The purpose of this investigation was to determine and characterise the anti-oxidant activity of the methanol extract of the leaves of Halleria lucida utilizing the DPPH (2,2-diphenyl-1-picrylhydrazyl) assay. The methanol extract of the leaves of H. lucida displayed promising anti-oxidant activity with an IC50 value of 8.49±0.12 μg/ml and was subsequently subjected to activity-guided fractionation resulting in the isolation of a flavone-type flavonoid and phenylpropanoid glycoside, namely luteolin-5-O-β-D-glucoside and verbascoside (acteoside), respectively. Both compounds displayed promising anti-oxidant activities with IC50 values of 6.12±0.40 and 7.18±0.08 μg/ml for luteolin-5-O-β-D-glucoside and verbascoside, respectively. Furthermore, isobologram construction was undertaken to determine pharmacological interactions between the isolated molecules resulting in a concentration-dependent additive and antagonistic interaction being recognised.

No description available.

The neuroprotective effects of AF4, a flavonoid-enriched extract derived from the peel of Northern Spy apples (containing quercetin-3-O-glucoside, quercetin-3-O-galactoside, quercetin-3-O-rhamnoside, quercetin-3-O-rutinoside, epicatechin, and cyanidin-3-O-galactoside) were examined by assessing neuronal loss and motor impairment resulting from hypoxic-ischemic (HI) brain injury in adult C57BL/6 mice. Relative to vehicle treatment (water, 10mL/kg/day), oral administration of AF4 (50 mg/kg/day) for 3 days reduces HI-induced neuronal loss in the striatum and hippocampus, motor impairments, and reduces the ability of LPS to stimulate the production of TNF-alpha in whole blood. Pretreatment with AF4 (1 ug/mL) decreased the death of mouse primary cortical neurons subjected to oxygen glucose deprivation (12 hours) in comparison to vehicle (DMSO) or the same concentration of quercetin or its metabolites. Taken together these findings indicate that AF4 reduces HI-induced brain injury and motor deficits by increasing the resistance of vulnerable neurons to ischemic cell death and decreasing the production of inflammatory cytokines.

The anthracycline quinone, doxorubicin (Adriamycin) is an antineoplastic agent that has substantial therapeutic activity against a broad variety of human cancers. Unfortunately, the use of this agent is limited by its cardiac toxicity, which is associated with free radical formation leading to apoptotic cell death. The goal of this work is to improve our understanding about doxorubicin induced cardiomyopathy and to identify compounds to limit doxorubicin induced cardiomyopathy. The knowledge gained here will have a generalized impact on all cardiac diseases involving oxidative stress and apoptosis. We show that doxorubicin induced apoptosis in primary neonatal rat cardiomyocytes can be attenuated by progesterone (PG). The anti-apoptotic action of PG was blocked by a progesterone receptor antagonist, Mifepristone (MF), indicating a progesterone receptor dependent pathyway. Affymetrix gene analyses found that PG treated cardiomyocytes increased the expression of 180 genes. Among the genes upregulated is NAD(P)H: Quinone Oxidoreductase-1 (NQO1) gene. NQO1 is a flavo-enzyme that can catalyze a two-electron reduction of Dox to a more stable hydroquinone, thereby acting as a defense mechanism against oxidative stress. The induction of NQO1 mRNA and NQO1 activity in cardiomyocytes was observed in a dose and time-dependent manner with PG treatment and was blocked by MF. Induction of NQO1 by b-naphoflavone, an inducer of NQO1, resulted in a decrease in caspase-3 activity. However, inhibition of NQO1 by dicoumarol did not attenuate the cytoprotective effect of PG. This data indicates that although induction of NQO1 can decrease Dox induced apoptosis, this is not the primary mechanism of cytoprotection induced by PG. Microarray analyses revealed that PG induced an increase of Bcl-XL mRNA. Inhibiting the expression of Bcl-XL using siRNA reduced the anti-apoptotic effect of PG, suggesting that Bcl-XL is a key player in PG induced cytoprotection. Western blot analyses indicated that PG induced the expression of Bcl-XL in a dose and time dependent manner consistent with the protective effect of PG. Induction of Bcl-XL by PG was blocked by cyclohexamide, but was not blocked by Actinomycin D indicating that a transcriptionally independent mechanism is responsible for the induction of Bcl-XL by PG. The activity of a bcl-x 3'UTR reporter was induced by PG and blocked by MF. These data suggest that PG may induce stabilization of the Bcl-X mRNA. We further explored the mechanism of PG induced Bcl-XL gene expression by comparing the effect of PG to two other steroids: corticosterone (CT) and retinoic acid (RA). Both CT and RA attenuate Dox induced apoptosis in cardiomyocytes. CT, but not RA or PG induced the activity of a GRE reporter plasmid. Analysis of the 5' region of the Bcl-XL promoter indicated that RA and CT, but not PG induced the activity of the 0.9kb region of the Bcl-XL promoter. The induction of the 0.9kb reporter plasmid by CT was glucocorticoid receptor dependent, since it was inhibited by MF. The Bcl-XL promoter does not contain any glucocorticoid or retinoid response elements, but does have AP-1 and NFkB response elements. CT, but not RA or PG induced the activity of an AP-1 reporter plasmid. RA, but not CT or PG induced the activity of an NFkB reporter plasmid. The induction of the 0.9kb Bcl-XL reporter plasmid by CT was blocked by expression of a dominant negative c-jun, TAM67 as well SB202190 indicating a nongenomic effect of CT in activating the Bcl-XL promoter through a p38 MAPK mediated AP-1 mechanism. Therefore although all three types of nuclear receptor ligands induce bcl-xL expression, the effect of CT is mediated by transcriptional activation by AP-1 signaling while NF-kB transcription factor appears to be involved in RA indced bcl-xL transcription.

<p> Introduction: 17β-estradiol (E2) has proposed anti-oxidant and membrane stabilizing properties that may attenuate exercise-induced damage, inflammation and alter gene expression. The purpose of this thesis was to determine if acute E2 supplementation would affect the oxidative stress, membrane damage, inflammation and global mRNA expression induced by eccentric exercise. Methods: 18 healthy young males were randomly assigned to 8 days of placebo (CON) or E2 (EXP) supplementation. Blood and muscle samples were collected at baseline (BL), following supplementation (PS), +3 hours (3H) and +48 hours (48H) after 150 single-leg eccentric contractions. Blood samples were analyzed for hormone concentration, creatine kinase (CK) activity and total antioxidant capacity (T AC). Inflammation was quantified by neutrophil and macrophage infiltration. Genes selected a priori for oxidative stress defense, membrane homeostasis and growth were analyzed with real-time RT-PCR. High density oligonucleotide based microarrays were screened for novel differences in mRNA expression. Results: A primary finding was that increased serum E2 did not affect anti-oxidant capacity, creatine kinase efflux or mRNA content of genes related to oxidative stress defence and membrane homeostasis. E2 did attenuate neutrophil infiltration into muscle but did not affect macrophage density. Microarray analysis revealed that exercise induced differential expression of 611 genes at 3H and confirmed that E2 did not affect mRNA content. Genes were manually clustered into biological categories and from this dataset the signaling pathways for RhoA and NF AT were identified as transcriptionally active. Both pathways regulate hypertrophic signaling through the AP-1 transcription factor complex. Conclusions and significance: A major contribution ofthis thesis is that E2 may affect exercise induced inflammation through mechanisms that that do not affect oxidative stress or membrane stability. Additionally, the transcriptional activation ofSTARS/RhoA/APl and NFAT/APl indicates that both are important for early repair and remodelling signaling after a single bout of unaccustomed eccentric exercise. </p> / Thesis / Doctor of Philosophy (PhD)

Faculty of Sciences School of Pharmacy and Pharmacology 0000073k moollaaneesa@yahoo.com / As part of an investigation of the biological activities of South African plants and due to their extensive traditional use and lack of scientific evidence, a phytochemical and pharmacological investigation was performed on 17 indigenous Agathosma species (19 samples). The chemical composition of the essential oils was determined using gas chromatography coupled to mass spectroscopy (GC-MS). Analysis resulted in the identification of 333 compounds. To evaluate the chemical similarities and differences, cluster analysis was used to assess the essential oil composition of the samples. The results showed qualitative and quantitative differences amongst the taxa. The essential oils of Agathosma hirsuta and A. zwartbergense are particularly rich in citronellal, hence they are tightly clustered in the dendrogram obtained from the cluster analysis. Linalool, myrcene and limonene are the major constituents of both A. capensis (Gamka) and A. capensis (Besemfontein). Qualitative and quantitative differences are noted in the chemical compositions of the leaf oils of Agathosma capensis (Gamka) and A. capensis (Besemfontein). Agathosma arida and A. lanata are united in a single cluster due to the compounds β-pinene, linalool and spathulenol being major components in both species. The presence of 1,8-cineole in large quantities in both Agathosma namaquensis (23.5%) and A. ovalifolia (9.7%), unites them in a single cluster. A wide chemical variability for the essential oils of indigenous Agathosma species has been demonstrated. There was considerable variation in the percentage oil yield of the essential oils. Agathosma hirsuta produced the highest yield (1.15%) whilst A. ovalifolia produced the lowest yield (0.16%). vi Previous studies have revealed that the coumarin and flavonoid components of Agathosma species are responsible for their biological activities. High performance liquid chromatography (HPLC) was used to document the non-volatile composition of Agathosma species and to establish if phenolic patterns were present amongst the species. All species were found to be rich in flavonoids (i.e. flavones and flavonols). Many of the compounds detected were common to most of the species. A pure coumarin, puberulin, was identified in the diethyl ether extract of Agathosma ovata (round-leaf) and detected in the dichloromethane and methanol (1:1) extract of A. namaquensis. Agathosma species have been used traditionally to treat a wide variety of infections. They has been used as a cough remedy, for the treatment of colds and flu, kidney and urinary tract infections, for the treatment of cholera and other stomach ailments. Based on the extensive use and lack of scientific evidence, a study was embarked upon to determine its bioactivity. Using the disc diffusion assay as a preliminary screening and thereafter the minimum inhibitory concentration (MIC) assay, the antimicrobial activity of the essential oils and non-volatile compounds was assessed on two Gram-positive bacteria, Staphylococcus aureus and Bacillus cereus, one Gram-negative bacterium, Klebsiella pneumoniae, and one yeast, Candida albicans. All of the extracts proved to be active against the four pathogens tested with the exception of Agathosma bathii which showed poor activity against Klebsiella pneumoniae (MIC value of 32mg/ml). The extracts exhibited stronger activity against the pathogens as compared to the essential oils. Both the essential oils and extracts exhibited higher activity towards the Gram-positive bacteria than the Gram-negative bacterium, with the extract of Agathosma ovata (round-leaf) displaying the greatest vii activity against Staphylococcus aureus (MIC value of 0.156mg/ml) and Bacillus cereus (MIC value of 0.125mg/ml). The extract of Agathosma parva displayed the greatest activity against Candida albicans and Klebsiella pneumoniae (MIC value of 1.5mg/ml). Amongst the essential oils, Agathosma pungens proved to be the most active against the Gram-positive pathogen, Bacillus cereus (MIC value of 3mg/ml). Agathosma collina was the most active against Candida albicans (MIC value of 3mg/ml) whilst A. zwartbergense proved to be the least active against most of the tested pathogens. The antimicrobial activity of the essential oils may be ascribed to oxygenated constituents, such as 1,8-cineole, linalool and carvacrol. The activity of the extracts may be ascribed to constituents such as flavonoids, coumarins and alkaloids. Due to the availability and accessibility of Agathosma ovata, a seasonal variation study was performed on the chemical composition of the essential oils and how this may impact on the antimicrobial activity. Furthermore, this species has recently been earmarked for commercial development by the flavour and fragrance industry and information on variability is required to establish the harvesting protocol. Ten samples were harvested in total. There was a substantial variation in the oil yield throughout the year, ranging from 0.23% in early Spring to 0.85% in late Autumn. A higher yield was observed during the flowering season as compared to the non-flowering season. Oil yields were low during Summer (0.44%-0.48%) which may have been due to the low oil content in stems and higher proportion of stems after flowering. The proportion of oil-rich green leaves also decreased markedly, hence affecting the yield. Overall the yields were dependant on the season harvested and proportion of plant parts distilled. viii The chemical composition of the essential oils was determined using GC-MS and resulted in the identification of 145 compounds in 10 of the samples. All samples contained a large number of common monoterpenes and had very similar compositions, with minor quantitative variation. Some components common to all samples include: sabinene, p-cymene, β-pinene, α-pinene, α-thujene, myrcene, limonene, linalool and terpinen-4-ol. Sabinene was found to be the most dominant component in all samples, ranging between 25.6% and 44.4%. Myrcene levels dropped sharply between the beginning of Spring and end of Summer, from 14.9% to 1.0%. β-pinene followed a similar trend, peaking during Spring and decreasing during the Summer months. The lowest levels of linalool (4.3%), myrcene (1.0%), β-pinene (3.9%), limonene (1.9%) and sabinene (25.6%), occurred during the Summer months when the temperatures were high. There was a Springtime increase in the levels of β- pinene, terpinen-4-ol, linalool, sabinene, limonene and p-cymene in the non-flowering Agathosma ovata. These changes may have been due to the higher proportion of young leaves during Spring, which may have oil compositions slightly different to those of mature leaves. A rare thiol derivative (tr) that could not be identified was detected in the March sample. Many of the changes were associated with flowering and the results obtained reveal that the chemical composition of the essential oil of Agathosma ovata is subject to seasonal variation. Using the MIC assay, the antimicrobial activity of the essential oils was assessed on two Gram-positive bacteria, Staphylococcus aureus and Bacillus cereus, one Gramnegative bacterium, Klebsiella pneumoniae, and one yeast, Candida albicans. The study demonstrated differences in the potency of antimicrobial activity of the essential oils distilled each month. The Winter samples were more active against Bacillus cereus, Staphylococcus aureus and Klebsiella pneumoniae. Activity in mid Spring ix was greater against Staphylococcus aureus (MIC value of 3mg/ml) and Klebsiella pneumoniae (MIC value of 3mg/ml), whilst activity decreased in Summer. There was a correlation between the concentrations of the active compounds each month and the oils antimicrobial activity. The results reveal that the antimicrobial activity of the essential oil of Agathosma ovata may not depend on the level of one component but rather the ratio of several components. ‘Buchu’ has been used traditionally as a general tonic and medicine. Tonics generally have a high anti-oxidant content in order to promote the overall well-being of the user. The anti-oxidant properties of the essential oils and non-volatile compounds was investigated using the 2, 2-diphenyl-1-picrylhydrazyl (DPPH) and 2, 2'-azinobis(3- ethylbenzothiazoline-6-sulfonic acid) (ABTS) assays. Only the non-volatile compounds exhibited activity. Their activities may be ascribed to the flavonoid components. Most of the species portrayed moderate to poor activity in the DPPH assay with the exception of Agathosma capensis (Gamka) (IC50 value of 24.08 + 4.42μg/ml) and A. pubigera (IC50 value of 35.61 + 0.86μg/ml) which were two of the most active species, although their activities were inferior when compared to vitamin C. The results from the ABTS assay differed from that of the DPPH assay. All extracts showed greater activity in this assay with Agathosma namaquensis (IC50 value of 15.66 ± 4.57μg/ml) and A. capensis (Besemfontein) (IC50 value of 19.84 ± 0.09μg/ml) being the most active species. This may be due to the ABTS assay having an additional reaction system. ‘Buchu’ has been used traditionally as an antipyretic, topically for the treatment of burns and wounds and for the relief of rheumatism, gout and bruises. The antix inflammatory activity of the essential oils and non-volatile compounds was assessed using the 5-lipoxygenase (LOX) assay. Only the essential oils exhibited activity. All proved to be active with the exception of Agathosma stipitata which was UV active and caused interference. This was due to its major compounds neral (39.9%) and geranial (10.1%) which absorbed strongly at 234 nm and hence rendered its spectrophotometric measurement impossible. The essential oil of Agathosma collina displayed the most promising activity (IC50 value of 25.98 ± 1.83μg/ml). It is well known that many herbal medicines can have adverse effects, in which case it is necessary to evaluate the benefit-risk profile. The toxic effects of Agathosma species have been poorly studied and no information is available in this regard. Hence the toxicity profile of the non-volatile compounds and essential oils was assessed on transformed human kidney epithelium (Graham) cells using the microculture tetrazolium (MTT) cellular viability assay. The extracts of Agathosma lanata (IC50 value of 26.17 ± 9.58μg/ml) and A. ovata (round-leaf) (IC50 value of 25.20 ± 6.30μg/ml) proved to be the most toxic, whilst the extracts of Agathosma bathii, A. capensis (Besemfontein), A. betulina, A. crenulata and A. namaquensis did not prove to be toxic at the concentrations tested. Serial dilutions displayed different inhibitions of cell growth and the species proved to be toxic in a dose-dependant manner. The essential oils of all 19 species proved to be much more toxic (IC50 values < 0.0001μg/ml) than a plant-derived compound that is considered relatively safe, namely quinine (IC50 value of 136.06 ± 4.06μg/ml). The toxicities of the essential oils may be due to compounds like methyl chavicol, eugenol, methyl eugenol, pulegone and methyl salicylate whilst the toxicities of the extracts may be due to the alkaloid and coumarin components.

Carbon/Carbon (C/C) composite is defined as a carbon fiber reinforced carbon matrix. Since 1958 research has been carried out on the C/C composites. The main reason for the development of new C/C composites is the number of advantages it has to offer when compared with the regular materials. The areas where C/C composites are being used extensively are aerospace, military, etc. These C/C composites have better physical, mechanical, thermal properties when compared to steel. That is the reason C/C brakes made a huge impact in the aerospace industry. The main drawback associated with the C/C brakes which are used in aerospace applications is the oxidation of the composite at higher temperatures. Also other problem linked with the C/C brake is the migration of the inhibitors on to the friction surface of the brake which can eventually decrease the friction coefficient of the brake material. So, characterizing the commercially available Anti-Oxidant(A/O) system, developing a new A/O system which can not only provide better oxidation protection, but also an improved anti-oxidant migration resistance will be our main goal of this project.

Plants have been utilized as a source of medicines since ancient times. They contain a vast range of secondary metabolites which play important roles in different diseases. The Scope of this study is to define the function of secondary metabolites from Andrographis paniculata (Kalmegh) and Silybum marianum (Milk Thistle) against DNA damage, which initiates many diseases. Sequential extraction of both plant materials was performed with different polarity solvents. Qualitative analysis was performed with Gas (GC) and High Performance Liquid Chromatography (HPLC). Primary extracts screening studies were performed against cytotoxicity, antioxidant and soluble collagen assays (Sircol dye). Further bioactivity was confirmed using the single-cell gel electrophoresis (Comet assay) to estimate levels of DNA damage. Fourier Transform Infrared analysis of bioactive extracts was performed to identify the functional groups present in them . Subsequently, bioactive extracts were further separated into acid, base, phenol and neutral fractions. These fractions and bioactive extracts were screened with the free radical assays to identify the scavenging activity. Chemical mapping of the bioactive fraction was performed with High Performance Liquid Chromatography (HPLC). Preparative-HPLC was performed to separate the compounds which were present in the bioactive fractions. MTT assay, Hydroxyl radical and nitric oxide radical scavenging activity assays were performed to screen the fractions and DNA protective activity of the bioactive fractions was confirmed with single-cell gel electrophoresis. These bioactive fractions were de-replicated with hyphenated techniques like LCMS and LCMS/MS to identify the molecular weight of the compounds and Quadrupole time-of-flight mass spectrometry was performed to identify the accurate mass of the compounds. Sequential extraction separates the non-polar and polar compounds present in the plant material. Qualitative analysis confirmed the presence of fatty acids in the non-polar extracts of both plants using GC and the presence of standard constituents in the polar extracts of both plants using HPLC. It also helps in chemical mapping of the extracts. Acetone, Methanol1 and Methanol2 extracts from either plant are non-cytotoxic. The high antioxidant activity is observed in methanol extracts from Andrographis paniculata and in acetone/methanol2 extracts from Silybum marianum. Extracts that protect against UVA and UVB damage also increased soluble collagen production in Human Dermal Fibroblast (HDF) in culture. Primary Screening helped to select six extracts out of twelve extracts for further analysis. Comet assay confirmed DNA protective activity in Methanol1 extract of Milk thistle and Acetone, Methanol2 extracts from Kalmegh. These three extracts were further fractionated into 38 fractions out of which three fractions that are F1, F13 and F31 fractions confirm the DNA protection activity. De-replication of the bioactive fractions was performed with LC-ESI-MS/MS which confirm twenty one compounds and accurate mass of fifteen compounds was determined using Q-tof mass spectrometry.

Indiscriminate use of pesticides appears to alter immune response in non-target organisms such as humans and other animals. Thus, immune modulation is considered as one of the potential risks and consequences following exposure to these chemicals. Because of the widespread usage, exposure to mixtures of pesticides during the lifetime of individuals is unavoidable and can result in potentiation of the toxic effects. Because immune cells are more susceptible to toxic insults at a lower dose than most other cell types, the effects of pesticides and their mixtures on murine splenocytes were evaluated. C57BL/6 male mouse splenocytes, in vitro, were exposed to permethrin and endosulfan, individually and in-combination (25-200 µM). The immunotoxic potential of these pesticides was monitored using a flow cytometric technique in combination with 7-Amino Actinomycin D (7-AAD) staining. Endosulfan exposures (25-150 µM) resulted in time- and dose-dependent increase in apoptotic and necrotic cell death in murine splenocytes, in vitro. Permethrin exposure (50-200 µM) resulted in neither a time-dependent/dose-dependent loss of splenocyte viability nor induction of apoptosis in splenocytes. With mixtures of permethrin and endosulfan, depressed viability and enhanced early apoptosis and late apoptosis/necrosis were observed. Exposure to mixtures of 50 µM endosulfan with 50 or 100 µM permethrin increased late apoptosis/necrosis compared to exposure to either chemical alone. DNA fragmentation, a hall mark of apoptosis was observed by DNA ladder technique, confirming the occurrence of apoptosis. Morphological observation using cytospun slides was also carried out to further confirm the presence of apoptosis and necrosis. These findings suggest that the immunotoxicity of endosulfan both individually and in mixtures with permethrin is associated with the occurrence of apoptotic and necrotic processes. Further, the ability of these pesticides to alter the oxidative status of the cells, via reactive oxygen species (ROS) generation and modulation of intracellular antioxidant enzymes levels, was investigated. We monitored the generation of ROS such as hydrogen peroxide (H₂O₂) with 2´, 7´- dichlorofluorescin diacetate (DCFH-DA) assay and superoxide anion (O₂⁻) with hydroethidine (HE) assay in combination with flow cytometry. Spectrophotometric techniques were used to measure antioxidant enzymes such as catalase (CAT), superoxide dismutase (SOD), glutathione reductase (GR) and glutathione peroxidase (GPX). Results of the analyses revealed that individual pesticides increased the production of H₂O₂ in a time and dose-dependent manner. Both time and dose-dependent increases in O₂⁻ production were caused by permethrin; whereas endosulfan exposure resulted in only a dose-dependent increase. However, exposure to mixtures of these pesticides had little or no effect on the generation of H₂O₂ and O₂⁻ radicals as compared to individual pesticides. The levels of SOD and GPX in pesticide-treated splenocytes were found to be not different from solvent control. An increase in GR and CAT levels in cells was noticed with permethrin (100 µM) exposure. These findings suggest that permethrin and endosulfan have the ability to affect the cellular oxidative status and can cause toxicity in immune cells, in vitro. / Master of Science

Ischaemia-reperfusion (IR) injury makes a major contribution to graft damage during kidney transplantation and increases the risks of primary non-function, delayed graft function and rejection. Oxidative damage to mitochondria is a key early event in IR injury. The aim of this project was to examine the safety and efficacy of the mitochondria-targeted antioxidant MitoQ in reducing pig and human kidney IR injury using an ex vivo normothermic perfusion (EVNP) system. Over a range of 500 nM to 250 µM using a 150 pig kidneys and 80 declined deceased human kidneys, MitoQ was successfully taken up by pig and human kidneys in a concentration-dependent manner, resulting in stable tissue concentrations over 24 hours of cold storage followed by 6 hours of EVNP. The uptake of MitoQ was increased approximately 2-fold when MitoQ was administered to warm (rather than cold) kidneys and when kidneys were preserved using hypothermic machine perfusion (rather than cold static storage). 50 µM MitoQ, administered to pig kidneys at the end of warm ischaemia, significantly increased renal blood and urine output flow at the end of 6 h EVNP compared to the control group. Creatinine clearance was numerically higher in the 50 µM MitoQ group compared to the control group but the difference did not reach statistical significance. To test the safety and efficacy of MitoQ in human kidney IRI, pairs of declined deceased human kidneys were used, with one kidney in each pair used as control. The total urine output, creatinine clearance and percentage fall of serum creatinine were numerically higher in the 50 µM MitoQ group compared to the control group, although the differences did not reach statistical significance during 3 h of EVNP. There was a significant difference in the renal blood flow between the 50 µM MitoQ group and the control group at the end of the first hour of EVNP. The renal blood flow remained relatively stable during the first hour of EVNP in the 50 µM MitoQ group compared to a significant decrease in renal blood flow in the control group. There was no effect on fractional excretion of sodium or oxidative injury markers (protein carbonyl formation, lipid peroxidation) in pig or human kidneys, which is consistent with previous studies that demonstrated the requirement of >24 hour after reperfusion for manifestation of changes in these parameters. In this thesis, I was able to successfully demonstrate the safety and potential efficacy of MitoQ in ameliorating renal IRI using pig kidneys. While more declined deceased human kidneys need to be analysed to fully explore the potential efficacy of MitoQ in ameliorating renal IRI, this study provides important data that will help inform future studies and ultimately a clinical trial for assessing the efficacy of the mitochondria-targeted antioxidant MitoQ in human kidney transplantation. My findings suggest that MitoQ has the potential to increase the use of marginal kidneys and to improve graft and patient outcomes.