Facile method for stiff, tough, and strong nanocomposites by direct exfoliation of multilayered graphene into native nanocellulose matrix

Nanofibrillated cellulose (NFC) is a natural fibrillar material with exceptionally high mechanical properties. It has, however, been exceedingly difficult to achieve nanocomposites with drastically improved mechanical properties by dispersing NFC as random networks to polymer matrices, even using compatibilization. We show nanocomposites consisting of aligned assemblies of multilayered graphene and NFC with excellent tensile mechanical properties without any surface treatments. The optimum composition was found at 1.25 wt % graphene multilayers, giving a Young's modulus of 16.9 GPa, ultimate strength of 351 MPa, strain of 12%, and work-of-fracture of 22.3 MJ m(-3). This combines high strength with relatively high toughness and is obtained by direct exfoliation of graphite within aqueous hydrogels of NFC where an optimum sonication power is described. The results suggest the existence of an attractive interaction between multilayered graphene flakes and cellulose. Aligned assemblies are obtained by removal of water by filtration. The concept can be beneficial for applications because it results in high mechanical properties by a simple and environmentally green process.     

From interfacial ring-opening polymerization to melt processing of cellulose nanowhisker-filled polylactide-based nanocomposites

In the present work, cellulose nanowhiskers (CNWs), extracted from ramie fibers, were incorporated in polylactide (PLA)-based composites. Prior to the blending, PLA chains were chemically grafted on the surface of CNW to enhance the compatibilization between CNW and the hydrophobic polyester matrix. Ring-opening polymerization of l-lactide was initiated from the hydroxyl groups available at the CNW surface to yield CNW-g-PLA nanohybrids. PLA-based nanocomposites were prepared by melt blending to ensure a green concept of the study thereby limiting the use of organic solvents. The influence of PLA-grafted cellulose nanoparticles on the mechanical and thermal properties of the ensuing nanocomposites was deeply investigated. The thermal behavior and mechanical properties of the nanocomposites were determined using differential scanning calorimetry (DSC) and dynamical mechanical and thermal analysis (DMTA), respectively. It was clearly evidenced that the chemical grafting of CNW enhances their compatibility with the polymeric matrix and thus improves the final properties of the nanocomposites. Large modification of the crystalline properties such as the crystallization half-time was evidenced according to the nature of the PLA matrix and the content of nanofillers.     

Clay nanopaper composites of nacre-like structure based on montmorrilonite and cellulose nanofibers—Improvements due to chitosan addition

Clay nanopaper are nanocomposites with nacre-like structure and multifunctional characteristics including high modulus, significant strength and toughness as well as fire retardancy and low oxygen transmission rate (OTR). Montmorrilonite (MTM) and nanofibrillated cellulose (NFC) hydrocolloids are combined with a chitosan (CS) solution to form high MTM content nanopaper structures by the use of a previously developed papermaking approach. Chitosan functions as flocculation agent and decreases dewatering time to less than 10% compared with MTM-NFC clay nanopaper. The effect of chitosan on the clay nanopaper structure was studied by X-ray diffraction (XRD), scanning electron microscopy (SEM) and Fourier transform infrared (FTIR) spectroscopy. Properties were measured by uniaxial tensile testing, thermogravimetric analysis (TGA), OTR and moisture adsorption experiments. A nacre-like multilayered structure was confirmed and the chitosan-clay nanopaper showed favorable mechanical properties at clay contents as high as 44-48 wt%.     

Fibrous cellulose nanocomposite scaffolds prepared by partial dissolution for potential use as ligament or tendon substitutes

Fibrous cellulose nanocomposites scaffolds were developed and evaluated for their potential as ligament or tendon substitute. The nanocomposites were prepared by partial dissolution of cellulose nanofiber networks using ionic liquid at 80 °C for different time intervals. Scanning electron microscopy study indicated that partial dissolution resulted in fibrous cellulose nanocomposites where the dissolved cellulose nanofibers formed the matrix phase and the undissolved or partially dissolved nanofibers formed the reinforcing phase. Mechanical properties of the composites in simulated body conditions (37 °C and 95% RH) after sterilization using gamma rays was comparable to those of natural ligaments and tendons. Stress relaxation studies showed stable performance towards cyclic loading and unloading, further confirming the possibility for using these composites as ligament/tendon substitute. In vitro biocompatibility showed a positive response concerning adhesion/proliferation and differentiation for both human ligament and endothelial cells. Prototypes based on the cellulose composite were developed in the form of tubules to be used for further studies.     

Preparation of bionanoreactor based on core-shell structured polyion complex micelles entrapping trypsin in the core cross-linked with glutaraldehyde

Recently, the polyion complex (PIC) micelle has been suggested as a promising carrier system for peptide and proteins. However, its utilities are limited by its sensitivity to the environment such as dilution and ionic strength of the solution. In this study, to overcome these obstructions, PIC micelles prepared from an anionic block copolymer, poly(ethylene glycol)-poly(alpha,beta-aspartic acid), and a cationic protein, trypsin, were cross-linked with glutaraldehyde through the Schiff base formation. On the basis of a light scattering technique, the results revealed an efficient resistance of the cross-linked PIC micelle to a high salt concentration, which was a key parameter controlling the structure of the PIC micelles. Moreover, the stability of trypsin after cross-linking was remarkably improved. Evidently, as a bionanoreactor and/or bionanoreservoir, the PIC micelles entrapping protein molecules in the cross-linked core reveal an improved stability, allowing their wide application in the fields of biotechnology and pharmaceutical sciences.     

Strong and tough cellulose nanopaper with high specific surface area and porosity

In order to better understand nanostructured fiber networks, effects from high specific surface area of nanofibers are important to explore. For cellulose networks, this has so far only been achieved in nonfibrous regenerated cellulose aerogels. Here, nanofibrillated cellulose (NFC) is used to prepare high surface area nanopaper structures, and the mechanical properties are measured in tensile tests. The water in NFC hydrogels is exchanged to liquid CO2, supercritical CO2, and tert-butanol, followed by evaporation, supercritical drying, and sublimation, respectively. The porosity range is 40-86%. The nanofiber network structure in nanopaper is characterized by FE-SEM and nitrogen adsorption, and specific surface area is determined. High-porosity TEMPO-oxidized NFC nanopaper (56% porosity) prepared by critical point drying has a specific surface area as high as 482 m(2) g(-1). The mechanical properties of this nanopaper structure are better than for many thermoplastics, but at a significantly lower density of only 640 kg m(-3). The modulus is 1.4 GPa, tensile strength 84 MPa, and strain-to-failure 17%. Compared with water-dried nanopaper, the material is softer with substantiallly different deformation behavior.     

Immobilization-stabilization of proteins on nanofibrillated cellulose derivatives and their bioactive film formation

In a number of different applications for enzymes and specific binding proteins a key technology is the immobilization of these proteins to different types of supports. In this work we describe a concept for protein immobilization that is based on nanofibrillated cellulose (NFC). NFC is a form of cellulose where fibers have been disintegrated into fibrils that are only a few nanometers in diameter and have a very large aspect ratio. Proteins were conjugated through three different strategies using amine, epoxy, and carboxylic acid functionalized NFC. The conjugation chemistries were chosen according to the reactive groups on the NFC derivatives; epoxy amination, heterobifunctional modification of amino groups, and EDC/s-NHS activation of carboxylic acid groups. The conjugation reactions were performed in solution and immobilization was performed by spin coating the protein-NCF conjugates. The structure of NFC was shown to be advantageous for both protein performance and stability. The use of NFC allows all covalent chemistry to be performed in solution, while the immobilization is achieved by a simple spin coating or spreading of the protein-NFC conjugates on a support. This allows more scalable methods and better control of conditions compared to the traditional methods that depend on surface reactions.     

Effects of the incorporation of a hydrophobic middle block into a PEG-polycation diblock copolymer on the physicochemical and cell interaction properties of the polymer-DNA complexes

One-component homopolymers of cationic monomers (polycations) and diblock copolymers comprising poly(ethylene glycol) (PEG) and a polycation block have been the most widely used types of polymers for the formulation of polymer-based gene delivery systems. In this study, we incorporate a hydrophobic middle block into the conventional PEG-polycation architecture and investigate the effects of this hydrophobic modification on the physicochemical and cell-level biological properties of the polymer-DNA complexes that are relevant to gene delivery applications. The ABC-type triblock copolymer used in this study consists of (A) PEG, (B) hydrophobic poly( n-butyl acrylate) (PnBA), and (C) cationic poly(2-(dimethylamino)ethyl methacrylate) (PDMAEMA) component polymers. The properties of the triblock copolymer/DNA complexes are compared with those of two other more conventional DNA carriers derived, respectively, using a PDMAEMA homopolymer and a PEG-PDMAEMA diblock copolymer that had comparable molecular weights for individual blocks. In aqueous solution, the PEG-PnBA-PDMAEMA polymer forms positively charged spherical micelles. The electrostatic complexation of these micelles with plasmid DNA molecules results in the formation of stable small-sized DNA particles that are coated with a micelle monolayer, as confirmed by agarose gel electrophoresis, dynamic light scattering (DLS), and cryogenic transmission electron microscopy (cryo-TEM). Proton nuclear magnetic resonance ( (1)H NMR) spectroscopy measurements indicate that the whole micelle-DNA assembly (named "micelleplex" for convenience) is shielded predominantly by the PEG chains. DLS and optical microscopy imaging measurements indicate that compared with PDMAEMA-DNA polyplexes, the micelleplexes have a significantly lower tendency to aggregate under physiological salt concentrations and show reduced interactions with negatively charged components in serum such as albumin and erythrocytes. While the micelleplexes are comparable to the PEG-PDMAEMA-based DNA polyplexes in terms of their stability against aggregation under high salt concentrations and in the presence of the albumin protein, they have a slightly higher tendency to interact with erythrocytes than the diblock copolymer polyplexes. Agarose gel electrophoresis measurements indicate that relative to the PEG-PDMAEMA polyplexes, the micelleplexes provide better protection of the encapsulated DNA from enzymatic degradation and also exhibit greater stability against disintegration induced by polyanionic additives; in these respects, the PDMAEMA homopolymer-based polyplexes show the best performance. In vitro studies in HeLa cells indicate that the PDMAEMA polyplexes show the highest gene transfection efficiency among the three different gene delivery systems. Between the micelleplexes and the PEG-PDMAEMA polyplexes, a higher gene transfection efficiency is observed with the latter system. All three formulations show comparable levels of cytotoxicity in HeLa cells.     

Bacterial synthesis of PHA block copolymers

Polyhydroxyalkanoates (PHA) containing block copolymers were synthesized in Cupriavidus necator using periodic substrate addition. Poly(3-hydroxybutyrate) (PHB) segments were formed during fructose utilization. Pulse feeds of pentanoic acid resulted in the synthesis of 3-hydroxyvalerate monomers, forming poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) random copolymer. PHA synthesis was controlled using analysis of oxygen uptake and carbon evolution rates from the bioreactor off-gas. A combination of characterization techniques applied to the polymer batches strongly suggests the presence of block copolymers: (i) Thermodynamically stable polymer samples obtained by fractionation and analyzed by differential scanning calorimetry (DSC) and nuclear magnetic resonance spectroscopy (NMR) indicate that some fractions, representing approximately 30% of the total polymer sample, exhibit melting characteristics and nearest-neighbor statistics indicative of block copolymers, (ii) preliminary rheology experiments indicate additional mesophase transitions only found in block copolymer materials, (iii) dynamic mechanical analysis shows extension of the rubbery plateaus in block copolymer samples, and (iv) uniaxial extension tests result in differences in mechanical properties (modulus and elongation at failure) expected of similarly prepared block copolymer and single polymer type materials.     

Shell-cross-linked micelles containing cationic polymers synthesized via the RAFT process: toward a more biocompatible gene delivery system

Block copolymers poly(2-(dimethylamino) ethyl methacrylate)-b-poly(polyethylene glycol methacrylate) (PDMAEMA-b-P(PEGMA)) were prepared via reversible addition fragmentation chain transfer polymerization (RAFT). The polymerization was found to proceed with the expected living behavior resulting in block copolymers with varying block sizes of low polydispersity (PDI <1.3). The resulting block copolymer was self-assembled in an aqueous environment, leading to the formation of pH-responsive micelles. Further stabilization of the micellar system was performed in water using ethylene glycol dimethacrylate and the RAFT process to cross-link the shell. The cross-linked micelle was found to have properties significantly different from those of the uncross-linked block copolymer micelle. While a distinct critical micelle concentration (CMC) was observed using block copolymers, the CMC was absent in the cross-linked system. In addition, a better stability against disintegration was observed when altering the ionic strength such as the absence of changes of the hydrodynamic diameter with increasing NaCl concentration. Both cross-linked and uncross-linked micelles displayed good binding ability for genes. However, the cross-linked system exhibited a slightly superior tendency to bind oligonucleotides. Cytotoxicity tests confirmed a significant improvement of the biocompatibility of the synthesized cross-linked micelle compared to that of the highly toxic PDMAEMA. The cross-linked micelles were taken up by cells without causing any signs of cell damage, while the PDMAEMA homopolymer clearly led to cell death.     

Cationic amphiphilic star and linear block copolymers: synthesis, self-assembly, and in vitro gene transfection

A series of amphiphilic star and linear block copolymers were synthesized using ATRP. The core consisted of either polystyrene (PS) or poly(n-butyl acrylate) (PBuA), having different glass-transition (T(g)) values. These polymers were used as macroinitiators in the polymerization of the cationic 2-(dimethylamino)ethyl methacrylate (DMAEMA). The polymers were used to study the effects of polymer architecture and flexibility on the self-assembling properties, DNA complexation, and transfection. All polymers formed core-shell micelles in aqueous solutions and condensed plasmid DNA. Linear PDMAEMA-PBuA-PDMAEMA has transfection efficiency comparable to PEI25K in ARPE19 cell line. Glassy state of the micellar core and star-shaped architecture decreased the DNA transfection compared with the rubbery and linear polymer structures. The polymers showed low cellular toxicity at low nitrogen/phosphate (n/p) ratios.     

Novel biobased nanocomposites from soybean oil and functionalized organoclay

Novel biobased nanocomposites have been prepared by the cationic polymerization of conjugated soybean oil (CSOY) or conjugated LoSatSoy oil (CLS) with styrene (ST) and divinylbenzene (DVB), and a reactive organomodified montmorillonite (VMMT) clay as a reinforcing phase. This filler has been prepared by the cationic exchange of sodium montmorillonite with (4-vinylbenzyl)triethylammonium chloride in aqueous solution. The nanostructures of the nanocomposites have been determined by using wide-angle X-ray diffraction (WAXD) and transmission electron microscopy (TEM), respectively. The results from WAXD and TEM indicate that a heterogeneous structure consisting of intercalation and partial exfoliation or an intercalation structure exists in the nanocomposites, depending on the amount of VMMT in the polymer matrix. The thermal, mechanical, and organic vapor barrier properties of the nanocomposites have been evaluated by dynamic thermal analysis, thermogravimetric analysis, mechanical testing, and toluene absorption. A significant improvement is observed in the thermal stability, the dynamic bending storage modulus, the compressive modulus, the compressive strength, the compressive strain at failure, and the vapor barrier performance for the CSOY-- and CLS-based nanocomposites with 1-2 wt % VMMT loading, where some individual exfoliated silicate platelets occur. For example, the CLS-based nanocomposite with 1-2 wt % VMMT exhibits increases of 100-128%, 86-92%, and 5-7% in compressive modulus, compressive strength, and compressive strain at failure, respectively. CLS with higher unsaturation and reactivity affords nanocomposites with higher thermal stability and higher mechanical properties than CSOY.     

Surface modification of cellulose fibers: towards wood composites by biomimetics

A biomimetic approach was taken for studying the adsorption of a model copolymer (pullulan abietate, DS 0.027), representing the lignin-carbohydrate complex, to a model surface for cellulose fibers (Langmuir-Blodgett thin films of regenerated cellulose). Adsorption results were assayed using surface plasmon resonance spectroscopy (SPR) and atomic force microscopy (AFM). Rapid, spontaneous, and desorption-resistant surface modification resulted. This effort is viewed as a critical first step towards the permanent surface modification of cellulose fibers with a layer of molecules amenable to either enzymatic crosslinking for improved wood composites or thermoplastic consolidation.     

Controlled thermoresponsive hydrogels by stereocomplexed PLA-PEG-PLA prepared via hybrid micelles of pre-mixed copolymers with different PEG lengths

The stereocomplexed hydrogels derived from the micelle mixture of two enantiomeric triblock copolymers, PLLA-PEG-PLLA and PDLA-PEG-PDLA, reported in 2001 exhibited sol-to-gel transition at approximately body temperature upon heating. However, the showed poor storage modulus (ca. 1000 Pa) determined their insufficiency as injectable implant biomaterials for many applications. In this study, the mechanical property of these hydrogels was significantly improved by the modifications of molecular weights and micelle structure. Co-micelles composed of block copolymers with two sizes of PEG block length were shown to possess unique and dissimilar properties from the micelles composed of single-sized block copolymers. The stereomixture of PLA-PEG-PLA comicelles showed a controllable sol-to-gel transition at a wide temperature range of 4 and 80 °C. The sol-gel phase diagram displays a linear relationship of temperature versus copolymer composition; hence, a transition at body temperature can be readily achieved by adjusting the mixed copolymer ratio. The resulting thermoresponsive hydrogels exhibit a storage modulus notably higher (ca. 6000 Pa) than that of previously reported hydrogels. As a physical network solely governed by self-reorganization of micelles, followed by stereocomplexation, this unique system offers practical, safe, and simple implantable biomaterials.     

Oxime linkage: a robust tool for the design of pH-sensitive polymeric drug carriers

Oxime bonds dispersed in the backbones of the synthetic polymers, while young in the current spectrum of the biomedical application, are rapidly extending into their own niche. In the present work, oxime linkages were confirmed to be a robust tool for the design of pH-sensitive polymeric drug delivery systems. The triblock copolymer (PEG-OPCL-PEG) consisting of hydrophilic poly(ethylene glycol) (PEG) and hydrophobic oxime-tethered polycaprolactone (OPCL) was successfully prepared by aminooxy terminals of OPCL ligating with aldehyde-terminated PEG (PEG-CHO). Owing to its amphiphilic architecture, PEG-OPCL-PEG self-assembled into the micelles in aqueous media, validated by the measurement of critical micelle concentration (CMC). The MTT assay showed that PEG-OPCL-PEG exhibited low cytotoxicity against NIH/3T3 normal cells. Doxorubicin (DOX) as a model drug was encapsulated into the PEG-OPCL-PEG micelles. Drug release study revealed that the DOX release from micelles was significantly accelerated at mildly acid pH of 5.0 compared to physiological pH of 7.4, suggesting the pH-responsive feature of the drug delivery systems with oxime linkages. Flow cytometry and confocal laser scanning microscopy (CLSM) measurements indicated that these DOX-loaded micelles were easily internalized by living cells. MTT assay against HeLa cancer cells showed DOX-loaded PEG-OPCL-PEG micelles had a high anticancer efficacy. All of these results demonstrate that these polymeric micelles self-assembled from oxime-tethered block copolymers are promising carriers for the pH-triggered intracellular delivery of hydrophobic anticancer drugs.     

Cellular internalization of poly(ethylene oxide)-b-poly(epsilon-caprolactone) diblock copolymer micelles

Poly(ethylene oxide)-b-poly(epsilon-caprolactone) (PEO-b-PCL) block copolymers self-assemble into micelles in aqueous solution. We have examined whether these micelles can internalize into P19 cells in vitro. Fluorescently labeled PEO(45)-b-PCL(23) block copolymer was prepared by conjugating a tetramethylrhodamine molecule to the end of the hydrophobic PCL block. Dynamic light scattering (DLS) and transmission electron microscopy (TEM) studies yielded 24 +/- 2 and 25 +/- 2 nm, respectively, for the diameters of the micelles. The studies also showed that chemical labeling did not effect the morphology or size. When the rhodamine-labeled PEO(45)-b-PCL(23) block copolymer micelles were tested in vitro, time-, concentration-, and pH-dependence of the internalization process suggested that internalization proceeded by endocytosis. The results from these studies provide the first direct evidence for the internalization of PEO(45)-b-PCL(23) micelles. Future studies will utilize multiple labeling of these micelles, allowing questions to be addressed related to the fate of internalized micelles as drug carriers, the destination of the incorporated drugs or fluorescent probes released from micelles, and the identification of the subcellular localization of the whole drug-carrier system within cells, both in vitro and in vivo.     

Towards Biomimicking Wood: Fabricated Free-standing Films of Nanocellulose,Lignin, and a Synthetic Polycation

Woody materials are comprised of plant cell walls that contain a layered secondary cell wall composed of structural polymers of polysaccharides and lignin. Layer-by-layer (LbL) assembly process which relies on the assembly of oppositely charged molecules from aqueous solutions was used to build a freestanding composite film of isolated wood polymers of lignin and oxidized nanofibril cellulose (NFC). To facilitate the assembly of these negatively charged polymers, a positively charged polyelectrolyte, poly(diallyldimethylammomium chloride) (PDDA), was used as a linking layer to create this simplified model cell wall. The layered adsorption process was studied quantitatively using quartz crystal microbalance with dissipation monitoring (QCM-D) and ellipsometry. The results showed that layer mass/thickness per adsorbed layer increased as a function of total number of layers. The surface coverage of the adsorbed layers was studied with atomic force microscopy (AFM). Complete coverage of the surface with lignin in all the deposition cycles was found for the system, however, surface coverage by NFC increased with the number of layers. The adsorption process was carried out for 250 cycles (500 bilayers) on a cellulose acetate (CA) substrate. Transparent free-standing LBL assembled nanocomposite films were obtained when the CA substrate was later dissolved in acetone. Scanning electron microscopy (SEM) of the fractured cross-sections showed a lamellar structure, and the thickness per adsorption cycle (PDDA-Lignin-PDDA-NC) was estimated to be 17 nm for two different lignin types used in the study. The data indicates a film with highly controlled architecture where nanocellulose and lignin are spatially deposited on the nanoscale (a polymer-polymer nanocomposites), similar to what is observed in the native cell wall.     

Supramolecular copolymer micelles based on the complementary multiple hydrogen bonds of nucleobases for drug delivery

Novel supramolecular copolymer micelles with stimuli-responsive abilities were successfully prepared through the complementary multiple hydrogen bonds of nucleobases and then applied for rapid intracellular release of drugs. First, both adenine-terminated poly(ε-caprolactone) (PCL-A) and uracil-terminated poly(ethylene glycol) (PEG-U) were synthesized. The supramolecular amphiphilic block copolymers (PCL-A:U-PEG) were formed based on multiple hydrogen bonding interactions between PCL-A and PEG-U. The micelles self-assembled from PCL-A:U-PEG were sufficiently stable in water but prone to fast aggregation in acidic condition due to the dynamic and sensitive nature of noncovalent interactions. The low cytotoxicity of supramolecular copolymer micelles was confirmed by MTT assay against NIH/3T3 normal cells. As a hydrophobic anticancer model drug, doxorubicin (DOX) was encapsulated into these supramolecular copolymer micelles. In vitro release studies demonstrated that the release of DOX from micelles was significantly faster at mildly acid pH of 5.0 compared to physiological pH. MTT assay against HeLa cancer cells showed DOX-loaded micelles had high anticancer efficacy. Hence, these supramolecular copolymer micelles based on the complementary multiple hydrogen bonds of nucleobases are very promising candidates for rapid controlled release of drugs.