The metabolism of 1- [2-(4-carbamoylphenoxy)ethylamino]-3-(2-methylphenoxy)propan-2-ol ('tolamolol'), a new beta-adrenoceptor blocking agent, has been studied in the mouse, rat, guinea pig, rabbit, dog and man. For these studies tolamolol was labelled with tritium and carbon-14. In all six species there was evidence for good absorption after oral administration, but the routes for excretion of radioactivity showed considerable variation between the species. The major route for excretion in mice, rats and guinea pigs was the faeces, but in rabbits and man the major route was the urine; the dog showed intermediate behaviour. Tolamolol was extensively metabolised, little unchanged drug being excreted. Two main metabolic pathways were identified. The most important pathway was hydroxylation of the 2-methylphenoxy ring, and this was the major biotransformation in all species except the dog. The second metabolic pathway, and the major route in the dog, was hydrolysis of the aromatic carbamoyl group. Significant hydrolysis also occurred in the rabbit but in other species this was a minor route. Glucuronide and sulphate conjugates of the hydroxylated metabolite were excreted in the urine of rabbits and man, and accounted for most of the excreted radioactivity in these two species. In the rat and guinea pig, and probably the mouse, conjugates were excreted in the bile rather than the urine, subsequently appearing in the faeces as the aglycones. The species difference in excretory routes is therefore due to differences in the extent of biliary excretion. A polar metabolite in dog urine has been tentatively identified as a sulphate ester, the aglycone being a secondary metabolite in which both aromatic hydroxylation and hydrolysis of the carbamoyl group have occurred. Cleavage of the alkyl chain did not occur to a significant extent in any of the species investigated.

This thesis is concerned with the search, on an empirical basis, for quantitative structure/activity relationships (QSARs) relating the drug activity of a given type of molecule to indices of its biochemical reactivity. The utility of such relationships lies in their ability both to ellucidate the mechanism of drug action, and to predict the drug activity of an untested molecule of the same type. Most of the reactivity indices used herein are electronic properties of the molecule, obtained from CNDO/2 MO calculations. In Chapter 1, the basis for using the form of QSAR known as the linear free energy relationship (LFER) to explain in vivo as well as in vitro drug activities is discussed. Section 1 of Chapter 2 is a review of the Huckel MO reactivity indices and their use in QSARs. Section 2 introduces SCF MO theory and the ZDO approximation. The types of index which may be calculated from SCF wavefunctions are discussed, along with their pharmacological applications. The indices used in the present work are propounded in Section 3. An MO and regression program (MOREG), which calculates these indices and uses them to produce QSARs, is described. MOREG calculations on series of antitumour phenyl triazenes and a-formylthiosemicarbazone (N)-heterocycles are described in Chapters 3 and 4, respectively. In Chapter 5, similar calculations are described for two series of CNS depressant 1,4-benzodiazepines. The extension of the CNDO/INDO program to a form permitting calculations on the antitumour PtCl2(NH3)2 analogues is described in Chapter 6.

The ability of a compound to induce the rat hepatic microsomal drug metabolizing enzymes was investigated using a series of commonly prescribed barbiturates. The biological half-life of these barbiturates was also determined in the rat using a sensitive gas-liquid chromatographic method. It was found that inductive ability depends on rate of metabolism of the compound and therefore on its chemical properties. Evidence is presented supporting the theory that ethanol is not a substrate of the enzyme system known to detoxify a diversity of compounds. The effect of chronic and acute ethanol administration on the mixed-function oxidase is investigated. The metabolic interaction of ethanol with barbiturates is studied both in vivo and in vitro. Ethanol is shown to potentiate the inductive effect of the barbiturates. Possible mechanisms of interaction are discussed. The interaction of barbiturates with imipramine is examined in vivo and imipramine is shown to inhibit the metabolism of pentobarbitone. The importance of the allyl group in the chemical induction of porphyria is considered using a variety of allyl group containing compounds. Porphyrogenicity of a compound is compared with its ability to induce drug metabolism. It is shown that compounds increasing the level of cytochrome P-450 in the microsomes do not necessarily induce delta-aminolaevulinic acid synthetase and vice versa. Finally the use of formic acid vapour in the carrier gas in gas liquid chromatography is critically considered. Formic acid is shown to reduce adsorption of polar compounds on stationary phase of the column and thus improves the limit of quantitation. Evidence is presented for possible mechanisms.

The present investigations were primarily aimed at defining the detoxicating capacity of the placenta, and then to a more general study of steroid and fatty acid metabolism. Concomitant studies were undertaken with foetal and maternal liver, plus certain other steroidogenic organs. Cytochrome P-450 was demonstrated both in the mitochondrial and microsomal fractions of human and rabbit placentae, but associated only with the microsomes in rat placenta. Evidence for rat and ferret testicular cytochrome P-450 has been presented. The placental CO-binding pigment responded similarly to that of maternal liver with regard to storage, deoxycholate treatment and the capacity of the reduced pigment to bind metyrapone. It did not prove possible to elicit binding spectra with various endogenous and foreign substrates normally metabolized by the hepatic microsomal mixed function oxidase system. Whereas enzyme inducing agents elevated foetal and maternal hepatic cytochrome P-450, they did not significantly alter such levels in the placenta. The ability of placentae from humans, rats, rabbits and pigs to metabolize a variety of anutrients, including hydroxylation of biphenyl, benzo[a]pyrene and zoxazolamine, N-demethylation of ethylmorphine and p-chloro-N-methylaniline, O-demethylation of p-nitroanisole, reduction of p-nitrobenzoic acid, hydrolysis of phenolphthalein glucuronide and conjugation of 4-methylumbelliferone, was examined. This activity was either not detectable or present at a rate no greater than 30% of that maternal liver (excluding B-glucuronidase activity). Foetal liver and certain steroidogenic organs studied exhibited comparable drug-metabolizing activity to that of placenta. The routes of detoxication in the placenta were usually distinct from those of maternal liver, and were not mediated through cytochrome P-450. Pretreatment of pregnant animals with enzyme inducing agents resulted in a proliferation of endoplasmic reticulum and drug-metabolizing activity of the maternal liver, but in the placenta the response was comparatively very small. The detoxicating capacity of the maternal liver did not appear markedly reduced during pregnancy. Unlike placentae of rats and rabbits, those of humans and pigs demonstrated a steroid ring-A aromatizing system involved in oestrogen biosynthesis. In the human placenta, the activity was largely associated with the microsomes. Nicotinamide was required in the homogenizing media for optimal activity, and in certain cases there was a need for Mg[++]. The multienzyme complex proved resistant to solubilization by treatment with deoxycholate, Triton WR-1339, papain, lipase and sonication. No inhibition of activity was observed in the presence of up to 90% CO with O[2] maintained at 5%. The studies strongly indicated that the steroid ring-A aromatizating system studied in the placenta was not mediated by a cytochrome P-450 mixed function oxidase system. Incubations in the presence of a variety of steroids, fatty acids and xenobiotics characterized the enzymes involved. The placentae of rats, rabbits and humans were found not to o-hydroxylate lauric acid, unlike the enzyme system of liver which is capable of oxygenating drugs and steroids also.

Carbenoxolone has been labelled with [14]C and [3]H. [[14]C]Carbenoxolone administered as a positioned-release capsule to duodenal ulcer patients was shown to be readily absorbed. The fate of [[14]C]- and [[3]H]carbenoxolone administered orally to human volunteers has been investigated. Most of the dose was recovered as carbenoxolone in the faeces, although a portion (< 15%) was present as carbenoxolone-30-glucuronide. Peak plasma concentrations indicated at least 50% absorption of the dose. Less than 10% of the dose underwent hydrolysis to enoxolone and succinate, in contrast to the rat where extensive hydrolysis occurred after oral administration. [1,4-[14]C[2]] Succinate was rapidly and almost completely metabolised to respiratory [14]CO[2] in a human volunteer. No significant hydrolysis of [[14]C]carbenoxolone occurred after administration to intact or biliary-cannulated squirrel monkeys. Carbenoxolone was excreted in the bile mainly as carbenoxolone-30-glucuronid Conjugation appeared to be a pre-requisite for biliary excretion and conjugates were excreted against a concentration gradient. A species difference in metabolism between primates (man and squirrel monkey) and the rat was attributed to the gastrointestinal microflora. Carbenoxolone has been found to be highly bound to the plasma proteins of man and common laboratory species. The binding was investigated by a variety of methods to obtain quantitative and qualitative data on the interaction. Albumin had a very high affinity (k > 10[7]) for carbenoxolone and was found to be the principal binding protein in human plasma, although binding to globulins also occurred; carbenoxolone also induced a conformational change in the albumin. The distribution of [[14]C]carbenoxolone in the rat showed that it was confined largely to the gastrointestinal tract, liver and plasma. Pretreatment of rats with carbenoxolone had no significant effect on various hepatic microsomal enzymes concerned with the metabolism of drugs and steroids. Neither did such pretreatment influence the biliary and urinary excretion of [[3]H] cortisol and its metabolites.

The development of JAFS, a new computational method to study the binding geometries of small fragment molecules to protein cavities, estimate their binding affnities and analyse how they compete for a common protein binding site, all in the context of Fragment Based Drug Discovery, is presented in this thesis. Fragment Based Drug Discovery is an approach to drug development which studies the binding of small ligands (fragments) forming high quality interactions with their target. Further optimization of these fragments into drug-like molecules, adding functionalities to increase affnity while controlling other relevant properties such as toxicity and absorption then takes place. JAFS studies the binding of fragments to their target proteins. The JAFS method consists of the execution and analysis of Monte Carlo simu-lations of fragments (and waters) in the binding cavities of proteins with an added degree of freedom which accounts for the scaling of the interaction energy of the fragment (and water). Sampling of states at very low interaction energies gives a boost in fragment con?gurational sampling while competition between di?erent fragments to remain at unscaled (high) interaction energies at a given binding site provides information on their relative binding a?nities. JAFS is built on the JAWS formulation for water binding to protein cavities. The performance of the JAFS method on a range of different test cases (T4 Lyzozyme, Major Urinary Protein I, Cyclin Dependent Kinase 2 and Heat Shock Protein 90) was studied. JAFS is divided in two protocols to rank fragments by affnity and locate binding geometries, respectively. The ranking of fragments by affnity to a common protein target was satisfactory (as compared to experimen-tal data) for the simpler systems (T4 Lyzozyme and Major Urinary Protein I). However, more demanding systems proved problematic, where the ranking of nine different ligands to the binding site of Cyclin Dependent Kinase 2 provided results unrelated to experimental binding affnities. Studying pose generation in sets of five repeats per simulation, the crystal binding geometry of every fragment studied was found in at least one of the re-peats, without providing any previous information on the system (such as the presence or location of water mediated interactions or the hydration state of the cavity). Consistency between repeats was however found to be problematic and no method is currently able to select the optimal binding geometry among all the gen-erated poses. Suggestions are given for further developments which would provide a methodology to rank poses.

This thesis explores online medicine purchasing and provides insight into how people account for this activity via the application of the concept of respectable deviance. Drawing together established deviance theories; respectable deviance considers the construction of online medicine purchasing, the justifications presented to challenge how it is labelled, and how the behaviour is managed. Those purchasing medicine online are not necessarily criminalised, however, the behaviour has been constructed as risky. This is because people can buy medicines that traditionally require prescription from registered practitioners. These new opportunities to purchase illicit medicines have implications for the pharmaceutical marketplace, regulation and governance, and healthcare expertise. The specific risks associated with online medicine purchasing, namely counterfeit medicines, criminal activity, and health implications, merge with the challenges to the marketplace, the challenges to regulation and governance, and the challenges to healthcare expertise. People purchasing medicines online acknowledge these ‘risks’, and redefine them in terms of justifications. Utilising an interpretivist mixed- methods study encompassing forum observations, online survey, and interviews, this research allows an understanding into how those engaging in‘risky’ behaviour breaking with accustomed practices (i.e. purchasing prescription/unauthorised medicine online), manage their performances with techniques of neutralizations, specifically challenging governance and medical expertise. At the same time, as the Web provides a space for deviance, it also provides a space for people to manage how their actions are perceived. Respectable deviance highlights how people respond to the unique risks and opportunities afforded in online medicine purchasing.

Lipidic formulations (LFs) are increasingly utilised for the delivery of poorly–water soluble drugs to improve oral bioavailability. In vitro lipolysis is capable of mimicking the lipid digestion process and therefore it is a suitable method for assessing the fate of drugs administered in LFs. Intestinal micellar solubilisation and first–pass metabolism are the main contributors to the oral bioavailability of drugs that belong to class II of the Biopharmaceutics Classification System (BCS). The intraluminal solubility of BCS II drugs in LFs can be estimated with the in vitro lipolysis model, whereas the first–pass extraction ratio can be assessed by performing microsomal stability assays. This thesis work proposes, for the first time, the combination of in vitro lipolysis and microsomal metabolism studies for the quantitative prediction of human oral bioavailability of BCS II drugs administered in LFs. Marinol® (Δ9–tetrahydrocannabinol dissolved in sesame oil) and Neoral® (a lipidic self–emulsifying drug delivery system of cyclosporin A), were selected as model LFs. The observed oral bioavailability (Fobserved) values were obtained from published clinical studies that described the oral administration of the selected LFs to human subjects. Two different lipolysis buffers, differing in the level of surfactant concentrations, were used for digestion of the LFs. The predicted fraction of absorbed dose (Fabs) was calculated by measuring the drug concentration in the micellar phase, obtained after ultra–centrifugation of the lipolysis medium. To determine the fraction of drug dose that escapes metabolism in the gut wall and in the liver (Fg∙Fh), microsomal metabolism stability studies with human intestinal and hepatic microsomes were performed. Clearance values were determined by applying the “in vitro half–life approach”, which is based on the measurement of the first–order rate depletion constant of a drug substrate. The estimated Fabs and Fg∙Fh values were combined for the calculation of the predicted oral bioavailability (Fpredicted). For the model LFs tested, results showed there was a correlation between Fobserved and Fpredicted values only when Fabs was calculated with the buffer characterised by more bio–relevant (lower) surfactant levels. The general accuracy of the predicted values, and the strong correlation shown with the clinical ones, suggests the novel in vitro lipolysis/metabolism approach could satisfactory quantitatively estimate the oral bioavailability of BCS II drugs administered in LFs.

Sustained release microparticles used for parenteral drug delivery must be well characterized in terms of their size range, morphology and function. It is widely understood that the chemistry and morphology of microparticles have a degree of interdependence which strongly affects drug release behaviour from microparticles. This thesis investigates, for the first time, the use of mid-IR imaging along with the development and optimisation of relevant multivariate image analysis methods for studying the real-time degradation of pharmaceutically relevant biodegradable polymer microparticles and the real-time release of protein based drugs from such microparticle systems. The application of attenuated total reflection - Fourier transform infrared spectroscopic(ATR-FTIR) imaging and analysis to monitor the degradation of a single microparticle is optimised and the developed methodology is detailed. A series of time resolved images of a PLGA microparticle undergoing hydrolysis at 70 °C are obtained using ATR-FTIR imaging for the first time. A novel partially supervised non-linear curve fitting (NLCF) tool is developed and the output from the NLCF is evaluated by direct quantitative comparison with a traditional peak height (PH) data analysis approach and multivariate curve resolution alternating least squares (MCR-ALS) analysis for the same images, in order to develop an image analysis strategy. The NLCF method is shown to facilitate the calculation of hydrolysis rate constants for both the glycolic (kG)and lactic (kL) segments of the PLGA copolymer. This results in improved spatial resolution on time-resolved microparticle images, so providing better insight into the dimensions of hydration layers and particle dimension changes during hydrolysis when compared to images derived from both PH measurements and MCR-ALS. The MCRALS routine is shown to be faster than NLCF and its images are found to provide sufficient contrast to be used for qualitative comparison. The optimised mid-IR-ATR procedures are then applied to investigate several factors influencing the hydrolytic degradation of a family of PLGA microparticles. Degradation rate constants for glycolic and lactic units are shown to increase (whilist maintaining a ~1.3 ratio between each other) with increasing initial glycolic content of the copolymer,temperature or γ-radiation exposure. Differential scanning calorimetry (DSC) and gel permeation chromatography (GPC) results indicate a chain scission based degradation in PLGA upon γ exposure. The distribution of lactic acid is probed with IR during the hydrolysis of a PLA microparticle for the first time, showing a diffusional pathway from the degrading microparticle outwards into surrounding water. Utilising the chemical selectivity of the infrared methodology, ATR-FTIR imaging is applied for the first time to monitor the redistribution and release of human growth hormone (hGH) from a range of CriticalMixTM processed PLGA/PLA microparticles during a set of dissolution experiments at 37 °C in D2O. Increasing the γ dose is shown to have a profound influence on the release mechanism, with higher γ doses leading to a dramatic increase in the initial burst release followed by retardation in the sustained release and a lower total level of hGH release over the dissolution experiment. These changes are shown to be the result of: (i) protein aggregation as a function of applied γ-dose as studied by size exclusion chromatography; (ii) decrease in overall porosity as studied by SEM; (iii) decrease in Mw of all of the component polymers post γ irradiation indicating a chain scission mechanism as studied by GPC and DSC; and (iv) the increase in the number of oxygenated components in the Poloxamer 407 excipient, thereby increasing the strength of interaction between the microparticle and the entrapped hGH. These findings suggest that any γ sterilisation dose should be less than 25 kGy and that other sterilisation methods may need to be considered, due to the stability of the studied formulations.

In recent years, dry granulation using roll compaction (DGRC) attracts considerable interest of engineers and researchers, especially in the pharmaceutical industry, due to its distinct feature that no liquid binder is needed. It is generally anticipated that as a size enlarge process, DGRC would improve properties of feed powders (such as flowability and bulk density), but it was also reported that DGRC could cause a reduction in powder compactibility. A wide range of powder properties, such as size, shape, flowability, compactibility and compressibility, were analysed for several pharmaceutical excipients using the state of art techniques. Elastic-plastic properties of single component powders and mixtures were also determined using the Drucker Prager Cap (DPC) model and an example of FEM application was presented. All the properties determined were used to investigate: 1) the prediction of ribbon milling from friability tests and 2) the effect of granule size on die filling and die compaction behaviour of pharmaceutical powders. A new and easy method was developed for predicting fines produced during ribbon milling. An exponential relation between the filling ratio and the shoe speed was found. Furthermore, it is shown that flowability is strongly influenced by the granule size, and there is a decrease in the tensile strength with the increase of the granule size. Additionally, for all the materials analysed a strong correlation between the flow indexes and the critical filling speed was observed and an empirical equation is obtained.