The in Vitro Metabolism of Estrogens by the Liver Studied by Means of a Colorimetric Method 12

Abstract

Since the work of Zondek (1) the in vitro action of liver slices and liver pulp on natural estrogens is usually called inactivation because the transformation undergone by these compounds results in loss of estrogenic activity as measured by bioassay methods (2). Using cc. of toluene were added to the mixture. All these operations were carried out in an ice bath.5 The colored material was extracted into toluene by shaking for 10 minutes in a shaking machine. After centrifuging for two minutes at 2,500 r.p.m., the toluene layer was pipetted off and read in a Beckman quartz spectrophotometer (Model DU) at 425 m#* in a 1 cm. cell against the blank.The blank was treated as the experimental sample except that no estrogen was present.Quantitative measurements of 17-j estradiol, estrone and estriol were obtained by this procedure between 10 and 150 micrograms, as indicated on Figure 1. The relation between optical densities and concentrations followed Beer's law between these two values. The slope of the curve was found to be similar for estrone and estradiol but different for estriol (Figure 1).The absorption spectrum of the colored compound showed a plateau-like peak between 400 mAi' and 450 miA with the maximum at 425 my approximately (Figure 2). 4 Kindly supplied by Drs. E. Schwenk and E. B. Hirshbirg, Schering Corporation, Bloomfield, New Jersey. 5 The tetrazotized dianisidine solution was prepared as follows: 25 mg. of dianisidine dihydrochloride (recrystallized from ethanol) were dissolved in 3 cc. of water. Three tenths cc. of concentrated hydrochloric acid, followed by 0.6 cc. of freshly prepared 5% sodium nitrite solution were added. After thorough mixing and a five minute waiting period, 0.66 cc. of a 5% urea solution was added. The solution was prepared in an ice bath, and made freshly for each experiment.341

Artificial Estrogens

“…Partial hepateetomy impairs the ability of the liver to inactivate "«''-estradiol (393). Liebermann, Tagnon, and Schulman (262) recorded high inactivating ability in vitro to rat liver with "«''-estradiol, estrone, and estriol. The average human liver was found to be capable of metabolizing 11.5 g. of "«''-estradiol in 25 hr.…”

The Metabolism of Estrogens by the Human Liver Studied in Vitro12

“…In the previous paper (1) recovery experiments were carried out with boiled rat liver slices in order to test the accuracy of this method. Similar experiments with boiled human liver slices were carried out here.…”