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United States Department of Agriculture

Agricultural Research Service

Research Project: DETERMINE NATURAL PRODUCT INDUCTION IN LEGUMES AND THE PHARMACOLOGIC CONSEQUENCES IN HUMAN MODEL SYSTEMS
2011 Annual Report


1a.Objectives (from AD-416)
Identify stress factors and modify biochemical feed stocks that can influence the production of legume phytochemicals. Test individual combinations of phytochemicals in human model systems. Characterize procedures that can enhance the levels of natural products having estrogenic, anticancer, or potential cancer preventative activities. Perform computer modeling of potential phytoalexin structures for PPAR agonist activity.


1b.Approach (from AD-416)
Soybean cultivars will initially be grown under well-controlled test tube conditions. After various growth periods, roots will be harvested and their natural products extracted. Selected compounds will be purified and identified using HPLC and various instrumental methods applicable to organic structural characterization such as MS, NMR, IR, and UV. These experiments will then be repeated in the presence of various stress factors such as cyst root nematodes, and in the presence of modified biochemical feed stocks such as aryl-substituted phenylalanine and cinnamic acid derivatives, in order to determine useful elicitation methods and to produce altered phytochemical materials, respectively. Interesting components will again be purified and identified, after which the pharmacologic properties for all materials will be assessed as either individual or combinations of phytochemicals using in vitro models for estrogenic, anticancer, and potential cancer preventative activities. The most promising natural products will be produced in larger quantities by scaling-up the appropriate controlled-growth conditions and by conducting chemical synthesis, as necessary, to support further pharmacological assessments in an expeditious manner. The latter will involve the use of in vivo models indicative of potential estrogenic activity, anticancer therapy, or cancer prevention in humans. Useful growing conditions connected to the most pharmacologically promising compounds or mixtures of components, will be extended to greenhouse settings and eventually to field environments.


3.Progress Report

Two approaches were developed to chemically synthesize soy glyceollin I (GLY I), which has been shown to be the active anticancer component of the soy glyceollin mixture. The first is a follow-up to our previous total synthesis and then multi-gram preparation of this precious material wherein we are now trying to devise an alternative chemical route that does not require the use a metal catalyst. This will alleviate all concerns about the potential toxicity when metals found even in only trace amounts within the final product material. Our second approach toward producing large-scale quantities of GLY I has involved attempts to obtain material from local soybean crops that have been stressed by attack from cyst root nematodes. As a follow-up to our previous total syntheses of GLY I, several synthetic approaches remain ongoing to obtain glyceollins II and III (the remaining 2 components of the soy glyceollin mixture). Importantly, a biomimetic synthesis, which mimics the natural biosynthesis in soy, of GLY I was also achieved. Coupled with results from University of Toledo (UT’s) biological testing activities, the biomimetic route has just been published. Another important contribution from these types of chemical efforts that occurred during this reporting period, is the successful production of the dehydrated forms of GLY I and glycinol. To better understand how the GLYs interact with estrogen receptors at the molecular level, we are conducting computational chemistry studies. Specifically, we are addressing the ability of each GLY to bind inside the estrogen receptor cavity. As a follow-up to our prior papers, studies completed successfully during this reporting period were recently published. The most noteworthy finding is that GLY I is not stable at the acidic pH normally found in the empty stomach such that neutralization to pH greater than 3 will be required for oral studies in rodents. Similarly, eventual consumption of these materials as part of the diet is likely to require them to be taken as part of a neutralizing meal, if not pre-formulated so as to avoid decomposition altogether. Research progress was monitored through teleconferencing, frequent email communications and reports.


Last Modified: 12/18/2014
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