The role of stable nitroxyl radical precursors as antioxidants in polyolefins

Various 2,2,6,6-tetramethyl piperidines and their N-alkyl derivatives of stable nitroxyl radical precursors containing acrylic(s) and methacrylic(s) groups were reactively processed in the presence of a peroxide as bound-antioxidant masterbatches for polyolefin stabilisation. It was found that grafting of the antioxidant monomers onto the polymer backbone was inevitably in competition with homopolymerisation of the monomers as well as melt degradation of the polymer and other side reactions. As previously reported, binding efficiency of bisacrylic nitroxyl precursor was maximum due to formation of unextractable homopolymer of the antioxidant. On the other hand, the binding efficiency of monoacrylic derivatives was low and the homopolymers were found extractable, which suggests that the bound monoacrylic derivatives are entirely grafted onto the polyolefin backbone. Application of bis and tri-functional coagents gave improved binding efficiency of the monoacrylic monomers. This may be due to copolymerisation of the antioxidants with the coagents and grafting of the copolymers onto the polymer backbone. Comparison of photostabilising activity of the fully extracted bound antioxidants to those of the corresponding unbound analogous showed lower results for the former. However, thermal stabilising activity of the bound antioxidants was higher than that of the unbound analogous due to better substantivity. Analysis using physical techniques and GPC for molecular weight distribution of masterbatches containing the bound monoacrylic antioxidants showed formation of high molecular weight products. Model reaction of a secondary amine derivative in liquid hydrocarbon and analysis of the product using FTIR and NMR spectroscopy indicated a possibility of side reaction, i.e. involvement of the amine active group (>N-H) of the antioxidant in the binding process to form the high molecular weight product. Implementation of various N-alkylated derivatives did not inhibit the side reaction. The photostabilising activity of the bound-antioxidants can be improved when applied in conjunction with small amounts of a benzophenone uv-stabiliser. The synergistic stabilising activity, however, was diminished when the uv-stabiliser was removed from the system during the service time. Nitroxyl precursors containing methacrylic group(s) gave lower binding efficiency than the corresponding acrylic derivatives. Reversible deploymerisation of the grafted methacrylic antioxidants may be responsible for this. Bis and tris-acrylic coagents improved the binding efficiency, and the presence of methacrylic group improved stabilising activity of the antioxidants. N-methyl derivatives were found to exhibit better stabilising activity than their parent secondary amine derivatives.

Mechanisms of antifatigue agents used in natural rubber

A large number of compounds containing quinonoid or hindered phenol functions were examined for their roles as antifatigue agents. Among the evaluated quinones and phenols expected to have macroalkyl radical scavenging ability, BQ, αTOC, γTOC and GM showed relatively good performance for fatigue resistance (although their performance was slightly less effective than the commercial aromatic amine antioxidants, IPPD and 6PPD). The compounds which were shown to have higher reactivity with alkyl radicals (via calculated reactivity indices) showed better fatigue resistance. This fact supports the suggestion that strong alkyl radical scavengers should be also effective antifatigue agents. Evidence produced based on calculation of reactivity indices suggests that the quinones examined react with alkyl radicals on the meta position of the quinone rings producing phenoxyl radicals. The phenoxyl radicals are expected either to disproportionate, to recombine with a further alkyl radical, or to abstract a hydrogen from another alkyl radical producing an olefine. The regeneration of quinones and formation of the corresponding phenols is expected to occur during the antifatigue activity. The phenol antioxidant, HBA is expected to produce a quinonoid compound and this is also expected to function in a similar way to other quinones. Another phenol, GM, which is also known to scavenge alkyl radicals showed good antifatigue performance. Tocopherols had effective antifatigue activity and are expected to have different antifatigue mechanisms from that of other quinones, hence αTOC was examined for its mechanisms during rubber fatiguing using HPLC analysis. Trimers of αTOC which were produced during vulcanisation are suggested to contribute to the fatigue activity observed. The evidence suggests that the trimers reproduce αTOC and a mechanism was proposed. Although antifatigue agents evaluated showed antifatigue activity, most of them had poor thermoxidative resistance, hence it was necessary to compensate for this by using a combination of antioxidants with the antifatigue agents. Reactive antioxidants which have the potential to graft on the polymer chains during reactive processing were used for this purpose. APMA was the most effective antioxidant among other evaluated reactive antioxidants. Although high ratio of grafting was achieved after optimisation of grafting conditions, it is suggested that this was achieved by long branches of APMA due to large extent of polymerisation. This is expected to cause maldistribution of APMA leading to reducing the effect of CB-D activity (while CB-A activity showed clear advantages for grafting). Further optimisation of grafting conditions is required in order to use APMA more effectively. Moreover, although synergistic effects between APMA and antifatigue agents were expected, none of the evaluated antifatigue agents, BQ, αTOC, γTOC and TMQ, showed significant synergism both in fatigue and thermoxidative resistance. They performed just as additives.

The study and characterisation of water-based latices used for can coating interiors

Water-based latices, used in the production of internal liners for beer/beverage cans, were investigated using a number of analytical techniques. The epoxy-graft-acrylic polymers, used to prepare the latices, and films, produced from those latices, were also examined. It was confirmed that acrylic polymer preferentially grafts onto higher molecular weight portions of the epoxy polymer. The amount of epoxy remaining ungrafted was determined to be 80%. This figure is higher than was previously thought. Molecular weight distribution studies were carried out on the epoxy and epoxy-g-acrylic resins. A quantitative method for determining copolymer composition using GPC was evaluated. The GPC method was also used to determine polymer composition as a function of molecular weight. IR spectroscopy was used to determine the total level of acrylic modification of the polymers and NMR was used to determine the level of grafting. Particle size determinations were carried out using transmission electron microscopy and dynamic light scattering. Levels of stabilising amine greatly affected the viscosity of the latex, particle size and amount of soluble polymer but the core particle size, as determined using TEM, was unaffected. NMR spectra of the latices produced spectra only from solvents and amine modifiers. Using solid-state CP/MAS/freezing techniques spectra from the epoxy component could be observed. FT-IR spectra of the latices were obtained after special subtraction of water. The only difference between the spectra of the latices and those of the dry film were due to the presence of the solvents in the former. A distinctive morphology in the films produced from the latices was observed. This suggested that the micelle structure of the latex survives the film forming process. If insufficient acrylic is present, large epoxy domains are produced which gives rise to poor film characteristics. Casting the polymers from organic solutions failed to produce similar morphology.

Degradation and stabilisation of diisocyanate cured polybutadiene

Hydroxyl terminated polybutadiene (HTPB) has been used as a rocket propellant binder which is required to be stored for at least twenty years. It is found that the excellent stress-strain characteristics of this propellant can be totally lost, during this long storage, due to the deterioration of the polybutadiene chains. As a result, the propellant can not stand the service loads, which may lead to a catastrophe. The study of the HTPB binder degradation, below 80°C, has been carried out by investigating the environmental factors and the changes which occur along the macromolecular chains. Results have shown that oxygen is the main factor which causes the crosslinking and chain scission reactions. The former is the predominant reaction and proceeds rapidly under oxygen sufficient environment. The unsaturation of polymer chain, which provides the desired physical properties to the binder, was lost with the increase in crosslink density. At the same time hydroperoxides were found to form and decompose along the polymer chains. Therefore, the deterioration of the binder results from the oxidation of polymer chains. Since the oxidation reaction occurred at higher rate than oxygen diffusion rate and oxygen diffusion rate is inversely proportional to the crosslink density, the binder, below the surface layer in a thick section container, could be naturally protected under an oxygen deficient condition for a long time. Investigation of the effectiveness of antioxidants in HTPB binder has shown that the efficiency of an antioxidant depends on its ability to scavenge radicals. Generally, aromatic amines are the most effective binder antioxidants. But when a peroxide decomposer is combined with an aromatic amine at the appropriate ratio, a synergistic effect is obtained, which gives the lowest binder gel increase rate.

Interaction of peptides, peptidomimetics and other drug candidates with the DI/tripeptide transport system in intestinal epithelial cells, using the in vitro caco-2 cell culture system

An uptake system was developed using Caco-2 cell monolayers and the dipeptide, glycyl-[3H]L-proline, as a probe compound. Glycyl-[3H]L-proline uptake was via the di-/tripeptide transport system (DTS) and, exhibited concentration-, pH- and temperature-dependency. Dipeptides inhibited uptake of the probe, and the design of the system allowed competitors to be ranked against one another with respect to affinity for the transporter. The structural features required to ensure or increase interaction with the DTS were defined by studying the effect of a series of glycyl-L-proline and angiotensin-converting enzyme (ACE)-inhibitor (SQ-29852) analogues on the uptake of the probe. The SQ-29852 structure was divided into six domains (A-F) and competitors were grouped into series depending on structural variations within specific regions. Domain A was found to prefer a hydrophobic function, such as a phenyl group, and was intolerant to positive charges and H+ -acceptors and donors. SQ-29852 analogues were more tolerant of substitutions in the C domain, compared to glycyl-L-proline analogues, suggesting that interactions along the length of the SQ-29852 molecule may override the effects of substitutions in the C domain. SQ-29852 analogues showed a preference for a positive function, such as an amine group in this region, but dipeptide structures favoured an uncharged substitution. Lipophilic substituents in domain D increased affinity of SQ-29852 analogues with the DTS. A similar effect was observed for ACE-NEP inhibitor analogues. Domain E, corresponding to the carboxyl group was found to be tolerant of esterification for SQ-29852 analogues but not for dipeptides. Structural features which may increase interaction for one series of compounds, may not have the same effect for another series, indicating that the presence of multiple recognition sites on a molecule may override the deleterious effect of anyone change. Modifying current, poorly absorbed peptidomimetic structures to fit the proposed hypothetical model may improve oral bioavailability by increasing affinity for the DTS. The stereochemical preference of the transporter was explored using four series of compounds (SQ-29852, lysylproline, alanylproline and alanylalanine enantiomers). The L, L stereochemistry was the preferred conformation for all four series, agreeing with previous studies. However, D, D enantiomers were shown in some cases to be substrates for the DTS, although exhibiting a lower affinity than their L, L counterparts. All the ACE-inhibitors and β-lactam antibiotics investigated, produced a degree of inhibition of the probe, and thus show some affinity for the DTS. This contrasts with previous reports that found several ACE inhibitors to be absorbed via a passive process, thus suggesting that compounds are capable of binding to the transporter site and inhibiting the probe without being translocated into the cell. This was also shown to be the case for oligodeoxynucleotide conjugated to a lipophilic group (vitamin E), and highlights the possibility that other orally administered drug candidates may exert non-specific effects on the DTS and possibly have a nutritional impact. Molecular modelling of selected ACE-NEP inhibitors revealed that the three carbonyl functions can be oriented in a similar direction, and this conformation was found to exist in a local energy-minimised state, indicating that the carbonyls may possibly be involved in hydrogen-bond formation with the binding site of the DTS.

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.

Photochemical degradation of polystyrene

In this work the oxidative degradation of pure polystyrene, polybutadiene and butadiene-modified polystyrene (normally called high impact polystyrene or HIPS) have been studied using a variety of physical and chemical techniques. The changes in dynamic-mechanical properties occurring during the ultra-violet light accelerated weathering of these polymers were followed by a visco-elastometric technique (Rheovibron) in the solid phase over a wide temperature range. Selective cross-linking of the polybutadiene in high-impact polystyrene caused the depression of the low temperature damping peak (tan d) with a corresponding sharp peak in tan d at ambient temperature accompanied by an integral rise in complex modulus. During the same period of photoxidation, the hydroperoxide concentration and gel content increased rapidly, reaching a maximum before decomposing photolytically with the destruction of unsaturation and with the formation of stable oxidation products. Infra-red spectroscopy showed the formation of carbonyl and hydroxyl groups. a,ß-unsaturated carbonyl was also identified and was formed by decomposition of both allylic hydroperoxide and initial peroxidic gel by ß-scission of the graft between polybutadiene and polystyrene. With further photoxidation a more stable ether gel was formed involving the destruction of the conjugating double bond of a,ß-unsaturated carbonyl. Addition of saturated and unsaturated ketones which are potential sensitisers of photoxidation to high-impact polystyrene and polybutadiene failed to photo-initiate the oxygen absorption of the polymers. A prior thermal oxidative treatment on the other hand eliminated the auto- accelerating stage leading to linear kinetics as the concentration of thermally-produced hydroperoxide approached a maximum. Antioxidants which act by destroying hydroperoxide lengthened the induction period to rapid oxygen absorption, whilst a phenolic antioxidant behaved as a weak photo-activator initially and a retarder later. Prior photolysis of high-impact polystyrene photo-activated the unsaturated component and caused similar changes in dynamic-mechanical properties to those found during photoxidation although at a much lower rate. Polybutadiene behaves as a photo-pro-oxidant for the destruction of polystyrene in high-impact polystyrene.

Design, synthesis and evaluation of cyclothialidine analogues as DNA gyrase inhibitors

Cyclothialidine, a natural product isolated from Streptomyces .filipinensis NR0484, has been proven to be a potent and selective inhibitor of the bacterial enzyme DNA gyrase. Gyrase inhibition results in cell death, the enzyme being the target of several currently used antibiotics. Cyclothialidine showed poor activity against whole bacterial cells, highlighting scope for improvement regarding cell membrane pemeability in order for the full potential of this new class of antibiotics to be realised, Structurally, cyclothialidine contains a 12-membered lactone ring which is partly integrated into a pentapeptide chain, with a substituted aromatic moiety bordering the lactone, Retrosynthetically it can be traced back to cis-3-hydroxyproline, 3,5-dihydroxy-2,6-dimethylbenzoic acid and four commercially available amino acids; two serine, one cysteine and one alanine. In this work, a model of cyclothialidine was synthesised in order to establish the methodology for more complex compounds. Analogues with hydroxy, dihydroxy and dihydroxymethyl substituted aromatic moieties were then prepared to ensure successful protection methods could be performed and the pharmacophore synthesised. The key aromatic moiety, 2,6-dimethyl-3,5-dihydroxybenzoic acid was produced via two successive Mannich reaction/reduction steps. Acid protection using 4-nitrobenzyl bromide and TBDMS hydroxyl protection followed by bromination of one methyl afforded the desired intermediate. Reaction with a serine/cysteine dipeptide, followed by deprotection and cyclisation under Mitsunobu conditions lead to the 12-membered lactone. An amine substituted aromatic analogue and also replacement of the cysteine sulphur by oxygen were attempted but without success. In an effort to improve cell permeability, a conjugate was synthesised between the pharmacophore and a cholesterol moiety. It was hoped the steroid fragment would serve to increase potency by escorting the molecule through the lipid environment of the cell membrane. The pharmacophore and conjugate were tested against a variety of bacterial strains but the conjugate failed to improve activity.

Azidoprofen as a soft anti-flammatory agent for the topical treatment of Psoriasis

Azidoprofen {2-(4-azidophenyl)propionic acid; AZP}, an azido-substituted arylalkanoic acid, was investigated as a model soft drug candidate for a potential topical non-steroidal anti-inflammatory agent (NSAIA). Reversed-phase high performance liquid chromatography (HPLC) methods were developed for the assay of AZP, a series of ester analogues and their· degradation products. 1H-NMR spectroscopy was also employed as an analytical method in selected cases. Reduction of the azido-group to the corresponding amine has been proposed as a potential detoxification mechanism for compounds bearing this substituent. An in vitro assay to measure the susceptibility of azides towards reduction was developed using dithiothreitol as a model reducing agent. The rate of reduction of AZP was found to be base-dependent, hence supporting the postulated mechanism of thiol-mediated reduction via nucleophilic attack by the thiolate anion. Prodrugs may enhance topical bioavailability through the manipulation of physico-chemical properties of the parent drug. A series of ester derivatives of AZP were investigated for their susceptibility to chemical and enzymatic hydrolysis, which regenerates the parent acid. Use of alcoholic cosolvents with differing alkyl functions to that of the ester resulted in transesterification reactions, which were found to be enzyme-mediated. The skin penetration of AZP was assessed using an in vitro hairless mouse skin model, and silastic membrane in some cases. The rate of permeation of AZP was found to be a similar magnitude to that of the well established NSAIA ibuprofen. Penetration rates were dependent on the vehicle pH and drug concentration when solutions were employed. In contrast, flux was independent of pH when suspension formulations were used. Pretreatment of the skin with various enhancer regimes, including oleic acid and azone in propylene glycol, promoted the penetration of AZP. An intense IR absorption due to the azide group serves as a highly diagnostic marker, enabling azido compounds to be detected in the outer layers of the· stratum corneum following their application to skin, using attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR). This novel application enabled a non-invasive examination of the percutaneous penetration enhancement of a model azido compound in vivo in man, in the presence of the enhancer oleic acid.

Lipophilic antifolates as potential antipsoriatic agents

The lipophilic dihydrofolate reductase (DHFR) inhibitor m-azidopyrimethamine (MZP) was investigated for suitability for development as a topical antipsoriatic agent. The clinical features and treatments for psoriasis were reviewed. High performance liquid chromatography (HPLC) was employed as the main analytical method, with UV spectroscopy being used in some cases. Reduction of the azido-group was proposed as a potential detoxification mechanism for MZP. The rates of reduction of a series of substituted phenyl azide compounds by dithiothreitol were investigated and found to depend on the substitution pattern of the aryl azide molecular, with electron deficient azides exhibiting faster rates of reduction in the system studied. The rates of reduction of MZP and analogous compounds were also studied using this model. The skin penetration of MZP was assessed using an in vitro hairless mouse skin model. The rate of permeation (flux) of MZP across hairless mouse skin was found to be dependent on the quantity of propylene glycol used as cosolvent in the vehicle and the pH. The use of a pretreatment regime of oleic acid in propylene glycol was shown to greatly increase the penetration of MZP through the hairless mouse skin as compared to application without pretreatment, or pretreatment with other penetration enhancers. The metabolism of MZP was studied in in vitro models comprising skin homogenates, SV-K14 human keratinocyte cell cultures and skin commensal bacterial cultures. No conversion of MZP to the corresponding amine was detected in any of the models. The growth inhibitory properties of MZP were investigated in an in vitro SV-K14 human keratinocyte cell culture model and compared with those of other DHFR inhibitors. [14C]-pyrimethamine was shown to be taken up by the SV-K14 keratinocytes.

Molecular architecture influences on material properties of pharmaceutical compounds

Salt formation has extensively been studied as a strategy to improve drug solubility but it has not been explored as a strategy to improve mechanical properties. A better understanding of which factors of the solid state can have an influence in the mechanical properties of pharmaceutical powders can help to optimise and reduce cost of tablet manufacturing. The aim of this study was to form different series of amine salts of flurbiprofen, gemfibrozil and diclofenac and to establish predictive relationships between architectural characteristics and physicochemical and mechanical properties of the salts. For this purpose, three different carboxylic acid drugs were selected: flurbiprofen, gemfibrozil and diclofenac, similar in size but varying in flexibility and shape and three different series of counterions were also chosen: one with increasing bulk and no hydroxyl groups to limit the hydrogen bonding potential; a second one with increasing number of hydroxyl groups and finally a third series, related to the latter in number of hydroxyl groups but with different molecular shape and flexibility. Physico-chemical characterization was performed (DSC, TGA, solubility, intrinsic dissolution rate, particle size, true density) and mechanical properties measured using a compaction replicator. Strained molecular conformations produce weaker compacts as they have higher energy than preferred conformations that usually lie close to energy minimums and oppose plastic deformation. It was observed that slip planes, which correspond to regions of weakest interaction between the planes, were associated with improved plasticity and stronger compacts. Apart from hydrogen bonds, profuse van der Waals forces can result in ineffective slip planes. Salts displaying two-dimensional densely hydrogen bonded layers produced stronger compacts than salts showing one-dimensional networks of non-bonded columns, probably by reducing the attachment energy between layers. When hydrogen bonds are created intramolecularly, it is possible that the mechanical properties are compromised as they do not contribute so much to create twodimensional densely bonded layers and they can force molecules into strained conformations. Some types of hydrogen bonding network may be associated with improved mechanical properties, such as type II, or R (10) 3 4 using graph-set notation, versus type III, or R (12) 4 8 , columns. This work clearly demonstrates the potential of investigating crystal structure-mechanical property relationship in pharmaceutical materials.

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.

Isolation, characterisation and application of novel microbial protein cross-linking enzymes

Microbial transglutaminase is favoured for use in industry over the mammalian isoform, and hence has been utilized, to great effect, as an applied biocatalyst in many industrial areas including the food and textiles industries. There are currently only a limited number of microbial TGase sources known. A number of organisms have been screened for transglutaminase activity using biochemical assays directed towards TGase catalyzed reactions (amine incorporation and peptide cross-linking assay). Of those organisms screened, TGase was identified in a number of isolates including members of the Bacillus and Streptomyces families. In addition, a protein capable of performing a TGase-like reaction was identified in the organism Pseudomonas putida that was deemed immunologically distinct from previously described TGase isoforms, though further work would be required to purify the protein responsible. The genuses Streptoverticillium and Streptomyces are known to be closely related. A number of micro-organisms relating to Streptomyces mobaraensis (formerly Streptoverticillium mobaraensis) have been identified as harboring a TGase enzyme. The exact biological role of Streptomyces TGase is not well understood, though from work undertaken here it would appear to be involved in cell wall growth. Comparison of the purified Streptomyces TGase proteins showed them to exhibit marginally different characteristics in relation to enzymatic activity and pH dependency upon comparison with Streptomyces mobaraensis TGase. In addition, TGase was identified in the organism Saccharomonospora viridis that was found to be genetically identical to that from S. mobaraensis raising questions about the enzymes dissemination in nature. TGase from S. baldaccii was found to be most diverse with respect to enzymatic characteristics whilst still retaining comparable E(y-glutamyl) lysine bond formation to S. mobaraensis TGase. As such S. baldaccii TGase was cloned into an expression vector enabling mass production of the enzyme thereby providing a viable alternative to S. mobaraensis TGase for many industrial processes.

Effect of contact surfaces on the thermal and photoxidation of dehydrated castor oil

The effect of stainless steel, glass, zirconium and titanium enamel surfaces on the thermal and photooxidative toughening mechanism of dehydrated castor oil films deposited on these surfaces was investigated using different analytical and spectroscopic methods. The conjugated and non-conjugated double bonds were identified and quantified using both Raman spectroscopy and 1D and 2D NMR spectroscopy. The disappearance of the double bonds in thermally oxidised oil-on-surface films was shown to be concomitant with the formation of hydroperoxides (determined by iodometric titration). The type of the surface had a major effect on the rate of thermal oxidation of the oil, but all of the surfaces examined had resulted in a significantly higher rate of oxidation compared to that of the neat oil. The highest effect was exhibited by the stainless steel surface followed by zirconium enamel, titanium enamel and glass. The rate of thermal oxidation of the oil-on-steel surface (at 100 °C, based on peroxide values) was more than five times faster than that of oil-on-glass and more than 21 times faster than the neat oil when compared under similar thermal oxidative conditions. The rate of photooxidation at 60 °C of oil-on-steel films was found to be about one and half times faster than their rate of thermal oxidation at the same temperature. Results from absorbance reflectance infrared microscopy with line scans taken across the depth of thermally oxidised oil-on-steel films suggest that the thermal oxidative toughening mechanism of the oil occurs by two different reaction pathways with the film outermost layers, i.e. furthest away from the steel surface, oxidising through a traditional free radical oxidation process involving the formation of various oxygenated products formed from the decomposition of allylic hydroperoxides, whereas, in the deeper layers closer to the steel surface, crosslinking reactions predominate.

The structural and electrochemical consequences of hydrogenating Copper N2S2 Schiff base macrocycles

A series of cis and trans tetradentate copper macrocyclic complexes, of ring size fourteen - sixteen, which employ amine and thioether donor groups are reported. Apart from 5,6,15,16-bisbenzo-8,13-diaza-1,4-dithia-cyclohexadecane copper(I) (cis-[Cu(H4NbuSen)]+) all of the complexes are obtained in the copper(II) form. Crystallographic analysis shows that the copper(II) complexes all adopt a distorted planar geometry around the copper. In contrast, cis-[Cu(H4NbuSen)]+ is found to adopt a distorted tetrahedral geometry. The complexes were subjected to electrochemical analysis in water and acetonitrile. The effect of the solvent, positions of the donor atoms (cis/trans) on E1/2 is discussed as is the comparison of the electrochemical behaviour of these complexes with their parent Schiff base macrocycles.