Vincent Ball

Polydopamine and Eumelanin: From Structure–Property Relationships to a Unified Tailoring Strategy

CONSPECTUS: Polydopamine (PDA), a black insoluble biopolymer produced by autoxidation of the catecholamine neurotransmitter dopamine (DA), and synthetic eumelanin polymers modeled to the black functional pigments of human skin, hair, and eyes have burst into the scene of materials science as versatile bioinspired functional systems for a very broad range of applications. PDA is characterized by extraordinary adhesion properties providing efficient and universal surface coating for diverse settings that include drug delivery, microfluidic systems, and water-treatment devices. Synthetic eumelanins from dopa or 5,6-dihydroxyindoles are the focus of increasing interest as UV-absorbing agents, antioxidants, free radical scavengers, and water-dependent hybrid electronic-ionic semiconductors. Because of their peculiar physicochemical properties, eumelanins and PDA hold considerable promise in nanomedicine and bioelectronics, as they are biocompatible, biodegradable, and exhibit suitable mechanical properties for integration with biological tissues. Despite considerable similarities, very few attempts have so far been made to provide an integrated unifying perspective of these two fields of technology-oriented chemical research, and progress toward application has been based more on empirical approaches than on a solid conceptual framework of structure-property relationships. The present Account is an attempt to fill this gap. Following a vis-à-vis of PDA and eumelanin chemistries, it provides an overall view of the various levels of chemical disorder in both systems and draws simple correlations with physicochemical properties based on experimental and computational approaches. The potential of large-scale simulations to capture the macroproperties of eumelanin-like materials and their hierarchical structures, to predict the physicochemical properties of new melanin-inspired materials, to understand the structure-property-function relationships of these materials from the bottom up, and to design and optimize materials to achieve desired properties is illustrated. The impact of synthetic conditions on melanin structure and physicochemical properties is systematically discussed for the first time. Rational tailoring strategies directed to critical control points of the synthetic pathways, such as dopaquinone, DAquinone, and dopachrome, are then proposed, with a view to translating basic chemical knowledge into practical guidelines for material manipulation and tailoring. This key concept is exemplified by the recent demonstration that varying DA concentration, or using Tris instead of phosphate as the buffer, results in PDA materials with quite different structural properties. Realizing that PDA and synthetic eumelanins belong to the same family of functional materials may foster unprecedented synergisms between research fields that have so far been apart in the pursuit of tailorable and marketable materials for energy, biomedical, and environmental applications.

Mechanotransductive surfaces for reversible biocatalysis activation

Fibronectin, like other proteins involved in mechanotransduction, has the ability to exhibit recognition sites under mechanical stretch. Such cryptic sites are buried inside the protein structure in the native fold and become exposed under an applied force, thereby activating specific signalling pathways. Here, we report the design of new active polymeric nanoassembled surfaces that show some similarities to these cryptic sites. These nanoassemblies consist of a first polyelectrolyte multilayer stratum loaded with enzymes and capped with a second polyelectrolyte multilayer acting as a mechanically sensitive nanobarrier. The biocatalytic activity of the film is switched on/off reversibly by mechanical stretching, which exposes enzymes through the capping barrier, similarly to mechanisms involved in proteins during mechanotransduction. This first example of a new class of biologically inspired surfaces should have great potential in the design of various devices aimed to trigger and modulate chemical reactions by mechanical action with applications in the field of microfluidic devices or mechanically controlled biopatches for example.

Self-assembly of tetramers of 5,6-dihydroxyindole explains the primary physical properties of eumelanin: experiment, simulation, and design.

Eumelanin is a ubiquitous pigment in nature and has many intriguing physicochemical properties, such as broad-band and monotonous absorption spectrum, antioxidant and free radical scavenging behavior, and strong nonradiative relaxation of photoexcited electronic states. These properties are highly related to its structural and mechanical properties and make eumelanin a fascinating candidate for the design of multifunctional nanomaterials. Here we report joint experimental-computational investigation of the structural and mechanical properties of eumelanin assemblies produced from dopamine, revealing that the mass density of dry eumelanin is 1.55 g/cm³ and its Youngs modulus is ≈5 GPa. We also find that wet eumelanin has a lower mass density and Youngs modulus depending on the water-to-melanin ratio. Most importantly, our data show that eumelanin molecules tend to form secondary structures based on noncovalent π stacking in both dry and wet conditions, with an interlayer distance between eumelanin molecules of 3.3 Å. Corresponding transmission electron microscope images confirm the supramolecular organization predicted in our simulations. Our simulations show that eumelanin is an isotropic material at a larger scale when eumelanin molecules are randomly oriented to form secondary structures. These results are in good agreement with experimental observations, density functional theory calculations, and bridge the gap between earlier experimental and small-scale quantum mechanical studies of eumelanin. We use the knowledge acquired from the simulations to select a partner molecule, a cationic phthalocyanine, allowing us to produce layer-by-layer films containing eumelanin that display an electrical conductivity 5 orders of magnitudes higher than that of pure eumelanin films.

Buffer dependence of refractive index increments of protein solutions

The refractive index increment of a protein solution is a property not only of the protein, but also of the solvent. This is demonstrated theoretically and confirmed experimentally using analytical interferometry.

Embedded silver ions-containing liposomes in polyelectrolyte multilayers: Cargos films for antibacterial agents

A new antibacterial coating made of poly(L-lysine)/hyaluronic acid (PLL/HA) multilayer films and liposome aggregates loaded with silver ions was designed. Liposomes filled with an AgNO 3 solution were first aggregated by the addition of PLL in solution. The obtained micrometer-sized aggregates were then deposited on a PLL/HA multilayer film, playing the role of a spacer with the support. Finally, HA/PLL/HA capping layers were deposited on top of the architecture to form a composite AgNO 3 coating. Release of encapsulated AgNO 3 from this composite coating was followed and triggered upon temperature increase over the transition temperature of vesicles, found to be equal to 34 degrees C. After determination of the minimal inhibitory concentration (MIC) of AgNO 3 in solution, the antibacterial activity of the AgNO 3 coating was investigated against Escherichia coli. A 4-log reduction in the number of viable E. coli cells was observed after contact for 120 min with a 120 ng/cm (2) AgNO 3 coating. In comparison, no bactericidal activity was found for PLL/HA films previously dipped in an AgNO 3 solution and for PLL/HA films with liposome aggregates containing no AgNO 3 solution. The strong bactericidal effect could be linked to the diffusion of silver ions out of the AgNO 3 coating, leading to an important bactericidal concentration close to the membrane of the bacteria. A simple method to prepare antibacterial coatings loaded with a high and controlled amount of AgNO 3 is therefore proposed. This procedure is far superior to that soaking AgNO 3 or Ag nanoparticles into a coating. In principle, other small bactericidal chemicals like antibiotics could be encapsulated by this method. This study opens a new route to modify surfaces with small solutes that are not permeating phospholipid membranes below the phase transition temperature.

Protein adsorption on dopamine-melanin films: Role of electrostatic interactions inferred from (ζ-potential measurements versus chemisorption

We recently showed the possibility to build dopamine-melanin films of controlled thickness by successive immersions of a substrate in alkaline solutions of dopamine [F. Bernsmann, A. Ponche, C. Ringwald, J. Hemmerlé, J. Raya, B. Bechinger, J.-C. Voegel, P. Schaaf, V. Ball, J. Phys. Chem. C 113 (2009) 8234-8242]. In this work the structure and properties of such films are further explored. The zeta-potential of dopamine-melanin films is measured as a function of the total immersion time to build the film. It appears that the film bears a constant zeta-potential of (-39+/-3) mV after 12 immersion steps. These data are used to calculate the surface density of charged groups of the dopamine-melanin films at pH 8.5 that are mostly catechol or quinone imine chemical groups. Furthermore the zeta-potential is used to explain the adsorption of three model proteins (lysozyme, myoglobin, alpha-lactalbumin), which is monitored by quartz crystal microbalance. We come to the conclusion that protein adsorption on dopamine-melanin is not only determined by possible covalent binding between amino groups of the proteins and catechol groups of dopamine-melanin but that electrostatic interactions contribute to protein binding. Part of the adsorbed proteins can be desorbed by sodium dodecylsulfate solutions at the critical micellar concentration. The fraction of weakly bound proteins decreases with their isoelectric point. Additionally the number of available sites for covalent binding of amino groups on melanin grains is quantified.

Reversible loading and unloading of nanoparticles in "exponentially" growing polyelectrolyte LBL films.

The exponentially growing layer-by-layer (LBL) films made from poly(diallyldimethylammonium chloride) (PDDA) and poly(acrylic acid) (PAA) were used to load and unload the CdTe nanoparticles (NPs). The reversible loading of NPs were investigated through UV-vis studies and further confirmed by confocal microscopy. In addition the LBL films were also compared for the release kinetics for pH 9 and 7 and films capped with (PDDA-PSS)10 layers. The amount of released particles at pH 9 was found to be at least 2 orders of magnitude higher than those at pH 7 and with (PDDA-PSS)10 capped layers after 25 h. This variation in film response for CdTe-particle release presents a route for studies in which highly swollen exponentially growing LBL films can be loaded with functionalized NPs for biological applications and explored as carriers to hold the NPs inside the films for self-assembly.

Isothermal microcalorimetry to investigate non specific interactions in biophysical chemistry.

Isothermal titration microcalorimetry (ITC) is mostly used to investigate the thermodynamics of “specific” host-guest interactions in biology as well as in supramolecular chemistry. The aim of this review is to demonstrate that ITC can also provide useful information about non-specific interactions, like electrostatic or hydrophobic interactions. More attention will be given in the use of ITC to investigate polyelectrolyte-polyelectrolyte (in particular DNA-polycation), polyelectrolyte-protein as well as protein-lipid interactions. We will emphasize that in most cases these “non specific” interactions, as their definition will indicate, are favoured or even driven by an increase in the entropy of the system. The origin of this entropy increase will be discussed for some particular systems. We will also show that in many cases entropy-enthalpy compensation phenomena occur.

Effective embedding of liposomes into polyelectrolyte multilayered films: the relative importance of lipid-polyelectrolyte and interpolyelectrolyte interactions

The layer-by-layer (LbL) self-assembly of polyelectrolytes has emerged as a powerful and versatile strategy for engineering (bio)surfaces with active compounds. One possibility to engineer such films is to dope them with nanoparticles or with intact vesicles as carriers filled with the compounds of interest. In our previous studies we demonstrated that intact vesicles stabilized by poly-L-lysine (PLL) coating can be embedded in polyelectrolyte multilayer films made from the combination of either poly-L-glutamic acid (PGA) and poly(allylamine hydrochloride) (PAH) or from the combination of hyaluronic acid (HA) and PLL. These findings were empirical and not correlated to the interaction mode between PLL decorated vesicles (PLL-Lip) with the multilayer film. In order to produce robust and stable multi-layered nanocoatings containing intact vesicles, one has to understand how PLL-coated vesicles are integrated in the LbL architecture. To that aim, vesicle adsorption on three kinds of films, namely (PAH/PSS)12, where PSS stands for poly-(4-styrenesulfonate), (PAH/PGA)12, and (PLL/HA)12 was studied. PLL-Lip adsorption strongly depends on the interactions between PLL with both the vesicles and the polyanions used to form polyelectrolyte multilayers as was shown by direct evidence with confocal and atomic force microscopy as well as by the investigation of the interaction between PLL-Lip with the three polyanions in solution, namely HA, PGA and PSS. The latest was investigated by means of isothermal titration microcalorimetry. As a result, PSS in solution is able to desorb PLL from PLL coated liposomes, and on the other hand, PGA and HA can adsorb on PLL-stabilized vesicles without desorbing or inducing only partial desorption of the pre-adsorbed PLL. As a consequence, the (PLL/HA)12 film was found to be the best system among the three investigated combinations of polycations/polyanions for vesicle entrapment. The amount of entrapped vesicles is then proportional to the number of vesicle deposition steps and is increased for liposomes with higher charge. The release of a dye encapsulated in the entrapped vesicles could be induced by temperature changes which shows that for the HA-PLL combination of polyelectrolytes, the embedded vesicles retain their integrity below the main phase transition temperature. The temperature increase leads to fast leakage of liposome cargo while the film structure is not changed (up to 45 °C) suggesting that the lipidic bilayer is destabilized by ionic contacts with a polyelectrolyte network of the film. The latest is supported by the fact that within increase of vesicle negative charge, the stability of film-embedded vesicles is decreased whereas stability of these vesicles in solution is increased.

Hole formation induced by ionic strength increase in exponentially growing multilayer films

Polyelectrolyte multilayer (PEM) films consist of polyanion/polycation super-structures that are sensitive to various stresses like ionic strength changes. We investigate the swelling process of the exponentially growing poly(L-lysine)/hyaluronic acid (PLL/HA) films induced by changes of the ionic strength of the contact solution. We show that above a first critical ionic strength the swelling is accompanied by a release of both polyelectrolytes constituting the film, leading to its subsequent dissolution. At a second critical ionic strength, the swelling of the multilayer is so important that, in addition to this polyelectrolyte release, formation of spherical holes is observed inside the film. The presence of dissolved PLL and HA chains in these holes leads to an increase of the concentration of their counterions inside of them, and thus induces an extra osmotic pressure. This in turn favors the size increase of the holes before they coalesce. The release of both polyelectrolytes from the film into the supernatant ultimately allows a decrease of the osmotic pressure inside the PLL/HA film, which finally leads to the disappearance of the holes and concomitantly to a complete dissolution of the film. When the release of polyelectrolytes into the solution is prevented by a poly(diallyldimethyl ammonium chloride)/poly(styrene sulfonate) (PDADMAC/PSS) capping film, the holes appear at a smaller critical ionic strength compared to uncapped films. Here too the formation of the holes is attributed to an increase of the osmotic pressure inside the film. As soon as the capping barrier ruptures because of the swelling of the film, both PLL and HA chains can diffuse out of the film and the holes decrease in size and disappear, as does the film.