Plasma antioxidant status, immunoglobulin G oxidation and lipid peroxidation in demented patients:Relevance to Alzheimer disease and vascular dementia

A large body of evidence supports a role of oxidative stress in Alzheimer disease (AD) and in cerebrovascular disease. A vascular component might be critical in the pathophysiology of AD, but there is a substantial lack of data regarding the simultaneous behavior of peripheral antioxidants and biomarkers of oxidative stress in AD and vascular dementia (VaD). Sixty-three AD patients, 23 VaD patients and 55 controls were included in the study. We measured plasma levels of water-soluble (vitamin C and uric acid) and lipophilic (vitamin E, vitamin A, carotenoids including lutein, zeaxanthin, β-cryptoxanthin, lycopene, α- and β-carotene) antioxidant micronutrients as well as levels of biomarkers of lipid peroxidation [malondialdehyde (MDA)] and of protein oxidation [immunoglobulin G (IgG) levels of protein carbonyls and dityrosine] in patients and controls. With the exception of β-carotene, all antioxidants were lower in demented patients as compared to controls. Furthermore, AD patients showed a significantly higher IgG dityrosine content as compared to controls. AD and VaD patients showed similar plasma levels of plasma antioxidants and MDA as well as a similar IgG content of protein carbonyls and dityrosine. We conclude that, independent of its nature - vascular or degenerative - dementia is associated with the depletion of a large spectrum of antioxidant micronutrients and with increased protein oxidative modification. This might be relevant to the pathophysiology of dementing disorders, particularly in light of the recently suggested importance of the vascular component in AD development. Copyright © 2004 S. Karger AG, Basel.

P4-235 protein oxidation, lipid peroxidation and antioxidant status are similarly altered in Alzheimer disease and vascular dementia

Background: A large body of evidence supports a role of oxidative stress in Alzheimer disease (AD) and in cerebrovascular disease. A vascular component might be critical in the pathophysiology of AD. Objective(s): To evaluate the simultaneous behavior of a broad spectrum of peripheral antioxidants and biomarkers of oxidative stress in AD and vascular dementia (VaD). Methods: Sixty-three AD patients, 23 VaD patients and 55 controls were included in the study. We measured plasma levels of water-soluble (vitamin C and uric acid) and lipophilic (vitamin E, vitamin A, carotenoids including lutein, zeaxanthin, [3-cryptoxanthin, lycopene, c~- and [3-carotene) antioxidant micronutrients as well as levels of biomarkers of lipid peroxidation [malondialdehyde (MDA)] and of protein oxidation [immunoglobniin G (Ig G) levels of protein carbonyls and dityrosine] in patients and controls. Results: AD and VaD patients showed significantly decreased plasma levels of the water-soluble vitamin C and uric acid, of the lipophilic vitamin Eand vitamin A, and of the carotenoids lutein, zeaxanthin, 13-cryptoxanthin, lycopene and (x-carotene as compared to controls; among biomarkers of oxidative stress, only the content of dityrosine in Ig G was found to be significantly higher (p < 0.01) in AD patients as compared to controls; although a trend towards higher levels of dityrosine was also observed in VaD subjects compared to controls (6.3 4- 1.7 ~M in VaD patients vs. 5.1 4- 1.6 IxM in controls; p = 0.06), it did not reach statistical significance. In a cumulative analysis of all patient samples, a significant inverse association was found between plasma lycopene and MDA levels (r = -0.53, p < 0.0001). Conclusions: Independent of its nature-vascular or degenerativedementia is associated with the depletion of a large spectrum of antioxidant micronutrients and with increased protein oxidative modification. This might be relevant to the pathophysiology of dementing disorders, particularly in light of the recently suggested importance of the vascular component in AD development.

Non-aqueous silicone elastomer gels as a vaginal microbicide delivery system for the HIV-1 entry inhibitor maraviroc

Aqueous semi-solid polymeric gels, such as those based on hydroxyethylcellulose (HEC) and polyacrylic acid (e.g. Carbopol®), have a long history of use in vaginal drug delivery. However, despite their ubiquity, they often provide sub-optimal clinical performance, due to poor mucosal retention and limited solubility for poorly water-soluble actives. These issues are particularly pertinent for vaginal HIV microbicides, since many lead candidates are poorly water-soluble and where a major goal is the development of a coitally independent, once daily gel product. In this study, we report the use of a non-aqueous silicone elastomer gel for vaginal delivery of the HIV-1 entry inhibitor maraviroc. In vitro rheological, syringeability and retention studies demonstrated enhanced performance for silicone gels compared with a conventional aqueous HEC gel, while testing of the gels in the slug model confirmed a lack of mucosal irritancy. Pharmacokinetic studies following single dose vaginal administration of a maraviroc silicone gel in rhesus macaques showed higher and sustained MVC levels in vaginal fluid, vaginal tissue and plasma compared with a HEC gel containing the same maraviroc loading. The results demonstrate that non-aqueous silicone gels have potential as a formulation platform for coitally independent vaginal HIV microbicides.

Dehydration at the lens surface:surface energy and surfactant persistence

Purpose: Most published surface wettability data are based on hydrated materials and are dominated by the air-water interface. Water soluble species with hydrophobic domains (such as surfactants) interact directly with the hydrophobic domains in the lens polymer. Characterisation of relative polar and non-polar fractions of the dehydrated material provides an additional approach to surface analysis. Method: Probe liquids (water and diiodomethane) were used to characterise polar and dispersive components of surface energies of dehydrated lenses using the method of Owens and Wendt. A range of conventional and silicone hydrogel soft lenses was studied. The polar fraction (i.e. polar/total) of surface energy was used as a basis for the study of the structural effects that influence surfactant persistence on the lens surface. Results: When plotted against water content of the hydrated lens, polar fraction of surface energy (PFSE) values of the dehydrated lenses fell into two rectilinear bands. One of these bands covered PFSE values ranging from 0.4 to 0.8 and contained only conventional hydrogels, with two notable additions: the plasma coated silicone hydrogels lotrafilcon A and B. The second band covered PFSE values ranging from 0.04 to 0.28 and contained only silicone hydrogels. Significantly, the silicone hydrogel lenses with lowest PFSE values (p<0.15) are found to be prone to lipid deposition duringwear. Additionally, more hydrophobic surfactants were found to be more persistent on lenses with lower PFSE values. Conclusions: Measurement of polar fraction of surface energy provides an importantmechanistic insight into surface interactions of silicone hydrogels.

An Investigation of the destruction of ion exchange resins

Ion exchange resins are used for many purposes in various areas of science and commerce. One example is the use of cation exchange resins in the nuclear industry for the clean up of radioactively contaminated water (for example the removal of 137Cs). However, during removal of radionuclides, the resin itself becomes radioactively contaminated, and must be treated as Intermediate Level Waste. This radioactive contamination of the resin creates a disposal problem. Conventionally, there are two main avenues of disposal for industrial wastes, landfill burial or incineration. However, these are regarded as inappropriate for the disposal of the cation exchange resin involved in this project. Thus, a method involving the use of Fenton's Reagent (Hydrogen Peroxide/soluble Iron catalyst) to destroy the resin by wet oxidation has been developed. This process converts 95% of the solid resin to gaseous CO2, thus greatly reducing the volume of radioactive waste that has to be disposed of. However, hydrogen peroxide is an expensive reagent, and is a major component of the cost of any potential plant for the destruction of ion exchange resin. The aim of my project has been to discover a way of improving the efficiency of the destruction of the resin thus reducing the cost involved in the use of hydrogen peroxide. The work on this problem has been concentrated in two main areas:-1) Use of analytical techniques such as NMR and IR to follow the process of the hydrogen peroxide destruction of both resin beads and model systems such as water soluble calixarenes. 2) Use of various physical and chemical techniques in an attempt to improve the overall efficiency of hydrogen peroxide utilization. Examples of these techniques include UV irradiation, both with and without a photocatalyst, oxygen carrying molecules and various stirring regimes.

Solid dispersions: formulation, characterisation, permeability and genomic evaluation

Poor water solubility is characterised by low dissolution rate and consequently reduced bioavailability. Formulation of solid dispersion of the drug has attracted considerable interest as a means of improving dissolution process of a range of poorly water soluble drugs. This current study investigates the formulation of solid dispersion for a range of poorly water soluble drugs with varying physicochemical properties including paracetamol, sulphamethoxazole, phenacetin, indomethacin, chloramphenicol, phenylbutazone and succinylsulphathiazole. Solid dispersions were prepared using various drugs to polymer ratios. PEG 8000 was selected as a carrier in the solid dispersions. The study revealed that inclusion of drug within the polymeric matrix, ratio of drug to polymer and physicochemical properties of the drug molecules enhance the dissolution rate. Characterisations of the solid dispersions were performed using DSC, FTIR and SEM. These studies revealed that all seven drugs were present in the amorphous form within the solid dispersions and there was a lack of interaction between the PEG 8000 and drug. Stability studies for solid dispersions showed that all seven drugs studied were unstable at accelerated conditions (40°C±2°C/75%RH±5%RH) whereas, they were found to be stable for 12 months at room conditions. Permeability of indomethacin, phenacetin, phenylbutazone and paracetamol were higher for solid dispersions as compared to drug alone across Caco-2 cell monolayers. From the cell uptake studies it was shown that PEG 8000 enhanced rhodamine123 uptake which suggested that PEG 8000 may increase the permeability of these drugs in solid dispersions. Gene expression profiles analyzing the expression changes in the ABC and solute carrier transporter during permeability studies.ABCA10, ABCB4, ABCC12, SLC12A6, MCT13, SLC22A12 and SLC6A6 gene expression were increased by indomethacin alone whereas solid dispersion of indomethacin resulted in a slight increase in expression. ABCC12 and SAMC gene expression was increased in case of paracetamol alone but slightly increased when exposed to solid dispersion of paracetamol.

Iron in the substrates and sporophores of Agaricus bisporus (Lange) Pilat during growth and development

The changes of the concentration of iron in the growth substrates and the sporophores of Agaricus bisporus (Lange) Pilat that occurred during culture under standard commercial conditions, were observed using atomic absorption spectrophotometry and iron-59 radiotracing techniques. The routes of translocation and sites of iron accumulation within the sporophore were shovn to alter during development and by the use of novel, pelletised substrates the concentration of iron in the mycelium of the substrates and in developing sporophores was observed during culture. Findings indicated that the compost was the major source of iron and that the concentration of iron in the compost mycelium varied cyclically in relation to the periodic appearance of sporophores. In the casing layer the mycelium is organised into strands which are responsible for the movement of iron from the compost into developing sporophores. A photographic technique for estimating sporophore growth rates showed that the accumulation of iron was not concomitant with sporophore growth and this was attributable to a declining quantity of available iron in the compost mycelium during sporophore growth. Variations in the quantity of iron in sporophores resulted primarily from differences in the quantity of water soluble iron in the compost but, the productivity of the crop, the type of casing layer and differences in watering also influenced sporophore composition. Changes in the concentration of extractable iron in the compost and casing layer throughout culture were related to mycelial activity and to a lesser extent were influenced by watering and the bacterial populations of the casing layer. Thus, the findings of this study give some indication of the relative importance that different cultural conditions exert over sporophore composition together with demonstrations of the movement of a single material within the sporophores and substrates during the cultivation of Agaricus bisporus.

Cyclodextrin complexes of antimicrobial agents

Poorly water-soluble drugs show an increase in solubility in the presence of cyclodextrins (CyD) due to the formation of a water-soluble complex between the drug and dissolved CyD. This study investigated the interactions of -Cyd and hydroxypropyl--CyD (HP--CyD, M.S. = 0.6) with antimicrobial agents of limited solubility in an attempt to increase their microbiological efficacy. The agents studied were chlorhexidine dihydrochloride (CHX), p-hydroxybenzoic acid esters (methyl, ethyl, proply and butyl) and triclosan. The interactions between the antimicrobials and CyDs were studied in solution and solid phases. Phase solubility studied revealed an enhancement in the aqueous drug solubility in the presence of the CyD and also gave an indication of the complex stability constant (Ks). The temperature-dependence of the stability constant of the complex was modelled by the van't Hoff plot which yielded the thermodynamic parameters for complexation. Further confirmation of the inclusion of the antimicrobials within the cavity of the CyDs in aqueous solution was obtained from proton magnetic resonance and ultraviolet absorption spectroscopies. The former method indicated that the chlorophenyl moiety of the CHX was included within the -CyD cavity and the stoichiometry of the complex formed was 1:1. The solid-phase complexes were prepared by freeze-drying. The inclusion complex of triclosan with HP--CyD was obtained from aqueous solution with the addition of ammonia. Evidence to confirm complex formation was obtained from DSC, IR and X-ray powder diffraction studies. Dissolution studies of the solid inclusion complexes using the dispersed powder technique illustrated their superior solubilities as compared to the equimolar physical mix of the guest and CyD. It was shown that these solutions of the complex were supersaturated with respect to the free guest. This was further demonstrated by diffusion studies which showed the flux of free drug from donor solutions of the antimicrobial-CyD complex to be significantly greater than the flux from donor suspensions of drug alone.

Development of plasmid dna formulations for application as microspheric dna vaccines

The advent of DNA vaccines has heralded a new technology allowing the design and elicitation of immune responses more adequate for a wider range of pathogens. The formulation of these vaccines into the desired dosage forms extends their capability in terms of stability, routes of administration and efficacy. This thesis describes an investigation into the fabrication of plasmid DNA, the active principle of DNA vaccines, into microspheres, based on the tenet of an increased cellular uptake of microparticulate matter by phagocytic cells. The formulation of plasmid DNA into microspheres using two methods, is presented. Formulation of microspheric plasmid DNA using the double emulsion solvent evaporation method and a spray-drying method was explored. The former approach involves formation of a double emulsion, by homogenisation. This method produced microspheres of uniform size and smooth morphology, but had a detrimental effect on the formulated DNA. The spray-drying method resulted in microspheres with an improved preservation of DNA stability. The use of polyethylenimine (PEI) and stearylamine (SA) as agents in the microspheric formulation of plasmid DNA is a novel approach to DNA vaccine design. Using these molecules as model positively-charged agents, their influence on the characteristics of the microspheric formulations was investigated. PEI improved the entrapment efficiency of the plasmid DNA in microspheres, and has minimal effect on either the surface charge, morphology or size distribution of the formulations. Stearylamine effected an increase in the entrapment efficiency and stability of the plasmid DNA and its effect on the micropshere morphology was dependent on the method of preparation. The differences in the effects of the two molecules on microsphere formulations may be attributable to their dissimilar physico-chemical properties. PEI is water-soluble and highly-branched, while SA is hydrophobic and amphipathic. The positive charge of both molecules is imparted by amine functional groups. Preliminary data on the in vivo application of formulated DNA vaccine, using hepatitis B plasmid, showed superior humoral responses to the formulated antigen, compared with free (unformulated) antigen.

Incorporation and release of macromolecules from biodegradable polymer vehicles

The initial objective of this work was to evaluate and introduce fabrication techniques based on W/0/W double emulsion and 0/W single emulsion systems with solvent evaporation for the incorporation of a surrogate macromolecule (BSA) into microspheres and microcapsules fabricated using P(HB-HV}, PEA and their blends. Biodegradation, expressed as changes in the gross and ultrastructural morphology of BSA loaded microparticulates with time was monitored using SEM concomitant with BSA release. Spherical microparticulates were successfully fabricated using both the W/0/W and 0/W emulsion systems. Both microspheres and microcapsules released BSA over a period of 24 to 26 days. BSA release from P(HB-HV)20% PCL 11 microcapsules increased steadily with time, while BSA release from all other microparticulates was characterised by an initial lag phase followed by exponential release lasting 6-11 days. Microcapsules were found to biodegrade more rapidly than microspheres fabricated from the same polymer. The incubation of microparticulates in newborn calf serum; synthetic gastric juice and pancreatin solution showed that microspheres and microcapsules were susceptible to enzymatic biodegradation. The in vitro incubation of microparticulates in Hank's buffer demonstrated limited biodegradation of microspheres and microcapsules by simple chemical hydrolysis. BSA release was thought to ocurr as a result of the macromolecule diffusing through either inherent micropores or via pores and channels generated in situ by previously dissolved BSA. However, in all cases, irrespective of percentage loading or fabrication polymer, low encapsulation efficiencies were obtained with W/0/W and 0/W techniques (4.2±0.9%- 15.5±0.5%,n=3), thus restricting the use of these techniques for the generation of microparticulate sustained drug delivery devices. In order to overcome this low encapsulation efficiency, a W/0 single emulsion technique was developed and evaluated in an attempt to minimise the loss of the macromolecule into the continuous aqueous phase and increase encapsulation efficiency. Poly(lactide-co-glycolide) [PLCG] 75:25 and 50:50, PEA alone and PEA blended with PLCG 50:50 to accelerate biodegradation, were used to microencapsulate the water soluble antibiotic vancomycin, a putative replacement for gentamicin in the control of bacterial infection in orthopaedic surgery especially during total hip replacement. Spherical microspheres (17.39±6.89~m,n=74-56.5±13.8~m,n=70) were successfully fabricated with vancomycin loadings of 10, 25 and 50%, regardless of the polymer blend used. All microspheres remained structurally intact over the period of vancomycin release and exhibited high percentage yields( 40. 75±2 .86%- 97.16±4.3%,n=3)and encapsulation efficiencies (47.75±9.0%- 96.74±13.2%,n=12). PLCG 75:25 microspheres with a vancomycin loading of 50% were judged to be the most useful since they had an encapsulation efficiency of 96.74+13.2%, n=12 and sustained therapeutically significant vancomycin release (15-25μg/ml) for up to 26 days. This work has provided the means for the fabrication of a spectrum of prototype biodegradable microparticulates, whose biodegradation has been characterised in physiological media and which have the potential for the sustained delivery of therapeutically useful macromolecules including water soluble antibiotics for orthopaedic applications.

The application of monolayer studies in the understanding of liposomal formulations

The study of surfactant monolayers is certainly not a new technique, but the application of monolayer studies to elucidate controlling factors in liposome design remains an underutilised resource. Using a Langmuir-Blodgett trough, pure and mixed lipid monolayers can be investigated, both for their interactions within the monolayer, and for interfacial interactions with drugs in the aqueous sub-phase. Despite these monolayers effectively being only half a bilayer, with a flat rather than curved structure, information from these studies can be effectively translated into liposomal systems. Here we outline the background, general protocols and application of Langmuir studies with a focus on their application in liposomal systems. A range of case studies are discussed which show how the system can be used to support its application in the development of liposome drug delivery. Examples include investigations into the effect of cholesterol within the liposome bilayer, understanding effective lipid packaging within the bilayer to promote water soluble and poorly soluble drug retention, the effect of alkyl chain length on lipid packaging, and drug-monolayer electrostatic interactions that promote bilayer repackaging.

Interactions between organic polymers and cement hydration products

Organic substances, particularly polymers, are finding increasing use in modifying the properties of cements and concrete. Although a significant amount of research has been conducted into the modification of the mechanical properties of cements by polymers, little is known about the nature of the interface and interactions taking place between the two phases. This thesis addresses the problem of elucidating such interactions. Relevant literature is reviewed, covering the general use of polymers with cements, the chemistry of cements and polymers, adhesion and known interactions between polymers and both cements and related minerals. Although several polymer systems were studied, two in particular were selected, as being well characterized. These were: - 1) polymethyl methacrylate (PMMA), the polymer derived from methyl methacrylate (MMA), and 2) an amine-cured epoxy resin system. By this approach, a methodology was developed for the examination of other polymer/cement interactions. Experiments were conducted in five main areas:- 1) polymer-cement adhesion and the feasibility of revealing interfacial regions mechanically, 2) chemical reactions between polymers and cements, 3) characterization of cement adhesion surfaces, 4) interactions affecting overall polymerisation rates, and 5) studies of polymer impregnated cements. The following conclusions were reached:- 1) The PMMA/cement interface contains calcium methacrylate as an interfacial reaction product, water being a reactant. Calcium methacrylate is detrimental to the properties of PMMA/cement composites, being highly water-soluble. 2) The pore surface of cement accelerates the polymerisation of MMA, leading to an increased molecular weight compared to polymerisation of pure MMA, minerals in hydrated cement powders having the opposite effect. 3) The investigation of reaction products presents a number of experimental problems, selection of appropriate techniques depending upon the system studied. For the two systems examined in detail, ion chromatography proved particularly useful; DTA, IRS and XPS indicated reactions, though the data was hard to interpret; XRD proving inconclusive. 4) It is impractical to reveal interfacial regions mechanically, but may be accomplished by chemical means.

Indwelling catheters and medical implants with FXIIIa inhibitors:a novel approach to the treatment of catheter and medical device-related infections

Central venous catheters (CVCs) are being utilized with increasing frequency in intensive care and general medical wards. In spite of the extensive experience gained in their application, CVCs are related to the long-term risks of catheter sheath formation, infection, and thrombosis (of the catheter or vessel itself) during catheterization. Such CVC-related-complications are associated with increased morbidity, mortality, duration of hospitalization, and medical care cost [1]. The present study incorporates a novel group of Factor XIIIa (FXIIIa, plasma transglutaminase) inhibitors into a lubricious silicone elastomer in order to generate an optimized drug delivery system whereby a secondary sustained drug release profile occurs following an initial burst release for catheters and other medical devices. We propose that the incorporation of FXIIIa inhibitors into catheters, stents, and other medical implant devices would reduce the incidence of catheter sheath formation, thrombotic occlusion, and associated staphylococcal infection. This technique could be used as a local delivery system for extended release with an immediate onset of action for other poorly aqueous soluble compounds. © 2012 Elsevier B.V. All rights reserved.

New hybrid system:poly (ethylene glycol) hydrogel with covalently bonded pegylated nanotubes

Hydrogels containing carbon nanotubes (CNTs) are expected to be promising conjugates because they might show a synergic combination of properties from both materials. Most of the hybrid materials containing CNTs only entrap them physically, and the covalent attachment has not been properly addressed yet. In this study, single-walled carbon nanotubes (SWNTs) were successfully incorporated into a poly(ethylene glycol) (PEG) hydrogel by covalent bonds to form a hybrid material. For this purpose, SWNTs were functionalized with poly(ethylene glycol) methacrylate (PEGMA) to obtain water-soluble pegylated SWNTs (SWNT–PEGMA). These functionalized SWNTs were covalently bonded through their PEG moieties to a PEG hydrogel. The hybrid network was obtained from the crosslinking reaction of poly(ethylene glycol) diacrylate prepolymer and the SWNT–PEGMA by dual photo-UV and thermal initiations. The mechanical and swelling properties of the new hybrid material were studied. In addition, the material and lixiviates were analyzed to elucidate any kind of SWNT release and to evaluate a possible in vitro cytotoxic effect. © 2010 Wiley Periodicals, Inc. J Appl Polym Sci, 2011.

Chitosan-modified dry powder formulations for pulmonary gene delivery

Spray-drying is an effective process for preparing micron-dimensioned particles for pulmonary delivery. Previously, we have demonstrated enhanced dispersibility and fine particle fraction of spray-dried nonviral gene delivery formulations using amino acids or absorption enhancers as dispersibility-enhancing excipients. In this study, we investigate the use of the cationic polymer chitosan as a readily available and biocompatible dispersibility enhancer. Lactose-lipid:polycation:pDNA (LPD) powders were prepared by spray-drying and post-mixed with chitosan or spray-dried chitosan. In addition, the water-soluble chitosan derivative, trimethyl chitosan, was added to the lactose-LPD formulation before spray-drying. Spray-dried chitosan particles, displaying an irregular surface morphology and diameter of less than 2 mu m, readily adsorbed to lactose-LPD particles following mixing. In contrast with the smooth spherical surface of lactose-LPD particles, spray-dried trimethyl chitosan-lactose-LPD particles demonstrated increased surface roughness and a unimodal particle size distribution (mean diameter 3.4 mu m), compared with the multimodal distribution for unmodified lactose-LPD powders (mean diameter 23.7 mu m). The emitted dose and in vitro deposition of chitosan-modified powders was significantly greater than that of unmodified powders. Moreover, the inclusion of chitosan mediated an enhanced level of reporter gene expression. In summary, chitosan enhances the dispersibility and in vitro pulmonary deposition performance of spray-dried powders.