Inflammation is increasingly thought to play an important role in the pathogenesis of a wide variety of acute and chronic diseases including ageing in general. To date, pharmacotherapy of inflammatory conditions is based on the use of non-steroidal anti-inflammatory drugs (NSAIDs). However, NSAIDs can cause serious gastrointestinal toxicity such as gastric bleeding and the formation of stomach ulcers. Even more ominously, some NSAIDs, particularly COX-2 inhibitors, have been linked to increased blood pressure, greatly increased risk of congestive heart failure, occurrence of thrombosis and myocardial infarction. Together, these findings provide the motivation for the development of anti-inflammatory treatments with fewer adverse effects. Reactive oxygen species (ROS) are produced mainly during oxidative phosphorylation and by activated phagocytic cells during oxidative burst. The excessive production of ROS can damage lipids, protein, membrane and nucleic acids. They also serve as important intracellular signalling molecules that are involved in the production of pro-inflammatory mediators such as cytokines and Nitric oxide. In chapter 2, hydrogen peroxide showed that it is not only intracellular signalling molecule, but travels through the extracellular space and activate adjacent cells. This hypothesis is supported by adding catalase (that converts hydrogen peroxide to water and oxygen in the extracellular space) and therefore the pro-inflammatory signal does not reach the next cell, leading to an overall reduction of the readouts, NO and TNF-I±. This concept can revolutionize drug development because hydrogen peroxide scavengers do not necessarily have to penetrate the cell membrane to exert their action - and even extreme hydrophilic molecules would be beneficial in diseases with chronic pro-inflammatory conditions. Antioxidants can protect against the damage induced by free radicals acting at various levels. Dietary and other components of plants form major sources of antioxidants. As ROS are signalling molecules, antioxidants which scavenge ROS, would therefore inhibit inflammation. In chapter 3, 115 food samples were prepared by a generic food-compatible processing method involving heating and tested for their anti-inflammatory activity in murine N11 microglia and RAW 264.7 macrophages, using nitric oxide (NO) and tumour necrosis factor-I± (TNF-I±) as proinflammatory readouts. Ten food samples including lime zest, English breakfast tea, honey-brown mushroom, button mushroom, oyster mushroom, cinnamon and cloves inhibited NO production in N11 microglia, with IC50 values below 0.5 mg/ml. The most active samples were onion, oregano and red sweet potato, exhibiting IC50 values below 0.1 mg/ml. When these ten food preparations were retested in RAW 264.7 macrophages, they all inhibited NO production similar to the results obtained in N11 microglia. In addition, English breakfast tea leaves, oyster mushroom, onion, cinnamon and button mushroom preparations suppressed TNF-I± production, exhibiting IC50 values below 0.5 mg/ml in RAW 264.7 macrophages. Considering the stability of activity during processing stages (patented processing methods of the CSIRO plant and food library), anti-inflammatory activity in both the cell lines and suppression of both NO and TNF-I± without cytotoxicity, cinnamon was been selected as most interesting extract for further chemical analysis "" chapter 4 and 5. In chapter 4-5, we examined the anti-inflammatory activities of C. zeylanicum and C. cassia extracts and their chemical constituents using RAW 264.7 macrophages. When extracts were tested in LPS and IFN-I³ activated RAW 264.7 macrophages, most of the anti-inflammatory activity, measured by downregulation of nitric oxide and TNF-I± production, was observed in the organic extracts. The most abundant compounds in these extracts were E-cinnamaldehyde and o-methoxycinnamaldehyde. When these and other constituents were tested for their anti-inflammatory activity in RAW 264.7 and J774A.1 macrophages, the most potent compounds were E-cinnamaldehyde and omethoxycinnamaldehyde, and they accounted for most of the inflammatory activity in either cinnamon species. Among the 115 samples, mushrooms were also showed significant anti-inflammatory activity and that leads to chapter 6. Edible mushrooms are attracting more and more attention as functional foods since they are rich in bioactive compounds, but their anti-inflammatory properties and the effect of food processing steps on this activity has not been systematically investigated. In chapter 6, White Button and Honey Brown (both Agaricus bisporus), Shiitake (Lentinus edodes), Enoki (Flammulina velutipes) and Oyster mushroom (Pleurotus ostreatus) preparations were tested for their anti-inflammatory activity in lipopolysaccharide (LPS) and interferon-I³ (IFN-I³) activated murine RAW 264.7 macrophages. Potent anti-inflammatory activity (IC50 < 0.1 mg/ml), measured as inhibition of NO production, could be detected in all raw mushroom preparations, but only raw Oyster (IC50 = 0.035 mg/ml) and Shiitake mushrooms (IC50 = 0.047 mg/ml) showed potent inhibition of TNF- I± production as well. Not only known traditional foods and plants but unknown Australian bush foods and plants might also have those properties. In our quest for the search of novel potent anti-inflammatory compounds involved the screening of 72 Australian tropical rainforest plant extracts from Northern Queensland. This chapter highlights the bioassay guided isolation and characterization of new antiinflammatory compounds from one of the most active plant, Cryptocarya mackinoniana The aim of this study (Chapter 7) was the identification of novel anti-inflammatory compounds present in Cryptocarya mackioniana young leaves. From this study we were able to identify a new compound, 12 and a known metabolite cryptocaryone from the DCM extract. The anti-inflammatory activities of both compounds were evaluated against RAW264.7 macrophages and compounds 12 and cryptocaryone showed significant inhibition of nitric oxide with IC50 6.36 ± 0.85 I¼g/mL and 1.26 ± 0.18 I¼g/mL respectively. In addition cryptocaryone showed significant inhibition of TNF-I± with IC50 3.58 ± 0.64 I¼g/mL. Isolation and identification of the remaining compounds is a work in progress.
| Date of Award | 2015 |
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| Original language | English |
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