Formulation of photocleavable liposomes and the mechanism of their content release

In pursuit of designing photocleavable liposomes as drug delivery vehicles, we synthesized several amphiphilic lipids by connecting stearyl amine (as the non-polar tail) and charged amino acids (as polar heads) via the o-nitrobenzyl derivatives. The lipids containing Glu, Asp, and Lys amino acids were subjected to photocleavage reaction by UV light, and the overall spectral changes of the chromophoric o-nitrobenzyl conjugates were determined as a function of time. The experimental data revealed that the feasibility of the cleavage reaction, nature and magnitude of the spectral changes during the course of the cleavage reaction, and their overall kinetic profiles were dictated by the type of amino acid constituting the polar head groups. The cleavage reactions of the Asp and Glu containing lipids were found to be more facile than that of the lysine-containing lipid. Using these lipids, we formulated photocleavable liposomes, and investigated the photo-triggered release of an encapsulated (within the liposomal lumen) dye as a function of time. The kinetic data revealed that the release of the liposomal content conformed to a two-step mechanism, of which the first (fast) step involved the photocleavage of lipids followed by the slow release of the liposomal content during the second step. The overall mechanistic features intrinsic to the photocleavage of Asp, Glu and Lys containing o-nitrobenzyl conjugated lipids, and their potential applications in formulating liposomes (whose contents can be "unloaded" by the UV light) as drug delivery vehicles are discussed.     

Eicosapentaenoic acid release from the red alga Pachymeniopsis lanceolata by enzymatic degradation

Forty-eight species of seaweeds from Japanese waters were screened for the valuable polyunsaturated fatty acids eicosapentaenoic acid (EPA). The eight species that contained the highest levels of these compounds were analyzed in detail. Of all species tested the red alga Pachymeniopsis lanceolata contained the highest EPA concentration, and it was present as both the free and bound forms. EPA constituted 38.7% of total fatty acids, and polar lipids were the main constituent of the total lipids in P. lanceolata. EPA was obtained from the marine algae P. lanceolata by enzymatic hydrolysis of the total lipids extract using phospholipase A₂(PLA₂). The release of EPA reached a plateau after 10 min of enzymatic treatment. These results suggest that P. lanceolata is a useful natural source of EPA and that PLA₂ treatment is a convenient method for obtaining EPA from the red alga.     

Interaction of lipoplexes with anionic lipids resulting in DNA release is a two-stage process

We propose a mechanism for DNA release from lipoplexes in cells that accounts for various observations of lipoplex-anionic lipid interactions. We examined the structural evolution of lipoplexes upon interaction with cellular lipids by synchrotron small-angle X-ray diffraction (SAXD), and the extent of DNA release from lipoplexes was determined by gel electrophoresis. We find that the interaction of lipoplexes with anionic cellular lipids is a two-stage process. In the first step, anionic lipids laterally diffuse into the complex and neutralize the cationic lipids. As a result, the membrane charge density of lipoplexes decreases and interactions between cationic lipids and DNA become weaker, but DNA is extremely poorly released. Only after the cationic charge of lipoplex membranes is completely neutralized by anionic lipids does DNA starts to be released significantly.     

Epoxy and hydroxy acids of the seed oil ofGaleopsis bifida

The lipids of the seeds of the hemp nettle, which is toxic for ruminants have been studied. The composition of the fatty acids of five classes of lipids and the position-species composition of the triacylglycerols have been determined. The presence of epoxyacyl- and hydroxyacylglycerols in the oil and the structures of the acids of these lipids have been established. Squalene has been isolated from the oil.     

Redox-triggered contents release from liposomes

An exciting new direction in responsive liposome research is endogenous triggering of liposomal payload release by overexpressed enzyme activity in affected tissues and offers the unique possibility of active and site-specific release. Bringing to fruition the fully expected capabilities of this new class of triggered liposomal delivery system requires a collection of liposome systems that respond to different upregulated enzymes; however, a relatively small number currently exist. Here we show that stable, approximately 100 nm diameter liposomes can be made from previously unreported quinone-dioleoyl phosphatidylethanolamine (Q-DOPE) lipids, and complete payload release (quenched fluorescent dye) from Q-DOPE liposomes occurs upon their redox activation when the quinone headgroup possesses specific substituents. The key component of the triggerable, contents-releasing Q-DOPE liposomes is a "trimethyl-locked" quinone redox switch attached to the N-terminus of DOPE lipids that undergoes a cleavage event upon two-electron reduction. Payload release by aggregation and leakage of "uncapped" Q-DOPE liposomes is supported by results from liposomes wherein deliberate alteration of the "trimethyl-locked" switch completely deactivates the redox-destructible phenomena (liposome opening). We expect that Q-DOPE liposomes and their variants will be important in treatment of diseases with associated tissues that overexpress quinone reductases, such as cancers and inflammatory diseases, because the quinone redox switch is a known substrate for this group of reductases.     

Detection of bilayer packing stress and its release in lamellar-cubic phase transition by time-resolved fluorescence anisotropy

An introduction of nonlamellar-forming lipids into planar bilayers generates packing stress, which is important for the biological functions of plasma membranes and is a driving force for the lamellar-nonlamellar phase transition. We have investigated the phase behavior of a binary system consisting of egg yolk phosphatidylcholine and monoolein (MO) and the changes in the local orientation order of lipids in a lamellar-bicontinuous cubic phase transition. Small-angle X-ray scattering has revealed that the lamellar-bicontinuous cubic phase transition occurs at an MO molar fraction (X(MO)) between 0.6 and 0.7. These phases were dispersed to form liposomes and cubosomes to monitor the anisotropy of the incorporated fluorescence probe, in which Pluronic F127, used as a dispersion stabilizer of the cubic phase, has been proven not to alter the cubic structure and the location of the probes. Time-resolved fluorescence anisotropy measurements on these dispersions have revealed that the order parameter of the probe in the lamellar phase increases with increasing X(MO), and that it decreases during the transition to the cubic phase. This observation suggests that packing stress generated by the addition of the nonlamellar-forming lipid is released by the phase transition.     

Structured emulsion-based delivery systems: Controlling the digestion and release of lipophilic food components

There is a need for edible delivery systems to encapsulate, protect and release bioactive and functional lipophilic constituents within the food and pharmaceutical industries. These delivery systems could be used for a number of purposes: controlling lipid bioavailability; targeting the delivery of bioactive components within the gastrointestinal tract; and designing food matrices that delay lipid digestion and induce satiety. Emulsion technology is particularly suited for the design and fabrication of delivery systems for lipids. In this article we provide an overview of a number of emulsion-based technologies that can be used as edible delivery systems by the food and other industries, including conventional emulsions, nanoemulsions, multilayer emulsions, solid lipid particles, and filled hydrogel particles. Each of these delivery systems can be produced from food-grade (GRAS) ingredients (e.g., lipids, proteins, polysaccharides, surfactants, and minerals) using relatively simple processing operations (e.g., mixing, homogenizing, and thermal processing). The structure, preparation, and utilization of each type of delivery system for controlling lipid digestion are discussed. This knowledge can be used to select the most appropriate emulsion-based delivery system for specific applications, such as encapsulation, controlled digestion, and targeted release.     

Lipids and alkaloids from <Emphasis Type="Italic">Heliotropium lasiocarpum</Emphasis>

Lipids of seeds with pericarp and an admixture of the air-dried aerial part (AP) of Heliotropium lasiocarpum Fisch. et Mey. (Boraginaceae) were studied. The contents and component and fatty-acid compositions of neutral lipids, glycolipids, phospholipids, and strongly bound lipids were established. It was found that part of the alkaloids was extracted together with the lipids from these plant organs. The chromatographic mobility of the heliotrope alkaloids was determined under the lipid separation conditions.     

MINOR SURFACE-ACTIVE LIPIDS OF OLIVE OIL AND VISCOELASTICITY OF PROTEIN FILMS AT THE OLIVE OIL-WATER INTERFACE

The influence of the minor surface-active lipids of olive oil on the viscoelastic parameters of bovine serum albumin, sodium caseinate and egg yolk films, formed following adsorption at the olive oil-protein solution interface, was studied. All the parameters substantialy decreased in the case of bovine serum albumin when surface-active lipids were removed from the olive oil while the absence of these components resulted in purely viscous films in the cases of sodium caseinate and egg yolk. Protein film viscoelasticity is probably influenced by olive oil surface-active lipids as a result of their interactions with adsorbed and unfolded protein molecules at the oil-protein solution interface.     

Epoxy and hydroxy acids of the seed oil ofGaleopsis bifida

The lipids of the seeds of the hemp nettle, which is toxic for ruminants have been studied. The composition of the fatty acids of five classes of lipids and the position-species composition of the triacylglycerols have been determined. The presence of epoxyacyl- and hydroxyacylglycerols in the oil and the structures of the acids of these lipids have been established. Squalene has been isolated from the oil.Institute of the Chemistry of Plant Substances, Academy of Sciences of the Uzbek SSR, Tashkent. Translated from Khimiya Prirodnykh Soedinenii, No. 3, pp. 288–295, May–June, 1981.     

Oxidation of monoglyceride, diglyceride, and triglyceride monolayers by aqueous potassium permanganate solution

Studies were carried out on the oxidative degradation of monoolein, monolinolein, diolein, dilinolein, triolein and trilinolein monolayers by injecting KMnO₄ solution under the monolayers. The effect of the initial surface pressure on oxidative degradation was also studied by measuring changes in the surface pressure and surface potential with time. The surface shear viscosities of these six lipids were measured in order to predict their molecular interactions at the air/water interface. The rates of oxidation for these lipids were found to be in the following order: dilinolein> monolinolein> monoolein> trilinolein> diolein> triolein. Interestingly, the surface shear viscosities of these six lipids were found to decrease in the same order. In the present study, an attempt has been made to correlate the effect of the initial surface pressure, the number of double bonds and the number of hydroxyl groups with the oxidation of these lipids by potassium permanganate solution.     

Bicellar systems as modifiers of skin lipid structure

The characterization of different bicellar aggregates and the effects of these systems on the stratum corneum (SC) microstructure have been studied. Dynamic light scattering (DLS) and freeze fracture electron microscopy (FFEM) techniques showed that both of the systems studied, dimyristoyl-phosphatidylcholine/dihexanoyl-phosphocholine (DMPC/DHPC) and dipalmitoyl-phosphocholine (DPPC)/DHPC, were formed by small discoidal aggregates at room temperature (20°C). Treating skin with DMPC/DHPC bicelles does not affect the SC lipid microstructure, whereas bicellar systems formed by DPPC and DHPC can promote the formation of new structures in the SC lipid domains. This indicates the passage of lipids from bicelles through the SC layers and also a possible interaction of these lipids with the SC lipids. Given the absence of surfactant in the bicellar composition and the small size of these structures, the use of these smart nano-systems offers great advantages over other lipid systems for dermatological purposes. Bicelles could be promising applications as drug carriers through the skin. This contribution, based on the new biological use of bicelles, may be useful to scientists engaged in colloid science and offers a new tool for different applications in skin and cosmetic research.     

Inner membrane lipids of Escherichia coli form domains

In prokaryotic cells, the hypothesis of the existence of lipid domains was considered. In order to test this hypothesis and study the organization of lipids in the inner membrane of Escherichia coli, we elaborated Langmuir films mimicking the inner leaflet of this membrane by considering lipids extracted from the inner membrane of E coli by Folch protocol.

Lipid monolayers were elaborated by using these extracts (Langmuir technique); the organization of the resulting films was studied at the air–water interface by Brewster angle microscopy and after transfer onto muscovite by atomic force microscopy. The existence of domains was demonstrated for different interfacial pressures of biological interest, and their stability was studied.     

Compositions of the lipids of the whitefishCoregonus peled from different parts of the body

The composition of the total, the neutral, and the polar lipids of the whitefishCoregonus peled (Gmelin) from different parts of the body — the white muscles of the back and abdomen, the red muscles, the brain, the internal organs, and the fish as a whole — have been studied. The amounts of lipids in these sections were 6.6, 16.1, 20.1, 13.8, 40.0, and 8.4%, respectively. It has been shown that the total and neutral lipids are similar with respect to the main groups of lipids, the bulk of them being represented by triacylglycerols (76.0–82.6%) and phospholipids (2.8–5.8%). The composition of the phospholipids depended substantially on their localization. In the lipids of the white muscles of the back, with a low level of choline-containing lipids, phosphatidylethanolamine and cardiolipin predominated. The lipids of the viscera were distinguished by a high content of sphingomyelin. A high level of choline-containing lipids and cerebrosides was found in the tissues of the brain.Tumen' State Medical Institute. Translated from Khimiya Prirodnykh Soedinenii, No. 1, pp. 24–27, January–February, 1985.     

Starch-lipid interactions studied by differential scanning calorimetry

The amylose-lipid complex shows an endothermic transition around 100 °C in excess water. Complexes were prepared by adding lipids to an amylose-solution, and the precipitated complex was studied in the DSC during a heating and cooling sequence. The thermal stability of the complex depends on the lipid part, and the reversibility during cooling depends on presence of excess lipids.The influence of lipids on the gelatinization of starch was studied by adding lipids to wheat and potato starch, respectively, before the DSC-analysis. Depending on the lipid, an earlier as well as a delayed gelatinisation could be obtained.     

Ionization behavior of amino lipids for siRNA delivery: determination of ionization constants, SAR, and the impact of lipid pKa on cationic lipid-biomembrane interactions

Ionizable amino lipids are being pursued as an important class of materials for delivering small interfering RNA (siRNA) therapeutics, and research is being conducted to elucidate the structure-activity relationships (SAR) of these lipids. The pK(a) of cationic lipid headgroups is one of the critical physiochemical properties of interest due to the strong impact of lipid ionization on the assembly and performance of these lipids. This research focused on developing approaches that permit the rapid determination of the relevant pK(a) of the ionizable amino lipids. Two distinct approaches were investigated: (1) potentiometric titration of amino lipids dissolved in neutral surfactant micelles; and (2) pH-dependent partitioning of a fluorescent dye to cationic liposomes formulated from amino lipids. Using the approaches developed here, the pK(a) values of cationic lipids with distinct headgroups were measured and found to be significantly lower than calculated values. It was also found that lipid-lipid interaction has a strong impact on the pK(a) values of lipids. Lysis of model biomembranes by cationic lipids was used to evaluate the impact of lipid pK(a) on the interaction between cationic lipids and cell membranes. It was found that cationic lipid-biomembrane interaction depends strongly on lipid pK(a) and solution pH, and this interaction is much stronger when amino lipids are highly charged. The presence of an optimal pK(a) range of ionizable amino lipids for siRNA delivery was suggested based on these results. The pK(a) methods reported here can be used to support the SAR screen of cationic lipids for siRNA delivery, and the information revealed through studying the impact of pK(a) on the interaction between cationic lipids and cell membranes will contribute significantly to the design of more efficient siRNA delivery vehicles.     

Analysis of lipids with desorption atmospheric pressure photoionization‐mass spectrometry (DAPPI‐MS) and desorption electrospray ionization‐mass spectrometry (DESI‐MS)

In this article, the effect of spray solvent on the analysis of selected lipids including fatty acids, fat‐soluble vitamins, triacylglycerols, steroids, phospholipids, and sphingolipids has been studied by two different ambient mass spectrometry (MS) methods, desorption electrospray ionization‐MS (DESI‐MS) and desorption atmospheric pressure photoionization‐MS (DAPPI‐MS). The ionization of the lipids with DESI and DAPPI was strongly dependent on the spray solvent. In most cases, the lipids were detected as protonated or deprotonated molecules; however, other ions were also formed, such as adduct ions (in DESI), [M‐H]⁺ ions (in DESI and DAPPI), radical ions (in DAPPI), and abundant oxidation products (in DESI and DAPPI). DAPPI provided efficient desorption and ionization for neutral and less polar as well as for ionic lipids but caused extensive fragmentation for larger and more labile compounds because of a thermal desorption process. DESI was more suitable for the analysis of the large and labile lipids, but the ionization efficiency for less polar lipids was poor. Both methods were successfully applied to the direct analysis of lipids from pharmaceutical and food products. Although DESI and DAPPI provide efficient analysis of lipids, the multiple and largely unpredictable ionization reactions may set challenges for routine lipid analysis with these methods. Copyright © 2012 John Wiley & Sons, Ltd.     

Influence of lipid composition on membrane activity of antimicrobial phenylene ethynylene oligomers

Host defense peptides (HDPs), part of the innate immune system, selectively target the membranes of bacterial cells over that of host cells. As a result, their antimicrobial properties have been under intense study. Their selectivity strongly depends on the chemical and mostly structural properties of the lipids that make up different cell membranes. The ability to synthesize HDP mimics has recently been demonstrated. To better understand how these HDP mimics interact with bilayer membranes, three homologous antimicrobial oligomers (AMOs) 1-3 with an m-phenylene ethynylene backbone and alkyl amine side chains were studied. Among them, AMO 1 is nonactive, AMO 2 is specifically active, and AMO 3 is nonspecifically active against bacteria over human red blood cells, a standard model for mammalian cells. The interactions of these three AMOs with liposomes having different lipid compositions are characterized in detail using a fluorescent dye leakage assay. AMO 2 and AMO 3 caused more leakage than AMO 1 from bacteria membrane mimic liposomes composed of PE/PG lipids. The use of E. coli lipid vesicles gave the same results. Further changes of the lipid compositions revealed that AMO 2 has selectively higher affinity toward PE/PG and E. coli lipids than PC, PC/PG or PC/PS lipids, the major components of mammalian cell membranes. In contrast, AMO 3 is devoid of this lipid selectivity and interacts with all liposomes with equal ease; AMO 1 remains inactive. These observations suggest that lipid type and structure are more important in determining membrane selectivity than lipid headgroup charges for this series of HDP mimics.     

Design of liposomes containing photopolymerizable phospholipids for triggered release of contents

We describe a novel class of light-triggerable liposomes prepared from a photo-polymerizable phospholipid DC_8,9PC (1,2-bis (tricosa-10,12-diynoyl)-sn-glycero-3-phosphocholine) and DPPC (1,2-Dipalmitoyl-sn-Glycero-3-Phosphocholine). Exposure to UV (254 nm) radiation for 0–45 min at 25 °C resulted in photo-polymerization of DC_8,9PC in these liposomes and the release of an encapsulated fluorescent dye (calcein). Kinetics and extents of calcein release correlated with mol% of DC_8,9PC in the liposomes. Photopolymerization and calcein release occurred only from DPPC/DC_8,9PC but not from Egg PC/DC_8,9PC liposomes. Our data indicate that phase separation and packing of polymerizable lipids in the liposome bilayer are major determinants of photo-activation and triggered contents release.     

Controlling liposomal drug release with low frequency ultrasound: mechanism and feasibility

The ability of low-frequency ultrasound (LFUS) to release encapsulated drugs from sterically stabilized liposomes in a controlled manner was demonstrated. Three liposomal formulations having identical lipid bilayer compositions and a similar size ( approximately 100 nm) but differing in their encapsulated drugs and methods of drug loading have been tested. Two of the drugs, doxorubicin and methylpredinisolone hemisuccinate, were remote loaded by transmembrane gradients (ammonium sulfate and calcium acetate, respectively). The third drug, cisplatin, was loaded passively into the liposomes. For all three formulations, a short exposure to LFUS (<3 min) released nearly 80% of the drug. The magnitude of drug release was a function of LFUS amplitude and actual exposure time, irrespective of whether irradiation was pulsed or continuous. Furthermore, no change in liposome size distribution or in the chemical properties of the lipids or of the released drugs occurred due to exposure to LFUS. Based on our results, we propose that the mechanism of release is a transient introduction of porelike defects in the liposome membrane, which occurs only during exposure to LFUS, after which the membrane reseals. This explains the observed uptake of the membrane-impermeable fluorophore pyranine from the extraliposomal medium during exposure to LFUS. The implications of these findings for clinical applications of controlled drug release from liposomes are discussed.