<OL> <LI> Identify microbial interactions in legumes influencing biosynthesis of isoflavonoids with potential phytoestrogenic activities. <LI>Characterize microbial elicitors of isoflavonoid production and develop procedures to manipulate isoflavonoid levels. <LI>Test individual or combinations of isoflavonoid compounds induced by microbial elicitors for phytoestrogenic effects in animal systems.
Soybean varieties will be grown under greenhouse conditions. After various periods, plants will be harvested and phytoestrogenic isoflavonoids will be extracted. The compounds will be purified and identified using HPLC coupled with UV detection. In order to determine elicitation of phytoestrogenic isoflavonoids, soybean plants will be exposed to various phytopathogens and known elicitors derived from phytopathogens. After exposure to these elicitors, isoflavonoids will be purified and identified as above. Estrogenic activity of isoflavonoids isolated from soybean tissues/organs will be analyzed by determining their ability to support in vitro growth of an estrogen dependent cell line. Compounds will be assayed for estrogen activity in assays using a yeast hybrid system having an estrogen dependent promoter coupled with a glucuronidase reporter gene. Estrogen specific induction of the genes will be determined by colorimetric determination. Compounds will be assayed in order to determine synergistic effects and to measure estrogenic potency.