2010 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.
In collaboration, we previously have shown that the soybean plant compounds called glyceollins inhibit the growth of breast and ovarian cancer cells in vitro and in vivo (in laboratory test animals). During the last year, both the Agricultural Research Service (ARS) and University of Toledo (UT) have demonstrated beneficial effects of in vitro studies that suggest that the glyceollins may be additionally useful for the potential treatment of prostate cancer. In order to further study the glyceollins’ activity in animal models, UT previously developed a chemical synthetic scheme to produce two types of glyceollins, namely glyceollin I and glyceollin II. A synthetic route is desirable since the glyceollins (a mixture of glyceollins I, II, and III) remain difficult to purify from soy material as individual components. During the last year, a one-gram supply of glcyeollin I was forwarded to the ARS for efficacy and side-effect toxicity studies in animals. A second, one-gram supply was also prepared and it is being utilized by UT for pharmacokinetic (PK) studies. The PK studies will ascertain the stability, oral bioavailability, and half-life after administration to animals. For these assessments, UT has developed both an high-performance liquid chromatography (HPLC) and a highly sensitive HPLC-MS/MS (mass spectrometry) bioanalytical method. Synthetic chemistry efforts have turned toward producing glyceollin III as an analytical standard along with several other phytochemical intermediates and closely related structural analogues associated with the glyceollin family. Lastly, UT is continuing to analyze the effect of root cyst nematode infections on soybean plants after having detected glyceollins in this type of stressed root tissue. The major impact of this research could lead to alternative or value-added uses of induced phytoestrogen containing food and health products. Progress by cooperators was monitored through routine teleconferencing, meetings, and scientific presentations of information relating to the project at professional society meetings and conferences.