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ARS Home » Northeast Area » Beltsville, Maryland (BHNRC) » Beltsville Human Nutrition Research Center » Diet, Genomics and Immunology Laboratory » Research » Research Project #436279

Research Project: Elucidating Phytonutrient Bioavailability, Health Promoting Effects and Mechanisms of Existing/Emerging Foods and Beverages

Location: Diet, Genomics and Immunology Laboratory

2021 Annual Report

Objective 1: Determine the accumulation and variability of important bioactive compounds in commonly consumed food crops as affected by factors such as stage of maturity (leafy greens harvested at various stages of maturity) and variety/processing (coffee products derived from different sources). [NP107, C1, PS1A] Objective 2: Determine bioavailability and cellular uptake of potential compounds in coffee products; investigate their effects on subclinical inflammation and its associated events related to chronic metabolic diseases; elucidate mechanisms of action on subclinical inflammation and related events. [NP107, C3, PS3B] Objective 3: Determine effects of brassica vegetables harvested at different stages of maturity (e.g., sprout, microgreen, baby green, mature plant) on high fat diet-induced inflammation, and adipose uncoupling protein 1 (UCP1) as mechanisms for their attenuation of high fat diet-induced weight gain. Elucidate the role of the microbiome in mediating changes in inflammation and liver lipid metabolism. [NP107, C3, PS3B]

Objective 1: Amounts of bioactive compounds in selected model plant products (e.g., coffee beans grown in different regions/conditions, coffee products, brassica vegetables harvested at different stages of maturity) will be determined using established HPLC, MS/MS and NMR methods. Objective 2: Both in-vivo and in-vitro models will be used for this objective. (1) We will determine bioavailability and cellular uptake of coffee compounds using cell culture (e.g., Caco-2, HepG2, monocytic THP-1) models. HPLC, metabolomic and lipidomic analytical technologies will be used to measure the compounds and associated metabolites. (2) We will determine potential effects of coffee/coffee chemicals (javamide-I/-II) on subclinical inflammation markers using a rodent model. Obesity will be induced in animals (e.g., rats) fed a high-fat diet, and the potential effects of coffee and coffee compounds (e.g., javamide-I/-II) on obesity-associated subclinical inflammation and biological changes will be determined to elucidate the effects. (3) We will determine the cellular/molecular mechanisms responsible for the biological effects using cell culture models. Cell will be treated with the compounds and effects of compounds on cellular pathways related to signal transduction pathways, inflammatory cytokines, adhesion molecular, transcriptional factors will be assessed at protein and message levels using western blots, ELISA and RT-PCR. Objective 3: We will determine effects of brassica vegetables harvested at different stages of maturity (e.g., sprout, microgreen, baby green, mature plant) on high fat diet-induced rodent model. Inflammatory marker and adipose uncoupling protein 1 (UCP1) will be assessed using ELISA at protein level and RT-PCR at message level as mechanisms for their attenuation of high fat diet-induced weight gain. Biochemical and marker genes analysis will be performed in liver and adipose tissues to assess the effects on lipid and energy metabolism. Metagenomic analysis using next generation sequencing technology will be performed to elucidate the role of the microbiome in mediating changes in inflammation and liver lipid metabolism.

Progress Report
This report is for a NP107 project entitled "Elucidating Phytonutrient Bioavailability, Health Promoting Effects and Mechanisms of Existing/Emerging Foods and Beverages." We focus on two foods, coffee and kale at different growth/maturation states. The following describes the current progress of the project. For Objective 1, HPLC analysis on coffee beans (Arabica and Robusta) was performed and the bioactives javamide-I/-II were found to be in the range of 0.03-5.0 mg/g. The amounts of javamide-I/-II, as well as other coffee compounds (chlorogenic acids and caffeine), were further analyzed in several coffee products in the market. Javamide-I/-II in a cup of coffee (236 mL) were found to be 0.25±0.06 and 1.89±0.39 mg, respectively. Also, one cup of coffee contains 22.8±0.7 to 71.5±2.21 mg chlorogenic acids (3-CQA, 4-CQA and 5-CQA) and 49.7±4.2 to 113.8±11.0 mg caffeine in the cup of coffee. This is the first demonstration of significant variability of Javamide-I/-II in coffee products available in the market. For Objective 2, the effects of coffee containing javamide-I/-II on bodyweight, low density liprotein (LDL), high density lipoprotein (HDL), total cholesterols, triglycerides, adipokines (adiponectin and leptin) and other obesity-related inflammatory/cardiovascular risk factors (C-reactive protein, sE-selectin, TNF-alpha, MCP-1) were examined in a non-obese animal model. Coffee consumption is often blamed for raising some metabolic factors (e.g., bodyweight, cholesterols) in non-obese individuals. There was no significant difference in the body weights between the control and coffee drinking group of rats. Additionally, there were no significant difference in plasma LDL, HDL and total cholesterol levels or adiponectin and leptin levels between the groups. Likewise, no significant difference in plasma levels of C-reactive protein and sE-selectin was observed. Hence, coffee containing javamide-I/-II are not likely to exert significant effects on bodyweight, LDL, HDL, total cholesterol, adiponectin, leptin, C-reactive protein, sE-selectin, TNF-alpha and MCP-1 in animals fed a normal diet. For Objective 3, composition analysis of kale grown at different growing stage were performed. Kale microgreen's (11 days after planting) bioactive composition (glucosinolates and fiber) was significantly different from that of baby kale (28 days after planting) and mature kale (57 and 87 days after planting). There was little difference between baby kale and mature kale. Given the large differences in composition between microgreen and mature stage, an animal experiment was then conducted to elucidate the effects of kale microgreen and mature kale on health using the diet-induced obesity rodent model. Six groups of mice were fed with following diets: 1) Low fat diet, 2) High fat diet, 3) Low fat diet + kale microgreen, 4) High fat diet + kale microgreen, 5) Low fat diet + mature Kale, 6) High fat diet + mature kale, for 8 weeks. Body weight and food intake were monitored. There were no differences in food intake between the diet groups, suggesting that the diets were well-received by the animals. Supplementation with either kale microgreen or mature kale significantly attenuated the high fat diet induced growth rate. Preliminary analysis of obesity-related health risks markers was also conducted. Supplementing kale microgreen and mature kale in high fat diet significantly attenuated high fat diet-induced increase in the lipoproteins HDL and LDL levels, but not very low density lipoprotein (VLDL). These results indicate different growing stages of kale appeared to elicit similar protective effects in a diet induced-obesity model. Transcriptomic, metabolomics, and metagenomics analysis on collected samples are in progress. Additionally, an ARS scientist, in collaboration with a scientist from Romania, investigated potential effects of climatic conditions and soil tillage systems on isoflavone content of soy. Climatic conditions significantly influenced both the soybean yields and the total isoflavone for the three years studied: 1473 kg/ha and 6944 µg/g (2014); 1544 kg/ha and 7989 µg/g (2015); 2109 kg/ha and 8101 µg/g (2016) (P < 0.05). There were also variations in the individual isoflavone contents (daidzin, genistin, glycitin, daidzein, genistein) year by year (P < 0.05). However, two conservative soil tillage systems (minimum tillage and no-tillage) did not affect total isoflavone content (µg/g), compared to the conventional system each field year. These data suggest that climatic conditions and no tillage systems may have more impact on total isoflavone content in soybeans.

1. Climatic conditions but not soil tillage systems affect isoflavone contents grown in Romania. Soybean is an important natural source for health-promoting bioactive isoflavones, but its content is likely to be influenced by external factors, such as climatic conditions and soil tillage systems. An ARS scientist, in collaboration with a scientist from Romania investigated the potential impacts of three different soil tillage systems, climatic conditions on crop yields, and isoflavone contents of soybeans. Climatic conditions significantly influenced both the soybean yields and the total isoflavone for the three years. However, compared to the conventional system each field year, two conservative soil tillage systems (minimum tillage and no-tillage) did not have adverse effects on total isoflavone content. These data showed that climatic conditions but not tillage-type soil systems are the primary effector of total isoflavone content in soybeans.

2. Cruciferous vegetable-derived compound as alternative to antibiotics. Antibiotic resistance is a critical health care problem, but alternatives to antibiotics remain scarce. ARS scientists at Beltsville, Maryland, in collaboration with the National Cancer Institute and University of Maryland scientists, seek to address this problem. The protective efficacies of bioactive phytochemicals from cruciferous vegetables to Citrobacter rodentium infection in mice, an E coli-like food-borne disease model, were studied. The cruciferous vegetable glucosinolate-derived compound indole-3-carbinol was identified to be protective against Cr infection through enhancing immune responses, increased production of antibodies against Cr. These results support the potential use of cruciferous vegetables and their bio-actives as an alternative to antibiotics for protection against food-borne bacterial infection.

3. Increased investments in food systems now could counteract the projected impact of climate change on mortality and disability by 2050. The interaction between climate change and mortality and disability is complex and needs elucidation. A modeling study by an ARS scientist and collaborators at the International Food Policy Research Institute (IFPRI) and RTI International (RTI) projects that by 2050 compared to 2010, chronic and hidden hunger will increase the overall years of life lost due to premature mortality and years lived with disability, also known as disability-adjusted life years (DALYs), by over 30 million globally. Expected impacts of climate change on the availability and access to nutritious food will exacerbate this change in DALYs by almost 10 percent. Fortunately, the data also show that strengthening food systems through investments in agricultural research and development, irrigation systems, market access, and infrastructure can significantly mitigate the effects of climate change and population growth on global hunger. This study provides critical information that delivering healthy diets to everyone through these investments can reduce the projected years lost, but action is urgently needed.

Review Publications
Muresan, L., Clapa, D., Borsal, O., Teodor, R., Wang, T.T., Park, J.B. 2020. Potential impacts of soil tillage system on isoflavone concentration of soybean as functional food ingredients. Land. 9(10):386.
Weaver, C., Fukagawa, N.K., Liska, D., Mattes, R.D., Matuszek, G., Nieves, J.W., Shapses, S.A., Snestelaar, L.G. 2020. U.S. documentation and regulation of human nutrition randomized controlled trials. Advances in Nutrition.
Park, J.B. 2020. Kahweol found in coffee inhibits IL-2 production via suppressing the phosphorylations of ERK and c-Fos in lymphocytic jurkat cells. Journal of Dietary Supplement. 18{1-11).
Cao, H., Sethumadhavan, K., Cao, F., Wang, T.T.Y. 2021. Gossypol decreased cell viability and down-regulated the expression of a number of genes in human colon cancer cells. Scientific Reports. 11:5922.
Liu, F., Liu, J., Wang, T.T., Changhu, X., Ziangzhao, M., Qingjuan, T., Li, R.W. 2020. Molecular and microbial signatures predictive of prebiotic action of neoagarotetraose in a dextran sulfate sodium-induced murine colitis model. Microorganisms. 8(7):995.
Liu, F., Smith, A.D., Solano Aguilar, G., Wang, T.T., Pham, Q., Tang, Q., Urban Jr, J.F., Xue, C., Li, R.W. 2020. Mechanistic insights into the attenuation of intestinal inflammation and modulation of the gut microbiome by krill oil using in vitro and in vivo models. Microbiome. 8:83.
Huang, G., Wu, Y., Liu, M., Sun, X., Lu, W., Gao, B., Wang, T.T., Yu, L. 2020. Potential biomarkers for early detection of 3-MCPD dipalmitate exposure in Sprague Dawley rats. Journal of Agricultural and Food Chemistry. 68(35):9594-9602.
Oliveira De Andrade, F., Liu, F., Zhang, X., Mariana, R., Dani, C., Cruz, I., Wang, T.T., Helferiche, W., Li, R.W., Hilakivi-Clarke, L. 2021. Genistein reduces the risk of local mammary cancer recurrence and ameliorates alterations in the gut microbiota in the offspring of obese dams. Nutrients. 12(1):201.
Wu, Y., Wang, J., He, Q., Yu, L., Pham, Q., Cheung, L., Zhang, Z., Kim, Y.S., Smith, A.D., Wang, T.T. 2020. Dietary Indole-3-carbinol alleviated spleen enlargement, enhanced IgG response in C3H/HeN mice infected with Citrobacter rodentium. Nutrients. 12(10)3148.
Yang, K., Xu, M., Cao, J., Qi, Z., Rahman, M., Holmen, B.A., Fukagawa, N.K., Zhu, J. 2021. Ultrafine particles altered gut microbial population and metabolic profiles in a sex-specific manner in an obese mouse model. Nature Scientific Reports. 11. Article 6906.
Fukagawa, N.K., Sulser, T.B., Beach, R., Dunston, S., Wiebe, K. 2021. Disability-adjusted life years (DALYs) due to chronic and hidden hunger under food system evolution with climate change and adaptation to 2050. American Journal of Clinical Nutrition.