A new direct radioimmunoassay measuring ovarian steroid metabolites in small aliquots of urine has been evaluated. For this purpose urinary levels of estrone-3-glucuronide (E1G) and pregnanediol-3-glucuronide (PG) were correlated with plasma values of estradiol (E2) and progesterone (P), respectively, from days 5 to 24 of the cycle in 24 women with normal cycles. Urine samples were representative of 1) first morning voiding; 2) 24-hour collection; and 3) all voidings except for the first morning one. The correlation between urinary E1G and PG with plasma values of E2 and P was statistically significant for the 3 collections of urine. The first morning voiding is selected as the sample of choice to determine E1G and PG due to its ease of collection and its high correlation with plasma E2 and P levels (r = .60, P less than .001; r = .73, P less than .001, respectively).