Peripheral levels of plasma estrone (E1) and estradiol (E2), as measured by RIA, showed little correspondence in the cycling hamster. A diurnal pattern of Ex was apparent during the cycle with peak levels reached at 2200. Estradiol levels were low on the first 2 days of the cycle, correlated with the absence of antral follicles, but were higher at 2200 of day 2. A peak was reached at 1400 of day 3 (approximately 90 pg/ml) and the levels remained elevated for at least 8 hr. E2 was depressed at 0900 of day 4, but this was followed by an almost linear increase between 0900 and 1500 when the highest value for the estrous cycle was attained (186.6 pg/ml). A sharp decrease, down to baseline values, followed. The day 4 peak in E2 was temporally related to the preovulatory surge in LH and an abrupt increase in ovarian progesterone. Phenobarbital (Phen) injected to day 4 cycling animals prevented the E2 peak at 1500, whereas the addition of 5 μg of LH restored E2 to normal values. However, E1 was not affected by either Phen or LH. Injection of Phen to day 3 animals at 1300 lowered E1 and E2 levels, but this treatment does not affect the time of ovulation. LH administration on day 3 elevated E1 and E2 to levels comparable to day 4 cycling animals. Unilaterally ovariectomized animals displayed depressed E1 levels as compared to controls only on day 3 of the first cycle while E2 was significantly elevated on day 1 and lowered on day 3. By the second cycle E1 and E2 did not differ from intact control animals. Chronic hypophysectomy (for 8 days) yielded E1 values comparable to ovariectomized hamsters, but E2 values were significantly higher. LH administration did not increase either E1 or E2, indicating that antral follicles are necessary for optimal estrogen secretion. Plasma estrogen levels in the hamster were much higher than values previously reported for the rat, guinea pig and rabbit, and within the range found in man and rhesus. This seems to represent a true species difference since assays on rat and rabbit plasma, using the same RIA technique used with the hamster, yielded E1 and E2 values similar to those reported in the literature. (Endocrinology92: 805, 1973)