Dietary protein is extensively degraded in the rumen into ammonia some of which which is in turn used by rumen bacteria to synthesize microbial protein. Excess ammonia is converted to urea in the liver and excreted in urine and milk. Microbial protein (MCP) is digested and absorbed as amino acids and nucleic acids. Nucleic acids are degraded in the liver into allantoin and excreted in the urine and milk. The ratio of urea to allantoin therefore is an indicator of efficiency of utilization of protein in the feed. Our aim was to investigate the efficiency of protein utilization by determining the urea:allantoin ratio in the milk of cows in the 1, 2, 3 and 6+ months of lactation. Herd test milk was collected from 44 HF and HF-Jersey cross cows (HF 75% to 100%) on a commercial dairy herd in Northern Victoria, Australia. The herd consisted of 2 cohorts, one in early lactation and one in the second half of lactation. All cows in the herd were offered the same ration (flat-feeding), consisting of approximately one-third of DM as concentrate and two-thirds of DM as grass based forage. Part of the forage was grazed pasture consisting predominantly of perennial English ryegrass (Lolium perenne) with a scattering of clover and some weeds. The milk was analyzed for urea concentration by an enzymatic method. Allantoin concentration was determined using high performance liquid chromatography. The MUN:MAC ratio was significantly different (P < 0.05) between cows in 1st month of lactation (27.92 ± 2.28) and cows in 2nd, 3rd or 6+ months in lactation (34.84 ± 1.68, 43.89 ± 5.35, 45.60 ± 3.42, respectively). This ratio was also significantly different (P < 0.05) between cows in 2nd month of lactation and 6+ months of lactation. Cows in the first month of lactation had a significantly higher MAC and lower MUN (P < 0.05) than cows 3 or more months in lactation. These results indicate that cows in early lactation are more efficient in the utilization of dietary protein