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Urine was collected from 55 cows via indwelling urinary catheters for periods of 12 h on different days (28 days prepartum to 60 days postpartum). Excretion of urinary creatinine (mg/h per kg bodyweight) among Holsteins increased from .94 on day 28 prepartum to 1.14 on day .5 postpartum and then decreased to .82 on days 30 to 45 of lactation. Excretion of creatinine among 12-h collections of urine on different days did not differ for groups of cows within periods prepartum and postpartum, and coefficients of variation within cows were 12 to 13%. Ratio of urinary estradiol (-17 alpha) to urinary creatinine was correlated (.93) more highly with its excretion based on volume of urine excreted than was its urinary concentration (.79) within periods prepartum and postpartum. Ratios of urinary metabolites to urinary creatinine rather than their concentrations should be used to express rates of excretion in cows' urine when urine excreted per hour is unknown. Moreover, daily rates of excretion of estradiol in urine can be estimated [ng estradiol/day = A X ng/mg urinary creatinine x kg bodyweight x 24 h, where A is average excretion of urinary creatinine (mg/h per kg bodyweight)] for respective days prepartum and postpartum.