The C-17 fatty acid esters of estradiol are naturally occurring estrogens which have been shown to circulate in blood. They are long-acting estrogens, analogous to the synthetic alkyl and aryl esters of estrogens which have been used pharmacologically for decades. To determine the mechanisms involved in the prolonged stimulation evoked by these nonpolar estrogens, several C-17 alkyl esters were synthesized and labeled with 3H at C-17 alpha, and their metabolism and clearance were studied and compared to those of estradiol in rats. The conversion of the C-17-3H to 3H2O was used as a marker of metabolism. While the clearance of the long chain esters from blood is somewhat slower than that of estradiol (t 1/2 = approximately 16 vs. 2 min, respectively), the rates of metabolism are dramatically different. For example, the t 1/2 of metabolism for two representative esters, estradiol-17-stearate and arachidonate, are 580 and 365 min, respectively, while the t 1/2 of metabolism for estradiol is about as fast as its clearance from blood (approximately 2 min). When the effect of chain length was studied, it was found that for the smaller esters, there was an inverse relationship between the size of the acyl group and the clearance from blood, i.e. the longer carboxylic acids were cleared more slowly. However, when the acyl group was lengthened from C12 to C14, the rate of clearance increased and was even faster with C18. Nevertheless, with all of the esters tested, the rate of metabolism steadily decreased as the chain length increased. These results are interpreted as indicating that the control point or rate-limiting step in the metabolism of the estradiol esters is the esterase that hydrolyzes the ester to estradiol. Thus, the prolonged estrogenic action of the C-17-alkyl esters is due to the slow release of estradiol from this hydrophobic reservoir.