Showing papers in "Macromolecular Bioscience in 2004"

An Overview of Polylactides as Packaging Materials

Abstract: Polylactide polymers have gained enormous attention as a replacement for conventional synthetic packaging materials in the last decade. By being truly biodegradable, derived from renewable resources and by providing consumers with extra end-use benefits such as avoiding paying the "green tax" in Germany or meeting environmental regulations in Japan, polylactides (PLAs) are a growing alternative as a packaging material for demanding markets. The aim of this paper is to review the production techniques for PLAs, summarize the main properties of PLA and to delineate the main advantages and disadvantages of PLA as a polymeric packaging material. PLA films have better ultraviolet light barrier properties than low density polyethylene (LDPE), but they are slightly worse than those of cellophane, polystyrene (PS) and poly(ethylene terephthalate) (PET). PLA films have mechanical properties comparable to those of PET and better than those of PS. PLA also has lower melting and glass transition temperatures than PET and PS. The glass transition temperature of PLA changes with time. Humidity between 10 and 95% and storage temperatures of 5 to 40 degrees C do not have an effect on the transition temperature of PLA, which can be explained by its low water sorption values (i.e. <100 ppm at Aw = 1). PLA seals well at temperatures below the melting temperature but an appreciable shrinking of the films has been noted when the material is sealed near its melting temperature. Solubility parameter predictions indicate that PLA will interact with nitrogen compounds, anhydrides and some alcohols and that it will not interact with aromatic hydrocarbons, ketones, esters, sulfur compounds or water. The CO2, O2 and water permeability coefficients of PLA are lower than those of PS and higher than those of PET. Its barrier to ethyl acetate and D-limonene is comparable to PET. The amount of lactic acid and its derivatives that migrate to food simulant solutions from PLA is much lower than any of the current average dietary lactic acid intake values allowed by several governmental agencies. Thus, PLA is safe for use in fabricating articles for contact with food.

Bone Tissue Engineering: State of the Art and Future Trends

Abstract: Although several major progresses have been introduced in the field of bone regenerative medicine during the years, current therapies, such as bone grafts, still have many limitations. Moreover, and in spite of the fact that material science technology has resulted in clear improvements in the field of bone substitution medicine, no adequate bone substitute has been developed and hence large bone defects/injuries still represent a major challenge for orthopaedic and reconstructive surgeons. It is in this context that TE has been emerging as a valid approach to the current therapies for bone regeneration/substitution. In contrast to classic biomaterial approach, TE is based on the understanding of tissue formation and regeneration, and aims to induce new functional tissues, rather than just to implant new spare parts. The present review pretends to give an exhaustive overview on all components needed for making bone tissue engineering a successful therapy. It begins by giving the reader a brief background on bone biology, followed by an exhaustive description of all the relevant components on bone TE, going from materials to scaffolds and from cells to tissue engineering strategies, that will lead to "engineered" bone. Scaffolds processed by using a methodology based on extrusion with blowing agents.

The sustainability of NatureWorks polylactide polymers and Ingeo polylactide fibers: an update of the future.

Abstract: Cargill Dow LLC produces a new packaging and fiber material – polylactide (PLA) – from annually renewable resources. The presence of the company and the PLA polymer on the industrial scene signals the emergence of a new model for industrial development in the twenty-first century. During the nineteenth century people relied predominantly on a wide range of natural materials, such as wood, hides, wool and starch, to provide the essentials of then-modern life. This picture changed significantly during the twentieth century, when people in developed countries experiencing the industrial revolution became almost totally dependent on fossil materials to produce the fuels, polymers and chemicals required for modern life. With exponential growth in the demand for fossil raw materials in both developing and developed countries today, the question increasingly posed is how we will derive the materials we will need in the twenty-first century. An increasingly broad range of experts and analysts have concluded that new – or in some very important ways, ‘‘old’’ – raw materials will become the foundation of packaging materials and fibers in a world challenged by the interrelated problems of depletion of fossil resources and of proliferation of global climate changing emissions, pollutants and solid wastes. A consensus remains that fossil resources will be required and used for quite some time, but it is also hardly doubted that maintaining and enhancing quality of life for a growing populationaround theglobe compels the developmentofnew technologies to produce packaging materials and fibers from new ‘‘old’’ resources like traditional agricultural crops (e.g., corn, wheat, sugar beets) and other grown biomass materials. It is in this new and renewed reality of renewable rawmaterials reliance that Cargill Dow LLC emerges. The objective of this paper is to answer the question: ‘‘What makes NatureWorks PLA a more sustainable polymer?’’ To answer this question the article addresses applications and marketing of PLA, costs, today’s and future renewable raw material resources, reduction of fossil fuels and the associated emissions of green house gases, waste management options, and manufacturing processes.

Peptide/protein hybrid materials: enhanced control of structure and improved performance through conjugation of biological and synthetic polymers.

Abstract: The conjugation of peptides/proteins and synthetic polymers is a useful strategy to overcome some of the limitations related to the use of the individual components. This review will highlight two aspects: enhanced structural control at the nanometer level and improved performance, in particular with respect to biomedical applications. In the former case, peptide sequences are mainly used to mediate self-assembly of synthetic polymers. In the latter case, conjugation of an appropriate synthetic polymer to a pharmaceutically active peptide/protein can, for example, prevent premature enzymatic degradation and enhance blood circulation times, which is therapeutically advantageous.

Production and characterization of chitosan fibers and 3-D fiber mesh scaffolds for tissue engineering applications.

Abstract: Summary: This study reports on the production of chitosan fibers and 3-D fiber meshes for the use as tissue engineering scaffolds. Both structures were produced by means of a wet spinning technique. Maximum strain at break and tensile strength of the developed fibers were found to be 8.5% and 204.9 MPa, respectively. After 14 d of immersion in simulated body fluid (SBF), scanning electron microscopy (SEM), energy dispersive spectroscopy (EDS), and inductively coupled plasma emission (ICP) spectroscopy analyses showed that a bioactive Ca-P layer was formed on the surface of the fibers, meaning that they exhibit a bioactive behavior. The samples showed around 120% max. swelling in physiological conditions. The pore sizes of 3-D chitosan fiber mesh scaffolds were observed to be in the range of 100–500m mb y SEM. The equilibrium-swelling ratio of the developed scaffolds was found to be around 170% (w/w) in NaCl solution at 378C. Besides that, the limit swelling strain was less than 30%, as obtained by mechanical spectroscopy measurements in the same conditions. The viscoelastic properties of the scaffolds were also evaluated by both creep and dynamic mechanical tests. By means of using short-term MEM extraction test, both types of structures (fibers and scaffolds) were found to be non-cytotoxic to fibroblasts. Furthermore, osteoblasts directly cultured over chitosan fiber mesh scaffolds presented good morphology and no inhibition of cell proliferation could be observed.

Polymer Alloys of Nodax Copolymers and Poly(lactic acid)

Abstract: Properties of polymer alloys comprising poly (lactic acid) and Nodax copolymers are investigated. Nodax is a family of bacterially produced polyhydroxyalkanoate (PHA) copolymers comprising 3-hydroxybutyrate (3HB) and other 3-hydroxyalkanoate (3HA) units with side groups greater than or equal to three carbon units. The incorporation of 3HA units with medium-chain-length (mcl) side groups effectively lowers the crystallinity and the melt temperature, T m , of this class of PHA copolymers, in a manner similar to that of alpha olefins controlling the properties of linear low density polyethylene. The lower T m makes the material easier to process, as the thermal decomposition temperature of PHAs is then relatively low. The reduced crystallinity provides the ductility and toughness required for many plastics applications. When a small amount of ductile PHA is blended with poly(lactic acid) (PLA), a new type of polymer alloy with much improved properties is created. The toughness of PLA is substantially increased without a reduction in the optical clarity of the blend. The synergy between the two materials, both produced from renewable resources, is attributed to the retardation of crystallization of PHA copolymers finely dispersed in a PLA matrix as discrete domains.

Biomimetic Polymer/Apatite Composite Scaffolds for Mineralized Tissue Engineering

Abstract: The material surface must be considered in the design of scaffolds for bone tissue engineering so that it supports bone cells adhesion, proliferation and differentiation. A biomimetic approach has been developed as a 3D surface modification technique to grow partially carbonated hydroxyapatite (the bonelike mineral) in prefabricated, porous, polymer scaffolds using a simulated body fluid in our lab. For the rational design of scaffolding materials and optimization of the biomimetic process, this work focused on various materials and processing parameters in relation to apatite formation on 3D polymer scaffolds. The apatite nucleation and growth in the internal pores of poly(L-lactide) and poly(D,L-lactide) scaffolds were significantly faster than in those of poly(lactide-co-glycolide) scaffolds in simulated body fluids. The apatite distribution was significantly more uniform in the poly(L-lactide) scaffolds than in the poly(lactide-co-glycolide) scaffolds. After incubation in a simulated body fluid for 30 d, the mass of poly(L-lactide) scaffolds increased approximately 40%, whereas the mass of the poly(lactide-co-glycolide) scaffolds increased by about 15% (see Figure). A higher ionic concentration and higher pH value of the simulated body fluid enhanced apatite formation. The effects of surface functional groups on apatite nucleation and growth were found to be more complex in 3D scaffolds than on 2D films. Surprisingly enough, it was found that carboxyl groups significantly reduced the apatite formation, especially on the internal pore surfaces of 3D scaffolds. These findings are critically important in the rational selection of materials and surface design of 3D scaffolds for mineralized tissue engineering and may contribute to the understanding of biomineralization as well.SEM micrograph of a poly(L-lactide) scaffold.

Enzymatic degradation of poly(L-lactide) and poly(epsilon-caprolactone) electrospun fibers.

Abstract: Poly(L-lactide) (PLLA) and poly(epsilon-caprolactone) (PCL) ultrafine fibers were prepared by electrospinning. The influence of cationic and anionic surfactants on their enzymatic degradation behavior was investigated by measuring weight loss, molecular weight, crystallinity, and melting temperature of the fibers as a function of degradation time. Under the catalysis of proteinase K, the PLLA fibers containing the anionic surfactant sodium docecyl sulfate (SDS) exhibited a faster degradation rate than those containing cationic surfactant triethylbenzylammonium chloride (TEBAC), indicating that surface electric charge on the fibers is a critical factor for an enzymatic degradation. Similarly, TEBAC-containing PCL fibers exhibited a 47% weight loss within 8.5 h whereas SDS-containing PCL fibers showed little degradation in the presence of lipase PS. By analyzing the charge status of proteinase K and lipase PS under the experimental conditions, the importance of the surface charges of the fibers and their interactions with the charges on the enzymes were revealed. Consequently, a "two-step" degradation mechanism was proposed: (1) the enzyme approaches the fiber surface; (2) the enzyme initiates hydrolysis of the polymer. By means of differential scanning calorimetry and wide-angle X-ray diffraction, the crystallinity and orientation changes in the PLLA and PCL fibers during the enzymatic degradation were investigated, respectively.

Asymmetric ABC-triblock copolymer membranes induce a directed insertion of membrane proteins.

Abstract: Asymmetric molecules and materials provide an important basis for the organization and function of biological systems. It is well known that, for example, the inner and outer leaflets of biological membranes are strictly asymmetric with respect to lipid composition and distribution. This plays a crucial role for many membrane-related processes like carrier-mediated transport or insertion and orientation of integral membrane proteins. Most artificial membrane systems are, however, symmetric with respect to their midplane and membrane proteins are incorporated with random orientation. Here we describe a new approach to induce a directed insertion of membrane proteins into asymmetric membranes formed by amphiphilic ABC triblock copolymers with two chemically different water-soluble blocks A and C. In a comparative study we have reconstituted His-tag labeled Aquaporin 0 in lipid, ABA block copolymer, and ABC block copolymer vesicles. Immunolabeling, colorimetric, and fluorescence studies clearly show that a preferential orientation of the protein is only observed in the asymmetric ABC triblock copolymer membranes.

Environmentally degradable bio-based polymeric blends and composites.

Abstract: Blends and composites based on environmentally degradable-ecocompatible synthetic and natural polymeric materials and fillers of natural origin have been prepared and processed under different conditions. Poly (vinyl alcohol (PVA) was used as the synthetic polymer of choice by virtue of its capability to be processed from water solution or suspension as well as from the melt by blow extrusion and injection molding. Starch and gelatin were taken as the polymeric materials from renewable resources. The fillers were all of natural origin, as waste from food and agro-industry consisted of sugar cane bagasse (SCB), wheat flour (WF), orange peels (OR), apple peels (AP), corn fibres (CF), saw dust (SD) and wheat straw (WS). All the natural or hybrid formulations were intended to be utilized for the production of: a) Environmentally degradable mulching films (hydrobiomulching) displaying, in some cases, self-fertilizing characteristics by in situ spraying of water solutions or suspensions; b) Laminates and containers to be used in agriculture and food packaging by compression and injection molding followed by baking. Some typical prototype items have been prepared and characterized in relation to their morphological and mechanical properties and tested with different methodology for their propensity to environmental degradation and biodegradation as ultimate stage of theirservice life. A relationship between chemical composition and mechanical properties and propensity to biodegradation has been discussed in a few representative cases.

Evolution of polyhydroxyalkanoate (PHA) production system by "enzyme evolution": successful case studies of directed evolution.

Abstract: Biotechnological studies towards the biosynthesis of polyhydroxyalkanoates (PHAs) biopolyesters have extensively progressed through the development of various metabolic engineering strategies. Historically, efficient PHA production has been achieved using the fermentation technology of naturally occurring PHA-producing bacteria based on external substrate manipulation (1st generation), and subsequent reinforcement with recombinant gene technology (2nd generation). More recently, "enzyme evolution" is becoming the 3rd generation approach for PHA production. A break-through in the chemical synthesis of macromolecules with desirable properties was achieved by the development of prominent chemical catalysts via "catalyst evolution", as represented by a series of Ziegler-Natta catalysts. Thus, one can easily accept the concept that the molecular evolution of the biocatalysts (enzymes) relevant to PHA synthesis will provide us with a chance to create novel PHA materials with high performance. The first trial of an in vitro enzyme evolution in PHA biosynthesis was reported by our group in 2001. The following literature data, as well as our own experimental results devoted to this new approach, have been accumulated over a short time. This review article focuses specifically on the concept and current case studies of the application of "enzyme evolution" to PHA biosynthesis.

Fermentative production of chemicals that can be used for polymer synthesis.

Abstract: The fermentative production of chemicals that can be used as monomers for the synthesis of polymers has become an important topic in biotechnology research because of the limited nature of petroleum and environmental issues. In particular, the fermentative production of metabolites such as dicarboxylic acids, amino acids, and diols, which are suitable as building blocks for subsequent polymerization, has attracted much attention. Various wild-type and metabolically engineered microorganisms have been developed for the efficient production of these chemicals from renewable resources. In addition, the development of fermentation strategies to achieve the highest possible productivities has been another focus of research, considering that these monomers should be produced at costs low enough to compete with petroleum-derived ones. In this paper, the metabolic pathways leading to the synthesis of such important monomers including succinic acid, lactic acid, fumaric acid, propan-1,2-diol, and propan-1,3-diol are reviewed. In addition, the metabolic engineering and fermentation strategies for their production are reviewed.

Amphiphilic block copolymers based on poly(2-acryloyloxyethyl phosphorylcholine) prepared via RAFT polymerisation as biocompatible nanocontainers.

Abstract: Amphiphilic block copolymers composed of poly(butyl acrylate) and poly(2-acryloyloxyethyl phosphorylcholine) have been prepared using reversible addition fragmentation transfer (RAFT) polymerisation. The conversion of the polymerisation was determined using online FT NIR spectroscopy. NMR spectroscopy was used not only to support the results obtained from FT NIR spectroscopy but also prove the formation of micelles. Due to the strong aggregation tendency of these block copolymers and the resulting difficulties concerning the molecular weight analysis test experiments were carried out replacing poly(2-acryloyloxyethyl phosphorylcholine) with poly(2-hydroxyethyl acrylate). Micelle size and the aggregation behaviour were investigated using dynamic light scattering. The sizes of the nanocontainers obtained were found to be influenced by the block length as well as the solvent leading to micelles in the range between 40 and 160 nm. The toxicity of the RAFT agent used was then analysed by cell growth inhibition tests.

Production of D-lactic acid by bacterial fermentation of rice starch.

Abstract: D-Lactic acid was synthesized by the fermentation of rice starch using microorganisms. Two species: Lactobacillus delbrueckii and Sporolactobacillus inulinus were found to be active in producing D-lactic acid of high optical purity after an intensive screening test for D-lactic acid bacteria using glucose as substrate. Rice powder used as the starch source was hydrolyzed with a combination of enzymes: alpha-amylase, beta-amylase, and pullulanase to obtain rice saccharificate consisting of maltose as the main component. Its average gross yield was 82.5%. Of the discovered D-lactic acid bacteria, only Lactobacillus delbrueckii could ferment both maltose and the rice saccharificate. After optimizing the fermentation of the rice saccharificate using this bacterium, pilot scale fermentation was conducted to convert the rice saccharificate into D-lactic acid with a D-content higher than 97.5% in a yield of 70%. With this yield, the total yield of D-lactic acid from brown rice was estimated to be 47%, which is almost equal to the L-lactic acid yield from corn. The efficient synthesis of D-lactic acid can open a way to the large scale application of high-melting poly(lactic acid) that is a stereocomplex of poly(L-lactide) and poly(D-lactide). Schematic representation of the production of D-lactic acid starting from brown rice as described here.

Polypyrrole Thin Films Formed by Admicellar Polymerization Support the Osteogenic Differentiation of Mesenchymal Stem Cells

Abstract: The objective of this study was to evaluate the attachment, proliferation, and differentiation of rat mesenchymal stem cells (MSC) toward the osteoblastic phenotype seeded on polypyrrole (PPy) thin films made by admicellar polymerization. Three different concentrations of pyrrole (Py) monomer (20, 35, and 50 x 10(-3) M) were used with the PPy films deposited on tissue culture polystyrene dishes (TCP). Regular TCP dishes and PPy polymerized on TCP by chemical polymerization without surfactant using 5 x 10(-3) M Py, were used as controls. Rat MSC were seeded on these surfaces and cultured for up to 20 d in osteogenic media. Surface topography was characterized by atomic force microscopy, X-ray photoelectron spectroscopy, and static contact angle. Cell attachment, proliferation, alkaline phosphatase (ALP) activity, and calcium content were measured to evaluate the ability of MSC to adhere and differentiate on PPy-coated TCP. Increased monomer concentrations resulted in PPy films of increased thickness and surface roughness. PPy films generated by different monomer concentrations induced drastically different cellular events. A wide spectrum of cell attachment characteristics (from excellent cell attachment to the complete inability to adhere) were obtained by varying the monomer concentration from 20 m to 50 x 10(-3) M. In particular the 20 x 10(-3) M PPy thin films demonstrated superior induction of MSC osteogenicity, which was comparable to standard TCP dishes, unlike PPy films of similar thickness prepared by chemical polymerization without surfactant. Adhesion of mesenchymal stem cells on tissue culture plates (TCP) coated with polypyrrole thin films made by admicellar polymerization.

Supramolecular Hydrogel Formation Based on Inclusion Complexation Between Poly(ethylene glycol)‐Modified Chitosan and α‐Cyclodextrin

Abstract: Supramolecular hydrogels have been prepared on the basis of polymer inclusion complex (PIC) formation between poly(ethylene glycol) (PEG)-modified chitosans and alpha-cyclodextrin (alpha-CD). A series of PEG-modified chitosans were synthesized by coupling reactions between chitosan and monocarboxylated PEG using water-soluble carbodiimide (EDC) as coupling agent. With simple mixing, the resultant supramolecular assembly of the polymers and alpha-CD molecules led to hydrogel formation in aqueous media. The supramolecular structure of the PIC hydrogels was confirmed by differential scanning calorimetry (DSC), X-ray diffraction, and (13)C cross-polarized/magic-angle spinning (CP/MAS) NMR characterization. The PEG side-chains on the chitosan backbones were found to form inclusion complexes (ICs) with alpha-CD molecules, resulting in the formation of channel-type crystalline micro-domains. The IC domains play an important role in holding together hydrated chitosan chains as physical junctions. The gelation property was affected by several factors including the PEG content in the polymers, the solution concentration, the mixing ratio of host and guest molecules, temperature, pH, etc. All the hydrogels in acidic conditions exhibited thermo-reversible gel-sol transitions under appropriate conditions of mixing ratio and PEG content in the mixing process. The transitions were induced by supramolecular association and dissociation. These supramolecular hydrogels were found to have phase-separated structures that consist of hydrophobic crystalline PIC domains, which were formed by the host-guest interaction between alpha-CD and PEG, and hydrated chitosan matrices below the pK(a).The formation of inclusion complexes between alpha-cyclodextrin and PEG-modified chitosan leads to the formation of hydrogels that can undergo thermo-reversible supramolecular dissociation.

Structure and Properties of Novel Fibers Spun from Cellulose in NaOH/Thiourea Aqueous Solution

Abstract: Cellulose was dissolved rapidly in a NaOH/ thiourea aqueous solution (95:45 in wt-%) to prepare a transparent cellulose solution, which was employed, for the first time, to spin a new class of regenerated cellulose fibers by wet spinning The structure and mechanical properties of the resulting cellulose fibers were characterized, and compared with those of commercially available viscose rayon, cuprammonium rayon and Lyocell fibers The results from wide angle X-ray diffraction and CP/MAS 13 C NMR indicated that the novel cellulose fibers have a structure typical for a family II cellulose and possessed relatively high degrees of crystallinity Scanning electron microscopy (SEM) and optical microscopy images revealed that the cross-section of the fibers is circular, similar to natural silk The new fibers have higher molecular weights and better mechanical properties than those of viscose rayon This low-cost technology is simple, different from the polluting viscose process The dissolution and regeneration of the cellulose in the NaOH/ thiourea aqueous solutions were a physical process and a sol-gel transition rather than a chemical reaction, leading to the smoothness and luster of the fibers This work provides a potential application in the field of functional fiber manufacturing

Spherical molecularly imprinted polymers (SMIPs) via a novel precipitation polymerization in the controlled delivery of sulfasalazine.

Abstract: Spherical molecularly imprinted polymers (SMIPs) have been prepared via a novel precipitation polymerization using sulfasalazine (prodrug used in the diseases of the colon) as template. The sulfasalazine was incorporated into SMIPs and into a spherical non-imprinted polymer (control), and then the release rate of the bioactive agent at different pH values was evaluated. Considerable differences in the release characteristics between imprinted and non-imprinted polymers have been observed. This opens the possibility of the development of drug release systems capable of modulating the release of a specific molecule. Photomicrography of spherical molecularly imprinted polymers (SMIPs).

Lactic acid-based functionalized polymers via copolymerization and chemical modification.

Abstract: Poly(lactic acid) polymers (PLA) are presently the most attractive compounds in the field of artificial degradable and biodegradable polymers. In order to enlarge the family, and thus the range of accessible properties, stereocopolymers and copolymers with various co-monomers have been synthesized. However, very few are functionalized, i.e. include functional groups attached to the main chains or as part of the side chains. In the search for degradable PLA-type polymers bearing functional groups to serve as intermediates for further chemical modifications, we are exploring two different routes. The first one is copolymerization with a protected hydroxyl-bearing lactide-type monomer, namely 3-(1,2,3,4-tetraoxobutyldiisopropylidene)dioxane-2,5-dione. The second route consists of the formation of a carbanionic site in the alpha-position to intrachain carbonyl functions by using lithium N,N-diisopropylamide followed by the coupling of electrophiles. Recent advances in this search are presented using several examples. In particular, it is shown that OH-functionalized PLA-type macromolecules can be made fluorescent by chemical coupling. It is also shown that substituents can be attached to PLA-type macromolecules in solution or to the surface of PLA-based devices selectively.

Thermally produced biodegradable scaffolds for cartilage tissue engineering.

Abstract: A novel process was developed to fabricate biodegradable polymer scaffolds for tissue engineering applications, without using organic solvents. Solvent residues in scaffolds fabricated by processes involving organic solvents may damage cells transplanted onto the scaffolds or tissue near the transplantation site. Poly(L-lactic acid) (PLLA) powder and NaCl particles in a mold were compressed and subsequently heated at 180 degrees C (near the PLLA melting temperature) for 3 min. The heat treatment caused the polymer particles to fuse and form a continuous matrix containing entrapped NaCl particles. After dissolving the NaCl salts, which served as a porogen, porous biodegradable PLLA scaffolds were formed. The scaffold porosity and pore size were controlled by adjusting the NaCl/PLLA weight ratio and the NaCl particle size. The characteristics of the scaffolds were compared to those of scaffolds fabricated using a conventional solvent casting/particulate leaching (SC/PL) process, in terms of pore structure, pore-size distribution, and mechanical properties. A scanning electron microscopic examination showed highly interconnected and open pore structures in the scaffolds fabricated using the thermal process, whereas the SC/PL process yielded scaffolds with less interconnected and closed pore structures. Mercury intrusion porosimetry revealed that the thermally produced scaffolds had a much more uniform distribution of pore sizes than the SC/PL process. The utility of the thermally produced scaffolds was demonstrated by engineering cartilaginous tissues in vivo. In summary, the thermal process developed in this study yields tissue-engineering scaffolds with more favorable characteristics, with respect to, freedom from organic solvents, pore structure, and size distribution than the SC/PL process. Moreover, the thermal process could also be used to fabricate scaffolds from polymers that are insoluble in organic solvents, such as poly(glycolic acid). Cartilage tissue regenerated from thermally produced PLLA scaffold.

Biodegradable polyesters reinforced with surface-modified vegetable fibers

Abstract: Flax fibers are investigated as reinforcing agents for biodegradable polyesters (Bionolle and poly(lactic acid) plasticized with 15 wt.-% of acetyltributyl citrate, p-PLLA). The composites are obtained either by high temperature compression molding fiber mats sandwiched between polymer films, or by batch mixing fibers with the molten polymer. Fibers in composites obtained by the latter method are much shorter (140-200 microm) than those of the mats (5,000 microm). Flax fibers are found to reinforce both p-PLLA and Bionolle (i.e. tensile modulus and strength increase) when composites based on fiber mats are investigated. Conversely, analogous composites obtained by batch mixing show poor mechanical properties. The observed behavior is attributed to the combined effect of fiber length and fiber-matrix adhesion. If flax fibers with a modified surface chemistry are used, the strength of short fiber composites is seen to improve significantly because the interface strengthens and load is more efficiently transferred. Appropriate surface modifications are performed by heterogeneous acylation reactions or by grafting poly(ethylene glycol) chains (PEG, molecular weight 350 and 750). The highest tensile strength of p-PLLA composites is reached when PEG-grafted flax fibers are used, whereas in the case of Bionolle the best performance is observed with acylated fibers.

Studies on the homogeneous acetylation of cellulose in the novel solvent dimethyl sulfoxide/tetrabutylammonium fluoride trihydrate.

Abstract: Summary: The novel solvent dimethyl sulfoxide (DMSO)/tetrabutylammonium fluoride trihydrate (TBAF · 3H2O) was studied for acetylation of linters cellulose. In order to control the degree of substitution (DS), acetylation of the macromolecule was carried out at different reaction time and temperature, molar ratio of reactants, as well as under variation of the concentration of TBAF · 3H2O in solution. Cellulose acetate (CA) was accessible with DS ranging from 0.43 to 2.77. The change in concentration of TBAF · 3H2O in DMSO showed a strong influence on DS. The most appropriate reaction conditions for acetylation of linters cellulose regarding maximal DS were evaluated. The structure of the CA was characterized by means of FTIR and NMR spectroscopy. The solubility of the CA depends not only on the DS but also on the reaction conditions applied, indicating a different distribution of acetate moieties both within and between polymer chains.

Release of Chlorambucil from Poly(N‐isopropylacrylamide) Hydrogels with β‐Cyclodextrin Moieties

Abstract: The present work is focused on investigating the behavior of controlled drug release poly(N-isopropylacrylamide) (PNIPA) hydrogels in the presence of beta-cyclodextrin (beta-CD). For this purpose, three types of NIPA hydrogels with beta-CD moieties were synthesized with different architectures according to our previous studies. An anti-cancer drug (chlorambucil, CLB), which can form an inclusion complex with beta-CD, was selected for loading and in vitro release studies. The drug was loaded into hydrogels via a swelling method. DSC was used to study the interactions between the CLB molecules and the polymers. The results indicate that the CLB-polymer interactions are at the molecular level. Loading CLB into these polymers can result in an evident decrease in the glass transition temperature (T(g)), and the variation of T(g) (DeltaT(g)) depends on the structures of the polymers and their beta-CD content. The controlled release experiments show that the presence of beta-CD can markedly enhance CLB release from shrunken PNIPA hydrogels and increase the ratio of CLB released in total drug loading content. Release profile of CLB from hydrogels 1a-c and 4 at pH 1.4 and 7.4, at 37 degrees C.

Biosynthesis and compositional regulation of poly[(3-hydroxybutyrate)-co-(3-hydroxyhexanoate)] in recombinant ralstonia eutropha expressing mutated polyhydroxyalkanoate synthase genes.

Abstract: A new strategy for bacterial polyhydroxyalkanoate (PHA) production by recombinant Ralstonia eutropha PHB-4 harboring mutated PHA synthase genes (phaC Ac ) from Aeromona caviae was investigated. The strain harboring wild-type phiC Ac gene produced a PHA copolymer misting of (R)-3-hydroxybutyrate and (R)-3-hydroxyhexanoate [P(3HB-co-3HHx)] with 3.5 mol-% of 3HHx fraction from soybean oil. When the mutants of phaC Ac gene were applied to this production system, 3HHx fraction in copolymers was varied in the range of 0-5.1 mol-%. Thus, the regulation of PHA copolymer compositions has been achieved by the use of mutated PHA synthase genes.

Porous-conductive chitosan scaffolds for tissue engineering, 1. Preparation and characterization.

Abstract: Novel porous-conductive chitosan scaffolds were fabricated by incorporating conductive polypyrrole (PPy) particles into a chitosan matrix and employing a phase separation technique to build pores inside the scaffolds. Conductive polypyrrole particles were prepared with a microemulsion method using FeCl 3 as a dopant. The preparation conditions were optimized to obtain scaffolds with controlled pore size and porosity. The conductivity of the scaffolds was investigated using a standard four-point probe technique. It was found that several kinds of scaffolds showed a conductivity close to 10 -3 S.cm -1 with a low polypyrrole loading of around 2 wt.-%. The main mechanical properties, such as tensile strength, breaking elongation and Young's modulus of the scaffolds, were examined both in the dry and in the hydrated states. The results indicated that a few different kinds of scaffolds exhibited the desired mechanical strength for some tissue engineering applications. The miscibility of polypyrrole and chitosan was also evaluated using a dynamic mechanical method. The presence of significant phase separation was detected in non-porous PPy/chitosan scaffolds but enhanced miscibility in porous PPy/chitosan scaffolds was observed.

Effect of Drug-loading Methods on Drug Load, Encapsulation Efficiency and Release Properties of Alginate/Poly-L-Arginine/Chitosan Ternary Complex Microcapsules

Abstract: The drug-loaded alginate/poly-L-arginine/chitosan ternary complex microcapsules were prepared by mixing method, absorption method and the combined method of mixing and absorption, respectively. The effect of drug-loading methods on drug load, the encapsulation efficiency and the release properties of the complex microcapsules were investigated. The results showed that the absorption process is a dominating factor to greatly increase the drug load of Hb into microcapsules. Upon loading Hb into microcapsules by combined method of mixing and absorption, the drug load (19.9%) is up to the maximum value, and the encapsulation efficiency is 93.8%. Moreover, the drug release is a zero-order kinetics process for the ternary complex microcapsules made by mixing. For the complex microcapsules made by absorption, the drug release is a first-order kinetics. However, for the complex microcapsules made by combining the mixing and the absorption, the drug release obeys a first-order kinetics during the first eighteen hours, changing afterwards to a zero-order kinetics process. Effect of drug-loading methods on drug load and encapsulation efficiency of alginate/poly-L-arginine/chitosan ternary complex microcapsules.

Metabolic Engineering for the Production of Copolyesters Consisting of 3-Hydroxybutyrate and 3-Hydroxyhexanoate by Aeromonas hydrophila

Abstract: Aeromonas hydrophila 4AK4 was able to synthesize copolyesters consisting of 3-hydroxybutyrate (3HB) and about 15 mol-% 3-hydroxyhexanoate (3HHx) (PHBHHx) when grown in long chain fatty acids such as dodecanoate regardless of growth conditions. To regulate the unit fraction in PHBHHx, phbA and phbB genes encoding beta-ketothiolase and acetoacetyl-CoA reductase in Ralstonia eutropha, were introduced into A. hydrophila 4AK4. When gluconate was used as cosubstrate of dodecanoate, the recombinant produced PHBHHx containing 3-12 mol-% 3HHx, depending on the gluconate concentration in media. Vitreoscilla hemoglobin gene, vgb, was also introduced into the above recombinant, resulting in improved PHBHHx content from 38 to 48 wt.-% in shake flask study. Fermentor studies also showed that increased gluconate concentration in medium containing dodecanoate promoted the recombinant strain harboring phbA and phbB genes to incorporate more 3HB unit into PHBHHx, resulting in reduced 3HHx fraction. Recombinant A. hydrophila harboring phbA, phbB and vgb genes demonstrated better PHBHHx productivity and higher conversion efficiency from dodecanoate to PHBHHx than those of the recombinant without vgb in fermentation study. Combined with the robust growth property and simple growth requirement, A. hydrophila 4AK4 appeared to be a useful organism for metabolic engineering.

Tissue engineering : key elements and some trends

Abstract: It is the believe of many that tissue engineering (or as some scientists prefer – regenerative medicine) is and will increasingly be the focus of biomaterials research, justified by a very significant and well-known clinical need for the establishment of alternative therapies for the treatment of tissue loss or end-stage organ failure, as the transplantation of tissues or organs in these patients is often limited by donor scarcity and is highly associated to the risk of rejection and disease transfer. In this sense, tissue engineering will lead to a great impact on health care providing in the coming decades. This expected evolution would also create the need for the education of new scientists that are ‘‘hybrid’’ and can perform multidisciplinary research, combining materials and biotechnology. Nevertheless, even the basics of tissue engineering are not clear for many researchers working on the general field of biomaterials and biomedical engineering. Conventional biomaterials have been very useful in the past, and have improved the life quality of many patients. Good examples are many different prosthesis, such as for instances the knee and hip joints. However it is easy to recognize that there are still no materials available that can adequately replace several functional tissues, such as bones, cartilage, or large bone segments, not to speak on complex organs. Just as an example, in the world market for bone grafts the so-called synthetic biomaterials represent only 10%, while autografts still account for around 50%. Therefore, despite the enormous benefits the contemporary technology has brought, the outer limits have been reached and new breakthroughs can only be expected from a novel hybrid technology that will reduce the shortcomings of the current material technology. Such a combined, biology driven approach is referred to as ‘‘tissue engineering’’, by which biological tissues are engineered through combining material technology and biotechnology. Tissue engineering typically involves the culture of living human cells usually in polymeric (ceramic) scaffold materials, ex vivo, and subsequently allowing them to develop into a three dimensional tissue. Substantial gains are expected to be obtained both from a medical and economic standpoint as a result of this emerging technology. It is expected that, in the near future, tissue engineering can take advantage of the recent breakthroughs in the fields of stem cell research, genomics and materials technology. Tissue engineering involves several steps, that go from (i) the selection, isolation, and culturing of primary (progenitor or stem from different origins) cells, inducing their differentiation to specific phenotypes, (ii) to the way they are cultured (not pre-cultured and just seeded and immediately implanted, or cultured in static conditions or in an all range of different specifically designed bio-reactors), (iii) to the design of adequate scaffolds, including the selection of adequate materials and routes to process them, the respective porosity, interconnectivity, surface characteristics, hydrophilicity, etc., and (iv) to the use of adequate animal models that will allow to test the efficacy of different tissue engineering approaches and the potential of different constructs (distinct combinations of scaffolds/cells/in vitro culturing conditions). Finally, all the related ethical considerations, especially when considering the use of stem cells (and potentially embryonic stem cells) and animal studies, as well as the need for proper standards must be taken into account, and unfortunately tend to differ from country to country. This introductory essay will very briefly present the stateof-the-art on topics such as: cells for tissue engineering (stem cells, isolation, characterization, etc.), culturing conditions (static, media, bioreactors, etc.), scaffolds (design, processing, choice of materials, etc.) and needed animal models to test the developed tissue engineering strategies and respective constructs. The Special Topic on Tissue Engineering, that Macromolecular Bioscience invited me to organize as a Guest Editor, comprises a selection of papers that touch several relevant and emerging topics on tissue engineering, providing the reader with a feeling on what is the present status of the field.

Hydrogel Formation between Enantiomeric B‐A‐B‐Type Block Copolymers of Polylactides (PLLA or PDLA: A) and Polyoxyethylene (PEG: B); PEG‐PLLA‐PEG and PEG‐PDLA‐PEG

Abstract: A mixed suspension of the enantiomeric B-A-B triblock copolymers, polyoxyethylene-block-poly(L-lactide)-block-polyoxyethylene (PEG-PLLA-PEG) and polyoxyethylene-block-poly(D-lactide)-block-polyoxyethylene (PEG-PDLA-PEG), was found to induce reversible gel-to-sol transition depending on the polymer concentration and temperature. The storage and loss moduli of the gel formed at lower temperature were much higher than those of the gel prepared from the corresponding ABA-type triblock copolymers because of the higher polymer concentration in the former. Although the stereo-complexation of the PLLA and PDLA blocks occurred at higher temperature also in the B-A-B copolymers, it was not responsible for the gelation of the mixed suspension. The PEG chains, involved in the helix formation of the PLLA and PDLA, should form helices with opposite helical senses to aggregate and lead the gelation of the system.

Synthesis and characterization of polymeric linseed oil grafted methyl methacrylate or styrene

Abstract: Syntheses of wholly natural polymeric linseed oil (PLO) containing peroxide groups have been reported. Peroxidation, epoxidation and/or perepoxidation reactions of linseed oil, either under air or under oxygen flow at room temperature, resulted in polymeric peroxides, PLO-air and PLO-ofl, containing 1.3 and 3.5 wt.-% of peroxide, with molecular weights of 2 100 and 3 780 Da, respectively. PLO-air contained cross-linked film up to 46.1 wt.-% after a reaction time of 60 d, associated with a waxy, soluble part (PLO-air-s) that was isolated with chloroform extraction. PLO-ofl was obtained as a waxy, viscous liquid without any cross-linked part at the end of 24 d under visible irradiation and oxygen flow. Polymeric peroxides, PLO-air-s and PLO-ofl initiated the free radical polymerization of both methyl methacrylate (MMA) and styrene (S) to give PMMA-graft-PLO and PS-graft-PLO graft copolymers in high yields with Mw varying from 37 to 470 kDa. The polymers obtained were characterized by FT-IR, (1)H NMR, TGA, DSC and GPC techniques. Cross-linked polymers were also studied by means of swelling measurements. PMMA-graft-PLO graft copolymer film samples were also used in cell-culture studies. Fibroblast cells were well adhered and proliferated on the copolymer film surfaces, which is important in tissue engineering.