Camillo La Mesa

Dynamics of DNA Adsorption on and Release from SDS−DDAB Cat−Anionic Vesicles: a Multitechnique Study

DNA adsorption and release from cat-anionic vesicles made of sodium dodecylsulfate-dodecyldimethylammonium bromide (SDS-DDAB) in nonstoichiometric amounts was investigated by different electrochemical, spectroscopic, and biomolecular strategies. The characterization of the vesicular system was performed by dynamic light scattering, which allowed estimating both its size and distribution function(s). The interaction dynamics was followed by dielectric spectroscopy and zeta-potential, as well as by agarose gel electrophoresis, AGE. Also, circular dichroism, CD, measurements were carried out, to ascertain possible structural rearrangements of DNA, consequent to the interactions with the cat-anionic vesicles. CD demonstrates that vesicle-bound DNA retains its native conformation. The results obtained by the aforementioned techniques are consistent and indicate that binding saturation is obtained at a [DNA/vesicles] charge ratio close to 0.8, considering only the excess surface charges on the vesicles. This result is apparently in contradiction with a purely electrostatic approach and is tentatively ascribed to the distance between charges on the biopolymer and the vesicle surface, respectively. A possible interpretation is discussed. The nucleic acid can be completely retrieved from the vesicles upon addition of adequate amounts of SDS, which is the defective surfactant in the vesicular system. Precipitation of the poorly soluble SD-DDA salt results in an almost complete release of DNA.

Multi- to unilamellar transitions in catanionic vesicles.

Sodium dodecylsulfate (SDS) and cetyltrimethylammonium bromide (CTAB) dispersed in aqueous solution form catanionic vesicles. Depending on composition, such vesicles show different net charge, stability, and interaction capability, indicative of the strong impact that catanionic systems may have in gene therapy and drug delivery technologies. To reveal the interplay among composition, net charge, sensitivity to temperature changes, vesicle size, and inner structure, a series of experiments on catanionic vesicles prepared at different SDS/CTAB mole ratios was performed. Dynamic light scattering, small-angle X-ray scattering, and zeta-potential experiments allow one to characterize an unexpected critical phenomenon at the nanoscale level. On heating, vesicles increase in size, but at a critical temperature an abrupt vesicle size reduction has been observed, together with a transition from multi- to a unilamellar state. The critical temperature regularly depends on the SDS/CTAB mole ratio. The unilamellar state obtained upon heating is retained for weeks. These phenomena suggest a new way to produce stable unilamellar vesicles with tunable size and charge.

Biological activity of SDS-CTAB cat-anionic vesicles in cultured cells and assessment of their cytotoxicity ending in apoptosis

SDS-CTAB cat-anionic vesicles are supramolecular aggregates forming complexes with biopolymers and enter the cells via membrane fusion or endocytosis. Different applicative areas exist: gene therapy, drug delivery and nanotechnology. We previously examined the absorption/release of biopolymers from vesicles in solution. Here we evaluate their cytotoxicity in cultured cells; to this end we characterized the vesicles and analyzed their biological effects at cellular and molecular level. At low concentration these vesicles have scarce consequences on normal cell growth; at higher dosage they activate apoptotic death processes, due to membrane damage. In conclusion, the use of these particles in nano-biotechnology represents an actual possibility.

Alkyl glucopyranoside-based niosomes containing methotrexate for pharmaceutical applications: Evaluation of physico-chemical and biological properties

We designed novel niosomes based on alkyl glucopyranoside surfactants and containing methotrexate as anticancer drug, to be used in the pharmaceutical field. The effects of surfactants with chains of different length on niosome size and their distribution, drug entrapment efficiencies and in vitro drug release were determined. Systems made of alkyl glucopyranosides and cholesterol form vesicles whose average size scales with the alkyl chains length of such surfactants. Vesicles size ranges between 300 and 500 nm, with low polydispersity index. In addition, the hemolytic activity of alkyl glucopyranosides as surfactant solutions or vesicular formulations was studied and compared, to identify possible structure-activity relationships. High methotrexate entrapment efficiency was obtained, confirming significant interactions between the drug and the niosomal matrices. After 24h the amount of methotrexate released from niosomal formulations is effectively delayed, compared to the free drug in solution. Hemolytic tests show that sugar-based surfactants are more hemolytic the longer is their alkyl chain. When the surfactants are in vesicular form, the reverse behavior holds. It was also inferred that vesicle formation reduces the surfactant toxicity. These niosomal formulations can be used as methotrexate delivery systems in anticancer therapy.

Water—Trifluoroethanol Mixtures: Some Physicochemical Properties

The-physicochemical properties of water—2,2,2 trifluoroethanol mixtures havebeen investigated, at 25°C, by density, viscosity, surface tension, and relativepermittivity. An analysis based on the Kirkwood theory for the dielectric polarizationof mixtures has been performed and the gκ correlation factor has been relatedto the occurrence of hydrogen bonding networks. Data on the above systemhave been analyzed and compared to data on water—ethanol mixtures, to obtaininformation on the role that partial fluorination plays in the interactions of alcoholswith water molecules. As inferred from excess properties, nonideality contributionsare smaller than in the corresponding water—ethanol solutions. Accordingly,the extension of hydrogen bond networks and/or van der Waals contributions tothe overall system stability are less significant than in simple alcohols.

Gemini surfactant–water mixtures: some physical–chemical properties

Abstract The phase diagram of the water-Gemini 16-4-16 system has been investigated and the phase boundaries were determined. DSC and optical microscopy were used to define the region of existence of the different phases. No liquid crystalline phases have been observed, however, a two-phase region and a wide gel phase follow the solution region. The solution region can be highly viscous, depending on composition and temperature. Surface tension and electrical conductance experiments have been performed, to define micelle formation and counter-ion binding to micelles. Interactions and motions over short distances were studied by 1H-NMR relaxation experiments. The drastic decrease of spin–spin relaxation time, T2, with Gemini composition ( ≈2 wt.%) was explained in terms of particle growth. Pulsed field gradient spin-echo (PGSE) NMR experiments were used to determine water and surfactant self-diffusion. Some modifications in the micellar structure were inferred on increasing the Gemini content in the mixture. Dynamic rheological experiments were performed for probing the solution microstructure. The observed high solution viscosity and the shear relaxation processes were rationalized in terms of the presence of entangled threadlike aggregates at a moderate concentration (≈ 4 wt.%). According to the Bohlin theory of flow as a cooperative phenomenon, the number of the micellar aggregates correlated to each other, and the interaction strength between the micellar units was obtained as a function of Gemini concentration.

Catanionic vesicles formed with arginine-based surfactants and 1,2-dipalmitoyl-sn-glycero-3-phosphate monosodium salt.

We report on mixing an anionic diacyl phospholipid (1,2-dipalmitoyl-sn-glycero-3-phosphate monosodium salt, DPPA) with either monoacyl and diacyl arginine-based surfactants. These mixtures are part of the rich family of pseudo-triple-chain and pseudo-tetra-chain catanionic mixtures, respectively. Vesicle size and zeta-potential were measured at several mixing ratios. Additional information on counterion binding, vesicle size, and integrity was obtained from ion selective electrode and Cryo-TEM measurements. Addition of positively charged surfactants to DPPA results in an increase of vesicle size. However, zeta-potential shows different trends, depending on whether water or acid media are used as solvent. In the latter, zeta-potential values progressively approach 0 upon addition of amino acid based surfactants. In water, surprisingly, zeta-potential values become more negative. The results are discussed in terms of modifications in counterion binding and vesicle size.

Niosomes from α,ω-trioxyethylene-bis(sodium 2-dodecyloxy-propylenesulfonate): Preparation and characterization

The synthesis and characterisation of new surfactants with peculiar physical-chemical properties are amongst the most promising and expanding issues in pharmacological colloid science. The most used vesicular carriers are liposomes prepared from a wide variety of natural and synthetic phospholipids, but several ionic and non-ionic amphiphiles have been used to form multilamellar and/or unilamellar vesicles. In the present study the synthesis of alpha,omega-trioxyethylene-bis(sodium 2-dodecyloxy-propylenesulfonate), an anionic Gemini surfactant, and its ability to form niosomes are elucidated. The compound forms vesicles with and without added cholesterol. The vesicular systems were characterized by size, shape and drug entrapment efficiency. The compounds to be incorporated are beta-carotene and ferulic acid, as antioxidants, acetyl salicylic acid, as FANS, and the antineoplastic 5-flurouracil, widely used in dermatological disorders. The results of this study show that alpha,omega-trioxyethylene-bis(sodium 2-dodecyloxy-propylenesulfonate) can be used for the preparation of niosomes entrapping lypophilic, amphiphilic or hydrophilic substances. These niosomes may be promising candidates as percutaneous carriers for the aforementioned drugs.

Interaction of bovine serum albumin with gemini surfactants

The interactions between bovine serum albumin and cationic gemini surfactants were investigated as a function of concentration, under different pH conditions. The investigation deals with dielectric relaxation, dynamic light scattering, zeta-potential, circular dichroism, and UV spectroscopy. The interactive behavior of the anionic form is quite different from the cationic species. It indicates that protein-surfactant interactions are mostly electrostatic in nature and depend on the state of charge of bovine serum albumin. The results indicate the presence of both hydrophobic and electrostatic contributions in the interactions of gemini with bovine serum albumin. Comparison of dynamic light scattering, dielectric relaxation, electrophoretic mobility, and optical circular dichroism allows drawing some preliminary hypotheses on the different contributions to surfactant binding and supports former studies on the formation of complexes between the bovine serum albumin and the above species.

Delivery of RNA and Its Intracellular Translation into Protein Mediated by SDS-CTAB Vesicles: Potential Use in Nanobiotechnology

Catanionic vesicles are supramolecular aggregates spontaneously forming in water by electrostatic attraction between two surfactants mixed in nonstoichiometric ratios. The outer surface charges allow adsorption to the biomembrane by electrostatic interactions. The lipoplex thus obtained penetrates the cell by endocytosis or membrane fusion. We examined the possible cytotoxic effects and evaluated the transfection efficiency of one vesicle type as compared to known commercial carriers. We show that the individual components of two different vesicles types, CTAB (cetyltrimethylammonium bromide) and DDAB (didodecyldimethylammonium bromide) are detrimental for cell survival. We also assayed the cytotoxicity of SDS-DDAB vesicles and showed dose and time dependency, with the DDAB component being per se extremely cytotoxic. The transfection efficiency of exogenous RNA mediated by SDS-CTAB increases if vesicles assemble in the presence of the reporter RNA; finally, freezing abrogates the transfection ability. The results of our experimental strategy suggest that catanionic vesicles may be adopted in gene therapy and control of antiproliferative diseases.