The goal of this study was to measure the effects of level of feeding and the form of progesterone (P4) administration on the concentrations and yields of faecal P4 metabolites relative to differences in plasma P4 concentrations in non-lactating cows. Six non-lactating Holstein-Friesian cows were ovariectomised (OVX) and allocated to two groups: (i) P4 by subcutaneous injection (P4-s.c., n=3); and (ii) P4 administration per vaginum (P4-p.v., n=3). Each cow in the P4-s.c. group was injected subcutaneously once daily with 200mg P4. Each cow in the P4-p.v. group had a CIDR device inserted for 11 days when it was removed and replaced with a second device for further 11 days. Cows were fed a ration containing lucerne (33%) and oaten (66%) chaff at a maintenance level (M) in two portions in the first period of the study, and at a half-maintenance (1/2M) level during the second period. Chromic oxide capsules (Cr(2)O(3)) were administered twice daily to allow faecal output (FO) to be estimated. Plasma P4 and faecal P4 metabolites (FP4M; 20-oxo-pregnanes, 20alpha- and 20beta-OH-pregnanes) were measured during the treatment period. Daily FO declined after reducing the M diet to 1/2M (4.77 versus 2.61kg; P<0.01), whereas plasma P4 concentrations increased in the P4-s.c. group (4.2 versus 6.2ng/ml; P<0.05), but not in the P4-p.v. group (0.9 versus 1.0ng/ml; P>0.2). The mass of P4 released from a CIDR device during each 11-day period (M or 1/2M) was similar (0.66 versus 0.63g). Faecal 20-oxo-pregnanes (20-oxo-) concentrations were not affected by day or level of feeding, whereas faecal 20alpha-OH (20alpha-) and 20beta-OH (20beta-) concentrations were increased with the 1/2M diet in the P4-s.c. group (4.3 versus 5.6 microg/gDM; 2.2 versus 5.6 microg/gDM, respectively; P<0.05), but not in the P4-p.v. group (2.3 versus 2.7 microg/gDM; 1.7 versus 3.04 microg/gDM P>0.05). These changes in concentration only partly compensated for the reduced FO with the 1/2M diets as daily yields of FP4M (20-oxo- and 20alpha-) were greater during the M diet period (20-oxo-: 6.9 versus 4.1 microg/gDM; 2.7 versus 1.5 microg/gDM, for P4-s.c. and P4-p.v. groups, respectively; P<0.05, 20alpha-: 19.9 versus 13.6 microg/gDM; 10.9 versus 6.6 microg/gDM for P4-s.c. and P4-p.v. groups, respectively; P<0.05). The level of feeding and the route of P4 administration had key roles in controlling P4 concentrations in blood and daily FP4M yield.