Analysis of the potential for Rubus fruit ellagitannins to induce anti-inflammatory effects in in vitro models : a dissertation presented in partial fulfilment of the requirements for the degree of MPhil in Nutritional Sciences at Massey University, Manawatū

Abstract

Nutrigenomics is the study of interactions between human diets and the genome with the aim of discerning how food components influence gene and protein expression in individuals. Previous studies have established the potential for ellagitannins (ETs) isolated from various fruits, to induce anti-inflammatory responses that may be beneficial to those afflicted with Inflammatory Bowel Disease (IBD). The hypothesis of this thesis is that Rubus fruit extracts, rich in ETs, induce an anti-inflammatory effect in mammalian cells involved in inflammatory processes. Methods were developed to extract, purify, and quantify the phytochemical composition of three selected Rubus fruit cultivars (HB19, Wakefield and ZZ). Extracts (EX: phenolic extract, ETx: ET-enriched extract) were initially assessed using a secondary cell line: RAW 264.7 macrophages. Cell viability using the MTT assay, found no cytotoxic effect at polyphenolic concentrations up to 100 μg/mL. The Griess assay measured levels of nitric oxide (NO) production and found a decrease at 50 μg/mL of polyphenolic exposure for most extracts except ZZ EX. Cytokine assessment showed that the extract, Wakefield ETx, lowered production of Interleukin 6 (IL6) by macrophages. All extracts increased production of Tumour Necrosis Factor alpha (TNFα) by macrophages. Colonic intestinal epithelial cells (colonic IEC) and bone marrow derived macrophages (BMDM), were isolated from control C57BI/6J mice and Interleukin 10 gene-deficient mice (Il10-/-). Viability results suggested that the polyphenolic concentration of 50 μg/mL, was the highest tolerated concentration for NO and cytokine assays. NO production was decreased for BMDM (Il10-/-) with extracts Wakefield EX and ZZ EX, and for BMDM (C57) with most extracts except HB19 EX. IL6 production by colonic IEC (Il10-/-) was decreased for all extracts. The cytokine profile for BMDM (Il10-/-) mice showed a decrease in IL12 and IL6 production, and no change in TNFα production in response to co-incubation with all extracts. There are indications from research done in this thesis that the Rubus fruit extracts from “ZZ” and “Wakefield” cultivars reduce inflammatory immune responses or potentially having the ability to act on different pathways or induce a wound healing response.