Guillermo Toriz

Role of (1,3)(1,4)-β-Glucan in Cell Walls: Interaction with Cellulose

(1,3)(1,4)-β-D-Glucan (mixed-linkage glucan or MLG), a characteristic hemicellulose in primary cell walls of grasses, was investigated to determine both its role in cell walls and its interaction with cellulose and other cell wall polysaccharides in vitro. Binding isotherms showed that MLG adsorption onto microcrystalline cellulose is slow, irreversible, and temperature-dependent. Measurements using quartz crystal microbalance with dissipation monitoring showed that MLG adsorbed irreversibly onto amorphous regenerated cellulose, forming a thick hydrogel. Oligosaccharide profiling using endo-(1,3)(1,4)-β-glucanase indicated that there was no difference in the frequency and distribution of (1,3) and (1,4) links in bound and unbound MLG. The binding of MLG to cellulose was reduced if the cellulose samples were first treated with certain cell wall polysaccharides, such as xyloglucan and glucuronoarabinoxylan. The tethering function of MLG in cell walls was tested by applying endo-(1,3)(1,4)-β-glucanase to wall samples in a constant force extensometer. Cell wall extension was not induced, which indicates that enzyme-accessible MLG does not tether cellulose fibrils into a load-bearing network.

In situ forming spruce xylan-based hydrogel for cell immobilization.

An in situ forming spruce xylan-based hydrogel was synthesized in two steps with the intended use of cell encapsulation and in vivo delivery. First, bioconjugate was obtained through the reaction of glucuronic acid groups from xylan backbone with tyramine (TA). After that, the gelation process was enabled by enzymatic crosslinking of the phenol-containing TA-xylan conjugate. Exhibiting an exponential increase in the storage modulus, a 3D gel network was formed in about 20s. The designed gel showed extensive swelling and retained its mechanical integrity for more than two months. Mesenchymal stem cells were encapsulated in the hydrogel and cultured for one week. The cells retained their adipogenic differentiation capacity inside the gel, as verified by lipid accumulation. From these facts, we conclude that spruce xylan is a promising precursor for in situ forming hydrogels and should be evaluated further for tissue engineering purposes.

Corncob arabinoxylan for new materials

Corncob agricultural waste was used as a source of arabinoxylan for preparation of films. Three arabinoxylan samples were prepared: crude extract (CCAX), purified by a washing step, and purified by bleaching CCAX. Films prepared with untreated CCAX were water soluble, yellowish in color and had poor mechanical properties. After the purification processes the Youngs modulus increased from ∼ 293 MPa to ∼ 1400-1600 MPa, and strength was improved from ∼ 9 MPa to around 53 MPa, while the strain at break was kept at ∼ 8% both in untreated and purified CCAX. The contact angle was increased from ∼ 21.3° to 67-74° after washing or bleaching CCAX. Acetylation of bleached CCAX showed the highest thermal resistance (325 °C), had low Tg (125°C) and a high contact angle (80°), and its films were stronger (strength ∼ 67 MPa; Youngs modulus ∼ 2241 MPa) and more flexible (∼ 13%). These characteristics make purified CCAX a suitable material to be used as a matrix for film applications.

Assembly of Debranched Xylan from Solution and on Nanocellulosic Surfaces

This study focused on the assembly characteristics of debranched xylan onto cellulose surfaces. A rye arabinoxylan polymer with an initial arabinose/xylose ratio of 0.53 was debranched with an oxalic acid treatment as a function of time. The resulting samples contained reduced arabinose/xylose ratios significantly affecting the molecular architecture and solution behavior of the biopolymer. With this treatment, an almost linear xylan with arabinose DS of only 0.04 was obtained. The removal of arabinose units resulted in the self-assembly of the debranched polymer in water into stable nanoparticle aggregates with a size around 300 nm with a gradual increase in crystallinity of the isolated xylan. Using quartz crystal microbalance with dissipation monitoring, the adsorption of xylan onto model cellulose surfaces was quantified. Compared to the nonmodified xylan, the adsorption of debranched xylan increased from 0.6 to 5.5 mg m(-2). Additionally, adsorption kinetics suggest that the nanoparticles rapidly adsorbed to the cellulose surfaces compared to the arabinoxylan. In summary, a control of the molecular structure of xylan influences its ability to form a new class of polysaccharide nanoparticles in aqueous suspensions and its interaction with nanocellulose surfaces.

Bagasse hydrolyzates from Agave tequilana as substrates for succinic acid production by Actinobacillus succinogenes in batch and repeated batch reactor.

The aim of this work was to obtain fermentable sugars by enzymatic or acid hydrolyses of Agave tequilana Weber bagasse in order to produce succinic acid with Actinobacillus succinogenes. Hydrolyses were carried out with mineral acids (sulfuric and hydrochloric acids) or a commercial cellulolytic enzyme, and were optimized statistically by a response surface methodology, having as factors the concentration of acid/enzyme and time of hydrolysis. The concentration of sugars obtained at optimal conditions for each hydrolysis were 21.7, 22.4y 19.8g/L for H2SO4, HCl and the enzymatic preparation respectively. Concerning succinic acid production, the enzymatic hydrolyzates resulted in the highest yield (0.446g/g) and productivity (0.57g/Lh) using A. succinogenes in a batch reactor system. Repeated batch fermentation with immobilized A. succinogenes in agar and with the enzymatic hydrolyzates resulted in a maximum concentration of succinic acid of 33.6g/L from 87.2g/L monosaccharides after 5 cycles in 40h, obtaining a productivity of 1.32g/Lh.

Immobilization of Actinobacillus succinogenes by adhesion or entrapment for the production of succinic acid

The production of succinic acid was studied with entrapped and adsorbed Actinobacillus succinogenes. The adsorption of fermentation products (organic acids in the concentration range of 1-20 g/L) on different supports was evaluated. It was found that succinic acid was adsorbed in small quantities on diatomite and zeolite (12.6 mg/g support). The highest production of succinic acid was achieved with A. succinogenes entrapped in agar beads. Batch fermentations with immobilized cells were carried out with glucose concentrations ranging from 20 to 80 g/L. Succinic acid (43.4 g/L) was obtained from 78.3g/L glucose, and a high productivity (2.83 g/Lh) was obtained with a glucose concentration of 37.6g/L. For repeated batch fermentations (5 cycles in 72 h) with immobilized cells in agar, the total glucose consumed was 147.55 g/L, while the production of succinic acid was 107 g/L. Immobilized cells reduced significantly the fermentation time, yield, productivity and final concentration of succinic acid.

Highly Hydrophobic Wood Surfaces Prepared by Treatment With Atmospheric Pressure Dielectric Barrier Discharges

Atmospheric dielectric barrier discharge (DBD) treatments of wood were done to attain water repellency on wood surfaces. A specially designed frequency controlled parallel-plate DBD reactor was utilized to produce the discharges. Ethylene, methane, chlorotrifluoroethylene and hexafluoropropylene were used as DBD reagents. Contact angle, water absorption, X-ray photoelectron spectroscopy (XPS) and atomic force microscopy (AFM) measurements on the modified surfaces were performed. For methane and ethylene, XPS data showed an increased surface atomic concentration of carbon from 72.7% on untreated samples up to 80.7 and 96%, respectively, whereas nearly 50% fluorine concentration was observed with fluorinated reagents. The C1s spectrum of hexafluoropropylene-DBD-treated wood sample showed that the CF3 group was introduced in a relative amount of 19%. AFM images showed distinct features for each of the DBD treatments, such as a deposit of a thin uniform film in the case of ethylene-DBD treatment, whereas the hexafluoropropylene-DBD treatment resulted in the nucleation of plasma-derived entities at the fiber surface and the subsequent growth of a film. Under optimized conditions the water contact angle was in the range of 139°–145°. The combination of depositing a low surface energy polymer on an already rough surface gave the surface-treated wood a highly hydrophobic character.

Biomimetic Inks Based on Cellulose Nanofibrils and Cross-Linkable Xylans for 3D Printing

This paper presents a sustainable all-wood-based ink which can be used for 3D printing of constructs for a large variety of applications such as clothes, furniture, electronics, and health care products with a customized design and versatile gel properties. The 3D printing technologies where the material is dispensed in the form of liquids, so called inks, have proven suitable for 3D printing dispersions of cellulose nanofibrils (CNFs) because of their unique shear thinning properties. In this study, novel inks were developed with a biomimetic approach where the structural properties of cellulose and the cross-linking function of hemicelluloses that are found in the plant cell wall were utilized. The CNF was mixed with xylan, a hemicellulose extracted from spruce, to introduce cross-linking properties which are essential for the final stability of the printed ink. For xylan to be cross-linkable, it was functionalized with tyramine at different degrees. Evaluation of different ink compositions by rheology measurements and 3D printing tests showed that the degree of tyramine substitution and the ratio of CNFs to xylan-tyramine in the prepared inks influenced the printability and cross-linking density. Both two-layered gridded structures and more complex 3D constructs were printed. Similarly to conventional composites, the interactions between the components and their miscibility are important for the stability of the printed and cross-linked ink. Thus, the influence of tyramine on the adsorption of xylan to cellulose was studied with a quartz crystal microbalance to verify that the functionalization had little influence on xylans adsorption to cellulose. Utilizing xylan-tyramine in the CNF dispersions resulted in all-wood-based inks which after 3D printing can be cross-linked to form freestanding gels while at the same time, the excellent printing properties of CNFs remain intact.

Regular Motifs in Xylan Modulate Molecular Flexibility and Interactions with Cellulose Surfaces

The presence of regular glycosidic motifs in xylan modulates its molecular flexibility and the affinity toward hydrophobic and hydrophilic cellulose microfibril surfaces in secondary plant cell walls. Xylan is tightly associated with cellulose and lignin in secondary plant cell walls, contributing to its rigidity and structural integrity in vascular plants. However, the molecular features and the nanoscale forces that control the interactions among cellulose microfibrils, hemicelluloses, and lignin are still not well understood. Here, we combine comprehensive mass spectrometric glycan sequencing and molecular dynamics simulations to elucidate the substitution pattern in softwood xylans and to investigate the effect of distinct intramolecular motifs on xylan conformation and on the interaction with cellulose surfaces in Norway spruce (Picea abies). We confirm the presence of motifs with evenly spaced glycosyl decorations on the xylan backbone, together with minor motifs with consecutive glucuronation. These domains are differently enriched in xylan fractions extracted by alkali and subcritical water, which indicates their preferential positioning in the secondary plant cell wall ultrastructure. The flexibility of the 3-fold screw conformation of xylan in solution is enhanced by the presence of arabinofuranosyl decorations. Additionally, molecular dynamic simulations suggest that the glycosyl substitutions in xylan are not only sterically tolerated by the cellulose surfaces but that they increase the affinity for cellulose and favor the stabilization of the 2-fold screw conformation. This effect is more significant for the hydrophobic surface compared with the hydrophilic ones, which demonstrates the importance of nonpolar driving forces on the structural integrity of secondary plant cell walls. These novel molecular insights contribute to an improved understanding of the supramolecular architecture of plant secondary cell walls and have fundamental implications for overcoming lignocellulose recalcitrance and for the design of advanced wood-based materials.

Use of Agave tequilana-lignin and zinc oxide nanoparticles for skin photoprotection

The use of sunscreens is essential for preventing skin damage and the potential appearance of skin cancer in humans. Inorganic active components such as zinc oxide (ZnO) have been used commonly in sunscreens due to their ability to block UVA radiation. This ultraviolet (UV) protection might be enhanced to cover the UVB and UVC bands when combined with other components such as titanium dioxide (TiO2). In this work we evaluate the photoprotection properties of organic nanoparticles made from lignin in combination with ZnO nanoparticles as active ingredients for sunscreens. Lignin nanoparticles were synthesized from Agave tequilana lignin. Two different pulping methods were used for dissolving lignin from agave bagasse. ZnO nanoparticles were synthesized by the precipitation method. All nanoparticles were characterized by SEM, UV-Vis and FT-IR spectroscopy. Nanoparticles were mixed with a neutral vehicle in different concentrations and in-vitro sun protection factor (SPF) values were calculated. Different sizes of spherical lignin nanoparticles were obtained from the spent liquors of two different pulping methods. ZnO nanoparticles resulted with a flake shape. The mixture of all components gave SPF values in a range between 4 and 13. Lignin nanoparticles showed absorption in the UVB and UVC regions which can enhance the SPF value of sunscreens composed only of zinc oxide nanoparticles. Lignin nanoparticles have the added advantage of being of organic nature and its brown color can be used to match the skin tone of the person using it.