SUMMARYIn ruminants, the urinary excretion of purine derivatives (PD) reflects the absorption of microbial purines and can be used as an index of microbial protein supply. The objective of this study, carried out in Aberdeen, 1992, was to examine whether PD concentrations in spot urine or plasma samples vary diurnally during a given feeding regime and if they reflect differences in daily PD excretion induced by varying feed intake. Sixteen sheep were offered ad libitum one of four diets (fresh weight basis, the rest of each diet being minerals and vitamins): (1) 99·9% lucerne (pelleted); (2) 50% hay, 30% barley, 9% fishmeal and 10% molasses; (3) 72% straw, 7% molasses and 20% molassed sugarbeet pulp; and (4) 97% barley. Measurements were made for 1 week after a 2-week adaptation period. Urine was collected daily on days 1–4 and hourly on days 5–7. Hourly urine collection was achieved using a fraction collector. Plasma samples were collected hourly from 09·00 to 17·00 h on day 4. Feed intake varied considerably (347–1718 g DM/day) between diets and between animals. Daily excretion of PD (7·1–22·6 mmol/day) was linearly related to DM intake (r = 0·85, n = 16), and so was the microbial N supply (3·9–19·5 g N/day) estimated from daily PD excretion (r = 0·87). In hourly urine samples, the ratio of PD:creatinine concentrations showed no significant difference between sampling times, and was linearlycorrelated with the daily PD excretion (r = 0·92). Similarly, plasma PD concentration also showed little diurnal fluctuation. Glomerular filtration rate (GFR) increased with feed intake. Plasma PD was not well correlated with daily PD excretion in urine (r = 0·57). The tubular load of PD (plasma PD × GFR) was better correlated with the daily excretion (r = 0·80). It appears that when sheep are fed ad libitum, PD in spot urine may provide a practical indicator of microbial protein supply status.