This experiment was conducted to determine the effects of slow release urea (SRU) and its interaction with crude protein (CP) level in the diet on N metabolism in Holstein steers. Eight rumen-cannulated Holstein steers (body weight 265 ± 18 kg) were used in a replicated 4 × 4 Latin square design with a 2 × 2 factorial treatment structure. Treatment factors were the CP level in the diet, 10.9% versus 12.1% CP, and the non-protein nitrogen source used, urea versus SRU. Total collection of urine and faeces for 7 days allowed the estimation of N retention and diet digestibility. In addition, blood and rumen sampling allowed estimation of rumen fermentation and blood N profiles. Decreasing CP intake from 12.1% to 10.9% reduced urinary N output, but also reduced diet digestibility and N retention. When compared to urea, SRU did not alter N retention, but reduced ruminal ammonia and plasma urea concentrations. Although SRU did not improve N retention at either CP level, rumen ammonia and plasma urea concentrations were reduced, which may indicate that SRU may carry a lower risk for toxicity when compared to urea when fed at higher dietary concentrations.