Hyper-branched poly(poly(ethylene glycol)methacrylate)-grafted surfaces by photo-polymerization with iniferter for bioactive interfaces

A new hyper-branched surface in which three species of architectures were constructed as stem chain, branched stem and twig chain-grafted branched chain of poly(poly(ethylene glycol)methacrylate) (poly(PEGMA)) by photo-polymerization using dithiocarbamyl group (DC) as iniferter was prepared and characterized. For these surfaces, radical copolymerization of styrene and an iniferter-activated chain that was previously synthesized was performed for using as base materials for surface coating. On a DC-activated surface, hyper-branched poly(PEGMA) was introduced by photo-polymerization and dithiocarbamylation. All modified surfaces were analyzed by X-ray photoelectron spectroscopy (XPS) and water contact angle measurements. Our results demonstrated that a highly hyper-branched graft architecture of poly(PEGMA) can be constructed on PU surface by photo-polymerization using dithiocarbamyl group as iniferter, in which first, second and third generation gave stem chain, branched chain and twig chain of poly(PEGMA), respectively. Our hyper-branched surfaces could be regulated by photo-irradiation time and might be controlled by feed amounts or other reaction conditions. This highly dense architecture of PEG chain with hydrophilicity and chain mobility, grafted on surface, is expected to be effectively utilized in bio-implantable substrates or micro- or nano-patterned surfaces for immobilization of bioactive molecules in biomedical fields.     

Functionalization of poly(ethylene glycol) and monomethoxy-poly(ethylene glycol)

Simple methods are described for the substitution of poly(ethylene glycol) and monomethoxy-poly(ethylene glycol) substitution. Affinity ligands, coenzymes, or enzymes can be covalently attached to the substitution product or they can be used as liquid ionexchangers.     

Pulsatile peptide release from multi-layered hydrogel formulations consisting of poly(ethylene glycol)-grafted and ungrafted dextrans

Multi-layered hydrogel formulations consisting of poly(ethylene glycol)-grafted dextran (PEG-g-Dex) and ungrafted Dex were investigated as a model of pulsatile drug release. In these formulations, it is considered that the grafted PEG domains act as a drug reservoir dispersed in the Dex matrix based on aqueous polymer two-phase systems. The formulations exhibited surface-controlled degradation by dextranase, and insulin release was observed in a pulsatile manner because of the multi-layered structure: PEG-g-Dex hydrogel layers containing insulin and insulin-free Dex hydrogel layers. Thus, it is suggested that the multi-layered hydrogel formulations using PEG-g-Dex and Dex are feasible for chronopharmacological drug delivery systems.     

Pulsatile peptide release from multi-layered hydrogel formulations consisting of poly(ethylene glycol)-grafted and ungrafted dextrans

Multi-layered hydrogel formulations consisting of poly(ethylene glycol)-grafted dextran (PEG-g-Dex) and ungrafted Dex were investigated as a model of Pulsatile drug release. In these formulations, it is considered that the grafted PEG domains act as a drug reservoir dispersed in the Dex matrix based on aqueous polymer two-phase systems. The formulations exhibited surface-controlled degradation by dextranase, and insulin release was observed in a pulsatile manner because of the multi-layered structure, PEG-g-Dex hydrogel layers containing insulin and insulin-free Dex hydrogel layers. Thus, it is suggested that the multi-layered hydrogel formulations using PEG-g-Dex and Dex are feasible for chronopharmacological drug delivery systems.     

Preparation and characterization of ibuprofen-loaded poly(lactide-co-glycolide)/poly(ethylene glycol)-g-chitosan electrospun membranes

Ibuprofen-loaded composite membranes composed of poly(lactide-co-glycolide) (PLGA) and poly(ethylene glycol)-g-chitosan (PEG-g-CHN) were prepared by electrospinning. The electrospun membranes were characterized by scanning electron microscopy (SEM), differential scanning calorimetry (DSC), mechanical evaluation and contact angle measurements. Shrinkage behavior of the membrane in buffer at 37 degrees C was also evaluated. It was found that PLGA glass transition temperature (Tg) decreased with increasing PEG-g-CHN content in the composite membranes, which results in a decrease in tensile stress at break but an increase in tensile strain of the membranes. The degree of shrinkage of these composite membranes decreased from 76 to only 3% when the PEG-g-CHN content in the membranes increased from 10 to 30%. The presence of PEG-g-CHN significantly moderated the burst release rate of ibuprofen from the electrospun PLGA membranes. Moreover, ibuprofen could be conjugated to the side chains of PEG-g-CHN to prolong its release for more than two weeks. The sustained release capacity of the PLGA/PEG-g-CHN composite membranes, together with their compliant and stable mechanical properties, renders them ideal matrices for atrial fibrillation.     

Branched poly(ethylene glycol) linkers

Novel types of methoxy poly(ethylene glycol) (PEG) linkers (U-PEG linkers) have been synthesized. These PEG linkers are linear polymers that attach to bioactive agents via a functional group, derived from a 2° alcohol, located in the center of the polymer chain versus the traditional terminal attachment site. These new types of linkers can be prepared with different functional groups (e.g. active ester, succinimidyl carbonate, and carbazate) for selected point of attachment, including ethylene oxide oligomers to provide “stems” when steric factors need to be addressed. Conversion of p-nitrophenyl carbonates to the more desirable succinimidyl carbonates has also been accomplished by a novel nucleophilic displacement procedure. Modification of proteins with these reagents is easily accomplished and is illustrated by the conjugation of a U-PEG linker with L -asparaginase.     

In situ fabrication of self-transformable and hydrophilic poly(ethylene glycol) derivative-modified polysulfone membranes

A self-transformable sulfonated poly(ethylene glycol) acrylate diblock copolymer (PEG-SO3A/OA) entrapped into polysulfone membrane was studied. The asymmetric membrane structure was prepared by a phase inversion process. The induced hydrophilicity by reorientation of diblock copolymer at the interface was evaluated by contact angle measurement, platelet adhesion test, and electron spectroscopy for chemical analysis (ESCA) depth profiling with ion sputtering. Molecular dynamic (MD) simulations as a function of copolymer density were also performed to obtain optimum interfacial structure information. The dependency of water clustering behavior as a hydrophilicity parameter was described in terms of an atom-atom radial distribution function (RDF). The results showed that the sulfonated diblock copolymer enhanced the hydrophilicity and long-term stability more than the copolymer having no hydrophobic block. Also, according to the ESCA, oxygen composition significantly began to decrease along the membrane depth, indicating the reorientation of diblock chains. The copolymer-entrapped surfaces significantly induced the degree of water clustering, and the resulting equilibrium rearrangement of interfacial structure was distinctly dependent upon the density of the copolymer.     

Physicochemical and biological properties of poly(ethylene glycol)-coupled immunoglobulin G. Part II. Effect of molecular weight of poly(ethylene glycol)

In order to obtain a stable human immunoglobulin G (IgG) preparation for clinical use, the chemical coupling of different molecular weights of poly(ethylene glycol)s (PEGs) to IgG molecules was achieved. The abilities of PEG-coupled IgGs (PEG-IgG hybrids) to aggregate were examined when they were subjected to such physicochemical treatments as interfacial exposure, heating, lyophilization, and acid treatment. It was found that the higher the molecular weight of PEG coupled, the more stable was the PEG-IgG hybrid obtained concerning interfacial exposure and heating. The hybrid was stable against lyophilization and acid treatment and its stability was independent of the PEG molecular weight. The decrease in antigen binding ability was suppressed as much as possible by the use of a small amount of PEG of higher molecular weight. The PEG-IgG hybrids were further assessed as a stabilizer for IgG. A limited degree of PEG coupling was required for the hybrids to achieve the most efficient stabilization of IgG: the optimal PEG contents of the hybrid were > 20 wt% (interfacial exposure), about 5'wt,?o (heating), 20 wt% (lyophilization), and 10 wt% (acid treatment) for PEG 5600. It was also confirmed that the PEG-IgG hybrid was superior to PEG and human serum albumin as a stabilizer.     

Thermoresponsive degradable poly(ethylene glycol) analogues

Thermoresponsive polymers have many biomedical applications, but their nondegradability limits their in vivo applications. Herein, we report a new type of degradable thermoresponsive polymers-degradable poly (ethylene glycol) analogues (DPEGs) having lower critical solution temperatures (LCSTs) ranging 10-50 degrees C. DPEGs were synthesized by condensation polymerization of PEG-di(meth)acrylates (PEGDA or PEGDMA) with dithiols. Their LCSTs could be easily tuned by the PEG-chain length and the types of the double bond in the PEG monomers and dithiols. Long PEG chain and the presence of hydrophilic groups in the dithiol monomer increased the LCST of the resulting DPEG. Crosslinking DPEG chains produced thermoresponsive hydrogels. The hydrogels prepared by the end-capping method maintained the thermoresponsive properties of the linear DPEG. The degradable thermoresponsive DPEGs and their hydrogels have great potentials for in vivo biomedical applications.     

Tethering poly(ethylene glycol)s to improve the surface biocompatibility of poly(acrylonitrile-co-maleic acid) asymmetric membranes

To improve the surface biocompatibility, asymmetric membranes fabricated from poly(acrylonitrile-co-maleic acid)s (PANCMAs) synthesized by water-phase precipitation copolymerization were tethered (or immobilized) with poly(ethylene glycol)s (PEGs) by esterification reaction. Chemical changes on the membrane surface were characterized by Fourier transform infrared spectroscopy and elemental analysis to confirm the immobilization of PEG onto the PANCMA membranes. The hydrophilicity and blood compatibility of the PEG-tethered PANCMA membrane were investigated by water contact angle, water absorption, protein adsorption, plasma platelets adhesion and cell adhesion measurements, and the results were compared with the corresponding PANCMA membranes. It was found that, after the tethering of PEG, the hydrophilicity of the membrane can be improved significantly, and the protein adsorption, platelets adhesion and macrophage attachment on the membrane surface are obviously suppressed. Furthermore, not only the content of maleic acid in PANCMA, which influences the tethering density of PEG, but also the molecular weight of PEG has great effect on the surface modification of PANCMA membranes for biocompatibility.     

Characterizing the modification of surface proteins with poly(ethylene glycol) to interrupt platelet adhesion

Surface protein modification with poly(ethylene glycol) (PEG) can inhibit acute thrombosis on damaged vascular and biomaterial surfaces by blocking surface protein-platelet interactions. However, the feasibility of employing protein reactive PEGs to limit intravascular and biomaterial thrombosis in vivo is contingent upon rapid and extensive surface protein modification. To characterize the factors controlling this potential therapeutic approach, the model protein bovine serum albumin was adsorbed onto polyurethane surfaces and modified with PEG-carboxymethyl succinimidyl ester (PEG-NHS), PEG-isocyanate (PEG-ISO), or PEG-diisocyanate (PEG-DISO) in aqueous buffer at varying concentrations and contact times. It was found that up to 5 PEGs could be attached per albumin molecule within one min and that adsorbed albumin PEGylation approached maximal levels by 6min. The lability of reactive PEGs in aqueous buffer reduced total protein modification by 50% when the PEG solution was incubated for 7min prior to application. For fibrinogen PEGylation (performed in the solution phase), PEG-NHS was more reactive than PEG-ISO or PEG-DISO. The gamma peptide of fibrinogen, which contains several key platelet-binding motifs, was highly modified. A marked reduction in platelet adhesion was observed on fibrinogen-adsorbed polyurethane treated with PEG-NHS or PEG-DISO. Relative differences in platelet adhesion on PEG-NHS and PEG-DISO modified surfaces could be attributed to differences in reactivity towards fibrinogen and the size of the polymer backbone. Taken together, these findings provide insight and guidance for applying protein reactive PEGs for the interruption of acute thrombotic deposition.     

Physicochemical and biological properties of poly(ethylene glycol)-coupled immunoglobulin G. Part II. Effect of molecular weight of poly(ethylene glycol)

In order to obtain a stable human immunoglobulin G (IgG) preparation for clinical use, the chemical coupling of different molecular weights of poly(ethylene glycol)s (PEGs) to IgG molecules was achieved. The abilities of PEG-coupled IgGs (PEG-IgG hybrids) to aggregate were examined when they were subjected to such physicochemical treatments as interfacial exposure, heating, lyophilization, and acid treatment. It was found that the higher the molecular weight of PEG coupled, the more stable was the PEG-IgG hybrid obtained concerning interfacial exposure and heating. The hybrid was stable against lyophilization and acid treatment and its stability was independent of the PEG molecular weight. The decrease in antigen binding ability was suppressed as much as possible by the use of a small amount of PEG of higher molecular weight. The PEG-IgG hybrids were further assessed as a stabilizer for IgG. A limited degree of PEG coupling was required for the hybrids to achieve the most efficient stabilization of IgG; the optimal PEG contents of the hybrid were greater than 20 wt% (interfacial exposure), about 5 wt% (heating), 20 wt% (lyophilization), and 10 wt% (acid treatment) for PEG 5600. It was also confirmed that the PEG-IgG hybrid was superior to PEG and human serum albumin as a stabilizer.     

Physical characterization of thin semi-porous poly(L-lactic acid)/poly(ethylene glycol) membranes for tissue engineering

This study examines physical properties of solvent-cast poly(L-lactic acid) (PLLA): poly(ethylene glycol) PEG membranes as a function of PEG molecular weight (MW) and incubation in vitro for 6 weeks. PEGs of MW 400, 1450 and 8000 were used. The morphological, thermal, mechanical and permeability properties of the membranes were studied prior to and after 3 and 6 weeks of incubation in phosphate-buffered saline (PBS) at 37 degrees C. The membranes showed a thickness of about 35+/-5 microm and were found to be semi-porous, with a non-porous surface as well as a porous surface with pore-diameters of 0.5-5 microm. The surface pore size was found to be a function of PEG MW used. All membranes were mechanically strong, with elastic moduli and tensile strength of 150-440 MPa and 7-36 MPa, respectively, all through the 6-week incubation period. The lower-MW PEGs plasticized PLLA based on high initial percent elongation; however, the effect was lost after 3 weeks of incubation in PBS. All membranes except those fabricated with PEG 8000 were impermeable for up to 6 weeks of incubation in PBS. Permeability studies showed that only PLLA:PEG 8000 membranes were permeable to methylene blue after 3 weeks of degradation.     

Biodegradable and thermoreversible hydrogels of poly(ethylene glycol)-poly(epsilon-caprolactone-co-glycolide)-poly(ethylene glycol) aqueous solutions

The aqueous solutions of triblock copolymers of poly(ethylene glycol)-poly(epsilon-caprolactone-co-glycolide)-poly(ethylene glycol) [PEG-P(CL-GA)-PEG] undergoing sol-gel transition as the temperature increases from 20 to 60 degrees C were successfully prepared. The thermogelling block copolymers were synthesized by subtle control of the hydrophilic/hydrophobic balance and the chain microstructures. The amphiphilic block copolymer formed micelles in aqueous solution, and the micelle aggregated as the temperature increased. The sol-gel transition of the copolymer aqueous solutions was studied focusing on the structure-property relationship. GA was incorporated into the polymer chain to prevent crystallization of PCL component and increase the polymer degradation. It is expected to be a promising long-term delivery system for pH-sensitive drugs, proteins, and genes.     

Synthesis and polymerization of poly(ethylene glycol) fumarates

Fumaric esters of poly(ethylene glycol) ( 1a–d ) were prepared as macromonomers. The halfesters 1a and 1c were obtained by interaction of maleic anhydride with the monoethers of poly(ethylene glycol) ( 2a and 2b ) in the presence of 4-dimethylaminopyridine, and the diesters 1b and 1d by esterification of methyl hydrogen fumarate ( 13 ) with the monoethers of poly(ethylene glycol) ( 2a and 2b ). The macromonomers were found to homopolymerize and to copolymerize with styrene and methyl methacrylate following a radical polymerization mechanism.     

Dynamic behavior of glucose oxidase-containing microparticles of poly(ethylene glycol)-grafted cationic hydrogels in an environment of changing pH

Poly(diethylaminoethyl-g-ethylene glycol) microparticles were prepared by suspension polymerization of diethylaminoethyl methacrylate, poly(ethylene glycol) monomethacrylate and the crosslinking agent tetra(ethylene glycol) dimethacrylate in silicone oil using redox initiators. Particles of different sizes, crosslinking ratios and graft molecular weights were prepared. The changes in the swelling of the particles were studied as the pH was changed between 3.0 and 7.4. The particles showed rapid swelling/deswelling dynamics in response to changes in pH. It was evident that faster response could be obtained from smaller particles. Changing the crosslinking ratio resulted in changes in the extent of swelling, as well as the speed of response. It was also found that longer graft lengths were responsible for increasing the effect of relaxation of the swelling of the network.     

Synthesis and water-swelling of thermo-responsive poly(ester urethane)s containing poly(epsilon-caprolactone), poly(ethylene glycol) and poly(propylene glycol)

Thermo-responsive multiblock poly(ester urethane)s comprising poly(epsilon-caprolactone) (PCL), poly(ethylene glycol) (PEG), and poly(propylene glycol) (PPG) segments were synthesized. The copolymers were characterized by GPC, NMR, FTIR, XRD, DSC and TGA. Water-swelling analysis carried out at different temperatures revealed that the bulk hydrophilicity of the copolymers could be controlled either by adjusting the composition of the copolymer or by changing the temperature of the environment. These thermo-responsive copolymer films formed highly swollen hydrogel-like materials when soaked in cold water and shrank when soaked in warm water. The changes are reversible. The mechanical properties of the copolymer films were assessed by tensile strength measurement. These copolymers were ductile when compared to PCL homopolymers. Young's modulus and the stress at break increased with increasing PCL content, whereas the strain at break increased with increasing PEG content. The results of the cytotoxicity tests based on the ISO 10993-5 protocol demonstrated that the copolymers were non-cytotoxic and could be potentially used in biomedical applications.     

Preparation of poly(ethylene glycol) grafted poly(3-hydroxyalkanoate) networks

Poly(3-hydroxyalkanoate)-g-poly(ethylene glycol) crosslinked graft copolymers are described. Poly(3-hydroxyalkanoate)s containing double bonds in the side chain (PHA-DB) were obtained by co-feeding Pseudomonas oleovorans with a mixture of nonanoic acid and anchovy (hamci) oily acid (in weight ratios of 50/50 and 70/30). PHA-DB was thermally grafted with a polyazoester synthesized by the reaction of poly(ethylene glycol) with MW of 400 (PEG-400) and 4,4′-azobis(4-cyanopentanoyl chloride). Sol-gel analysis and spectrometric and thermal characterization of the networks are reported.     

Effect of poly(ethylene glycol) molecular weight on tensile and swelling properties of oligo(poly(ethylene glycol) fumarate) hydrogels for cartilage tissue engineering.

This study was designed to determine the effect of changes in poly(ethylene glycol) (PEG) molecular weight on swelling and mechanical properties of hydrogels made from a novel polymer, oligo(poly(ethylene glycol) fumarate) (OPF), recently developed in our laboratory. Properties of hydrogels made from OPF with initial PEG molecular weights of 860, 3900, and 9300 were examined. The PEG 3900 formulation had a tensile modulus of 23.1 +/- 12.4 kPa and percent elongation at fracture of 53.2 +/- 13.7%; the PEG 9300 formulation had similar tensile properties (modulus: 16.5 +/- 4.6 kPa, elongation: 76.0 +/- 26.4%). However, the PEG 860 gels had a significantly higher modulus (89.5 +/- 50.7 kPa) and a significantly smaller percent elongation at fracture (30.1 +/- 6.4%), when compared with other formulations. Additionally, there were significant differences in percent swelling between each of the formulations. Molecular weight between crosslinks (M(c)) and mesh size were calculated for each OPF formulation. M(c) increased from 2010 +/- 116 g/mol with PEG 860 to 6250 +/- 280 g/mol with PEG 9300. Mesh size calculations showed a similar trend (76 +/- 2 A for PEG 860 to 160 +/- 6 A for PEG 9300). It was also found that these hydrogels could be laminated if a second layer was added before the first had completely crosslinked. Mechanical testing of these laminated gels revealed that the presence of an interfacial area did not significantly alter their tensile properties. These results suggest that the material properties of OPF-based hydrogels can be altered by changing the molecular weight of PEG used in synthesis and that multilayered OPF hydrogel constructs can be produced, with each layer having distinct mechanical properties.     

Degradative properties and cytocompatibility of a mixed-mode hydrogel containing oligo[poly(ethylene glycol)fumarate] and poly(ethylene glycol)dithiol

Our laboratory is currently exploring synthetic oligo(poly(ethylene glycol)fumarate) (OPF)-based biomaterials as a means to deliver fibroblasts to promote regeneration of central/partial defects in tendons and ligaments. In order to further modulate the swelling and degradative characteristics of OPF-based hydrogels, OPF crosslinking via a radically initiated, mixed-mode reaction involving poly(ethylene glycol) (PEG)-diacrylate and PEG-dithiol was investigated. Results demonstrate that mixed-mode hydrogels containing OPF can be formed and that the presence of 20 wt.% PEG-dithiol increases swelling and decreases degradation time vs. 10 wt.% PEG-dithiol and non-thiol-containing hydrogels (20% thiol fold swelling 28.7+/-0.8; 10% thiol fold swelling 11.6+/-1.4; non-thiol 8.7+/-0.2; 20% thiol-containing hydrogels degrade within 15 days in vitro). After encapsulation, tendon/ligament fibroblasts remained largely viable over 8 days of static culture. While the presence of PEG-dithiol did not significantly affect cellularity or collagen production within the constructs over this time period, image analysis revealed that the 20% PEG-dithiol gels did appear to promote cell clustering, with greater values for aggregate area observed by day 8. These experiments suggest that mixed-mode OPF-based hydrogels may provide an interesting alternative as a cell carrier for engineering a variety of soft orthopedic tissues, particularly for applications when it is important to encourage cell-cell contact.