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International audience; We have studied the effect of different ions belonging to the extended Hofmeister series on the thermosensitive polymer poly(N-isopropylacrylamide) (PNIPAM), by combining Differential Scanning Calorimetry (DSC) and Dynamic Light Scattering (DLS). The variations in the lower critical solution temperature (TLCS) and enthalpy change during PNIPAM phase separation evidence the importance of considering both hydration and hydrophobicity to explain the interaction of ions with interfaces. The results obtained in the presence of inorganic ions can be explained by the tendency of water molecules to preferentially hydrate the PNIPAM chains or the ions, depending on the kosmotropic (highly hydrated) or chaotropic (poorly hydrated) character of the ions. On the contrary, tetraphenyl organic ions (Ph4B− and Ph4As+) interact with the hydrophobic moieties of PNIPAM chains, inducing a significant reduction of the TLCS. DLS results show that the aggregation state of PNIPAM above the TLCS is also strongly influenced by the presence of ions. While macroscopic phase separation (formation of a polymer-rich phase insoluble in water) was apparent in the presence of inorganic ions, we observed the formation of submicron PNIPAM aggregates at temperatures above the TLCS in the presence of the hydrophobic ions. Kinetically arrested monodisperse PNIPAM nanoparticles were formed in the presence of the Ph4B− anion, while a rather polydisperse distribution of particle sizes was observed in the presence of Ph4As+. These results show that ionic specificity influences both the static (thermodynamic) and dynamic (kinetically controlled aggregation) states of PNIPAM in an aqueous environment.

International audience; A full molecular-level understanding of protein adsorption in important situations such as the formation of protein films at solid/liquid interfaces or the formation of a protein corona over inorganic nanoparticles is still lacking. All-atomic implicit solvent molecular dynamics (MD) simulations, which are successfully employed in many related protein studies (such as protein folding for example) are emerging also as a useful tool to investigate proteins at surfaces. Implicit solvent simulations replace the detailed description of the solvent by a continuum media and effective atom–atom interactions retaining the atomistic detail in the description of the system of interest. This allows the simulation of larger systems and longer time scalesas compared with full MD simulations including explicit solvent. In this brief review, we present an overview of the current state of the application of this technique to the study of problems such as the interaction of proteins with solid surfaces and the structure of protein corona over inorganic nanoparticles. Limitations of the approach and future perspectives are also outlined.

Ionic specificity, also known as Hofmeister effects, is a widely studied phenomenon related with the ability of ions of the same valence to produce different effects over many interfaces. Anions and cations were arranged by Hofmeister in 1888 according to their capability of precipitating proteins. Ions are classified in these series as kosmotropes (strongly hydrated) or chaotropes (weakly hydrated). In addition, anions usually produce more intense effects than cations. Many authors have developed different models to explain ion specific phenomena considering ion characteristics as size, polarizability or ion hydration; and ion-surface interactions by dispersion forces, chemical bindings or water hydration layers. Nevertheless, in spite of the performed advances in this area, a detailed microscopic theory to completely understand ionic specificity is still absent. This thesis aims to elucidate the subjacent microscopic mechanisms in ionic specificity. For that, experiments, simulations and theoretical studies with different systems have been performed. We propose that ion accumulation or exclusion from interfaces is mediated by the interaction of the solvent with both, ions and surfaces. Special emphasis has been given to organic ions. The tetraphenyl ions are big hydrophobic ions recently added to the Hofmeister series as superchaotropes. These organic ions allow us to explore further in ion specific effects, since they have a deep impact on interfaces, in particular on hydrophobic soft systems as polymers or proteins. An attractive characteristic of the tetraphenyl ions is that the anion (Ph4B−) and the cation (Ph4As+) are nearly identical. We have demonstrated that the anion and the cation show different behaviors in solution and when they interact with other surfaces in spite of their similar chemical structure. Moreover, the anion always shows stronger specific effects than the cation. In addition, this difference is greater when ions interact with the thermoresponsive polymer PNIPAM (Poly (N-isopropylacrylamide)) instead of hard surfaces. This favorable interaction is mediated by the hydrophobic effect. PNIPAM undergoes a sharp and reversible transition from a soluble to an insoluble state in aqueous solution. The transition temperature can be modified by the ionic conditions of the medium. We have studied this transition temperature with inorganic and organic ions. We have observed that the transition depends on the capacity of the ions to compete with the water hydrating PNIPAM and also to the affinity of the poorly hydrated ions to accumulate on the hydrophobic moieties of the PNIPAM. The great ion accumulation in the insoluble state of PNIPAM produces important charge reversal of charged PNIPAM microparticles, when these last ions act as counterions. On the other hand, several proteins of biotechnological interest and different degree of hydrophobicity were studied by physical adsorption of the different proteins onto hydrophobic surfaces. We have observed that the accumulation of the tetraphenyl ions is greater on increasingly hydrophobic interfaces. In addition, these large ions interact so strongly with the soft biomolecules that proteins undergo conformational changes. Proteins films, in water, are swollen when ions enhance their electrostatic charge, while these layers are compressed when ions screen the electrostatic repulsion between molecules. Nevertheless, the ionic concentration needed to achieve similar states of structuring protein is one order of magnitude lower for Ph4B− than for Ph4As+ corroborating that the anion causes more intense effects than the cation. To carry out all this study we have used several experimental techniques, the most relevant of them have been: electrophoretic mobility, Atomic Force Microscopy (AFM), Quartz Crystal Microbalance (QCM), and Differential Scanning Calorimeter (DSC). In addition, Molecular Dynamics (MD) simulations were developed to analyze at microscopic level the interaction between ions and interfaces. La presente tesis tiene como objetivo dilucidar los mecanismos microscópicos que dan lugar a la especificidad iónica. Para ello, se han realizado experimentos, simulaciones y estudios teóricos con diferentes sistemas. Se propone que la acumulación o exclusión de los iones en las interfaces está mediada por la interacción del solvente tanto con iones como con superficies. Se ha hecho especial hincapié en los iones orgánicos. Los iones tetrafenil son iones hidrófobos de gran tamaño que han sido recientemente añadidos a las series de Hofmeister como súper caotrópicos. Estos iones orgánicos nos permiten explorar la especificidad iónica desde otro punto de vista. Los iones orgánicos interaccionan mucho más fuerte con las interfaces que los iones inorgánicos, sobre todo en sistemas hidrófobos de tipo soft como polímeros o proteínas. Una característica muy relevante de los iones tetrafenil es que el anión (Ph4B−) y el catión (Ph4As+) son prácticamente idénticos. A pesar de su similar estructura química, hemos demostrado que el anión y el catión muestran comportamientos diferentes en solución y cuando interactúan con otras superficies. De hecho, el anión siempre muestra efectos específicos más acusados que el catión., Tesis Univ. Granada. Programa Oficial de Doctorado en: Física y Ciencias del Espacio

Here, we study films of proteins over planar surfaces and protein-coated microspheres obtained from the adsorption of three different proteins ( β -casein, β -lactoglobulin and bovine serum albumin (BSA)). The investigation of protein films in planar surfaces is performed by combining quartz crystal microbalance (QCM) and atomic force microscopy (AFM) measurements with all-atomic molecular dynamics (MD) simulations. We found that BSA and β -lactoglobulin form compact monolayers, almost without interstices between the proteins. However, β -casein adsorbs forming multilayers. The study of the electrokinetic mobility of protein-coated latex microspheres shows substantial condensation of ions from the buffer over the complexes, as predicted from ion condensation theories. The electrokinetic behavior of the latex-protein complexes is dominated by the charge of the proteins and the phenomenon of ion condensation, whereas the charge of the latex colloids plays only a minor role., The authors wish to thank the financial support granted by the project CTS-6270 (Junta de Andalucía, Spain) and the Spanish “Ministerio de Economía y Competitividad (MINECO), Plan Nacional de Investigación, Desarrollo e Innovación Tecnológica (I + D + i)” (Project FIS2016-80087-C2-1-P). J.F.acknowledges financial support from the Spanish Ministry of Economy and Competitiveness, through the “Severo Ochoa” Programme for Centres of Excellence in R & D (SEV-2015-0496) awarded to ICMAB. L.P.-F. thanks the financial support provided by the COST Action MP1303: Understanding and Controlling Nano and Mesoscale Friction.

In this study we have investigated how different proteins interact with big organic ions. Two ions that are similar in size and chemical structure (Ph4B− anion and Ph4As+ cation) were studied. The proteins chosen are the two major allergenic proteins of cow's milk, β-lactoglobulin and β-casein, and bovine serum albumin, BSA, as the reference protein. First, a quantitative study to determine the hydrophobic degree of the proteins was performed. Then, electrokinetic and stability measurements on protein-coated polystyrene (PS) microspheres as a function of the tetraphenyl ion concentration were carried out. Our results show that the affinity of the organic ions depends on the hydrophobicity of the interface. Big charge inversions and re-stabilization patterns were observed at very low concentrations of tetraphenyl ions for the most hydrophobic protein studied (with β-casein). Besides, the ionic concentrations needed to destabilize these colloidal systems were roughly one order of magnitude lower for the anion than for the cation. In addition, we studied conformational changes of the adsorbed proteins with a quartz crystal microbalance. Proteins were adsorbed onto hydrophobic flat substrates and then exposed to the tetraphenyl ions. The protein films swelled or collapsed as a function of the accumulation of tetraphenyl ions. Similarly to the electrokinetic/stability studies, the ionic concentration necessary to trigger structural changes of the protein films was one order of magnitude larger for the cation than for the anion. All the results evidence that the accumulation of these organic ions on an interface depends directly on its degree of hydrophobicity. We attribute the different interactions of the anion and the cation with these interfaces to their dissimilar hydration, which makes the anion show a more hydrophobic behaviour than the cation.

Minute concentrations of big hydrophobic ions have the ability to induce substantial effects in soft matter systems, including novel phases in lipid layers, giant charge inversion in colloids and nanostructuration in polymer surfaces in contact with water. The effects are so strong that the term "soft matter disruptors" was coined to describe their deep impact on interfaces, which goes far beyond that found by using the classical ions considered in lyotropic (Hofmeister) sequences. In these effects, solvation thermodynamics plays a fundamental role. Interestingly, it is possible to obtain big hydrophobic cations and anions with an almost identical size and structure (e.g. Ph4B(-), Ph4As(+)), which only differ in their central atom. Here we employ different techniques (Molecular Dynamics (MD) simulations, electrophoretic mobility and Atomic Force Microscopy (AFM)) to demonstrate the dramatic differences in the interaction of Ph4B(-) and Ph4As(+) with poly(N-isopropylacrylamide) (PNIPAM), a thermoresponsive polymer with expanded (well hydrated) and collapsed (poorly hydrated) states. Although both ions interact strongly with neutral PNIPAM chains and cationic or anionic PNIPAM microgels in the collapsed states, the effects of Ph4B(-) on PNIPAM are always substantially stronger than the effects of Ph4As(+). MD simulations predict that ion-PNIPAM free energy of interaction is four times larger for Ph4B(-) than for Ph4As(+). Electrokinetic and AFM experiments show that, acting as counter-ions, both ions are able to invert the charge of anionic or cationic PNIPAM microgels at minute concentrations, but the charge inversion due to Ph4B(-) is much larger than that obtained with Ph4As(+). Therefore, even for big ions of identical size, shape and valence, the affinity of anions and cations for interfaces is intrinsically different.