The effects of model foods rich in bioactive compounds on brain-gut regulation and neurodegeneration : A thesis submitted in partial fulfilment of the requirements for the degree of Doctor of Philosophy at Lincoln University

A compelling interaction between the diet, gut and host physiology has emerged. Biologically active compounds particularly dietary fibre (DF) and polyphenols in a diet have long been appreciated due to their inverse relation with rising chronic diseases, particularly metabolic disorder, obesity, type 2 diabetes and cancers. Research has illustrated a complex interaction of multiple peptides (gut endocrine systems) and nutritional inputs physiologically affect appetite control and host immunity. Despite the potential health benefits of consuming a diet rich in bioactive compounds, little is known about the bio-functional interactions such as digestibility, effects of digested and fermented metabolites on the attenuation of gut endocrine system and immunity of the host. This study developed a cookie from wholemeal cereals which incorporated different proportions (0, 5, 10 and 15 %) of blackcurrant or astaxanthin-rich microalgae Haematococcus pluvialis powder as a replacement of wheat, barley and oat flours. The study evaluated the effects of blackcurrant/astaxanthin on cookie textural properties. The in-vitro carbohydrate digestibility of the cookies demonstrated the glycaemic control. Inclusion of blackcurrant/astaxanthin powder in wholemeal cereals cookies significantly (p < 0.05) decreased the rate of glucose release, when compared with wholemeal cereals cookie control. The 15 % blackcurrant and astaxanthin powder incorporated cookies had a significantly (p < 0.05) higher fibre fraction, compared to the control flour cookies. Blackcurrant powder contains about 25 % and astaxanthin-rich H. pluvialis powder contains more than 30 % DF. Using an in-vitro fermentation model, the prominent gut bacterial growth and fermentative properties of the indigestible cookie fraction were evaluated. The combination of four bacterial species Lactobacillus and Bifidobacterium was utilised using the isolated fibre fractions from different cookies as substrates. These fibre fractions significantly increased the microbial growth and their metabolite (short-chain fatty acids, SCFAs) production. The blackcurrant/astaxanthin incorporated substrates significantly increased the bacterial populations, lowered fermentation pH and increased optical density (OD600) at 6, 12, 24 and 48 h incubation when compared with negative (medium only) and positive control (wholemeal cereals cookies). The increased concentrations of SCFAs production from the in-vitro bacterial fermentation indicated selective fermentation capacity of blackcurrant/astaxanthin cookie fibre fractions. The simulated digestion and fermentation and cellular biological activity of the cookie digesta was studied and revealed possible links between metabolites and various physiological processes associated to health and diseases. Chemical extraction of free, bound and total phenolics of the blackcurrant/astaxanthin cookies exhibited significantly higher radical scavenging capacity in chemical and cell models, which is believed to have the inverse relationship with many chronic diseases such as cancer. The incorporation of blackcurrant/astaxanthin significantly (p < 0.05) increased the total phenolic content (by about 60 %), significantly improved oxygen radical absorbance capacity (by about 25 %) of the cookie. The higher ORAC and DPPH indicated that blackcurrant/astaxanthin incorporated cookie digesta/fermenta/extracts were strong antioxidants, which was in accordance with the higher total phenolic content. The major phenolic acids of the digesta/extracts were assessed by RP-HPLC, and were found to be ferulic acid, p-hydroxy benzoic acid and p-coumaric acid. However, quercetin-3-rutinoside, quercetin derivatives and kaemferol-3-glucoside were only found in blackcurrant incorporated cookie extracts. Whereas, the cinnamic and salicylic acid were found in astaxanthin incorporated cookie extracts. Major therapeutic target of cancer diseases, the inhibition of abnormal cell growth and/or proliferation activity and cellular oxidation were observed when using the cookie digesta/fermenta/extract on HepG2 cell model. Importantly, qRT-PCR exhibited significantly higher mRNA levels of the appetite regulatory cholecystokinin (CCK), glucagon-like peptide-1(GLP-1), gastrin-releasing peptide (GRP) and nucleobindin-2 (NUCB2)/nesfatin genes (ten to twelve-folds) expression. This significant interplay in genetic modulation reflected the positive link between signalling molecules SCFAs and/or bioactive phenolics and appetite control. The lower levels of inflammatory cytokines interleukin-1β (IL-1β), interleukin-6 (IL-6) and inflammation activator nuclear factor-kappa B (NF-kB) (half to two-fold) was demonstrated in HepG2 cells treated with blackcurrant/astaxanthin incorporated cookie. The anti-oxidative/inflammation response of digesta/fermenta/extract in dynamic biological changes could be a therapeutic alternative to facilitates metabolic homeostasis, counter inflammation and enhance immunity. Metabolites from gut microbiota interact with the brain physiology through signalling molecules SCFAs are starting to emerge. Interestingly, our study demonstrated that the bacterial metabolites SCFAs did not significantly inhibit amyloid-β (Aβ) aggregation in a cell model of Alzheimer’s disease. Enhanced functionalities including glycaemic control, antioxidation, anti-proliferation and genetic modulation of the bioactive compounds in cookie accelerates our knowledge to regulate appetite, oxidative stress and inflammation. Noteworthy, the study of cell model reveals the possible links between food intake and the manipulation of brain and gut health. The exerted synergistic effects of this study suggest that there may be a new and effective option to prevent and control chronic diseases in human.