Five multiparous, ruminally cannulated Holstein cows (two lactating and three dry) weighing (X +/- SD) 667 +/- 35 kg were used to study the effect of abomasal purine infusion on the excretion of purine derivatives. Cows were fed corn silage four times daily at 90% of ad libitum intake (X = 9.16 kg of dry matter/d). Purines were infused into the abomasum as brewer's yeast suspensions in five incremental amounts (0 to 380 mmol/d) during five experimental periods according to a 5 x 5 Latin square design. Periods were 7 d; purine infusions were conducted during the last 4 d, and urine was collected during the last 3 d of each period. Ruminal purine outflow in all cows was measured during an experimental period immediately preceding and immediately following the five infusion periods and in each cow during the 0-mmol/d infusion period of the experiment. The relationship between total (milk plus urine) daily excretion of purine derivatives (allantoin plus uric acid) and total (abomasal infusion plus ruminal outflow) daily purine flow was quantified by linear regression analysis and was described by the relationship: Y = 0.856X + 103 (r2 = 0.93). The slope (0.856) indicated that 86% of purines that reached the omasum were excreted as purine derivatives. In the two lactating cows, urinary purine derivatives accounted for 98.4% of the total purine derivatives that were excreted. Ruminal flow of microbial CP can be estimated from the CP:purine ratio of ruminal microorganisms and the excretion of purine derivatives.