Humidity Response of Cellulose Thin Films.

Cellulose-water interactions are crucial to understand biological processes as well as to develop tailor made cellulose-based products. However, the main challenge to study these interactions is the diversity of natural cellulose fibers and alterations in their supramolecular structure. Here, we study the humidity response of different, well-defined, ultrathin cellulose films as a function of industrially relevant treatments using different techniques. As treatments, drying at elevated temperature, swelling, and swelling followed by drying at elevated temperatures were chosen. The cellulose films were prepared by spin coating a soluble cellulose derivative, trimethylsilyl cellulose, onto solid substrates followed by conversion to cellulose by HCl vapor. For the highest investigated humidity levels (97%), the layer thickness increased by ca. 40% corresponding to the incorporation of 3.6 molecules of water per anhydroglucose unit (AGU), independent of the cellulose source used. The aforementioned treatments affected this ratio significantly with drying being the most notable procedure (2.0 and 2.6 molecules per AGU). The alterations were investigated in real time with X-ray reflectivity and quartz crystal microbalance with dissipation, equipped with a humidity module to obtain information about changes in the thickness, roughness, and electron density of the films and qualitatively confirmed using grazing incidence small angle X-ray scattering measurements using synchrotron irradiation.

Multilayer Density Analysis of Cellulose Thin Films.

An approach for the multilayer density analysis of polysaccharide thin films at the example of cellulose is presented. In detail, a model was developed for the evaluation of the density in different layers across the thickness direction of the film. The cellulose thin film was split into a so called "roughness layer" present at the surface and a "bulk layer" attached to the substrate surface. For this approach, a combination of multi-parameter surface plasmon resonance spectroscopy (SPR) and atomic force microscopy (AFM) was employed to detect changes in the properties, such as cellulose content and density, thickness and refractive index, of the surface near layer and the bulk layer. The surface region of the films featured a much lower density than the bulk. Further, these results correlate to X-ray reflectivity studies, indicating a similar layered structure with reduced density at the surface near regions. The proposed method provides an approach to analyse density variations in thin films which can be used to study material properties and swelling behavior in different layers of the films. Limitations and challenges of the multilayer model evaluation method of cellulose thin films were discussed. This particularly involves the selection of the starting values for iteration of the layer thickness of the top layer, which was overcome by incorporation of AFM data in this study.

Deposition of Cellulose-Based Thin Films on Flexible Substrates.

This study investigates flexible (polyamide 6.6 PA-6.6, polyethylene terephthalate PET, Cu, Al, and Ni foils) and, for comparison, stiff substrates (silicon wafers and glass) differing in, for example, in surface free energy and surface roughness and their ability to host cellulose-based thin films. Trimethylsilyl cellulose (TMSC), a hydrophobic acid-labile cellulose derivative, was deposited on these substrates and subjected to spin coating. For all the synthetic polymer and metal substrates, rather homogenous films were obtained, where the thickness and the roughness of the films correlated with the substrate roughness and its surface free energy. A particular case was the TMSC layer on the copper foil, which exhibited superhydrophobicity caused by the microstructuring of the copper substrate. After the investigation of TMSC film formation, the conversion to cellulose using acidic vapors of HCl was attempted. While for the polymer foils, as well as for glass and silicon, rather homogenous and smooth cellulose films were obtained, for the metal foils, there is a competing reaction between the formation of metal chlorides and the generation of cellulose. We observed particles corresponding to the metal chlorides, while we could not detect any cellulose thin films after HCl treatment of the metal foils as proven by cross-section imaging using scanning electron microscopy (SEM).

Lectins at Interfaces-An Atomic Force Microscopy and Multi-Parameter-Surface Plasmon Resonance Study.

Lectins are a diverse class of carbohydrate binding proteins with pivotal roles in cell communication and signaling in many (patho)physiologic processes in the human body, making them promising targets in drug development, for instance, in cancer or infectious diseases. Other applications of lectins employ their ability to recognize specific glycan epitopes in biosensors and glycan microarrays. While a lot of research has focused on lectin interaction with specific carbohydrates, the interaction potential of lectins with different types of surfaces has not been addressed extensively. Here, we screen the interaction of two specific plant lectins, Concanavalin A and Ulex Europaeus Agglutinin-I with different nanoscopic thin films. As a control, the same experiments were performed with Bovine Serum Albumin, a widely used marker for non-specific protein adsorption. In order to test the preferred type of interaction during adsorption, hydrophobic, hydrophilic and charged polymer films were explored, such as polystyrene, cellulose, N,-N,-N-trimethylchitosan chloride and gold, and characterized in terms of wettability, surface free energy, zeta potential and morphology. Atomic force microscopy images of surfaces after protein adsorption correlated very well with the observed mass of adsorbed protein. Surface plasmon resonance spectroscopy studies revealed low adsorbed amounts and slow kinetics for all of the investigated proteins for hydrophilic surfaces, making those resistant to non-specific interactions. As a consequence, they may serve as favorable supports for biosensors, since the use of blocking agents is not necessary.

Interaction of industrially relevant cationic starches with cellulose.

Industrially relevant, commercially available cationic starches have been investigated towards their interaction capacity with cellulose thin films derived from trimethylsilyl cellulose (TMSC). The starches used in this study stem from different sources (potato, pea, corn) and featured rather low degrees of substitution ranging from 0.030 to 0.062. The interaction of those starches with cellulose thin films was studied by surface plasmon resonance spectroscopy under flow conditions using concentrations of 1.0mgml-1 and a flow rate of 25µlmin-1. All the investigated starches employed in this study were capable to efficiently interact with the slightly negatively charged cellulose surface leading to irreversible deposition on the surface. As complementary techniques atomic force microscopy and x-ray photoelectron spectroscopy were used to confirm the presence of the starches on the cellulose film surface. Further, dynamic light scattering and size exclusion chromatography measurements were performed to correlate adsorbed amount, particle size and molecular weight of the starches to their interaction behavior.

How Bound and Free Fatty Acids in Cellulose Films Impact Nonspecific Protein Adsorption.

The effect of fatty acids and fatty acid esters to impair nonspecific protein adsorption on cellulose thin films is investigated. Thin films are prepared by blending trimethylsilyl cellulose solutions with either cellulose stearoyl ester or stearic acid at various ratios. After film formation by spin coating, the trimethylsilyl cellulose fraction of the films is converted to cellulose by exposure to HCl vapors. The morphologies and surface roughness of the blends were examined by atomic force microscopy revealing different feature shapes and sizes depending on the blend ratios. Nonspecific protein adsorption at the example of bovine serum albumin toward the blend thin films was tested by means of surface plasmon resonance spectroscopy in real-time. Incorporation of stearic acid into the cellulose leads to highly protein repellent surfaces regardless of the amount added. The stearic acid acts as a sacrificial compound that builds a complex with bovine serum albumin thereby inhibiting protein adsorption. For the blends where stearoyl ester is added to the cellulose films, the cellulose:cellulose stearoyl ester ratios of 3:1 and 1:1 lead to much lower nonspecific protein adsorption compared to pure cellulose, whereas for the other ratios, adsorption increases. Supplementary results were obtained from atomic force microscopy experiments performed in liquid during exposure to protein solution and surface free energy determinations.

Interaction of Tissue Engineering Substrates with Serum Proteins and Its Influence on Human Primary Endothelial Cells.

Polymer-based biomaterials particularly polycaprolactone (PCL) are one of the most promising substrates for tissue engineering. The surface chemistry of these materials plays a major role since it governs protein adsorption, cell adhesion, viability, degradation, and biocompatibility in the first place. This study correlates the interaction of the most abundant serum proteins (albumin, immunoglobulins, fibrinogen) with the surface properties of PCL and its influence on the morphology and metabolic activity of primary human arterial endothelial cells that are seeded on the materials. Prior to that, thin films of PCL are manufactured by spin-coating and characterized in detail. A quartz crystal microbalance with dissipation (QCM-D), a multiparameter surface plasmon resonance spectroscopy instrument (MP-SPR), wettability data, and atomic force microscopy are combined to elucidate the pH-dependent protein adsorption on the PCL substrates. Primary endothelial cells are cultured on the protein modified polymer, and conclusions are drawn on the significant impact of type and form of proteins coatings on cell morphology and metabolic activity.

Enzymes as Biodevelopers for Nano- And Micropatterned Bicomponent Biopolymer Thin Films.

The creation of nano- and micropatterned polymer films is a crucial step for innumerous applications in science and technology. However, there are several problems associated with environmental aspects concerning the polymer synthesis itself, cross-linkers to induce the patterns as well as toxic solvents used for the preparation and even more important development of the films (e.g., chlorobenzene). In this paper, we present a facile method to produce micro- and nanopatterned biopolymer thin films using enzymes as so-called biodevelopers. Instead of synthetic polymers, naturally derived ones are employed, namely, poly-3-hydroxybutyrate and a cellulose derivative, which are dissolved in a common solvent in different ratios and subjected to spin coating. Consequently, the two biopolymers undergo microphase separation and different domain sizes are formed depending on the ratio of the biopolymers. The development step proceeds via addition of the appropriate enzyme (either PHB-depolymerase or cellulase), whereas one of the two biopolymers is selectively degraded, while the other one remains on the surface. In order to highlight the enzymatic development of the films, video AFM studies have been performed in real time to image the development process in situ as well as surface plasmon resonance spectroscopy to determine the kinetics. These studies may pave the way for the use of enzymes in patterning processes, particularly for materials intended to be used in a physiological environment.

Exploring Nonspecific Protein Adsorption on Lignocellulosic Amphiphilic Bicomponent Films.

In this contribution, we explore the interaction of lignocellulosics and proteins aiming at a better understanding of their synergistic role in natural systems. In particular, the manufacturing and characterization of amphiphilic bicomponent thin films composed of hydrophilic cellulose and a hydrophobic lignin ester in different ratios is presented which may act as a very simplified model for real systems. Besides detailed characterizations of the films and mechanisms to explain their formation, nonspecific protein adsorption using bovine serum albumin (BSA) onto the films was studied using a quartz crystal microbalance with dissipation (QCM-D). As it turns out, the rather low nonspecific protein adsorption of BSA on cellulose is further reduced when these hydrophobic lignins are incorporated into the films. The lignin ester acts in these blend films as sacrificial component, probably via an emulsification mechanism. Additionally, the amphiphilicity of the films may prevent the adsorption of BSA as well. Although there are some indications, it remains unclear whether any kind of protein interactions in such systems are of specific nature.

Adsorption Studies of Organophosphonic Acids on Differently Activated Gold Surfaces.

In this study, the formation of self-assembled monolayers consisting of three organophosphonic acids (vinyl-, octyl-, and tetradecylphosphonic acid) from isopropanol solutions onto differently activated gold surfaces is studied in situ and in real time using multiparameter surface plasmon resonance (MP-SPR). Data retrieved from MP-SPR measurements revealed similar adsorption kinetics for all investigated organophosphonic acids (PA). The layer thickness of the immobilized PA is in the range of 0.6-1.8 nm corresponding to monolayer-like coverage and correlates with the length of the hydrocarbon chain of the PA molecules. After sintering the surfaces, the PA are irreversibly attached onto the surfaces as proven by X-ray photoelectron spectroscopy and attenuated total reflection infrared and grazing incidence infrared spectroscopy. Potential adsorption modes and interaction mechanisms are proposed.

Triggering protein adsorption on tailored cationic cellulose surfaces.

The equipment of cellulose ultrathin films with BSA (bovine serum albumin) via cationization of the surface by tailor-made cationic celluloses is described. In this way, matrices for controlled protein deposition are created, whereas the extent of protein affinity to these surfaces is controlled by the charge density and solubility of the tailored cationic cellulose derivative. In order to understand the impact of the cationic cellulose derivatives on the protein affinity, their interaction capacity with fluorescently labeled BSA is investigated at different concentrations and pH values. The amount of deposited material is quantified using QCM-D (quartz crystal microbalance with dissipation monitoring, wet mass) and MP-SPR (multi-parameter surface plasmon resonance, dry mass), and the mass of coupled water is evaluated by combination of QCM-D and SPR data. It turns out that adsorption can be tuned over a wide range (0.6-3.9 mg dry mass m(-2)) depending on the used conditions for adsorption and the type of employed cationic cellulose. After evaluation of protein adsorption, patterned cellulose thin films have been prepared and the cationic celluloses were adsorbed in a similar fashion as in the QCM-D and SPR experiments. Onto these cationic surfaces, fluorescently labeled BSA in different concentrations is deposited by an automatized spotting apparatus and a correlation between the amount of the deposited protein and the fluorescence intensity is established.