The purpose of this research is to identify metabolic pathways affected by intake of dietary bioactives and to determine their mechanisms of action. Bioactives of particular interest include polyphenols from berries, lycopene, beta-carotene and vitamin A, and isothiocyanates. Samples from previously conducted human studies will be used to support most of the objectives for the 2020 research activities. This Inter-Agency Agreement will capitalize on joint expertise of the NCI DCP NSRG and the USDA Beltsville Human Nutrition Research Center to conduct analyses on samples from human feeding interventions to better understand the role of diet in cancer prevention. More specifically, objectives of this research are: 1) to evaluate the metabolomics profile in urine and plasma using both NMR and LC-MS from the beta-carotene and lycopene tomato juice human feeding study conducted previously, 2) to analyze carotenoid absorption after consumption of freeze-dried broccoli, 3) to analyze isothiocyanates in urine after consumption of freeze dried broccoli, and 4) to evaluate pro-inflammatory cytokines in blood after consumption of brassica vegetables. Research on the reduction of cancer risk associated with the consumption of Brassica vegetables has been particularly promising. However, the unique biochemistry of the glucosinolate-myrosinase system in Brassica vegetables presents challenges that must be overcome. Hydrolysis of gluroraphanin leads to an unstable intermediate and subsequently to sulforaphane via a Lossen-type rearrangement, or in the presence of epithiospecifier protein (ESP), to sulforaphane nitrile. Studies have shown that a significant portion of glucoraphanin from broccoli is converted to sulforaphane nitrile due to ESP endogenous to broccoli. Importantly, sulforaphane has potent anticancer activity but sulforaphane nitrile has little or no such activity. Thus, it would be advantageous to prepare broccoli such that the formation of sulforaphane is favored over that of sulforaphane nitrile. Ideally, methods for optimizing sulforaphane production from cooked broccoli could be developed, given that most Americans preferentially consume cooked over fresh broccoli. Thus, we are also proposing to conduct a randomized crossover human trial to compare the net accumulation of urinary sulforaphane and other glucoraphanin metabolites following consumption of either fresh broccoli or cooked broccoli supplemented with a small amount of raw daikon radish. Typically, glucosinolate levels are maintained in steamed broccoli although myrosinase and ESP activity become negligible with heating. Daikon radish has the remarkable property that it contains myrosinase but no ESP. Therefore, this study may show that cooked broccoli served with raw daikon radish results in higher levels of sulforaphane than those resulting from an equivalent portion of fresh broccoli.
