The last two decades have seen an increased drive to administer parenteral nutrition (PN) to patients in their home environments, thereby reducing associated hospital costs and improving patient quality of life. The occurrence of deranged nutritional biochemistry results has baffled PN experts for years because PN additives are marketed for the general needs of patients and PN is tailored to each patient’s requirements (both formulation and regimen). This thesis documents the investigations into HPN population characteristics, the extent of nutritional abnormalities (deficiencies and excesses) in a cohort of LT PN patients in Wales. Both cross-sectional and longitudinal retrospective study designs were employed alongside small-scale laboratory efforts to investigate stability of vitamin D in PN additives using High Performance Liquid Chromatography (HPLC). Characteristics of the HPN population in Wales were shown to be variable in terms of PN requirements for a predominantly female sample population (2:1); in whom 78.6% of patients received PN for indications relating to short bowel syndrome (SBS). A database analysis of micronutrient test results revealed a high prevalence of deficiencies of vitamin D and selenium, as well as excesses of manganese and water-soluble vitamins; which can lead to clinically relevant effects in patients. The sample population was shown to have impaired bone health since first receiving PN; respective sites of the femoral neck and total hip presented 58% and 60.8% of patients had osteopenia, while 28% and 19.6% had osteoporosis. Evidence in the literature links these clinical outcomes of metabolic bone disease (MBD) to patients’ inadequate vitamin D status. A final study exploring the adequacy of the trace element (TE) preparation Additrace®, found it lacking in selenium and excessive in manganese for the general requirements of the PN population. Clinician-directed supplementation of PN outside of Additrace® was associated with better micronutrient status in patients and more test results within range.

For the past two decades, the development of protein therapeutics has significantly expanded with numerous biopharmaceutical and biosimilar products entering the medicine market every year, and even more queuing in the pipeline globally. Biologics are very complex molecules and therefore extremely sensitive to minor changes in the manufacturing process, which can result in heterogeneity and affect the stability, potency and immunogenicity of the final product. Public health organisations, such as EMA (European Medicines Agency), require that biological products should be extensively tested for their similarity to the original drug (in the case of a biosimilar) as well as to products from different batches (batch-to-batch comparison). The issued guidelines focus, among other tests, on physicochemical characterisation of these molecules. The suggested analytical techniques, however, are only vaguely named in the specifications, leaving the final decision to the manufacturers. The present work focuses on the use of different combinations of analytical techniques with an aim to demonstrate similarity or dissimilarity between two or more samples. The selected instrumental techniques are characterised by their simplicity and are able to detect structural differences and microheterogeneity of the active ingredient in different samples, aggregation, degradation and post-translational modifications (PTMs). Seven studies were completed in total, each one to a different extent, and these included protein therapeutics such as insulin and monoclonal antibodies. The applied techniques served for primary (MS),* secondary (far-UV CD, FT-IR) and tertiary structure (near-UV CD, fluorescence) comparison of the examined samples. Particle size comparability and detection of aggregation was achieved with DLS, and higher-order structure comparison with 1D 1 H-NMR. Coupling of the techniques with temperature-dependent measurements enabled further comparison on the thermal stability of the samples and provided confidence in the observed (at room temperature) results. The acquired empirical experience pointed out the advantages and disadvantages of each technique compared to the rest of the techniques, possible solutions to the encountered challenges, and the cases that one technique can be used instead of another or as complementary to it. Finally, a potential SOP is suggested, advising on which biophysical techniques should be used depending on the structure of the protein that is examined and its formulation.

Malaysia’s rainforests have a great biodiversity, a lot of which has yet to be studied. This presents researchers with the opportunity for the discovery of new chemical structures which may be new leads in drug discovery. With problems such as antibiotic resistance and the need for improved drugs for the treatment of diseases such as cancer and neurodegenerative diseases, there is need for more research into drug discovery. The focus of the research covered in this document is on Pericampylus glaucus, a common climbing plant for which there is limited literature available. The aim of this study is to determine its pharmacological activities, based on what is known of its traditional use, in the hope that it can be used in the development of pharmaceutical drugs in the future. The literature review of the traditional uses of P. glaucus suggests that the plant may be used in ethnomedicine for its anti-inflammatory and/or antibacterial activity as it is used to treat many conditions in which inflammation and infection is indicated. Components of P. glaucus extracts were investigated through a variety of phytochemical screening methods and were found to include polyphenols such as tannins and flavonoids, terpenes, sterols, alkaloids and saponins and similarities between the different extracts were observed. P. glaucus was found to possess antioxidant activity, with the stem chloroform extract possessing the greatest radical scavenging activity. The antibacterial assays revealed only mild antibacterial activity by P. glaucus extracts, making the use of the plant for treating infections seem unlikely, although this may be due to activity against specific strains of bacteria. The anti-inflammatory assays suggested a number of different mechanisms by which P. glaucus could exert anti-inflammatory activity, with many of the extracts and known compounds inhibiting COX-1 and COX-2 enzymes, and only the fruit hexane extract providing less than 50% inhibition of COX-1 and COX-2 at 500 μg/mL. The hexane extracts generally prevented protein denaturation to a greater extent compared to other extracts, as well as palmitic and stearic acid. The extracts showed a varying ability to prevent lysis of red blood cells, which were used as a model for lysosomes, depending on whether lysis was induced by hypotonicity or by heat. Some anticancer activity, cytotoxicity and growth promoting activity was recorded when cells were cultured with P. glaucus extracts. The ethanol extracts were found to exert the greatest cytotoxic activity, with the exception of the fruit ethanol extract. The root ethanol extract was the most toxic towards HK1 and MRC-5, while the stem hexane extract increased HK1 proliferation by over 200% and the root hexane extract increased MRC-5 proliferation by over 600%. Because of the growth promoting activity that was observed and the traditional use of P. glaucus for the treatment of wounds, the wound healing activity of the extracts was tested and found to be much less effective than the control of 10% FBS, although the greatest wound healing activity of around 20% wound coverage after 24 h was seen by the fruit extracts. Since P. glaucus is traditionally used for the treatment of venomous snake bites, its antivenom activity was assessed and found to have some antivenom activity, with the fruit hexane extract increasing cell proliferation by over 50% at 200 μg/mL and the alkaloid, periglaucine A, increasing proliferation by over 70% demonstrating significant antivenom activity. The antioxidant activity is most likely attributable to the polyphenols as the total polyphenol content correlated with the results of the DPPH radical scavenging activity of the extracts. The mild antibacterial activity suggested that P. glaucus may be used more for its anti-inflammatory activity in traditional medicine rather than antibacterial activity. The findings of toxicity towards cancer cells supports the traditional use of P. glaucus for the treatment of cancer. The results of the wound healing activity would indicate that P. glaucus would have a mild effect in the treatment of wounds due to its ability to promote cell proliferation and migration, however, its anti-inflammatory activity and mild antibacterial activity may be beneficial in treating wounds. It did possess antivenom activity, which supports its use for the treatment of envenomation by snakes, although the fruit extracts had stronger antivenom activity compared to the root, which is the plant part traditionally used. The mild antibacterial activity of P. glaucus may partly contribute to its medicinal effects, however, this study reported stronger evidence of anti-inflammatory, cytotoxic and antivenom activity which may be the properties that practitioners of traditional medicine have found valuable when selecting this plant for treatment. The pharmacological activities reported support a number of the traditional uses for the plant, P. glaucus, suggesting that the traditional use may have a scientific ground and could be investigated further, possibly leading to the discovery or development of new anti-inflammatory, chemotherapeutic or antivenom drugs.

Cancer is one of the leading causes of death worldwide with approximately 14 million new cases reported in 2012. Systemic chemotherapy is widely used to treat cancer and can often result in toxicity and adverse side effects. To reduce these adverse side effects and potentially increase efficacy, the focus of this thesis is to create a new intratumoural drug delivery platform for patients who are too ill to receive systemic chemotherapy or with inoperable tumours. Herein, an injectable formulation composed of a hydrogel and drug-loaded polymeric nanocapsules (NCs) was developed, to deliver the chemotherapeutic locally within the tumour and hence reduce the adverse side effects. The hydrogel is used to control the release of the NCs within the tumour, while small NCs (< 100 nm) aim to promote NC penetration throughout the tumour mass. Self-emulsification, a low energy and solvent-free method, was used to produce the polymeric NCs. The lipid core of the NCs was suitable for encapsulation of anticancer drugs: docetaxel (DCX) and 4N-myristoyl-gemcitabine (GEM C14). NCs with diameters of 40 and 80 nm and with zeta potential values of ~ -4 ± 1 to –19 ±1 mV were produced. Both anticancer drugs, DCX and GEM C14 were encapsulated with efficiencies of greater than 90%. GEM C14 was released from the NCs by diffusion with 63 ± 8 and 66 ± 10 % released in 30 days from 40 and 80 nm NCs respectively. Additionally, GEM C14-loaded NCs showed activity against various cancer cell lines in vitro where the following GI50 values were obtained: 15 ± 6, 10 ± 9, 13 ± 3 nM in HCT 116, MIA PaCa-2 and Panc-1 cell lines respectively. Although not as potent in a GEM-resistant pancreatic cancer cell line (Panc-1 GEM resistant), GEM C14 NCs (GI50= 410 ± 463 nM) were significantly more active than GEM C14 alone (GI50=8300 ± 5523 nM). Hydrogels were prepared using a low molecular weight (LMW) gelator - N4-octanoyl-2’-deoxycytidine. This amphiphilic molecule self-assembles in water forming a 3D nanofibre network with the ability to reform after the application of stress, making it suitable for injection through a syringe/needle. N4-octanoyl-2’-deoxycytidine was used to gel the NC suspension via a heat-cooling cycle. The strength of gel-NC composite was determined by oscillatory rheology and its 3D nanofibre structure containing spherical NCs was confirmed by TEM. The gel-NCs formulation reformed instantly into a gel after injection through the needle and formed a depot when injected in an in vitro simulation of intratumoural delivery i.e. chicken tissue. In summary, a novel NCs-gel composite has been designed demonstrating appropriate characteristics for intratumoural drug delivery.

VanS is an integral membrane protein, and a receptor histidine kinase representing on half of the VanSR two component system. In the presence of vancomycin, as well as certain other antibiotics, VanS undergoes autophosphorylation at a conserved histidine residue and then transfers this phosphoryl group to a conserved aspartate on VanR. The phosphorylated form of VanR binds to the promoter region of the vanHAX operon, inducing expression of these genes, which confer glycopeptide antibiotic resistance to the cell. Although the vancomycin resistance mechanism is mostly well-characterised, questions surrounding the structure and mode of activation of VanS remain unanswered. This thesis describes work towards the structural and ligand-binding characterisation of the extracellular \sensor" domain of VanS. Little is known of the structure of this domain, and while there exists evidence in the literature to support numerous theories surrounding the identity of the VanS ligand, no report has yet been made of a direct observation of a binding event involving any VanS protein and its ligand To identify the ligand to VanS, the catalytically active, full-length VanS proteins of Enterococcus faecium (VanSA) and Streptomyces coelicolor (VanSSC) were heterologously expressed, solubilised and purified and binding studies were carried out by solution-state NMR. We present evidence of interactions between vancomycin and both VanS proteins, indicating that direct binding of vancomycin may be the mechanism by which both proteins are activated. The VanSSC extracellular domain was isolated as a synthetic peptide, its structure characterised, and the binding interface between this domain and vancomycin was investigated. Evidence of binding at the N-terminal end of the extracellular domain, in agreement with the findings of Koteva et al. (2010), is presented here. An interaction between vancomycin and DPC detergent micelles was also observed and characterised, and the biological significance of such an interaction is discussed.

續斷傳統上具補肝腎強筋骨功效，為中醫骨傷科、婦科之常用要藥，臨床上除可用於腰膝酸軟、跌打損傷、骨折等症，亦用於治療胎漏、胎動不安、滑胎，常用於孕婦安胎之用。現代藥理研究亦表明續斷具有抗骨質疏作用、促進骨損傷癒合、降低在體和離體子宮的收縮幅度及緊張度、具免疫調節活性、具抗氧化抗衰老活性、保護神經元細胞作用、抗阿茲海默症作用、抗菌抗炎、抗腫瘤活性、對機體不同器官病變的保護作用、改善糖尿病等作用。 觀乎目前大部分中藥材的藥理研究，對於實驗所使用之中藥材往往只由個別學者進行經驗鑒別，並沒有對其進行嚴謹的鑒別。過往曾有學者提出藥食兩用的枸杞子含有毒性的阿托品類生物鹼的結論，令中藥界震驚並憂慮枸杞子的安全性；後來不少學者提出該學者的研究所指的“枸杞子”並不一定與中藥所使用的枸杞子品種一樣。由此可見，品種鑒別對中藥的藥理研究甚為重要，差若毫釐，缪以千里，使用不正確的原料作研究對像，有可能出現矛盾的結果，亦錯誤評價中藥的療效性、安全性等方面，窒礙中藥日後的研究發展。 續斷，在過往《中國藥典》中曾多次修訂藥材來源以及藥材原植物拉丁名，至使過往一些對續斷的藥理研究或综述可能並未包含真正的續斷，或取用了非續斷原藥材的植物進行研究及分析。目前香港並沒有法定的中藥材標準，大部份研究均是參考《中國藥典》，而2005年香港衛生署出版的《香港中藥材標準》則屬於指導性質，並無實際法律效力，只屬參考性質，導致研究人員往往從供應商採購時不一定能確保待研究藥材的質量。因此本研究認為在進行藥理研究以前，需要先為待研究藥材進行定性定量的鑒別檢測，以確保待研究藥材的質量符合標準。 目的：本研究通過比較《中國藥典》（2010年版)及《香港中藥材標準》（第五冊)中所採用的鑒別方法對續斷進行鑒別，觀察兩種標準在鑒別及定量層面上的異同，從而評價品種鑒別對藥理研究的重要性。 方法：根據《香港中藥材標準》，對續斷藥材進行的顯微鑒別(粉末、切片)方法及HPLC法檢測川續斷皂苷VI含量，並進行比對。 結果及結論：《中國藥典》及《香港中藥材標準》的顯微鑒別(粉末及切片)的要求及顯微鑒別特徵物相符；而HPLC法檢測，《中國藥典》與《香港中藥材標準》所使用的HPLC條件及操作方法略有不同，但含量標準均為以乾燥品計算，川續斷皂苷VI(C47H76O18)不少於2.0%。而藥材樣品含川續斷皂苷VI百分含量(%)為4.58%，均符合《中國藥典》原藥材及飮片，以及《香港中藥材標準》的含量要求。然而，《香港中藥材標準》較明確要求色譜系統適用性參數，並對成分峰的分離度有指明要求，在操作上能較容易理解，較適合於藥學研究者標準地執行藥材定性定量鑒别相關操作。本研究分別根據《中國藥典》及《香港中藥材標準》之鑒別方法對續斷進行鑒別及定性定量分析，比較兩個標準上在定性鑒別及定量分析層面上的異同，為日後續斷的藥理研究的藥材定性定量鑒别提供客觀依據，為合理評價續斷的藥效性、安全性研究做了有意義的基礎。 關鍵詞：續斷；中藥研究；鑒別；質量標準；比較

Dissertation submitted in partial compliance with the requirements of the Masters Degree in Technology: Homoeopathy, Durban Institute of Technology, 2003. / The purpose of this randomised double-blind placebo-controlled study was to evaluate the efficacy of Oioscorea vil/osa cream in the treatment of menopausal syndrome in terms of subjective and objective data; and to compare the subjective data with that obtained from a concurrent study of ProgestoNat\xAE cream (McTeer, 2003). / M

目的：國內中藥材品種混亂的現象普遍，而香港的中藥材大多是由國內進口，其中相當部分將轉口至世界各地，中藥材品種混亂的問題對本港乃至各地都有影響。本課題旨在對香港藥食兩用中藥材進行調研整理和鑒別研究。通過市場調查獲得香港市場流通過程中的藥食兩用的中藥材、中藥飲片品種的商品規格、價格、產地資訊。獲取藥食兩用中藥材和中藥飲片的代用品、偽劣品種和品種混淆的情況。通過品種考證鑒定香港市面流通中藥材混淆品種的基源。發現存在的問題並提出解決意見。 方法：通過實地走訪調查、詢問、填寫調查表等方式，對香港十八個行政區的中藥批發商、中藥零售企業、食品零食商進行調查，並拍攝被調查的藥材和飲片、替代品、混淆品的照片作為影像記錄。 結果：1）香港藥食兩用藥材價格差異大，質量參差不齊 2）中藥規格等級劃分混亂，價格透明度低。3）香港部份市售的中藥材小薊、枳椇子、金銀花、青果、昆布、薏苡仁、酸棗仁、鮮白茅根、枸杞子、山藥基原與藥典存在差異。4 ）根據大棗的歷史沿革，推斷案（大棗、黑棗）為鼠李科植物棗Ziziphus jujuba Mi11.的成熟果實。 討論：1）香港藥食兩用中藥材存在基原混亂、規格劃分不統一、質量不穩定等問題。2）為保證中藥材質量，建議《香港中藥材標準》強制實施。3）開展基礎研究，開發新資源，尋找中藥代用品。4）完善中醫藥相關法律法規，嚴格監控中藥生產、購進、銷售各環節。5）發展高質量、高附加值的中藥產業。 關鍵詞：藥食兩用、中藥材、調查整理、性狀鑒別。

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