In the first part of this thesis (chapters 1-8) the structure/activity (S/A) correlation studies on a class of anti-cancer drugs based on flavone acetic acid (FAA) by means of computer modelling techniques are reported. In particular, semiempirical and ab-initio quantum mechanical calculations have been performed on ten FAAs whose expeirmental anti-cancer activity was known. The results show that some calculated properties such as bond lengths, atomic charges, energies of the HOMO and the atomic orbitals involved in its formation, correlate with the anti-tumour activity. The correlations found were then used on another 38 molecules analogous to FAA whose anti-cancer activity had also been measured and of the 21 active molecules, 20 were predicted to be active by these SA correlations (95% success rate). From this study it also emerged that the pyrone ring may be directly involved in the anti-tumour mode of action of the FAA and it is suggested that vitamin-K may also play a role. The second part (chapter 9) is a study of the dependence of the molecular electrostatic potential on the basis set. From this study it emerged that GEOSMALL and MINI-1 minimal basis sets produce MEPs that are more similar to those obtained with the 6-31G** basis set than the MEP obtained with the STO-NG basis set. GEOSMALL and MINI-1 also give better energies and better properties than STO-NG and their use is recommended when properties of large organic molecules are of interest. Also, from this study it emerged that the use of Mulliken charges for the calculation of the MEP with the point charge approximation is not advisable for it may lead to very different pictures of the electrostatic potential calculated directly from the wave function.

Thermodynamic parameters have been obtained for the complexes formed between protons, first Transition Series metal ions and the asparaginate ion in aqueous solution at 25°C in an ionic background of 3.00 M (sodium) perchlorate. The metal ions under consideration are manganese(II), iron(II), cobalt(II), nickel(II), copper(II) and zinc(II). The techniques employed were potentiometry to obtain the formation constants (and hence ΔG°) for the complexes, and aqueous titration calorimetry to obtain the corresponding ΔH° (and hence Δs°) values. The results show that iron, cobalt, nickel and zinc are capable of binding up to three asparaginate ligands, whereas for manganese and copper the maximum number of bound ligands is two. All the complexes reported one simple AnB species: no hydroxy, protonated or polynuclear species were detected. The results are used (l) to discuss an unusual "homologue" effect whereby the trends among homologues for bonding to protons and to metal ions are opposite to each other, this apparently being a repercussion of solvation differences between homologues and (2) to show that quite a large proportion of asparaginate in blood plasma may be complexed to Zn(II), Fe(II), and Co(II), as well as the expected Cu(II). The techniques of potentiometry and calorimetry are further employed to determine the thermodynamic parameters for three ternary systems, one, copper(II)-histidinate-threoninate, which is known to exist in vivo, and two others, copper(II)-asparaginate-histidinate and copper(II)-asparaginate-threoninate, which have not yet been detected. Structures are suggested for the complexes Cu.asn.his,H+, Cu.asn.his.thr. , Cu.asn.thr. Cu.his.thr. and Cu.his.thr.H+ . In these complexes histidinate is tridentate to Cu(II), threoninate is bidentate and asparaginate is intermediate between bi and tridentate. The site of proton attachment in the protonated complexes is the primary amine site of the histidinate ligand. Throughout the work extensive use was made of computer programs, all of which are described in the chapter entitled "Computational Aspects".

Section A; Insoluble elastin was isolated from bovine aorta and ligaraentum nuchae by the use of guanidine and dithiothreitol in conjunction with collagenase, purified by affinity chromatography. The preparations were free from carbohydrate and exhibited amino acid compositions similar to that of alkali-purified elastin, with the exception of the concentration of some polar amino acid residues. N-terminal analyses indicated a very low level of polypeptide chain damage in the preparations. Upon mild alkaline treatment glycine was selectively liberated as amino-end group. Elastin from ligamentum nuchae was examined by electron microscopy, X-ray diffraction, optical polarisation analysis and dynamometry. It is suggested that elastin fibrils consist of a lateral array of primary filaments. Section B: The microfibrillar component of adult bovine ligamentum nuchae was isolated by solubilisation in guanidine-dithiothreitol followed by S-carboxymethylation and treatment with collagenase. The amino acid composition of the material was at variance to those previously reported for similar preparations. N-terminal analysis revealed glycine as the only end-group, at a concentration of 65.9 moles/106g of protein. This value, corresponding to a molecular weight of about 15,000 daltons, was in good agreement with the results of sedimentation equilibrium analyses.

No description available.

No description available.

Xylose, a sugar not metabolised by brain, and glucose may be transported into brain by a common mechanism (Gilbert, 1965). In the present studies the effects of some anticonvulsant drugs on the uptake of xylose by the non-raffinose compartment of cerebral cortex slices have been examined. The slices were pre-incubated for 30 minutes in oxygenated bicarbonate media, containing pyruvate as nutrient, and raffinose, before transfer to similar media containing xylose. Employing an incubation time of 9 minutes, and a xylose concentration in the medium of 50 mM, it was found that the anticonvulsant drugs acetazolamide, 20 μM, and ethosuximide, 500 μM, caused an increase in xylose uptake, while diphenylhydantoin, 100μM, was without effect in this respect,-SH group blocking agents (5,5- dithiobis (2~nitobenzoic acid), 100μM, or iodoacetamide, 100 μM), did not appear to affect 'basal' xylose uptake by the slices, but prevented stimulation of xylose uptake by acetazolamide or ethosuximide. Studies of the initial velocity of xylose uptake by the slices, over a range of medium xylose concentrations, indicated that the xylose uptake process appeared to conform to Michaelis-Menten kinetics; the apparent Km (under control conditions) for xylose being 87 mM, and the Vmax. being 30 millimole, 1iter of intracellular water-1minute-1. Acetazolamide increased both the Km and Vmax. of the xylose transport process. Both these parameters were decreased by ethosuximide. The effect of phenobarbitone on the kinetics of xylose uptake by cerebral cortex slices has been found (Gilbert, Ortiz, and Millichap, 1966) to resemble that recorded here for ethosuximide. The anticonvulsants which have been shown to be capable of increasing xylose uptake by brain slices also increased brain glucose content 'in vivo' (Gilbert, Gray, and Heaton, 1971), - probably as a consequence of a stimulation of glucose transport into brain rather than a depression of cerebral metabolism. The uptake of glucose may, under certain conditions, be the rate - limiting factor in its metabolism by brain (Joanny, Gorriol, a Hillman, 1969; Buschiazzo, Terrell, and Regen, 1970), Glucose may also have a direct stabilizing effect on brain cells (Goldman and Good, 1969)* The stimulation of sugar uptake 'brought about by the drugs might thus contribute to their anticonvulsant effect However, the interaction of these drugs with cerebral membranes may, in addition to affecting permeability to sugars, result in other effects more directly responsible for their anticonvulsant efficacy.

Nitric oxide is a recognised dilator of vascular smooth muscle and therefore is central in the control of blood flow. A lack of blood flow in humans can have very important implications in a number of disorders of both cutaneous tissue and internal circulation. In this thesis we look at the synthesis of new nitric oxide donors, their stabilities and their possible medicinal usage. These donors have been based on the S-nitrosothiol group, connected to sugar moieties, simple amino acids or linked glycoaminoacids. The donors prepared have been used to investigate the skin blood flow and localised responses to nitric oxide, proving that NO has an important role in the maintenance of healthy skin. These will be further investigated as possible treatments for disorders involving a lack of cutaneous blood flow, such as connective tissue disorders and the repeated ulceration often seen in diabetic patients. A set of clinical trials have been carried out comparing the responses of healthy patients and sufferers of Raynaud's Phenomenon to exogenous nitric oxide. In this we have highlighted a number of differences and have helped to determine a possible cause of the disorder. We have prepared a number of slow release NO donors which have been shown to produce a sustained vasodilatory response in blood vessels with removed or damaged endothelial cells. These show promise for use in the treatment of patients with circulatory disorders, especially for subjects following treatment for atherosclerosis. Basic studies investigating the stabilities of these compounds have been carried out, in order to aid our understanding of their mode of breakdown.

1) Transverse hippocampal slices were prepared from rats and maintained in a recording chamber. Intracellular recording techniques were used to measure the amplitudes of the resting potentials (mean -63.4mV), the action potentials (mean 72.5mV), and the input resistance (mean 16.5M?) of neurones in the CA1 region of the hippocampus. Impalements of neurones could be maintained for up to 9.5hrs. 2) Impaled neurones in the CA1 region of the hippocampus were identified as pyramidal neurones after injection of the fluorescent dye Lucifer Yellow CH and subsequent microscopical examination of the slices. The projections of the apical and basal dendrites were complete and in some cases the axon could be identified and traced toward the subiculum. Injections of pyramidal neurones in the CA3 region and of granule cells in the area dentata demonstrated the different morphologies of the three types of neurone. 3) CA1 neurones were hyperpolarised by up to 10mV by serotonin applied locally by iontophoresis. This response was slow on onset and lasted between 30s and 3min. In contrast, glutamic acid and acetylcholine depolarised these neurones. 4) The amplitude of the hyperpolarisation induced by serotonin was dependent on the amplitude and duration of the iontophoresis current. A decreased input resistance of the neurones was associated with and followed the same time course as the potential change induced by serotonin. 5) The serotonin response was reversed at membrane potentials more negative than the reversal potential which was between -81mV and -104mV. This high reversal potential indicated that the response might be mediated by efflux of potassium ions from the neurones. 6) In ion substitution experiments, the serotonin response was shown to be dependent on the extracellular potassium ion concentration, but was independent of the chloride ion concentration. 7) The serotonin response was blocked by the putative serotonergic antagonist methysergide (100μM). However, the responses of the neurones to serotonin were not affected by cyproheptadine, methergoline, mianserin, and 1-propranolol which have also been proposed to be antagonists of serotonin in the central nervous system. 8) The significance of these results is discussed with reference to previous investigations by other workers.

Tumour-associated arteries are a potential target for therapeutic strategies aimed at reducing or stopping tumour growth and metastases. However, the tumour vasculature exists in a state of 'near maximal vasodilation' and is markedly hyporesponsive to vasoactive agents generally. The role of nitric oxide (NO) in the growth and maintenance of solid tumours is unclear. Recent evidence suggests that it may be responsible for some of the characteristics of angiogenically- derived vessels found within the tumour and also for those of 'normal' vessels which become incorporated into the surrounding host tissue. A marked hyporesponsiveness to vasoactive agents also characterises vessels from animals in endotoxin-induced shock. One aim of the present study then was to compare the properties of arteries which formerly supplied an implanted solid tumour (tumour epigastric artery, or TEA) with arteries taken from experimental animals (rat tail artery, or RTA) injected with bacterial lipopolysaccaride (LPS). The extent of endotoxaemia resulting from LPS treatment was assessed by (a) monitoring arterial blood pressure, using the tail cuff method; (b) measuring accumulated plasma nitrite and nitrate; (c) monitoring the development of hyporesponsiveness of RTAs to phenylephrine (PE); (d) detecting expression of inducible nitric oxide synthase (iNOS) by Western blot and immunocytochemical analysis; and (e) monitoring the loss of sensitivity of pre-contracted vessels from LPS-treated rats exposed to the vasodilator S-nitroso-N-acetylpenicillamine. RTAs from LPS-treated rats and TEAs were less sensitive to PE than control arteries. This difference was abolished by non-selective NOS inhibitors (L-NAME and L-NMMA) and by the isoform-selective inhibitor aminoguanidine (AG). Furthermore, the protein synthesis inhibitor cycloheximide also reversed the hyporesponsiveness to PE of both types of vessel. The 'restoring' effect of cycloheximide was abolished when given after indomethacin, a specific cyclooxygenase inhibitor. Chronic, oral administration of L-NAME or AG to tumour-bearing rats (via the drinking water) significantly slowed tumour growth. When L-NAME treatment was halted, tumour growth resumed at pre-L-NAME (control) rates. A single i.p. injection of LPS also impaired tumour growth: this was dependent upon the time of injection, with no effect seen 24hrs prior to tumour implantation and the maximum delaying effect at 11 days post-implantation. LPS potentiated the growth retarding effects of L-NAME but abolished those of AG. These results provide evidence for the involvement of iNOS-derived NO in regulating the tone of arteries supplying a solid tumour and also of those from animals in septic shock. They suggest that NO may assist tumour growth by helping to maintain an adequate flow of blood into the tumour and they highlight the L-arginine/NO pathway as a potential target for the design of improved therapeutic interventions.

Ethanol production by Zymomonas mobilis CP4 under chemostatic and continuous transient reactor. Ethanol production by Zymomonas mobilis CP4 under chemostat and continuous transient operation has been investigated. Under carbon limitation with simple chemostatic operation, glucose, fructose and sucrose were tested at 35° C. At 2%(w/v) carbon source medium, glucose was more efficiently utilised. Ethanol production values for glucose, fructose and sucrose were respectively; volumetric productivity 2.5, 1.8 and 2.5 g/1/h; conversion to ethanol efficiency 96, 88 and 76% at values of dilution rate of 0.35, 0.21 and 0.32 h-1 which represent approximately 85% of Dm. With 5%(w/v) glucose values of volumetric productivity of 5 g/l/h and conversion to ethanol efficiency of 85% were achieved. However, under nitrogen limitation with glucose 5% (w/v), approximately 25% of feed glucose medium was being washed out while conversion to ethanol efficiency was around 60% at a dilution rate of 0.17 h-1 Under alternating glucose concentrations of 2 and 5%(w/v) medium and at a fixed dilution rate of 0.2 h-1, continuous transient operation achieved values of ethanol conversion higher than 80%. However, both continuous transient (utilising high alternating glucose concentration of 8 and 11% and 8 and 16%(w/v)) and simple chemostatic operation running with 9.5 and 12%(w/v) glucose medium were less satisfactory as conversion efficiency values were as less satisfactory as conversion efficiency values were as low as 40 and 29% for the former and 48 and 45% for the 1atter. Pulsing the culture with mineral salts (NH+4, Mg++,K+) and ethanol, the culture showed no effect with mineral salts and a strong inhibitory effect on growth for ethanol.