2013 Annual Report
1a.Objectives (from AD-416):
Objective 1. Fractionate fiber, water-soluble, alcohol-soluble and lipid-soluble fractions from the increasingly large amounts of fruit and vegetable byproducts including pomace of olives, grapes, and pomegranates and the bran of cereal grains in order to identify, condense, and test bioactive compounds in animal models of obesity related diseases.
Sub-Objective 1.1. Separate different fruit tissues (skin, pulp, juice) and cereal brans (germ, aleurone, fat) using physical processes, and fractionate followed by extraction with food approved solvent methods.
Sub-Objective 1.2. Assess bioactivity of extracts and residues of 1.1 by feeding mice or hamsters hypercholesterolemic diets and determining physiologic characteristics of obesity related metabolic syndrome.
Objective 2. Develop processing methods that disrupt food matrices or cell barriers in order to increase accessibility to cellular contents and increase the bioavailability of phytonutrients from fractions of fruit and vegetable based byproducts including pomace of olives, grapes, and pomegranates, as well as grain byproducts from Obj. 1.1 that contain complex mixtures of phenolic compounds that may help to prevent obesity related diseases and add value.
Sub-Objective 2.1. Evaluate processes such as enzymatic treatment (cellulase, esterase, phytase), sonication, ethanol extraction, and high pressure to
increase bioavailability of phytochemicals and fiber, and to improve consistency of composition and bioavailability of samples for in vivo studies.
Subobjective 2.2. Test samples identified as bioactive in 1.2 as well as those further processed in 2.1 for their bioavailability. Also characterize phytochemical composition of bioavailable fractions by HPLC.
Objective 3. Evaluate a variety of viscous and gelling soluble dietary fibers to produce breads and other cereal products that are high in fiber and micronutrients.
Sub-Objective 3.1. Breads and other baked products will be developed from non-gluten whole grains and bran such as corn, rice, oat, barley, rye, quinoa, amaranth in order to increase the diversity of whole grain products available to consumers. Viscous soluble dietary fibers are necessary to replace gluten, but also reduce postprandial glycemic response and plasma cholesterol.
Sub-Objective 3.2. Preliminary studies indicate difference in hepatic nutrient metabolism between wheat and barley flours was not accounted for by soluble fiber content. We propose to evaluate a variety of cereal grains and seeds for bioactivity by analyzing expression of hepatic genes representative of metabolism of sterols, bile acids, and fat, and indicators of inflammation.
1b.Approach (from AD-416):
Food processing technologies will be applied to release the high concentration of polyphenolic components from the matrices of the skin and bran byproducts of juice, wine, oil and flour production. Polyphenolics will be extracted from these processed matrices by systematic application of food grade solvents into broad classes and a fiber fraction. Mice and hamster models of obesity related diseases will be used to determine the potential of the polyphenolic or fiber fractions to reduce or prevent the characteristics of metabolic diseases. Mechanisms will be investigated through gene expression pathway analysis. Of particular interest is the ability of polyphenolics or fiber to reduce the absorption of lipopolysaccharides that trigger adipose inflammatory responses. Viscous soluble fibers will be used to develop breads from nontraditional whole grains and seeds to determine if they have potential to reduce metabolic syndrome in animal models.
Substantial progress was made on the three project objectives and subobjectives. In collaboration with Cooperative Research and Development Agreement (CRADA) partners, we confirmed the physiological effects of chardonnay grape seed flour by performing a dose response study of cholesterol lowering using hamsters fed high fat diets. We also fractionated the grape seed flour into water extractable and nonextractable fractions and found that both fractions had biological activity. We also found that there were significant changes in the species and numbers of bacteria in the feces of hamsters fed grape seed flour. These findings were shared with researchers at the Mayo Clinic to design a human trial of grape seed flour. This research was also accelerated by CRADA funding and focus on the supplementation of breads with protein byproduct from rice syrup production. An SBIR grant was applied for and awarded ($95,000) to the CRADA partner to further document the physiological effects of grape seed flour in animal models with USDA to support a future human study.
Cholesterol lowering by extractable and nonextractable components of grape seed. In collaboration with Cooperative Research Agreement Development Agreement partners, we extracted chardonnay grape seed flour and fed the extract and nonextractable fractions to hamsters on high fat diets. Both extractable and nonextractable fractions lowered plasma cholesterol. These results show the importance of nonextractable polyphenolic compounds that may be metabolized into bioactive compounds by gut bacteria. Rather than supplements, our research has resulted in development and sales of food products containing grape seed flours high in polyphenolics and fiber.
Determination of dose of grape seeds polyphenolics for human studies at Mayo Clinic. Almost all published studies of the cholesterol lowering properties of grape seeds were performed with alcoholic extracts. A meta-analysis of cholesterol lowering by grape seed showed no effect. We evaluated the effects of dose of grape seed extracts in hamsters. Using published dosage from human studies as well as dose based on our 2012-3 hamster studies, we found that dose in published human studies was too low to show physiological effects and accounts for their lack of effect. The results and a comprehensive review and mechanistic hypothesis of grape polyphenolic action were presented to researchers at the Mayo Clinic, Rochester, Minnesota. A positive human trial of the cholesterol lowering ability of grape seed flours will reduce risk factors for chronic disease in the United States and increase demand for a byproduct of wine making.
CRADA Partner is Awarded a Grant Based on Research Results. Based on Cooperative Research and Development Agreement (CRADA) research results, the CRADA partner, Sonomaceuticals, was awarded a Small Buiness Innovation Research (SBIR) grant for preclinical studies of grape seed flour. The $95,000 grant was awarded to the CRADA partner to support research at US Department of Agriculture, Western Regional Research Center, Albany, California, for preclinical studies in animal models of cholesterol lowering. The study will determine the cholesterol lowering and weight lowering effects of whole grape seed flours on obese and insulin resistant animal models. The study will also include the effects of whole grape seed flour on the population of gut microbes. The study results will be used to determine dose and treatment of grape seed flour for human studies and increase understanding of the mechanism of action. Human studies are necessary for Food and Drug Administration (FDA) health claims that will increase consumer acceptance of foods containing grape seed derived polyphenolic compounds.
Whole grain, gluten-free, egg-free pasta. Pasta of garbanzo cereal mixtures were developed and evaluated by consumer sensory panels. Whole grain pasta can be easily made at home using kitchen counter-top appliance as well as in the commercial production. They are low in fat, calories, good source of protein, dietary fiber and minerals. Acceptability determined by in house volunteers was for rice-garbanzo 84%, corn-garbanzo 70%, millet-garbanzo 54% and sorghum-garbanzo 48%. Whole grain, gluten-free, egg-free, high protein pasta would increase whole grain consumption and offer a healthy option to vegetarians as well as to gluten sensitive individuals.
Cholesterol lowering dose response of Chardonnay grape seed flour in hamsters. In collaboration with Cooperative Research Agreement Development Agreement (CRADA) partners, we found that plasma total and low density lipoprotein (LDL) cholesterol (risk factors for cardiovascular disease) were lowered in hamsters on high fat diets. Changes in liver genes for cholesterol metabolic pathways and fat metabolism support the physiological observations. Combining data from 2012, we found a linear relationship between a component of grape seed (catechin) and plasma cholesterol. Chardonnay grape seed flour contains the highest amount of polyphenolics of food ingredients and reduces blood pressure in humans and may assist in weight management and reducing blood cholesterol.
Kahlon, T.S., Milczarek, R.R., Chiu, M.M. 2013. Whole grain gluten-free egg-free pasta. Cereal Foods World. 58(1):4-7. DOI: 10.1094/CFW-58-1-0004.
Kahlon, T.S., Milczarek, R.R., Chiu, M.M. 2012. In Vitro bile acid binding of kale, mustard greens, broccoli, cabbage and green bell pepper improves with microwave cooking. Vegetos: An International Journal of Plant Research. 25(2):29-36.
Shao, D., Yokoyama, W.H., Bartley, G.E., Pan, Z., Zhang, H., Zhang, A. 2013. Plasma and hepatic cholesterol-lowering in hamsters by tomato pomace, tomato seed oil and defatted tomato seed supplemented in high fat diets. Food Chemistry. 139:589-596.
Hung, S., Yokoyama, W.H., Kim, H., Bartley, G.E., Anderson, W.K., Alberts, D.R., Langhorst, M.L., Williams, D.M., Stott, W., Turowski, M. 2012. HPMC supplementation reduces abdominal fat content, intestinal permeability, inflammation, and insulin resistance in diet-induced obese mice. Journal of Agricultural and Food Chemistry. 60(44):11149-11156.
Kim, H., Bartley, G.E., Young, S., Yokoyama, W.H. 2013. HPMC supplementation reduces fatty liver, intestinal permeability, and insulin resistance with altered hepatic gene expression in diet-induced obese mice. Journal of Agricultural and Food Chemistry. 61(26):6404-6411.
Li, Y., Yokoyama, W.H., Shoemaker, C.F., Wei, D., Ma, J., Zhu, S., Zhong, F. 2013. Properties of chitosan microencapsulated orange oil prepared by spray-drying and its stability to detergents. Journal of Agricultural and Food Chemistry. 61(13):3311-3319.