photos. It was confirmed by Raman spectroscopy that benzyl disulfide was loaded in the liposomal bilayer. The release degree of calcein enveloped in liposome incorporating benzyl disulfide was investigated for 24h at 25°C, 37°C, and 45°C, when the concentration of dl-dithiothreitol (DTT) was 0mM, 5mM, 10mM and 30mM. At all the DTT concentrations tested, the release degree was more extensive as the temperature was higher. At all the temperatures tested, the release degree became more extensive as the DTT concentration increased. For example, the maximum release degree at 37°C of calcein loaded in liposome of which egg PC to benzyl disulfide weight ratio was 20:5 increased from 2.9% to 14.8% when DTT concentration increased from 0mM to 10mM. DTT could reduce benzyl disulfide to benzyl thiols and the fragments of benzyl disulfide would re-orient due to its polarity change in the liposomal membrane, resulting in the membrane fluctuation and the promoted release. At all the temperatures and all the DTT concentrations tested, the release was more sensitive to the DTT concentration as the benzyl disulfide content in liposome was higher, indicating that the reduction of the disulfide compound was responsible for the promoted release.

Methylglyoxal as a scavenger for superoxide anion-radical.

Methylglyoxal at a concentration of 5 mM caused a significant inhibition of superoxide anion radical (O2 (·-)) comparable to the effect of Tirone. In the process of O2 (·-) generation in the system of egg phosphatidylcholine liposome peroxidation induced by the azo-initiator AIBN, a marked inhibition of chemiluminescence in the presence of 100 mM methylglyoxal was found. At the same time, methylglyoxal did not inhibit free radical peroxidation of low-density lipoprotein particles, which indicates the absence of interaction with methylglyoxal alkoxyl and peroxyl polyenoic lipid radicals. These findings deepen information about the role of methylglyoxal in the regulation of free radical processes.

Tripolyphosphate-sensitive egg phosphatidylcholine liposomes incorporating hydrophobically modified poly(ethylene imine)

ABSTRACTLiposomes, which release their contents in answer to tripolyphosphate (TPP, a penta-anion), were prepared by immobilizing hydrophobically modified poly(ethylene imine) (HmPEI) on the surface of egg phosphatidylcholine (egg PC) liposome. HmPEI was prepared by covalently attaching decanoyl chloride to PEI through a condensation reaction. According to the 1H NMR spectrum, the number of decanoyl chloride per one molecule of PEI was about 21, and HmPEI was air/water interface-active. HmPEI could readily complex with TPP in HEPES buffer (30 mM, pH 7.0), confirmed by Fourier transformed infrared spectrophotometer spectroscopy. The complexation increased with increasing the concentration of HmPEI and TPP, investigated through the measurement of optical density and light scattering intensity. Liposomes incorporating HmPEI were prepared by a film hydration and sonication method. The liposomes were multi-lamellar vesicles, observed on transmission electron microscope. Liposomes free of HmPEI did not release calcien when they were mixed with TPP. Liposomes whose egg PC/HmPEI was relatively low (e.g., 20:1 and 20:2) released calcein but not extensively (less than 10%) when mixed with TPP. Liposomes whose egg PC/HmPEI was relatively high (e.g., 20:4 and 20:20) released calcein extensively. For example, when the liposomes with lager amount of HmPEI were mixed with TPP so that HmPEI/TPP weight ratio was 8:1, the release degree in 60 sec was more than 70%. HmPEI can complex with TPP through electrostatic interaction and the complexation was thought to cause perturbation in the liposomal membranes and trigger the release.