1. Protein-bound, trichloroacetic acid- (TCA) insoluble 99Mo appeared in plasma a few hours after the introduction of 99Mo-labelled molybdate (30 mg Mo) into the rumen of sheep maintained on a basic diet supplemented with elemental sulphur (3 g S/d).

2. Most of the 99Mo could be displaced from its protein carrier in vitro and the labelled compounds displaced were identified by Sephadex G-25 chromatography as di- and trithiomolybdate. Tetrathiomolybdate was not detected.

3. In control experiments protein-bound, TCA-insoluble 99Mo predominated in plasma after the direct administration of [99Mo]tetrathiomolybdate, either into the rumen or intravenously. The 99 Mo could be displaced in vitro and [99Mo]tetrathiomolybdate identified, although [99Mo]trithiomolybdate was also present.The study provides direct evidence of thiomolybdate synthesis and absorption in ruminants in vivo.

1. Protozoa were labelled by incubating 100 ml rumen fluid with [14C]choline for 1 h. The protozoa were concentrated by centrifugation and then washed with rumen fluid. This reduced residual 14C in the fluid medium to insignificant amounts whilestill retaining the viability of the labelled protozoa. Washing procedures using formal saline (40 g formaldehyde/1 saline (9 g sodium chloride/1)) and saline were developed to isolate protozoa for estimation of specific radioactivity.

2. The protozoal pool in freshly-collected rumen fluid incubated in vitro retained 90%of the radioactivity for up to 6 h following addition of 14C-I a belled protozoa produced as indicated previously. The specific radioactivity of protozoa did not change during the incubation period.

3. Protozoa labelled with [14C]choline and then stored until they died rapidly lost 14C to methane when they were incubated in rumen fluid or were injected into the rumen. Some [14C]choline was salvaged under these conditions by the live protozoa present as they apparently incorporated up to 13% of the label from the dead protozoa. However, protozoal debris from the injected solution could also have been present in the isolated protozoa.

4. The in vitro results suggested that the protozoal preparations were viable, and that the incorporated choline did not have a turnover in excess of the turnover of nitrogen (i.e. specific radioactivity remained constant with time in vitro) suggesting that the dilution of specific radioactivity of protozoa following mixing of a 14C-labelled dose of protozoa represented the rate of irreversible loss and also replacement of protozoa in the rumen.

5. 14C-labelled protozoa had a half-life in the rumen which was greater than that of rumen fluid and in six animals the protozoal replacement rate was l-4·l mg N/min.

6. Losses of 14C from labelled protozoa in the rumen in methane or via abomasal digesta were 65 and 35% respectively.

7. The results suggest that protozoal growth may be as high as 32% of the total microbial protein synthesis in the rumen but that 65% of the protozoa die and are degraded in the rumen.

1. A method for estimating the proportions of bacterial- and protozoal-N in the total non-ammonia-N reaching the lower gut of the ruminant under steady-state conditions was evaluated. Three trials using two different diets were conducted with a Holstein steer equipped with a rumen cannula and duodenal re-entrant cannulas.

2. An intraruminal primed infusion of (15nh4)2so4 was administered for 68 h during each trial. Bacteria and protozoa samples were isolated from rumen fluid at approximately 6 h intervals during each infusion period. Total non-ammonia-N was isolated from duodenal digesta samples taken at approximately the same times. All of these samples were analysed for 15N enrichment. A computer program was used to fit equations to the 15N-enrichment curves of bacterial- and protozoal-N. Models of both bacterial- and protozoal-N kinetics consisted of a small pool which equilibrated rapidly with rumen NH3 and a large pool with a fractional turnover rate of 0.045–0.070/h for bacterial-N and 0.056–0.069/h for protozoal-N.

3. Abomasal fluid turnover was estimated by a single injection of polyethylene glycol (molecular weight 4000) into the rumen followed by sampling of rumen fluid and duodenal digesta.

4. Estimates of abomasal fluid turnover, bacterial-N turnover, and protozoal-N turnover were entered into an equation which was adjusted by computer iteration to fit the 15n-enrichment curve of duodenal digesta non-NH3-N generated from each (15nh4)2so4 infusion period. The computer fit of this equation to the observed results gave estimates of 0.39–0.45 and 0.22–0.41 for the proportion of duodenal non-NH3-N derived from bacterial-N and protozoal-N respectively.

5. This method is potentially useful in estimating microbial protein passage to the lower gut in ruminants. Sampling digesta from the omasum rather than the duodenum would simplify the method and possibly increase the reliability of the estimates.

1. The deposition of fat during pregnancy and its subsequent mobilization during lactation was studied in the rat. The utilization of protein during lactation was also investigated.

2. Fat accumulation was rapid and continuous from early pregnancy. Approximately one-third was deposited subcutaneously, the remainder being stored in central depots.

3. The volume of milk produced during lactation was varied by offering protein at two concentrations from day 2 post partum. Optimum lactation was achieved with a high-protein diet. The dams lost no body protein, but 60% of the total body fat was mobilized, despite a marked increase in food consumption. On a low-protein diet, food intake did not rise during lactation. Some body protein was catabolized and, again, 60% of the fat was mobilized although milk output, as judged by the gain in carcass energy of their pups, was reduced to approximately one-third of optimum.

4. Protein was used more efficiently by lactating rats, when compared with virgin controls.

5. We conclude that fat stored during pregnancy makes a major contribution to the energy costs of lactation, and that its mobilization is under hormonal rather than dietary control.