Potentiometric Study of Carbon Nanotube/Surfactant Interactions by Ion-Selective Electrodes. Driving Forces in the Adsorption and Dispersion Processes.

The interaction (adsorption process) of commercial ionic surfactants with non-functionalized and functionalized carbon nanotubes (CNTs) has been studied by potentiometric measurements based on the use of ion-selective electrodes. The goal of this work was to investigate the role of the CNTs' charge and structure in the CNT/surfactant interactions. Non-functionalized single- (SWCNT) and multi-walled carbon nanotubes (MWCNT), and amine functionalized SWCNT were used. The influence of the surfactant architecture on the CNT/surfactant interactions was also studied. Surfactants with different charge and hydrophobic tail length (sodium dodecyl sulfate (SDS), octyltrimethyl ammonium bromide (OTAB), dodecyltrimethyl ammonium bromide (DoTAB) and hexadecyltrimethyl ammonium bromide (CTAB)) were studied. According to the results, the adsorption process shows a cooperative character, with the hydrophobic interaction contribution playing a key role. This is made evident by the correlation between the free surfactant concentration (at a fixed [CNT]) and the critical micellar concentration, cmc, found for all the CNTs and surfactants investigated. The electrostatic interactions mainly determine the CNT dispersion, although hydrophobic interactions also contribute to this process.

Binding of cationic single-chain and dimeric surfactants to bovine serum albumin.

The interactions between bovine serum albumin (BSA) and the single-chain surfactants N-benzyl-N,N-dimethyl-N-(1-dodecyl)ammonium bromide (PH12) and N-cyclohexylmethyl-N,N-dimethyl-N-(1-dodecyl)ammonium bromide (CH12) and their two dimeric counterparts, N,N'-[1,3-phenylenebis(methylene)]bis[N,N-dimethyl-N-(1-dodecyl)]ammonium dibromide (12PH12) and N,N'-[cyclohexane-1,3-diylbis(methylene)]bis[N,N-dimethyl-N-(1-dodecyl)]ammonium dibromide (12CH12), respectively, have been investigated by surface tension, fluorescence, circular dichroism, ζ potential, and atomic force microscopy. The results obtained permit the examination of the way an increase in the number of hydrophobic chains and the substitution of a cyclohexyl ring by a phenyl ring, either in the headgroup of single-chain surfactants or in the spacer of dimeric surfactants, affect BSA-surfactant interactions. The comparison of the fluorescence results with those obtained by ζ potential measurements shows that the sites of binding of the surfactants with aromatic rings to the BSA are somewhat different from those of the surfactants with no aromatic rings.

A novel biocompatible polymer derived from D-mannitol used as a vector in the field of genetic engineering of eukaryotic cells.

The design and preparation of new vectors to transport genetic material and increase the transfection efficiency continue being an important research line. Here, a novel biocompatible sugar-based polymer derived from D-mannitol has been synthesized to be used as a gene material nanocarrier in human (gene transfection) and microalga cells (transformation process). Its low toxicity allows its use in processes with both medical and industrial applications. A multidisciplinary study about the formation of polymer/p-DNA polyplexes has been carried out using techniques such as gel electrophoresis, zeta potential, dynamic light scattering, atomic force microscopy, and circular dichroism spectroscopy. The nucleic acids used were the eukaryotic expression plasmid pEGFP-C1 and the microalgal expression plasmid Phyco69, which showed different behaviors. The importance of DNA supercoiling in both transfection and transformation processes was demonstrated. Better results were obtained in microalga cells nuclear transformation than in human cells gene transfection. This was related to the plasmid's conformational changes, in particular to their superhelical structure. It is noteworthy that the same nanocarrier has been used with eukaryotic cells from both human and microalga.

Single -and Multi-Walled Carbon Nanotubes as Nanocarriers for the Delivery of 7-Hydroxyflavone.

The research on flavonoids has exponentially grown since their first therapeutic evidence, in 1937. They are effective in vitro in a wide range of human diseases, particularly those mediated by free radicals, such as cancer, atherosclerosis, AIDS, or neuronal diseases. However, their applications have been reduced due to their low solubility, poor absorption, and rapid metabolism. Flavonoid encapsulation in nanocarriers significantly improves their oral absorption, protects the drug against degradation, decreases the first-pass hepatic effect, and makes absorption through the lymphatic system easier. In this work, carbon nanotubes were used as nanocarriers of 7-hydroxyflavone, 7-HF. The encapsulation of 7-HF into pristine single- and multi-walled carbon nanotubes, and into -COOH functionalized single-walled carbon nanotubes has been investigated. The equilibrium association constants were estimated. The structural backbone of 7-HF, two benzene rings linked through three carbon atoms that form a pyran heterocyclic ring containing a keto group, seems to play a key role in the 7-HF/CNT interactions, although other types of interactions are also at work. The in vitro release of 7-HF was studied at three pHs, 2.0, 7.4, and 9.2, mimicking the different biological barriers of the human organism.

Study of the reaction 2-(p-nitrophenyl)ethyl bromide + OH⁻ in dimeric micellar solutions.

The dehydrobromination reaction 2-(p-nitrophenyl)ethyl bromide + OH⁻ was investigated in several alkanediyl-α-ω-bis(dodecyldimethylammonium) bromide, 12-s-12,2Br⁻ (with s = 2, 3, 4, 5, 6, 8, 10, 12) micellar solutions, in the presence of NaOH 5 × 10⁻³ M. The kinetic data were quantitatively rationalized within the whole surfactant concentration range by using an equation based on the pseudophase ion-exchange model and taking the variations in the micellar ionization degree caused by the morphological transitions into account. The agreement between the theoretical and the experimental data was good in all the dimeric micellar media studied, except for the 12-2-12,2Br⁻ micellar solutions. In this case, the strong tendency to micellar growth shown by the 12-2-12,2Br⁻ micelles could be responsible for the lack of accordance. Results showed that the dimeric micelles accelerate the reaction more than two orders of magnitude as compared to water.

Cationic Single-Chained Surfactants with a Functional Group at the End of the Hydrophobic Tail DNA Compacting Efficiency.

The interaction between calf-thymus DNA, ctDNA, and various single-chained surfactants with different functional groups at the end of hydrophobic tail was studied with the goal of investigating the influence of the functional group nature on surfactant DNA compacting efficiency. The surfactants investigated were dodecyltriethylammonium bromide (DTEABr), triethyl(1-phenoxydodecyl)ammonium bromide (12PhBr), triethyl(2-naphthoxydodecyl)ammonium bromide (12NBr) and 11-(isonicotinoyloxy)-N,N,N-triethyl-1-undecanaminium bromide (11PyBr). Results made evident that the surfactants' tendencies to self-aggregation is the key factor determining their efficiency to compact the nucleic acid. Subsequently, DOPE/12NBr/pEGFP-C1 lipoplexes, with different cationic surfactant molar fractions (α) and mass ratios (L/D), were prepared and characterized. DOPE is a zwitterionic phospholipid 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine, and the plasmid pEGFP-C1 carries a GFP coding sequence with the necessary regulatory elements for constitutive expression of the gene in human cells. 12NBr was chosen because it was the most efficient DNA compacting agent among the surfactants investigated. Finally, the cytotoxicity and transfection efficiency (TE) of DOPE/12NBr/pDNA lipoplexes, with different compositions, were investigated.

Multivalent Calixarene-Based Liposomes as Platforms for Gene and Drug Delivery.

The formation of calixarene-based liposomes was investigated, and the characterization of these nanostructures was carried out using several techniques. Four amphiphilic calixarenes were used. The length of the hydrophobic chains attached to the lower rim as well as the nature of the polar group present in the upper rim of the calixarenes were varied. The lipid bilayer was formed with one calixarene and with the phospholipid 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine, DOPE. The cytotoxicity of the liposomes for various cell lines was also studied. From the results obtained, the liposomes formed with the least cytotoxic calixarene, (TEAC12)4, were used as nanocarriers of both nucleic acids and the antineoplastic drug doxorubicin, DOX. Results showed that (TEAC12)4/DOPE/p-EGFP-C1 lipoplexes, of a given composition, can transfect the genetic material, although the transfection efficiency substantially increases in the presence of an additional amount of DOPE as coadjuvant. On the other hand, the (TEAC12)4/DOPE liposomes present a high doxorubicin encapsulation efficiency, and a slow controlled release, which could diminish the side effects of the drug.

Concentration and medium micellar kinetic effects caused by morphological transitions.

The reaction methyl naphthalene-2-sulfonate + Br(-) was investigated in several alkanediyl-α-ω-bis(dodecyldimethylammonium) bromide, 12-s-12,2Br(-) (with s = 2, 3, 4, 5, 6, 8, 10, 12), micellar solutions in the absence and in the presence of various additives. The additives were 1,2-propylene glycol, which remains in the bulk phase, N-decyl N-methylglucamide, MEGA10, which forms mixed micelles with the dimeric surfactants, and 1-butanol, which distributes between the aqueous and micellar phases. Information about the micellar reaction media was obtained by using conductivity and fluorescence measurements. In all cases, with the exception of water-1,2-prop 12-5-12,2Br(-) micellar solutions, with 30% weight percentage of the organic solvent, a sphere-to-rod transition takes place upon increasing surfactant concentration. In order to quantitatively explain the experimental data within the whole surfactant concentration range, a kinetic equation based on the pseudophase kinetic model was considered, together with the decrease in the micellar ionization degree accompanying micellar growth. However, theoretical predictions did not agree with the experimental kinetic data for surfactant concentrations above the morphological transition. An empirical kinetic equation was proposed in order to explain the data. It contains a parameter b which is assumed to account for the medium micellar kinetic effects caused by the morphological transition. The use of this empirical equation permits the quantitative rationalization of the kinetic micellar effects in the whole surfactant concentration range.

Metallo-Liposomes of Ruthenium Used as Promising Vectors of Genetic Material.

Gene therapy is a therapeutic process consisting of the transport of genetic material into cells. The design and preparation of novel carriers to transport DNA is an important research line in the medical field. Hybrid compounds such as metallo-liposomes, containing a mixture of lipids, were prepared and characterized. Cationic metal lipids derived from the [Ru(bpy)3]2+ complex, RuC11C11 or RuC19C19, both with different hydrophobic/lipophilic ratios, were mixed with the phospholipid DOPE. A relation between the size and the molar fraction α was found and a multidisciplinary study about the interaction between the metallo-liposomes and DNA was performed. The metallo-liposomes/DNA association was quantified and a relationship between Kapp and α was obtained. Techniques such as AFM, SEM, zeta potential, dynamic light scattering and agarose gel electrophoresis demonstrated the formation of lipoplexes and showed the structure of the liposomes. L/D values corresponding to the polynucleotide's condensation were estimated. In vitro assays proved the low cell toxicity of the metallo-liposomes, lower for normal cells than for cancer cell lines, and a good internalization into cells. The latter as well as the transfection measurements carried out with plasmid DNA pEGFP-C1 have demonstrated a good availability of the Ru(II)-based liposomes for being used as non-toxic nanovectors in gene therapy.

Water-ethylene glycol cationic dimeric micellar solutions: aggregation, micellar growth, and characteristics as reaction media.

Effects of ethylene glycol (EG) addition on the micellization and on the micellar growth in two aqueous didodecyl dicationic dibromide surfactant, 12-s-12,2Br- (s=2, 6) solutions, with the weight percentage of EG up to 50%, have been investigated. An increment in the amount of EG makes the aggregation process less spontaneous due to the water-EG mixtures being better solvents for the cationic dimeric surfactant molecules than pure water (solvophobic effect). Results show that C*, the surfactant concentration where the sphere-to-rod transition occurs, increases when EG content in the bulk phase increases. The amount of the organic solvent influences C* principally through the decrease in the hydrocarbon/bulk phase interfacial tension (air/bulk phase surface tension) caused by its presence. Changes in the aggregation number, in the micropolarity, in the microviscosity, and in the rheological behavior accompanying micellar growth were studied in the water-EG micellar solutions. Kinetic studies provide information about the characteristics of the dimeric micelles as microreactors. Kinetic data also show that an increase in the surfactant concentration leads to micellar growth.

A Non-Viral Plasmid DNA Delivery System Consisting on a Lysine-Derived Cationic Lipid Mixed with a Fusogenic Lipid.

The insertion of biocompatible amino acid moieties in non-viral gene nanocarriers is an attractive approach that has been recently gaining interest. In this work, a cationic lipid, consisting of a lysine-derived moiety linked to a C12 chain (LYCl) was combined with a common fusogenic helper lipid (DOPE) and evaluated as a potential vehicle to transfect two plasmid DNAs (encoding green fluorescent protein GFP and luciferase) into COS-7 cells. A multidisciplinary approach has been followed: (i) biophysical characterization based on zeta potential, gel electrophoresis, small-angle X-ray scattering (SAXS), and cryo-transmission electronic microscopy (cryo-TEM); (ii) biological studies by fluorescence assisted cell sorting (FACS), luminometry, and cytotoxicity experiments; and (iii) a computational study of the formation of lipid bilayers and their subsequent stabilization with DNA. The results indicate that LYCl/DOPE nanocarriers are capable of compacting the pDNAs and protecting them efficiently against DNase I degradation, by forming Lα lyotropic liquid crystal phases, with an average size of ~200 nm and low polydispersity that facilitate the cellular uptake process. The computational results confirmed that the LYCl/DOPE lipid bilayers are stable and also capable of stabilizing DNA fragments via lipoplex formation, with dimensions consistent with experimental values. The optimum formulations (found at 20% of LYCl content) were able to complete the transfection process efficiently and with high cell viabilities, even improving the outcomes of the positive control Lipo2000*.

Influence of the degree of oligomerization of surfactants on the DNA/surfactant interaction.

The interaction between calf thymus DNA, ctDNA, and a series of oligomeric surfactants derived from N-benzyl-N,N-dimethyl-N-(1-dodecyl)ammonium chloride is investigated. The influence of the surfactants' degree of oligomerization (2, 3 and 4) on the ctDNA/surfactant interaction is studied, as well as the effect of the structure of the spacer group linking the individual surfactant fragments. In particular, the effect of the distance between the positive charges and the hydrophobic chains within the oligomers on these interactions was examined, by using the three positional isomers (i.e., ortho-, meta-, and para-) with the rigid xylidene moiety as spacer. Results show that the dimeric ("gemini") surfactants are much more efficient in the inversion of the nucleic acid charge than the single-chained (monomeric) surfactant. Whereas the ortho - isomer causes a partial condensation, the meta - and para - isomers can completely condense ctDNA. The meta - and para - isomers of the trimeric surfactants can also completely condense the polynucleotide. In contrast, the tetrameric surfactant investigated does not change the morphology of the nucleic acid from an elongated coil into a compacted form, in spite of effectively inverting the nucleic acid's charge in their complex. Accordingly, the capacity for ctDNA compaction of oligomeric surfactants is not simply correlated to their degree of oligomerization, but depends on a complex balance of the number and relative distance of cationic charges and/or hydrophobic tails in the surfactants for effectively interacting with the nucleic acid to form the appropriate complex. This information will help to design more effective cationic surfactants as non-viral vectors for gene therapy.

Preparation and Characterization of New Liposomes. Bactericidal Activity of Cefepime Encapsulated into Cationic Liposomes.

Cefepime is an antibiotic with a broad spectrum of antimicrobial activity. However, this antibiotic has several side effects and a high degradation rate. For this reason, the preparation and characterization of new liposomes that are able to encapsulate this antibiotic seem to be an important research line in the pharmaceutical industry. Anionic and cationic liposomes were prepared and characterized. All cationic structures contained the same cationic surfactant, N,N,N-triethyl-N-(12-naphthoxydodecyl)ammonium. Results showed a better encapsulation-efficiency percentage (EE%) of cefepime in liposomes with phosphatidylcholine and cholesterol than with 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE). The presence of cholesterol and the quantity of egg-yolk phospholipid in the liposome increased the encapsulation percentage. The bactericidal activity against Escherichia coli of cefepime loaded into liposomes with phosphatidylcholine was measured. The inhibitory zone in an agar plate for free cefepime was similar to that obtained for loaded cefepime. The growth-rate constant of E. coli culture was also measured in working conditions. The liposome without any antibiotic exerted no influence in such a rate constant. All obtained results suggest that PC:CH:12NBr liposomes are biocompatible nanocarriers of cefepime that can be used in bacterial infections against Escherichia coli with high inhibitory activity.

Optimized Preparation of Levofloxacin Loaded Polymeric Nanoparticles.

In this work, poly(lactic-co-glycolic acid) (PLGA) and chitosan (CS) nanoparticles were synthesized with the purpose of encapsulating levofloxacin (LEV). A thorough study has been carried out in order to optimize the preparation of LEV-loaded polymeric nanoparticles (NPs) suitable for parenteral administration. Changes in the preparation method, in the organic solvent nature, in the pH of the aqueous phase, or in the temperature were investigated. To the authors´ knowledge, a systematic study in order to improve the LEV nanocarrier characteristics and the yield of drug encapsulation has not been carried out to date. The physicochemical characterization of the NPs, their encapsulation efficiency (EE), and the in vitro release of LEV revealed that the best formulation was the emulsion-solvent evaporation method using dichloromethane as organic solvent, which renders suitable LEV loaded PLGA NPs. The morphology of these NPs was investigated using TEM. Their antimicrobial activities against several microorganisms were determined in vitro measuring the minimum inhibitory concentration (MIC). The results show that the use of these loaded LEV PLGA nanoparticles has the advantage of the slow release of the antibiotic, which would permit an increase in the time period between administrations as well as to decrease the side effects of the drug.

Mixtures of monomeric and dimeric surfactants: hydrophobic chain length and spacer group length effects on non ideality.

Critical micelle concentrations of the Cm TAB+12- s-12 (s=3, 4, 5 and m=10, 12, 14, 16) binary systems have been determined, through conductivity and fluorescence measurements, at 298 K. Application of different theoretical approaches to explain mixed micellization shows that non-ideality of the binary systems follows the trend C16TAB+12-3-12

Importance of hydrophobic interactions in the single-chained cationic surfactant-DNA complexation.

The goal of this work was to understand the key factors determining the DNA compacting capacity of single-chained cationic surfactants. Fluorescence, zeta potential, circular dichroism, gel electrophoresis and AFM measurements were carried out in order to study the condensation of the nucleic acid resulting from the formation of the surfactant-DNA complexes. The apparent equilibrium binding constant of the surfactants to the nucleic acid, Kapp, estimated from the experimental results obtained in the ethidium bromide competitive binding experiments, can be considered directly related to the ability of a given surfactant as a DNA compacting agent. The plot of ln(Kapp) vs. ln(cmc), cmc being the critical micelle concentration, for all the bromide and chloride surfactants studied, was found to be a reasonably good linear correlation. This result shows that hydrophobic interactions mainly control the surfactant DNA compaction efficiency.

Role of the solvophobic effect on micellization.

Experimental data of amphiphiles aggregation phenomena in water-organic solvent mixtures were considered with the idea of investigating the role of the solvophobic effect on micellization. Changes in the critical micelle concentration, in the micellar ionization degree (for ionic surfactants) and in the aggregation number accompanying variations in the composition of the bulk phase of the micellar solutions were examined with the scope of understanding which properties of the water-organic solvent mixtures are important in the micellization process. Results point out that the cohesive energy density, measured either through the Hildebrand-Hansen solubility parameter or the Gordon parameter, seems to play an important role in determining the contribution of the solvophobic effect on the Gibbs energy of micellization in water-organic solvents mixtures.

Micellar kinetic effects in gemini micellar solutions: influence of sphere-to-rod transitions on kinetics.

The reactions 4-nitrobenzenesulfonate+Br(-) and methyl naphthalene-2-sulfonate+Br(-) were studied in various water-ethylene glycol, EG, [C(12)H(25)(CH(3))(2)N(CH(2))(s)N(CH(3))(2)C(12)H(25)]Br(2) micellar solutions (12-s-12,2Br(-) with s=3-5 methylene groups). Results showed that the observed rate constant of the two reactions varied when a sphere-to-rod transition occurs. This morphological transition is accompanied by changes in the interfacial region water content and in its polarity. The micellar ionization degree is also altered, making the counterion interfacial concentration change. Finally, variations on the molar surfactant volume, V(m), also follow the sphere-to-rod transitions. A simple pseudophase kinetic model is inadequate for quantitatively discussing the kinetic micellar effects observed, since the changes accompanying micellar growth affect the second-order rate constant in the micellar pseudophase, the equilibrium binding constant and the surfactant molar volume, neither of them remaining constant in the whole surfactant concentration range. However, this simple model can be helpful in the treatment of kinetic data for surfactant concentrations below the morphological transitions, providing some interesting, although approximate, information.

Stoppering/unstoppering of a rotaxane formed between an N-hetorycle ligand containing surfactant: ß-cyclodextrin pseudorotaxane and pentacyanoferrate(II) ions.

The assembly of a surfactant-based rotaxane by adding the labile aquopentacyanoferrate(II) ion to the previously formed pseudorotaxane between the surfactant 11-(isonicotinoyloxy)-N,N,N-triethyl-1-undecanaminium bromide and ß-cyclodextrin was investigated by 1H NMR and kinetic measurements. NMR spectroscopy has showed that the rotaxane can be formed through two different mechanisms. The rotaxane can be unstoppered by using the pyridine ligand substitution reaction by the high-field cyanide ligand. In this work a new method is developed for the preparation of several new surfactant-based rotaxanes by changing the hydrophilic and hydrophobic regions of the surfactants and the nature of the macrocycle.

Host-guest interactions between cyclodextrins and surfactants with functional groups at the end of the hydrophobic tail.

The aim of this work was to investigate the influence of the incorporation of substituents at the end of the hydrophobic tail on the binding of cationic surfactants to α-, ß-, and γ-cyclodextrins. The equilibrium binding constants of the 1:1 inclusion complexes formed follow the trend K1(α-CD)>K1(ß-CD)≫K1(γ-CD), which can be explained by considering the influence of the CD cavity volume on the host-guest interactions. From the comparison of the K1 values obtained for dodecyltriethylammonium bromide, DTEAB, to those estimated for the surfactants with the substituents, it was found that the incorporation of a phenoxy group at the end of the hydrocarbon tail does not affect K1, and the inclusion of a naphthoxy group has some influence on the association process, slightly diminishing K1. This makes evident the importance of the contribution of hydrophobic interactions to the binding, the length of the hydrophobic chain being the key factor determining K1. However, the presence of the aromatic rings does influence the location of the host and the guest in the inclusion complexes. The observed NOE interactions between the aromatic protons and the CD protons indicate that the aromatic rings are partially inserted within the host cavity, with the cyclodextrin remaining close to the aromatic rings, which could be partially intercalated in the host cavity. To the authors' knowledge this is the first study on the association of cyclodextrins with monomeric surfactants incorporating substituents at the end of the hydrophobic tail.