Nanosized Water Channels Associated with Hydrophobic and Hydrophilic Fibrillar Arrangements Formed on Nafion Surfaces in Confined Regions.

Herein, the origin of interfacial water nanosized channel distributions attached onto Nafion surfaces is investigated. The surface fibrillary hydrophilic and hydrophobic arrangements were observed on AFM images scanned on Nafion surfaces immersed in water. Then, by analyzing the force vs separation curves, it is possible to map arrays of interfacial water channels and their locations. Nafion surface profiles and the water interfacial patterns are then combined using this AFM technique. As there are no reported experimental techniques to measure water nanochannel cross sections, presented measurements report on their dimensions. Water nanochannels characterized by ε < 7 attached to hydrophilic fibrillary sections form aggregated water domains, a highly organized water structure compared with bulk water. Channels are attached to Nafion surface hydrophilic fibrillary domains in confined sites.

Fibrillary Arrangement of Elongated, Almost Parallel Aggregates of Hydrophobic and Hydrophilic Domains Forming the Nafion Surface Structure Improved Contrast Atomic Force Microscopy Images.

A significant improvement in spatial resolution is reported in Nafion surface maps when compared to previous atomic force microscopy images of the Nafion surface scanned in air. The technique ability is to generate maps showing approximately few nanometer (∼2-5 nm) patterns to the long fiber length (>2 µm). Atomic force microscopy force vs separation curve profiles registered in water are used to characterize the surface hydrophobic and hydrophilic domains. Initially, Nafion surfaces were imaged in air for comparison and then immersed in water. Nafion surfaces immersed in water display a matrix of hydrophilic and hydrophobic regions with fibrillary structure dimensions of ∼40 nm formed by fiber pairs. Ribbons formed by two pairs with diameters of ∼83 nm are separated by larger channels.

Variable Interfacial Water Nanosized Arrangements Measured by Atomic Force Microscopy.

While there seems to be broad agreement that cluster formation does exist near solid surfaces, its presence at the liquid/vapor interface is controversial. We report experimental studies we have carried out on interfacial water attached on hydrophobic and hydrophilic surfaces. Nanosized steps in the measured force vs distance to the surface curves characterize water cluster profiles. An expansion of the interfacial structure with time is observed; the initial profile extent is typically ∼1 nm, and for longer times expanded structures of ∼70 nm are observed. Our previous results showed that the interfacial water structure has a relative permittivity of ε ≈ 3 at the air/water interface homogeneously increasing to ε ≈ 80 at 300 nm inside the bulk, but here we have shown that the interfacial dielectric permittivity may have an oscillating profile describing the spatial steps in the force vs distance curves. This low dielectric permittivity arrangements of clusters extend the region with ε ≈ 3 inside bulk water and exhibit a behavior similar to that of water networks that expand in time.

Imaging Ion Pairs Forming Structural Arrangements in Interfacial Regions.

A technique to image ion pairs in solution is reported. We investigated structural and dynamic properties of ion-pair distributions deposited on highly oriented pyrolytic graphite (HOPG) surfaces in electrolyte solutions. Atomic force microscopy images of HOPG immersed in NaCl and KCl solutions display regular arrangements on top of the hexagonal carbon rings forming the HOPG atomic structure. These arrangements are the result of the low value of the aqueous interfacial dielectric constant (εr ≈ 3-11). The measured ion-pair radius is a function of the salt present in the solution; for KCl, the ion-pair radius is equal or smaller than 0.42 nm; for NaCl, the ion-pair radius is 0.36 nm. A comparison of these values with their crystalline lattice dimensions indicates that both KCl and NaCl ion pairs in solution at the HOPG/solution interfacial region exist as tight contact ion pairs in quasistationary distributions. The NaCl ion-pair distribution forms an aligned arrangement, and the KCl distribution is formed by intercalated pairs.

Hydrated excess protons and their local hydrogen bond transport network as measured by translational, librational, and vibrational frequencies.

A clear molecular description of excess hydrated protons and their local hydrogen bond transport network remains elusive. Here, the hydrogen bond network of excess hydrated protons in water bridges was probed by measuring their Raman spectra and comparing them to the spectra of protons in ice and water. The proton vibrational spectrum and the hydrogen bond network translational and librational spectra were recorded. The spectra of the water bridge and water exhibit clear differences, indicating the presence of a structure in water bridges when subjected to an electric field of ∼106 V/m that has not been previously reported. The intermolecular Raman spectrum of the floating water bridge exhibits a hydrogen bond stretching band at 150-250 cm-1, librational bands within the 300-1000 cm-1 spectral range, and a large band at 1500-3000 cm-1, which corresponds to the vibrational signature of excess hydrated protons in the water bridge structure. The excess protons are shown to move predominantly at the air/water interface, and the effect of this distribution is a measurable change in the air/water interfacial tension from ∼80 to ∼32 N/m. Therefore, hydrated protons must have a unique water arrangement that enables them to propagate without sinking into bulk water. This local polarized hydrogen bond network in the interfacial water region is characterized by a translational spectrum similar to that of ice V.

Hydrated Excess Proton Raman Spectral Densities Probed in Floating Water Bridges.

Excess proton structures in water remain unclear. The motion and nature of excess protons in water were probed using a supported water bridge structure in electric field (E) with an intensity of ∼106 V/m. The experimental setup generated protons that exhibit a long lifetime. The effect of excess protons in water induced a ∼3% variation in the pH for a 300 V overvoltage at the cathode. The current versus voltage curves show a current space-charge-limited operation. By measuring the space-charge distribution in both the cathode and anode and by adjusting the Mott-Gurney law to the measured excess hydrated proton current and the voltage drop in the cationic space-charge region, the protonic mobility was determined to be ∼200 × 10-8 m2/(V·s) (E ≈ 4 × 106 V/m). This measured mobility, which is typically five times larger than the reported mobility for protons in water, is in agreement with the mechanism outlined by Grotthuss in 1805. The measured mid-Raman spectrum covering 1000-3800 cm-1 range indicates the species character. The hydrated excess proton spectral response through the mid-Raman at 1760 and 3200 cm-1 was attributed to the Zundel complex and the region at ∼2000 to ∼2600 cm-1 response is attributed to the Eigen complex, indicating a core structure simultaneously with a Eigen-like and Zundel-like character, suggesting a rapid fluctuation between these two structures or a new specie.

Chiral Asymmetric Structures in Aspartic Acid and Valine Crystals Assessed by Atomic Force Microscopy.

Structures of crystallized deposits formed by the molecular self-assembly of aspartic acid and valine on silicon substrates were imaged by atomic force microscopy. Images of d- and l-aspartic acid crystal surfaces showing extended molecularly flat sheets or regions separated by single molecule thick steps are presented. Distinct orientation surfaces were imaged, which, combined with the single molecule step size, defines the geometry of the crystal. However, single molecule step growth also reveals the crystal chirality, i.e., growth orientations. The imaged ordered lattice of aspartic acid (asp) and valine (val) mostly revealed periodicities corresponding to bulk terminations, but a previously unreported molecular hexagonal lattice configuration was observed for both l-asp and l-val but not for d-asp or d-val. Atomic force microscopy can then be used to identify the different chiral forms of aspartic acid and valine crystals.

Water exclusion layers probed by depth scan confocal Raman microscopy.

Depth scan confocal Raman microscopy was employed to map water and air spatial distributions in immersed superhydrophobic films. Due to the lack of visible nanobubbles on flat surfaces, we have probed heterogeneous surfaces where solid­liquid, liquid­vapor, and vapor­solid coexist. Depth scan profiles show liquid exclusion (vapor) layers inside the fiber arrangement and water in contact only at the fiber apex.

Anisotropic growth of water-puckered pentamers on a mica terrace.

Ice nucleation at mica terrace edges in air forms mounds of water molecules that grow larger as the step-edge height increases from a few Angstroms to hundreds of nanometers. The structures of the ice deposits at mica terrace edges were characterized by atomic force microscopy (AFM), and the edges were shown to act as nucleators for water pentamers, thereby forming a zigzag structure with lattice parameters of 0.72 ± 0.07 and 0.60 ± 0.06 nm. A three-dimensional arrangement of three pentamers of water molecules, which formed a parasol-like structure, was assembled to match the AFM images. Seven three-fused pentamers were clustered to form large hexamers that cover the entire surface. The nucleation at the edges reveals a substantially larger growth rate than that on the mica terraces; consequently, highly terraced mica slabs could be used as new and more efficient structures for seeding clouds and causing rain. On the basis of this finding, a new ice-condensation structure was designed with pyramidal features and steps of 100 nm in height and width.

Drainage kinetics of nanochannels fabricated in water films a few molecules thick on mica at room temperature.

Nanochannels of the order of 20 nm in diameter and forming arrangements that were a few micrometres wide were fabricated on nanometre-thick ice-like deposits on planar mica surfaces at room temperature. Because an atomic force microscopy tip can write lines on ice-like layers covering mica substrates in air that are stable under invariant conditions of humidity and temperature, the water films were modulated with nanochannels. By analysing the shape and morphology of the material removed after channel fabrication for various time intervals, the channel profile was shown to vary with a scale of a tenth of a second. In this configuration (hydrophobic tip and hydrophilic substrate, 65% RH), at the channel top region there were only aggregates of loose flakes formed after the film inscription but no liquid. Apparently, the Kelvin effect is responsible for the nanochannel profile variation with time, but the calculated and measured values of the drainage time constant are at variance by six orders of magnitude. This reduction of the mass transfer is associated with the small dimensions of the ∼ 10 nm-wide channels.

Imaging ice-like structures formed on HOPG at room temperature.

In this work, ice was viewed at the nanoscale by scanning an atomic force microscopy tip over a highly oriented pyrolytic graphite (HOPG) surface in air. At low scan velocities, the tip exhibited stick-slip motion with a period of 0.13 nm corresponding to the scanner step; at higher velocities, the HOPG lattice and the periodicity of the ice were visible. A hexagonal structure with a 0.45 ± 0.04 nm periodicity was observed in which the distance between the second neighbors of the HOPG coincided with the distance of the first neighbors for the ice-like arrangement. Small water clusters were also nucleated with an ice-Ic structure (0.34 ± 0.03 nm), and thus, the ice layers consisted of extensive sets composed of arrangements of hexamers and tetramers.

Long nanofibers arrays through surfactant self-assembly.

Self assembly of molecules can be a simple and versatile approach that may lead to nanostructures. Here we report the formation of arrangements of exceptionally long nanofibers of cationic surfactant hexadecyltrimethylammonium bromide molecules with highly defined spatial and parallel ordering. Arrangements of approximately 1 microm long nanofibers were observed by non-contact atomic force microscopy. The long nanofiber patterns form structures in approximately 5 x 5 microm2 regions and consist of approximately 6 nm wide lines. The formation mechanism of the fibers is shown to be the coalescence of isolated surfactant micelles (approximately 3.2 nm diameter in solution) in the convection stream of the surfactant solution drop close to the pinned contact-line region during drying. The size of micelles formed in solution determines the diameter of the deposited fibers. New deposition schemes of micelles forming nanorods on bare silicon previous to the formation of surfactant bilayers are now being investigated based on the data obtained in this work.

Effect of a highly concentrated lipopeptide extract of Bacillus subtilis on fungal and bacterial cells.

Lipopeptides produced by Bacillus subtilis are known for their high antifungal activity. The aim of this paper is to show that at high concentration they can damage the surface ultra-structure of bacterial cells. A lipopeptide extract containing iturin and surfactin (5 mg mL(-1)) was prepared after isolation from B. subtilis (strain OG) by solid phase extraction. Analysis by atomic force microscope (AFM) showed that upon evaporation, lipopeptides form large aggregates (0.1-0.2 microm(2)) on the substrates silicon and mica. When the same solution is incubated with fungi and bacteria and the system is allowed to evaporate, dramatic changes are observed on the cells. AFM micrographs show disintegration of the hyphae of Phomopsis phaseoli and the cell walls of Xanthomonas campestris and X. axonopodis. Collapses to fungal and bacterial cells may be a result of formation of pores triggered by micelles and lamellar structures, which are formed above the critical micelar concentration of lipopeptides. As observed for P. phaseoli, the process involves binding, solubilization, and formation of novel structures in which cell wall components are solubilized within lipopeptide vesicles. This is the first report presenting evidences that vesicles of uncharged and negatively charged lipopeptides can alter the morphology of gram-negative bacteria.

Quorum sensing detected by atomic force microscopy imaging of corrals surrounding multicellular arrangement of bacteria.

Connectivity of the glycocalyx covering of small communities of Acidithiobacillus ferrooxidans bacteria deposited on hydrophilic mica plates was imaged by atomic force microscopy. When part of the coverage was removed by water rinsing, an insoluble structure formed by corrals surrounding each individual bacterium was observed. A collective ring structure with clustered bacteria (>or=3) was observed, which indicates that the bacteria perceived the neighborhood in order to grow a protective structure that results in smaller production of exopolysaccharides material. The most surprising aspect of these collective corral structures was that they occur at a low bacterial cell density. The deposited layers were also analyzed by confocal Raman microscopy and shown to contain polysaccharides, protein, and glucoronic acid.

Extracellular polymeric bacterial coverages as minimal area surfaces.

Surfaces formed by extracellular polymeric substances enclosing individual and some small communities of Acidithiobacillus ferrooxidans on plates of hydrophobic silicon and hydrophilic mica are analyzed by means of atomic force microscopy imaging. Accurate nanoscale descriptions of such coverage surfaces are obtained. The good agreement with the predictions of a rather simple but realistic theoretical model allows us to conclude that they correspond, indeed, to minimal area (constant mean curvature) surfaces enclosing a given volume associated with the encased bacteria. This is, to the best of our knowledge, the first shape characterization of the coverage formed by these biomolecules, with potential applications to the study of biofilms.

Volume of extracellular polymeric substance coverage of individual Acidithiobacillus ferrooxidans bacterium measured by atomic force microscopy.

The Acidithiobacillus ferrooxidans response to stress associated with the drying process is known to be the production of extracellular polymeric substance (EPS) coverage. Here, samples of A. ferrooxidans suspensions grown in 1.8 pH and 3.0 pH and dried on mica and silicon are shown to form a structure of isolated bacteria. Individual bacteria coverage patterns were imaged by atomic force microscopy (AFM) on hydrophobic (silicon) and hydrophilic (mica) substrates. A comparison of images of covered and uncovered bacteria establish the volume of individual EPS coverage. The EPS production for bacteria on hydrophobic substrates shows a substantial decrease (a factor of 30) in the EPS volume per bacterium when compared with the one on hydrophilic substrates. Shape and volume determination of EPS structures on bacteria as a function of hydrophobicity or hydrophilicity of the substrate may help to determine the functions of EPS on bacterial aggregates.

Effects of the antimicrobial peptide PGLa on live Escherichia coli.

The activity of PGLa, an antimicrobial peptide isolated from hemocytes of frog skin and its secretions on living Escherichia coli, was investigated by imaging the cells with atomic force microscopy (AFM) in physiological conditions and by measuring its cellular stiffness. The treatment of bacteria with the antimicrobial peptide PGLa in the culture medium had two stages. The first was characterized by the loss of surface stiffness and consequent loss of bacteria topographic features and the formation of micelles probably originating from the disruption of the outer membrane. The formation of outer membrane originated micelles is in agreement with the carpet-like mechanism of action proposed for antimicrobial peptides of the magainin family. The peptide action also resulted in the removal of bacterial pili. In a second stage there was further damage which resulted in total cell rupture. The addition of Mg(2+) ions prior to peptide treatment partially inhibited the effects of PGLa on bacteria. This result suggests that PGLa interacts with the outer membrane by displacing Mg(2+) from LPS, inserting itself into the bilayer and cross-bridging the negative charges of LPS lipids as proposed in the self-promoted pathway mechanism. The peptide effect on the bacteria was compared to the activity of the chelating agent EDTA that damages the bacterial outer membrane by removing Mg(2+) ions.