Optimization of a yeast estrogen screen and its applicability to study therelease of estrogenic isoflavones from a soygerm powder

Here we describe a redesigned protocol of the yeast estrogen screen develop ed by Routledge and Sumpter. The redesigned test comprises two steps. First , a large a-mount of yeast with estrogenic compounds is incubated for 24 hr . Subsequently, a mixture of cycloheximide and the chromogenic substrate ch lorophenol red-beta -D-galactopyranoside (CPRG) is added. The cycloheximide stops protein synthesis and allows for an end-point measurement of P-galac tosidase activity generated during the first 24 hr. CPRG is converted to ch lorophenol red and reflects beta -galactosidase activity, which is indicati ve of the estrogenic activity. The modifications shorten the duration of th e assay at least I day and avoid interference of the estrogenic CPRG or chl orophenol red. The redesigned and the original protocol were used to study the estrogenic activity of bisphenol A, methoxychlor, p,p ' -DDT, and isofl avones (genistein, daidzein, and glycitein). Bisphenol A, methoxychlor, and genistein triggered higher levels of beta -galactosidase activity in the r edesigned protocol. Estrogenic activity of p,p ' -DDT could only be demonst rated with the redesigned protocol. Glycitein and daidzein failed to give a response with both protocols. We also studied deconjugation of beta -glyco sidic isoflavones present in soygerm powder. Treatment of the soygerm powde r with beta -glycosidase released isoflavones. The estrogenic response of t he samples was confirmed with the redesigned protocol and correlated with t he amount of genistein present. The release of isoflavones under conditions prevailing in the intestines was studied. Bacterial beta -glycosidase pres ent in the large intestine released isoflavones, and moderate estrogenic ac tivity could lie demonstrated.