Controlled release of heparin from poly(epsilon-caprolactone) electrospun fibers

Sustained delivery of heparin to the localized adventitial surface of grafted blood vessels has been shown to prevent the vascular smooth muscle cell (VSMC) proliferation that can lead to graft occlusion and failure. In this study heparin was incorporated into electrospun poly(epsilon-caprolactone) (PCL) fiber mats for assessment as a controlled delivery device. Fibers with smooth surfaces and no bead defects could be spun from polymer solutions with 8% w/v PCL in 7:3 dichloromethane: methanol. A significant decrease in fiber diameter was observed with increasing heparin concentration. Assessment of drug loading, and imaging of fluorescently labeled heparin showed homogenous distribution of heparin throughout the fiber mats. A total of approximately half of the encapsulated heparin was released by diffusional control from the heparin/PCL fibers after 14 days. The fibers did not induce an inflammatory response in macrophage cells in vitro and the released heparin was effective in preventing the proliferation of VSMCs in culture. These results suggest that electrospun PCL fibers are a promising candidate for delivery of heparin to the site of vascular injury. (C) 2005 Elsevier Ltd. All rights reserved.

Preparation of poly (alkylcyanoacrylate) nanoparticles by polymerization of water-free microemulsions

Phase diagrams of the pseudoternary systems ethyloleate, polyoxyethylene 20 sorbitan mono-oleate/sorbitan monolaurate and propylene glycol with and without butanol as a co-surfactant were prepared. Areas containing optically isotropic, one-phase systems were identified and samples therein designated as droplet, bicontinuous or solution type microemulsions using conductivity, viscosity and self-diffusion NMR. Nanoparticles were prepared by polymerization of selected microemulsions with ethyl-2-cyanoacrylate and the morphology of the particles was investigated. Addition of monomer to all types of microemulsions led to the formation of nanoparticles, which had an average size of 244 +/- 25 nm, an average polydispersity index of 0.15 +/- 0.04 and a zeta-potential of -17 +/- 3 mV. The formation of particles from water-free microemulsions of different types is surprising, particularly considering that polymerization is expected to occur at a water-oil interface by base-catalysed polymerization. It would appear that propylene glycol is sufficiently nucleophilic to initiate the polymerization. The use of water-free microemulsions as templates for the preparation of poly (alkylcyanoacrylate) nanoparticles opens up interesting opportunities for the encapsulation of bioactives which do not have suitable properties for encapsulation on the basis of water-containing microemulsions.

Solution properties of star and linear poly(N-isopropylacrylamide)

The LCST transitions of novel N-isopropylacrylamide ( NIPAM) star polymers, prepared using the four-armed RAFT agent pentaerythritoltetrakis(3-(S-benzyltrithiocarbonyl) propionate) (PTBTP) and their hydrolyzed linear arms were studied using H-1 NMR, PFG-NMR, and DLS. The aim was to determine the effect of polymer architecture and the presence of end groups derived from RAFT agents on the LCST. The LCST transitions of star PNIPAM were significantly depressed by the presence of the hydrophobic star core and possibly the benzyl end groups. The effect was molecular weight dependent and diminished once the number of repeating units per arm >= 70. The linear PNIPAM exhibited an LCST of 35 degrees C, regardless of molecular weight; the presence of both hydrophilic and hydrophobic end groups after hydrolysis from the star core was suggested to cancel effects on the LCST. A significant decrease in R-H was observed below the LCST for star and linear PNIPAM and was attributed to the formation of n-clusters. Application of a scaling law to the linear PNIPAM data indicated the cluster size n = 6. Tethering to the hydrophobic star core appeared to inhibit n-cluster formation in the lowest molecular weight stars; this may be due to enhanced stretching of the polymer chains, or the presence of larger numbers of n-clusters at temperatures below those measured.

Design and characterisation of novel blends of poly(lactic acid)

This thesis is concerned with the effect of polymer structure on miscibility of the three component blends based on poly(lactic acid) (PLA) with using blending techniques. The examination of novel PLA homologues (pre-synthesised poly(a-esters)), including a range of aliphatic and aromatic poly(a-esters) is an important aspect of the work. Because of their structural simplicity and similarity to PLA, they provide an ideal system to study the effect of polyester structures on the miscibility of PLA polymer blends. The miscibility behaviour of the PLA homologues is compared with other aliphatic polyesters (e.g. poly(e-caprolactone) (PCL), poly(hydroxybutyrate hydroxyvalerate) (P(HB-HV)), together with a series of cellulose-based polymers (e.g. cellulose acetate butyrate (CAB)). The work started with the exploration the technique used for preliminary observation of the miscibility of blends referred to as “a rapid screening method” and then the miscibility of binary blends was observed and characterised by percent transmittance together with the Coleman and Painter miscibility approach. However, it was observed that symmetrical structures (e.g. a1(dimethyl), a2(diethyl)) promote the well-packing which restrict their chains from intermingling into poly(L-lactide) (PLLA) chains and leads the blends to be immiscible, whereas, asymmetrical structures (e.g. a4(cyclohexyl)) behave to the contrary. a6(chloromethyl-methyl) should interact well with PLLA because of the polar group of chloride to form interactions, but it does not. It is difficult to disrupt the helical structure of PLLA. PLA were immiscible with PCL, P(HB-HV), or compatibiliser (e.g. G40, LLA-co-PCL), but miscible with CAB which is a hydrogen-bonded polymer. However, these binary blends provided a useful indication for the exploration the novel three component blends. In summary, the miscibility of the three-component blends are miscible even if only two polymers are miscible. This is the benefit for doing the three components blend in this thesis, which is not an attempt to produce a theoretical explanation for the miscibility of three components blend system.

Degradable poly(ethylene glycol)-based hydrogels:synthesis, physico-chemical properties and in vitro characterization

Designing degradable hydrogels is complicated by the structural and temporal complexities of the gel and evolving tissue. A major challenge is to create scaffolds with sufficient mechanical properties to restore initial function while simultaneously controlling temporal changes in the gel structure to facilitate tissue formation. Poly(ethylene glycol) was used in this work, to form biodegradable poly(ethylene glycol)-based hydrogels with hydrolyzable poly-l-lactide segments in the backbone. Non-degradable poly(ethylene glycol) was also introduced in the formulation to obtain control of the degradation profile that encompasses cell growth and new tissue formation. The dependence on polymer composition was observed by higher degradation profiles and decreased mechanical properties as the content of degradable segments was increased in the formulation. Based on in vitro tests, no toxicity of extracts or biomaterial in direct contact with human adipose tissue stem cells was observed, and the ultraviolet light treatment did not affect the proliferation capacity of the cells.

Bile-salt-induced aggregation of poly(N-isopropylacrylamide) and lowering of the lower critical solution temperature in aqueous solutions

The effect of sodium cholate (NaC; concentration 1-16 mM), a biological surfactant, on the aggregation behavior of 1% (w/v, 2.2 × 10(-3) M) poly(N-isopropylacrylamide) (PNIPAM) aqueous solutions was studied as a function of temperature. From turbidity, dynamic light scattering, viscosity, and fluorescence measurements, it was observed that (i) there is NaC-induced nanoscale aggregation of PNIPAM in its sol state and (ii) the lower critical solution temperature corresponding to sol-gel transition shifts to a lower temperature by about 2 °C.

Cleavage of caspases-1, -3, -6, -8 and -9 substrates by proteases in skeletal muscles from mice undergoing cancer cachexia

A prominent feature of several type of cancer is cachexia. This syndrome causes a marked loss of lean body mass and muscle wasting, and appears to be mediated by cytokines and tumour products. There are several proteases and proteolytic pathways that could be responsible for the protein breakdown. In the present study, we investigated whether caspases are involved in the proteolytic process of skeletal muscle catabolism observed in a murine model of cancer cachexia (MAC16), in comparison with a related tumour (MAC13), which does not induce cachexia. Using specific peptide substrates, there was an increase of 54% in the proteolytic activity of caspase-1, 84% of caspase-8, 98% of caspase-3 151% to caspase-6 and 177% of caspase-9, in the gastrocnemius muscle of animals bearing the MAC16 tumour (up to 25% weight loss), in relation to muscle from animals bearing the MAC13 tumour (1-5% weight loss). The dual pattern of 89 kDa and 25 kDa fragmentation of poly (ADP-ribose) polymerase (PARP) occurred in the muscle samples from animals bearing the MAC16 tumour and with a high amount of caspase-like activity. Cytochrome c was present in the cytosolic fractions of gastrocnemius muscles from both groups of animals, suggesting that cytochrome c release from mitochondria may be involved in caspase activation. There was no evidence for DNA fragmentation into a nucleosomal ladder typical of apoptosis in the muscles of either group of mice. This data supports a role for caspases in the catabolic events in muscle involved in the cancer cachexia syndrome. © 2001 Cancer Research Campaign.

Dipole moments and Kerr effect of poly(n-vinylcarbazole) and its complexes with trinitrofluorenone

N-vinylcarbazole was polymerised using the free radical catalyst (azo-bisisobutyronitrile) and cationic catalysts (boron-trifluoride etherate and aluminium chloride). The polymers produced were characterised by molecular weight measurements and powder x-ray diffraction. The tacticity of the polymer samples was determined using proton and carbon-13 nuclear magnetic resonance spectroscopy. Measurements of their static dielectric permittivity and electro-optical birefringence (Kerr effect) in solution in 1,4-dioxane were carried out over a range of temperatures. The magnitudes of the dipole moments and Kerr constants were found to vary with changes in the tacticity of poly(N-vinylcarbazole). The results of these measurements support the view that the stereostructure of poly(N-vinylcarbazole) is sensitive to the mechanism of polymerisation. These results, together with proton and carbon-13 N.M.R. data, are discussed in terms of the possible conformations of the polymer chains and the relative orientation of the bulky carbazole side groups. The dielectric and molecular Kerr effect studies have also been carried out on complexes formed between 2,4,7-trinitro-9-fluorenone (TNF) and different stereoregular forms of poly(N-vinylcarbazole) in solution in 1,4-dioxane. The differences in the molar Kerr constants between pure (uncomplexed) and complexed poly(N-vinylcarbazole) samples were attributed to changes in optical anisotropy and dipole moments. A molecular modelling computer program Desktop Molecular Modeller was used to examine the 3/1 helical isotactic and 2/1 helical syndiotactic forms of poly(N-vinylcarbazole). These models were used to calculate the pitch distances of helices and the results were interpreted in terms of van der Waal's radii on TNF. This study indicated that the pitch distance in 3/1 isotactic helices was large enough to accommodate the bulky TNF molecules to form sandwich type charge transfer complexes whereas the pitch distance in syndiotactic poly(N-vinylcarbazole) was smaller and would not allow a similar type of complex formation.

Poly (D,L-Lactide) based microspheres for pulmonary delivery of proteins

Initial work focused on the preparation, optimisation and characterisation of poly (D,L-lactide) (PLA) microspheres with the aim of optimising their formulation based on minimizing the particle size into the range suitable for pulmonary delivery to alveoli. In order to produce dry powders and to enhance their long-term physico-chemical stability, microspheres were prepared as a dry powder via freeze-drying. Optimisation studies showed that using appropriate concentrations of polymer 3% (w/v) in organic phase and emulsifier 10% (w/v) in external aqueous phase, the double solvent evaporation method produced high protein loading microspheres (72 ± 0.5%) with an appropriate particle size for pulmonary drug delivery. Combined use of trehalose and leucine as cyroprotectants (6% and 1% respectively, w/v) produced freeze-dried powders with the best aerosolisation profile among those tested. Although the freeze-dried PLA microsphere powders were not particularly respirable in dry powder inhalation, nebulisation of the rehydrated powders using an ultrasonic nebuliser resulted in improved aerosilisation performance compared to the air-jet nebuliser. When tested in vitro using a macrophage cell line, the PLA microspheres system exhibited a low cytotoxicity and the microspheres induced phagocytic activity in macrophages. However, interestingly, the addition of an immunomodulator to the microsphere formulations (4%, w/w of polymer) reduced this phagocytic activity and macrophage activation compared to microspheres formulated using PLA alone. This suggested that the addition of trehalose dibehenate may not enhance the ability of these microspheres to be used as vaccine delivery systems.

Reversible addition–fragmentation chain transfer polymerization of 2-chloro-1,3-butadiene

Controlled polymerization of 2-chloro-1,3-butadiene using reversible addition–fragmentation chain transfer (RAFT) polymerization has been demonstrated for the first time. 2-Chloro-1,3-butadiene, more commonly known as chloroprene, has significant industrial relevance as a crosslinked rubber, with uses ranging from adhesives to integral automotive components. However, problems surrounding the inherent toxicity of the lifecycle of the thiourea-vulcanized rubber have led to the need for control over the synthesis of poly(2-chloro-1,3-butadiene). To this end, four chain transfer agents in two different solvents have been trialed and the kinetics are discussed. 2-Cyano-2-propylbenzodithioate (CPD) is shown to polymerize 2-chloro-1,3-butadiene in THF, using AIBN as an initiator, with complete control over the target molecular weight, producing polymers with low polydispersities (Mw/Mn < 1.25 in all cases).

The performance of poly(styrene)-block-poly(2-vinyl pyridine)-block- poly(styrene) triblock copolymers as pH-driven actuators

Poly(styrene)-block-poly(2-vinyl pyridine)-block-poly(styrene) (PS-b-P2VP-b-PS) triblock copolymers were synthesised by anionic polymerisation. Thick films were cast from solution and their structure analysed by small angle X-ray scattering (SAXS). Longer annealing times led to more ordered structures whereas short evaporation times effectively "lock" the polymer chains in a disordered state by vitrification. Well-ordered structures not only provide an isotropic network, which reduces localised stress within the material, but are also essential for fundamental studies of soft matter because their activity on the molecular scale must be analysed and understood prior to their use in technological applications. Well-characterised PS-b-P2VP-b-PS materials have been coupled to a pH-oscillating reaction and their potential application as responsive actuators is discussed. This journal is © The Royal Society of Chemistry.

Electrografting of poly (carbazole-co-acrylamide) onto highly oriented pyroltic graphite. A cyclovoltammetric, atomic force microscopic and ellipsometric study

The electrocopolymerization of carbazole and acrylamide on highly oriented pyrolytic graphite (HOPG) from ACN solutions via cyclovoltammetry (CV) was studied in order to evaluate the possibility to deposit uniform and thin but pinhole-free and still reactive coatings onto graphite-like substrates. The morphology of the coatings was investigated using atomic force microscopy and the coating thicknesses and optical parameters were measured using ellipsometry. It was found that under the chosen conditions thin (coating thickness hf>180 nm) and relatively smooth (root mean square surface roughness RMS<150 nm) P(Cz-co-AAm)-coatings exhibiting a uniform globuoidal morphology can be deposited onto graphite. From a certain coating thickness (hf>50 nm) no pinholes could be detected. It was found that the thickness of the deposited coatings increases almost linearly with increasing number of CV-cycles while keeping all other experimental parameters (scan rate and comonomer concentration ratio) constant. No influence of the comonomer concentration ratio on the film thickness and coating appearance could be observed, however, at quite low initial concentrations. However, the CV-scanning rate has quite a significant influence on the thickness of the deposited coatings. Higher scan rates (100 mV/s) result in thin (hf≈22 nm) coatings whereas at lower scan rates (<50 mV/s) coatings with thicknesses of approximately 50 nm were obtained. The optical coating parameters (the refractive index n and extinction coefficient k) seem to be independent of the deposition parameters and therefore averaged values of n̄=1.54±0.03 and k̄=0.08±0.03 were obtained.

Direct observation of morphological development during the spin-coating of polystyrene-poly(methyl methacrylate) polymer blends

We present results of the direct observation, in real-space, of the phase separation of high molecular weight polystyrene and poly(methyl methacrylate) from ortho-xylene using our newly developed technique of high speed stroboscopic interference microscopy. Taking a fixed concentration (3 wt % in o-xylene) at a fixed composition (1:4 by weight) and by varying the rotational rate during the spin-coating process, we are able to observe the formation of a range of phase separated bicontinuous morphologies of differing length-scales. Importantly, we are able to show that the mechanism by which the final phase separated structure is formed is through domain coarsening when rich in solvent, before vitrification occurs and fixes the phase separated structure. The ability to directly observe morphological development offers a route toward controlling the length-scale of the final morphology through process control and in situ feedback, from a single stock solution. © 2013 Wiley Periodicals, Inc.

Poly(glycerol adipate-co-ω-pentadecalactone) spray-dried microparticles as sustained release carriers for pulmonary delivery

Purpose: The aim of this work was to optimize biodegradable polyester poly(glycerol adipate-co-ω-pentadecalactone), PGA-co-PDL, microparticles as sustained release (SR) carriers for pulmonary drug delivery. Methods: Microparticles were produced by spray drying directly from double emulsion with and without dispersibility enhancers (L-arginine and L-leucine) (0.5-1.5%w/w) using sodium fluorescein (SF) as a model hydrophilic drug. Results: Spray-dried microparticles without dispersibility enhancers exhibited aggregated powders leading to low fine particle fraction (%FPF) (28.79±3.24), fine particle dose (FPD) (14.42±1.57 μg), with a mass median aerodynamic diameter (MMAD) 2.86±0.24 μm. However, L-leucine was significantly superior in enhancing the aerosolization performance ( L-arginine:%FPF 27.61±4.49-26.57±1.85; FPD 12.40±0.99-19.54±0.16 μg and MMAD 2.18±0.35-2. 98±0.25 μm, L-leucine:%FPF 36.90±3.6-43.38±5. 6; FPD 18.66±2.90-21.58±2.46 μg and MMAD 2.55±0.03-3. 68±0.12 μm). Incorporating L-leucine (1.5%w/w) reduced the burst release (24.04±3.87%) of SF compared to unmodified formulations (41.87±2.46%), with both undergoing a square root of time (Higuchi's pattern) dependent release. Comparing the toxicity profiles of PGA-co-PDL with L-leucine (1.5%w/w) (5 mg/ml) and poly(lactide-co-glycolide), (5 mg/ml) spray-dried microparticles in human bronchial epithelial 16HBE14o-cell lines, resulted in cell viability of 85.57±5.44 and 60.66±6.75%, respectively, after 72 h treatment. Conclusion:The above data suggest that PGA-co-PDL may be a useful polymer for preparing SR microparticle carriers, together with dispersibility enhancers, for pulmonary delivery. © Springer Science+Business Media, LLC 2011.

Preparation and property testing of compatibilized poly(l-lactide)/thermoplastic polyurethane blends

Poly(l-lactide) (PLL) has been blended with a polycaprolactone-based thermoplastic polyurethane (TPU) elastomer as a toughening agent and a poly(l-lactide-co-caprolactone) (PLLCL) copolymer as a compatibilizer. Both 2-component (PLL/TPU) and 3-component (PLL/TPU/PLLCL) blends were prepared by melt mixing, characterized, hot-pressed into thin sheets and their tensile properties tested. The results showed that, although the TPU could toughen the PLL, the blends were largely immiscible leading to phase separation. However, addition of the PLLCL copolymer improved blend compatibility. The best all-round properties were found for the 3-component blend of composition PLL/TPU/PLLCL = 90/10/10 parts by weight.