Preparation of J-aggregate liposome dispersions and their chromic transformation

We report the preparation of aqueous liposome dispersions of J-aggregates formed by the amphiphilic merocyanine dye (MD). A series of liposome-forming lipids were dispersed together with MD J-aggregates at different molar ratios of MD to lipid. The MD J-aggregate dispersions prepared with 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) at the MD to DMPC ratio of 0.16 exhibit good dispersibility; that is, they can be readily redispersed without any flocculation even after their precipitation. By use of different counterions for the MD molecules, two types of J-aggregate dispersions, one that exhibits an absorption band (J-band) at 635 nm (type I) and the other at 600 nm (type II), were obtained. As an example of the use of MD J-aggregates liposome dispersions, the thermochromic transformation of MD J-aggregates was demonstrated. When the dispersions are heated, J-aggregates of type I transformed into type II at a certain temperature (T(disp)). The parameters that control the speed of the transformation and the value of T(disp) were determined.     

Dynamic processes in endocytic transformation of a raft-exhibiting giant liposome

The dynamic response of a raft-exhibiting giant liposome to external stimuli, such as the addition of Triton X-100 or osmotic stress, was studied. We observed that daughter vesicles are generated inside of the liposome through endocytic budding. It was found that the budding to generate daughter vesicles is classified into two different routes, simple budding through the invagination of a whole raft and budding from the boundary of a raft accompanied by waving motion. Smaller rafts show a preference for simple budding, whereas large rafts mainly adopt the other process. We discuss the mechanism of this difference in terms of the kinetic pathway of internalization by considering the line energy and bending energy of the membrane.     

Supramolecular assemblies of nucleoside phosphocholine amphiphiles

A family of new uridine phosphocholine amphiphiles that were prepared using a convenient four-step synthetic route is described. Physicochemical studies (differential scanning calorimetry, small-angle X-ray scattering, UV-vis and circular dichroism spectroscopies, light microscopy, transmission electronic microscopy, and scanning electron microscopy) show that these amphiphiles spontaneously assemble into supramolecular structures including vesicles, fibers, hydrogels, and organogels. In aqueous solution, the amphiphiles possessing saturated alkyl chains self-assemble into DNA-like helical fibers in the crystalline state below T(m) and compact bilayers above the melting temperature (T(m)). The transition from bilayers to fibers is thermally reversible. Above a threshold concentration (>6% w/w), a hydrogel is formed due to an entangled network of the fibers. A therapeutic agent such as DNA can be entrapped within the hydrogel structure. In addition to forming bilayer vesicles and hydrogels in aqueous solution, these nucleoside amphiphiles also form organogels in cyclohexane above T(m). Scanning electron microscopy shows a continuous multilamellar phase in the organogels.     

A synthetic receptor for phosphocholine esters

A bifunctional Zn-salen modified cavitand, reminiscent of the enzyme phospholipase C, shows high efficiency and synergic effect in the binding of the phospholipid DOPC.     

The structure of zwitterionic phosphocholine surfactant monolayers

The structure of a zwitterionic phosphocholine (PC) surfactant monolayer adsorbed on the surface of water has been determined using neutron reflectivity in combination with H/D isotopic substitution. The most significant results of this study are the level of hydration of the PC headgroup and the lack of dehydration with increasing temperature and salt addition. The fraction of the alkyl chain (f(c)) immersed in water for all three chain isomers studied was found to be around 0.15, suggesting that the PC headgroup geometries influenced not only the headgroup hydration but also the degree of immersion of the alkyl chain in water. At the critical micelle concentration (CMC), the number of water molecules associated with the PC headgroup in C(m)PC (m = 12, 14, 16) was on order of 15. This value was significantly greater than that obtained for nonionic and ionic surfactants with similar limiting area per molecule at the CMC (A(cmc)). However, the fraction of the chain immersed in water for the ionic and nonionic surfactants was much greater. This suggests that the unique surface biocompatibility of PC surfactants arises from their strong affinity for water, and the relatively low fraction of mixing with the alkyl chain arises from the higher structural order within the PC monolayer. As surface coverage decreased, the number of water molecules associated with each PC headgroup increased, but f(c) remained constant for all the surfactants. This observation was consistent with the small variation in the thickness of the headgroup region, and the entire layer changed little with surfactant concentration. This is attributed to the role of PC headgroup geometries to maintain the conformational order within the layer as packing density varies. Further structural analysis based on a kinematic approach showed that, as the chain length was increased from C12 to C14 to C16 at the CMC, the angle of tilt for the alkyl chain increased from 40 degrees to 48 degrees to 53 degrees , respectively, whereas the thickness of the whole layer and that of the PC head region was largely constant. The almost vertical projection of the PC headgroup from these single alkyl chain surfactants is in sharp contrast to its strongly tilted conformation, as reported for dichain phospholipids such as dipalmitoyl glycerol phosphocholine (DPPC).     

Acyl transfer from phosphocholine lipids to melittin

Transfer of fatty acyl groups from membrane phospholipids to melittin, a commonly studied membrane-active peptide, has been observed to occur over extended time periods. Transfer can be detected after 1-2 days and selectively targets amino groups at the N-terminal end of the peptide.     

Facile ring transformation from gluconolactone to cyclitol derivative via spiro sugar ortho ester

[reaction: see text] A short step preparation of cyclitol derivative 8 which is a versatile synthon for the synthesis of valiolamine and its related compounds is described. Key steps in this preparation are a novel enol ether formation from spiro sugar ortho esters with AlMe3 and an intramolecular Aldol condensation of alkyl enol ethers catalyzed by ZnCl2 in THF-H2O. With these reactions, gluconolactone derivative 1 was efficiently converted into 8 in short steps.     

Electrochemical transformation of cyanoacetic ester and alkylidenecyanoacetic esters into 3-substituted 1,2-dicyanocyclopropane-1,2-dicarboxylates

Electrolysis of cyanoacetic ester and alkylidenecyanoacetic esters in an undivided cell in the presence of mediators (alkali metal halides) gives rise to 3-substituted, 1,2-dicyanocyclopropane-1,2-dicarboxylates in 60–95% yields. Translated fromIzvestiya Akademii Nauk, Seriya Khimicheskaya, No. 6, pp. 1165–1168, June, 1998.     

A crystallization-induced asymmetric transformation to prepare (R)-4-chlorophenylalanine methyl ester

A second order asymmetric transformation of racemic 4-chlorophenylalanine methyl ester was achieved via salt formation with (2S,3S)-(−)-tartaric acid in the presence of salicylaldehyde to afford the desired (R)-enantiomer of 4-chlorophenylalanine methyl ester in good yield and high enantiomeric purity.     

Quantitative determination of phosphocholine and the activity of phospholipase C

Summary 1. The ester of dextran and pelentanic acid with a maximum value of 140 has been synthesized.2. The influence of some reaction conditions on the composition of the esters obtained has been investigated.Khimiya Prirodnykh Soedinenii, Vol. 1, No. 4, pp. 245–246, 1965     

Quantitative determination of phosphocholine and the activity of phospholipase C

Summary 1. A quantitative method for determining phosphocholine by means of thin-layer chromatography has been proposed.2. In the enzymatic cleavage of lecithins with phospholipase C, this method permits the amount of diglycerides formed to be determined as well as the phosphocholine.3. The agreement of the results of the analysis of phosphocholine and the diglycerides is a reliable criterion of the activity of the phospholipase C.Khimiya Prirodnykh Soedinenii, Vol. 2, No. 4, pp. 233–235, 1966     

C-Reactive protein-directed immobilization of phosphocholine ligands on a solid surface

The complexes of C-reactive protein (CRP) with its polymerizable phosphocholine ligands adsorbed at the styrene-water interface were polymerized. The molecularly imprinted polymer (MIP) exhibited a binding affinity for CRP comparable to that of immobilized anti-CRP antibody. The determination of human serum CRP using the MIP-based sandwich immunoassay has been demonstrated.     

On the hydration of the phosphocholine headgroup in aqueous solution

The hydration of the phosphocholine headgroup in 1,2-dipropionyl-sn-glycero-3-phosphocholine (C(3)-PC) in solution has been determined by using neutron diffraction enhanced with isotopic substitution in combination with computer simulation techniques. The atomic scale hydration structure around this head group shows that both the -N(CH(3))(3) and -CH(2) portions of the choline headgroup are strongly associated with water, through a unique hydrogen bonding regime, where specifically a hydrogen bond from the C-H group to water and a strong association between the water oxygen and N(+) atom in solution have both been observed. In addition, both PO(4) oxygens (P=O) and C=O oxygens are oversaturated when compared to bulk water in that the average number of hydrogen bonds from water to both X=O oxygens is about 2.5 for each group. That water binds strongly to the glycerol groups and is suggestive that water may bind to these groups when phosophotidylcholine is embedded in a membrane bilayer.     

Facile iodination of the vinyl groups in protoporphyrin IX dimethyl ester and subsequent transformation of the iodinated moieties

Iodination of protoporphyrin IX dimethyl ester using phenyliodine bis(trifluoroacetate) (PIFA) and I₂ was studied. Iodine added to both the C3- and C8-vinyl groups equally to afford the iodohydrin or iodoether in the presence of water or alcohol, respectively. Any meso-hydrogen atom was not substituted by an iodine atom under these conditions, although both the vinyl group and one of the meso positions of methyl pyropheophorbide-a bearing a chlorin π-system, a chlorophyll-a derivative, was modified with PIFA and I₂. The reaction intermediates derived from the porphyrin were more reactive than those from the chlorin and liable to form intermolecular linkages. The obtained 2-iodo-1-hydroxyethyl group was transformed into a formyl group by a mild treatment. The corresponding iodoether moiety was readily converted into the acetyl group under basic conditions. These transformations were also applicable to smaller olefins such as styrene.     

Lewis Acid-triggered selective zincation of chromones, quinolones, and thiochromones: application to the preparation of natural flavones and isoflavones

A Lewis acid-triggered zincation allows the regioselective metalation of various chromones and quinolones. In the absence of MgCl(2), a C(3) zincation is observed, whereas in the presence of MgCl(2) or a related Lewis acid, C(2) zincation occurs. Applications to a natural flavone, isoflavone, and quinolone are shown.     

Dimetallic functionalities in liposome bilayers

A dicopper(II) complex can be covalently linked to palmitate/palmitoyl-oleoyl-phosphatidylcholine (PA/POPC) liposomes using the following one-pot strategy: preformed [Cu₂(bpbp)(PA)](ClO₄)₂ (bpbp^?  = 2,6-bis((N,N′-bis(2-picolyl)amino)methyl)-4-tertbutylphenolato) was incorporated into POPC liposomes with a loading of up to 10 mol%. Despite its shape and charge, the decoration of PA/POPC liposomes with {Cu₂(bpbp)}³⁺ did not disrupt the liposome structure; however, the mean liposome diameter increased from about 130 nm (0 mol% dicopper complex) to about 150 nm (10 mol% dicopper complex). Single-crystal X-ray structures furnish ‘snapshots’ of the pH-dependent solution state derivatives of {Cu₂(bpbp)}³⁺, and model the structure of the [Cu₂(bpbp)(PA)]²⁺ head group at the surface of the liposomes. An impressive plasticity in the intramolecular non-bonded Cu….Cu distance for these ions, ranging from 3 to 4 Å, in [Cu₂(bpbp)OH]²⁺, [Cu₂(bpbp)(OAc)(H₂O)]²⁺ and [Cu₂(bpbp)(H₂O)₂]³⁺ allows for their utility as labile reagents in water. Remarkably, the flexible dicopper site is selective for a single carboxylate ligand, so that [Cu₂(bpbp)(PA)]²⁺ is favoured even in the presence of other chemically similar oxoanions, such as CO₃^2 ? , HCO₃^? , NO₃^? , ClO₄^? , ReO₄^?  and CF₃SO₃^? .     

Synthesis and antitumor activity of new alkylphospholipids containing modifications of the phosphocholine moiety

New antitumor alkylglycerophospholipids, in which primarily the phosphocholine moiety of the platelet activating factor (PAF) molecule was modified, were synthesized from 1-alkyl-2-substituted glycerols by introducing polar head phosphoryl groups having methylene bridges of various lengths (from 2 to 14 carbons). They were tested for PAF agonistic activity and antitumor properties. In a series of 1-octadecyl-2-acetoacetylglycerophospholipids (1a-f), an increase in the length of the methylene bridge separating the phosphate and trimethylammonio group in the polar head side chain at position 3 of the glycerol backbone resulted in a progressive decrease in PAF agonistic activity and a characteristic change in antitumor activity against human promyelocytic leukemia cells (HL-60). Maximal potency was obtained with the compound having a decamethylene bridge (1e, IC50 value = 1.5 microgram/ml). Thus, alkylphospholipids possessing a decamethylene bridge and a variety of substituents at position 2 (1g-n) were synthesized. They showed potent inhibitory activity with IC50 values ranging from 0.4 to 1.9 micrograms/ml, depending on the nature of the 2-substituent in the phospholipid molecule. In in vivo tests of the present series of alkylglycerophospholipids (1a--n), using mice bearing sarcoma 180 and mice with mammary carcinoma MM46 (both cells and compounds were given i.p.), 1-octadecyl-2-acetoacetyl-3-glyceryl omega-trimethylammoniodecyl phosphate (1e) showed the most potent life-prolonging effect. The structure-activity relationships are discussed.     

Luminescent dipyridophenazine-ruthenium probes for liposome membranes

Ruthenium complexes with dipyridophenazine (dppz) type ligands have several characteristics that make them good candidates for use as luminescence probes for hydrophobic environments. Most studies have concerned DNA intercalation, but also lipid membrane fluidity and liposome orientation have been assessed. We report here dipyridophenazine derivatives ([Ru(phen)2dppz]2+) substituted with one or two alkyl ether chains of different lengths aimed at finding the optimum substitution for a high quantum yield when bound to a phospholipid membrane bilayer. The orientation of membrane bound molecules is studied using flow linear dichroism (LD) with phospholipid vesicles as membrane models. LD, excitation anisotropy, steady state luminescence and excited-state lifetime measurements are used to quantitatively investigate the insertion and orientation of the complexes in the vesicles. All complexes are inserted with their long axis of the dppz moiety mainly parallel to the lipid chains, and the degree of orientation is comparable to that of the orientation probe retinoic acid. The ruthenium "head group" with its positive charge functions as a buoy at the water-membrane interface while the hydrophobic chain part embeds the complex down into the bilayer. The complex with two hexyl ether substituents (named D6) has the optimal chain length regarding membrane insertion and orientation, and together with the highest quantum yield, is the best luminescence membrane probe in the two series.     

Effect of leucinyl-phenylalanyl-valine on DMPC liposome membrane

Leucinyl-phenylalanyl-valine (LFV) is a hydrophobic tripeptide with a flat egg shaped structure with the long axis dimension of about 12 A. The effect of LFV on dimyristoylphosphatidylcholine (DMPC) liposome membrane has been studied by differential scanning calorimetry (DSC) and fluorescence spectroscopy. Calorimetric studies shows that incorporation of LFV completely abolishes the pretransition temperature with broadening of main transition temperature. Four conceptually different fluorescence probes, 1-naphthol (1-ROH) an excited state proton transfer probe, 8-anilino-1-naphthalenesulphonate (ANS) a solvent polarity probe, 1-6-diphenylhexatriene (DPH) an anisotropy probe and pyrene an excimer-forming probe have been used for fluorescence spectroscopic studies. For 1-ROH, ANS and DPH, a decreased partitioning with increasing mol.% of LFV was observed. Increasing LFV mol.% caused a decrease in the neutral form emission of 1-ROH, and a decrease in fluorescence intensity with red shift in ANS. The excimer formation ability of pyrene also decreased. The phase transition behavior of DMPC membrane in the presence of LFV was similar to the known effect of cholesterol on lipid bilayers. These results suggest that LFV cause an increased compactness of membrane.