| Summary: | Polysaccharides & oligosaccharides have a wide range of functions in both industry & the natural world. Saccharides have the potential to adopt an incredible number of conformations. The role of saccharides in cellular communication/recognition is beginning to be elucidated within the scientific comriiunity. The immune system especially uses saccharides to recognise 'self & 'non-self. The potential influence of saccharides upon the functioning of the immune system is enormous. The analysis of polysaccharides & oligosaccharide samples using an immune based method could provide an insight into the relationship between saccharide structure-function. Reactive oxygen species (ROS) were used.to monitor immune cell activity & therefore asses the immunomodulatory effect ofpolysaccharides & oligosaccharides. A fluorescent method was developed that provides a dynamic, reliable, efficient, non-operator assessed technique for the analysis of ROS. The technique uses the compound 2' 7'-dichlorofluorescein diacetate (DCFH-DA). DCFH-DA is a non-fluorescent compound that can pass through cell membranes. Once it is in the cytoplasm, esterases remove the acetates to produce 2', 7'-dichlorodihydrofluorescein (DCFH). DCFH is easily oxidised to 2', 7'-dichlorofluorescein (DCF) a highly fluorescent compound (excitation 485 nm; emission 530 nm). The polysaccharides gellan gum caused increase of up to 80% in ROS production compared to control immune cells. Tremellan also caused increases of up to 74% in ROS production by immune cells. Several yeast cell wall fractions were tested, all of which showed immunostimulatory effects on the production of ROS. An L-fucose-rich c1avan caused 22% inhibition of ROS production, whilst locust bean gum caused up to 48% inhibition ofROS production by immune cells. Monosaccharides & oligosaccharides with degrees of polymerisation (DP) ranging from 1 to 8 were also tested for their immunomodulatory properties on the production of ROS. Laminarihexaose (DP 6) and laminariheptaose (DP 7) gave differing results because of their DP &Jor structure, with laminarihexaose causing a 16% decrease in ROS production and laminariheptaose causing a 25% increase in ROS production from immune cells compared to control cells after 60 minutes. A series of arabino-oligosaccharide (DP 6, 7 & 8) were also tested. As their DP increased so did their inhibitory effect on ROS production by immune cells. Manno-oligosaccharides (DP 5-7), derived from locust bean gum, gave similar results to its parent polysaccharide. Many other poly~ccharides & oligosaccharides were tested, giving stimulatory, inhibitory and neutral results. These results have provided an insight into the relationship between size/structure-function of saccharides. The potential for further tests and manufacture ofdesigner oligosaccharides using these results is vast.
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