Accurate models of protein diffusion are important in a no. of applications, including liq.-liq. phase sepn. and growth of protein crystals for X-ray diffraction studies. In concd. multicomponent protein systems, significant deviations from pseudobinary behavior can be expected. Rayleigh interferometry is used to measure the four elements (Dij)v of the ternary diffusion coeff. matrix for the extensively investigated protein, hen egg-white lysozyme (component 1) in aq. NaCl (component 2) at pH 4.5 and 25 °C. These are the first multicomponent diffusion coeffs. measured for any protein system at concns. high enough to be relevant to modeling and prediction of crystal growth or other phase transitions, and the first for a system involving lysozyme at any concn. The four ternary diffusion coeffs. for the system lysozyme chloride/NaCl/water are reported for lysozyme chloride at 0.60 mM (8.6 mg/mL) and NaCl at concns. of 0.25, 0.50, 0.65, 0.90, and 1.30 M (1.4, 2.8, 3.7, 5.1, and 7.2 wt %), with the latter two compns. being supersatd. One cross-term, (D21)v, is 80-259 times larger than the main term (D11)v and 7-18 times larger than (D22)v. Std. interferometric diagnostic tests indicate that aggregation is unimportant in our expts. We also present binary diffusion coeffs. Dv for lysozyme chloride/water at concns. from 0.43 to 3.08 mM (6.2-44.1 mg/mL), at the same pH and temp. The precision of the results is about 0.1% for the binary diffusion coeffs. and diagonal ternary diffusion coeffs., and about 1-2% for the cross-terms. For the ternary systems investigated, we show that a single pseudobinary diffusion coeff. does not accurately describe diffusive transport, and predictions by simple models such as the Nernst-Hartley equations are inaccurate at the higher concns. considered here. Finally, dynamic light-scattering diffusion coeffs., differing from both our interferometrically measured (Dij)v and a theor. prediction of light-scattering diffusion coeffs. in multicomponent systems, are reported for the same solns. used for the ternary expts. at 1.30 M

Despite the growing technological interest of polydopamine (dopamine melanin)-based coatings for a broad variety of applications, the factors governing particle size, shape, and electronic properties of this bioinspired multifunctional material have remained little understood. Herein, we report a detailed characterization of polydopamine growth, particle morphology, and paramagnetic properties as a function of dopamine concentration and nature of the buffer (pH 8.5). Dynamic Light Scattering data revealed an increase in the hydrodynamic radii (Rh) of melanin particles with increasing dopamine concentration in all buffers examined, especially in phosphate buffer. Conversely, a marked inhibition of particle growth was apparent in Tris buffer, with Rh remaining as low as <100 nm during polymerization of 0.5 mM dopamine. Small angle neutron scattering data suggested formation of bidimensional structures in phosphate or bicarbonate buffers, while apparently three-dimensional fractal objects prevailed in Tris buffer. Finally, electron paramagnetic resonance spectra revealed a broader signal amplitude with a peculiar power saturation decay profile for polydopamine samples prepared in Tris buffer, denoting more homogeneous paramagnetic centers with respect to similar samples obtained in phosphate and bicarbonate buffers. Overall, these results disclose Tris buffer as an efficient modulator of polydopamine buildup and properties for the rational control and fine-tuning of melanin aggregate size, morphology, and free radical behavior.

For ternary systems, we present a method for using measured values of the four ternary diffusion coeffs. and the Onsager reciprocal relations to ext. derivs. of solute chem. potentials with respect to solute molar concns. The method is applicable to systems in which the molar concn. of one solute is very small compared to that of the other, and also small enough that an inverse concn. dependence dominates certain activity coeff. derivs. These conditions apply to a large no. of aq. systems involving macromols. of biol. interest. Unlike other techniques, the present method can be used to study undersatd. and supersatd. solns. The approach is illustrated for the lysozyme chloride-NaCl-H2O system at 25°, using data reported here for pH 6.0 at 0.60 mM (8.6 mg/mL) lysozyme chloride and 0.25, 0.50, 0.65, 0.90, and 1.30 M (1.4, 2.8, 3.7, 5.1, and 7.2 wt %) NaCl concns., and our earlier data for pH 4.5 at the same concns. We use these solute chem. potential derivs. to compute the protein cation charge approx., and to construct a function approximating the deriv. of the lysozyme chloride chem. potential with respect to NaCl concn., which we integrate over a range of NaCl concns. This provides the change of the lysozyme chloride chem. potential with NaCl concn. well into the supersatd. region, and hence provides the driving force for nucleation and crystal growth of lysozyme chloride as a function of the extent of supersatn. We also compute the diffusion Onsager coeffs. (Lij)0 for each compn. at pH 4.5 and 6.0. Binary diffusion coeffs. of aq. lysozyme chloride at 0.89 mM (12.7 mg/mL) for pH values from 4.0 to 6.0, and at pH 6.0 for concns. from 0.25 to 1.95 mM (3.6-27.9 mg/mL) are also reported

Establishing structure-property relationships in the black insoluble eumelanins, the key determinants of human pigmentation and skin photoprotective system, is a considerable conceptual and experimental challenge in the current drive for elucidation of the biological roles of these biopolymers and their application as advanced materials for organoelectronics. Herein, we report a new breakthrough toward this goal by the first detailed investigation on the nanoscale level of the oxidative polymerization of 5,6-dihydroxyindole (DHI), a model process of eumelanin synthesis. On the basis of a combined use of spectrophotometry, dynamic light scattering (DLS), and small-angle neutron scattering (SANS) investigations, it was possible to unveil the dynamics of the aggregation process before precipitation, the key relationships with visible light absorption and the shape of fundamental aggregates. The results indicated a polymerization mechanism of the type: Polymer(n) + DHI(x) = Polymer(n+x), where DHI(x) indicates monomer, dimer, or low oligomers (x ≤ 5). During polymerization, visible absorption increases rapidly, reaching a plateau. Particle growth proceeds slowly, with formation of 2-D structures ~55 nm thick, until precipitation occurs, that is, when large aggregates with a maximum hydrodynamic radius (R(h)) of ~1200 nm are formed. Notably, markedly smaller R(h) values, up to ~110 nm, were determined in the presence of poly(vinyl alcohol) (PVA) that was shown to be an efficient aggregation-preventing agent for polymerizing DHI ensuring water solubilization. Finally, it is shown that DHI monomer can be efficiently and partially irreversibly depleted from aqueous solutions by the addition of eumelanin suspensions. This behavior is suggested to reflect oxidant-independent competing pathways of polymer synthesis and buildup via monomer conversion on the active aggregate surface contributing to particle growth. Besides filling crucial gaps in DHI polymerization, these results support the attractive hypothesis that eumelanins may behave as a peculiar example of living biopolymers. The potential of PVA as a powerful tool for solution chemistry-based investigations of eumelanin supramolecular organization and for technological manipulation purposes is underscored.

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An efficient drug delivery strategy is presented for novel anticancer amphiphilic ruthenium anionic complexes, based on the formation of stable nanoparticles with the cationic lipid 1,2-dioleyl-3-trimethylammoniumpropane chloride (DOTAP). This strategy is aimed at ensuring high ruthenium content within the formulation, long half-life in physiological media, and enhanced cell uptake. An in-depth microstructural characterization of the aggregates obtained mixing the ruthenium complex and the phospholipid carrier at 50/50 molar ratio is realized by combining a variety of techniques, including dynamic light scattering (DLS), small angle neutron scattering (SANS), neutron reflectivity (NR), electron paramagnetic resonance (EPR), and zeta potential measurements. The in vitro bioactivity profile of the Ru-loaded nanoparticles is investigated on human and non-human cancer cell lines, showing IC(50) values in the low μM range against MCF-7 and WiDr cells, that is, proving to be 10-20-fold more active than AziRu, a previously synthesized NAMI-A analog, used for control. Fluorescence microscopy studies demonstrate that the amphiphilic Ru-complex/DOTAP formulations, added with rhodamine-B, are efficiently and rapidly incorporated in human MCF-7 breast adenocarcinoma cells. The intracellular fate of the amphiphilic Ru-complexes was investigated in the same in vitro model by means of an ad hoc designed fluorescently tagged analog, which exhibited a marked tendency to accumulate within or in proximity of the nuclei.

The structural organization of matter in poly(vinyl alcohol) (PVA) hydrogels obtained by repeatedly freezing and thawing dilute solutions of PVA in D 2 O is investigated by use of small-angle neutron scattering measurements (SANS). This study is the first systematic and quantitative investigation in the medium range of length scales on PVA hydrogels obtained by freezing and thawing techniques. The studied gels have a complex hierarchical structure, extending over a wide range of length scales. The structural organization on the micron length scale originates from the presence of two separated phases constituted by polymer-rich and polymer-poor regions. The network structure may be interpreted in terms of the connectivity of the regions occupied by the polymer-rich phase, which extend over the macroscopic dimensions of the sample. The organization on the medium length scale is provided by the presence of small crystallites, fringed micelle-like, within the polymer-rich phase. In these regions, the crystals are highly connected by swollen amorphous tie chains. The presence of these tie chains ensures the connectivity of the macroscopic network. The structural organization on the short length scale is essentially provided by the relative arrangement of chains within the crystallites and in the swollen amorphous zones. The PVA hydrogel structure has been modeled as a collection of polydisperse and homogeneous spherical crystallites, interacting via hard-spheres potential. SANS experiments permitted us to obtain values of the crystallite size of about 33 Å, of the volume fraction of polymer-rich phase of the order of 1% and a value of the average crystallite-crystallite correlation distances of the order of a few tens of nanometers, independent of the imposed number of freeze/thaw cycles (n), for n > 1. The present analysis also indicates that the structure of the gel obtained imposing a single freeze/thaw cycle is somehow intermediate between the structure of the homogeneous starting solution and the structure of the already well-formed sample obtained by imposing two consecutive cycles.