Location: Healthy Processed Foods Research
2020 Annual Report
Objectives
Objective 1: Resolve how novel single or in combination bioactive phytochemicals may enable the prevention of obesity and/or obesity related metabolic dysfunction in animal models.
• Sub-objective 1A: Investigate the interactions of polyphenols on bile acids, proteins, and enzymes in intestinal lumen and their effect on intestinal permeability and FXR activation.
• Sub-objective 1B: Evaluate ingredients containing two or more bioactive compounds such as polyphenols encapsulated for synergism in obese animal models.
• Sub-objective 1C: Evaluate polyphenols encapsulated and/or immobilized in plant or microbial cell ghosts or gel bioactive fiber matrices such as HPMC, alginate, or glucomannan.
Objective 2: Following gut fermentation of phytochemicals, determine the metabolites and/or peptide products that prevent inflammation in cell culture or animal models.
• Sub-objective 2A: Identify major metabolites and proteins in extracts from food processing byproducts fermented by gut bacteria.
• Sub-objective 2B: Culture 3T3-L1 adipocytes and lipopolysaccharide-activated RAW 264.7 macrophages with metabolites and/or peptides identified in 2A to assess anti-inflammatory response and lipid accumulation.
Objective 3: Integrate measurable allergenic properties with methods to mitigate food allergens in nuts and dairy.
• Sub-objective 3A: Develop methods for investigating conformational allergenic determinants of food allergens and assess their prevalence.
• Sub-objective 3B: Define, characterize, and develop methods to detect novel nut allergens.
Approach
Hypothesis 1A: Polyphenols complexed by proteins modulate bile acid interactions with gut microbe. Specific research procedures: A high fat diet induced obesity (DIO) mouse (C57BL/6J) will be used. Polyphenols, conjugated and free bile acids in the intestinal lumen, colon, and feces will be measured by HPLC and intestinal permeability by FITC dextran. The expression of FGF15 and FXR expression will be measured by RT-PCR. Inflammatory and diabetic biomarkers by BioPlex multiplex ELISA.
Hypothesis 1B: Synergistic interactions exist between polyphenols and other phytochemicals. Specific research procedures: The DIO mice will be fed single compounds or combinations. Polyvinylalcohol removal of polyphenolics and loss of activity would suggest synergistic compound is a polyphenol. Efflux pump inhibitors will be fed with grape seed proanthocyanidins and bile acids. Gut microbiome will be analyzed.
Hypothesis 1C: Polyphenols beneficial health effects may be due to reducing the rapid influx of sugar, fat and other nutrients from rapidly digested foods that overload organs by binding to and inhibiting digestive enzymes. Specific research procedures: Yeast, bacteria, and plant cell ghosts infused with grape seed polyphenols, resveratrol or other bioactive phytochemicals will be provided by UC Davis. The encapsulated polyphenols will be tested in the DIO model described in Objective 1A.
Hypothesis 2A: Fermentation produces absorbable phytochemicals from plant materials and bacterial protein/peptides. Specific research procedures: Extracts of food processing byproducts from apples, red beets, eggplants, grapes, olives, and cereal brans will be fermented with B fragilis, L plantarum, C perfringens, Bifidobacterium longum. The extract will be analyzed for total protein content, nucleic acids, and total polyphenolics. Specific phytochemicals will be analyzed by LC/MS.
Hypothesis 2B: Products of fermentation and/ or bacterial derived peptides inhibit inflammation or lipid accumulation in adipose tissue. Specific research procedures: Glucose metabolism and lipid accumulation of 3T3-L1adipocytes and inflammatory responses in RAW264.7 macrophages will be measured with or without additions of substances from 2A.
Hypothesis 3A: Structural information can be used to develop a new method that is applicable to map conformational IGE epitopes. Specific research procedures: An alternative ELISA format would use His-tagged allergens immobilized to N2+ coated microtiter plates. Final reverse mutant will have a His-tag. The pTNS2 vector will be used to construct plasmids for making the C-terminal His-tagged ßLG and its mutant by omitting the stop codon. To map the conformational IgE epitopes in ßLG, sera from 20 subjects with IgE antibodies against milk allergens will be used.
Hypothesis 3B: PRU DU 8 orthologs in other plants are also food allergens. Specific research procedures: To characterize Pru du 8, we will use the recombinant allergen as antigen and make polyclonal antibodies in rabbits commercially. We will isolate the coding sequence for the new protein from the corresponding tree nuts to deduce the translated protein sequence.
Progress Report
This project has just been certified following NP306 Office of Scientific Quality Review, and replaced expired project 2030-41440-007-00D, "Adding Value to Plant-Based Waste Materials through Development of Novel, Healthy Ingredients and Functional Foods." During the reporting period, evaluation of the uptake of a metabolite produced by gut bacteria was measured in blood of mice in collaboration with the University of Florida. Three manuscripts were submitted for publication.
Accomplishments
