1. A mathematical treatment is given of the marker technique for studying water balances in the rumen. The treatment is extended to cover non-steady-state conditions and emphasis is placed on the underlying assumptions and practical limitations of the technique.

2. The 51Cr complex of ethylenediamine tetra-acetic acid (51Cr EDTA) was used as a marker in the rumen of sheep.

3. The 51Cr EDTA entered 95–98% of the water in the rumen. Mixing in the rumen, though adequate, was usually not quite complete even 1·5 h after injection of the dose. The techniques of injecting the dose and of withdrawing the sample were found to be important; some techniques often led to poor mixing.

4. Provided that the time of injection of the dose was suitably selected, reliable estimates of the volume of water in the rumen could readily be made, but reliable estimates of flow rates needed stringent attention to details of technique.

1. The metabolic responses to daily treatment with 1 μg cyanocobalamin were studied in sixteen patients with megaloblastic anaemia due to cobalamin deficiency.

2. The fall in serum iron was sluggish in most patients. The pattern of reticulocyte responses varied widely, the most common being a plateau with suboptimal maximal value.

3. Urinary propionic acid excretion before treatment was normal in two out of six patients. A transient rise during treatment was observed in two patients, possibly owing to demands on coenzyme stores. The fall in urinary propionic acid excretion during treatment was slow.

4. Serum vitamin B12 levels rose during treatment. In two patients an abrupt rise was found, possibly owing to release of stored cobalamin into the circulation.

1. The same six high-protein diets that were fed to rats (Carpenter & Anantharaman, 1968) have now been fed to chicks.

2. The net nitrogen retention by chicks, per 100 kcal consumed, was for each diet greater by 20–50% than the retention by rats. For a diet based on a mixture of commercial protein concentrates the NDpCal % was 19·1 (± 0·18); this value greatly exceeded the theoretical maximum of 14·6 obtained from the equations of Miller & Payne (1963).

3. Although groundnut protein plus lysine has a calculated chemical score of only 56, chicks receiving this at a high level retained N at the same rate (NDpCal % of 17·5–17·9) as those receiving a diet which included egg protein at the level (26% of the dietary ME) predicted to be optimal for them.

4. The ‘endogenous + metabolic’ losses of N were in almost the same proportion to metabolic size for chicks as for rats.

1.Eight groups of weanling rats were maintained for 60 days on diets containing calcium and phosphorus in the ratio 0·1 at four different levels of mineral, namely 0·08, 0·12, 0·23 and 0·32% Ca, and in the presence and absence of added ergocalciferol.

2. Provision of vitamin D increased final body-weight, whereas at the highest mineral intake body-weight was reduced.

3. At each level of dietary mineral the serum Ca concentration was raised by vitamin D, whilst the serum P concentration was lowered except when the mineral intake was lowest.

4. With increasing mineral intake the serum Ca increased until at the highest intake it declined. Serum P was increased only at the highest mineral intake.

5. Measurements were made of bone weights, ash content and volume. Provision of vitamin D increased the fresh weight of bone at all levels of dietary mineral, and the dry weight and ash content were increased except at the lowest level of mineral intake. In general, the higher the dietary mineral the greater was the bone weight and its ash content.

6. The presence of vitamin D resulted in an increase in tooth mass and ash content at each level of mineral intake. Tooth mass and ash increased with increasing dietary mineral except at the highest level when there was a decline.

7. At the lower levels of mineral intake the ratio of tooth mass to bone mass was greater than at the higher levels of intake.

8. Histological examination of the bones revealed no increase in the width of the epiphyseal cartilage in any group. In the group with the lowest levels of Ca and P without vitamin D (0·08% Ca, 0·8% P) the shafts of the femurs appeared thinner and porotic, and the trabeculae were thin; in the group fed the same diets supplemented with vitamin D the shafts were even thinner but less porotic; in all other groups the histological appearance of the bone was within normal limits.

9. Changes in the appearance of the dentine were demonstrated in all groups.

1. Examination of the digesta from all regions of the avian digestive tract showed that volatile fatty acids (VFAs) were present in greatest concentration in the caeca and that they comprised mainly acetic, propionic and butyric acids.

2. All droppings contained VFAs but they were present in highest concentration in those of caecal origin. Caecectomy was followed by a marked reduction in the total output of VFAs.

3. Birds 14–20 weeks of age had similar concentrations of VFAs along the tract and similar numbers and distribution of micro-organisms.

4. Portal blood contained all the VFAs found in the digestive tract whilst peripheral blood contained only acetic and formic acids.

5. The almost complete absence of VFAs from the tract contents of germ-free birds showed that the VFAs normally present in the tract were of microbial origin.

6. The presence of similar levels of acetate in the peripheral blood of conventional and germfree birds indicated that circulating acetate was mainly of endogenous and not microbial origin.

7. The significance of VFAs as an energy source is discussed.

1. In three experiments sheep were offered a choice of two rations: one high in NaCl (7.5 or 15%) and one low in NaCl. The intake of each ration was recorded when fresh water was freely available, when increasing amounts of NaCl were added to the drinking water and when the amount of fresh water available was restricted.

2. Low levels of NaCl in the drinking water or mild restriction of available water were without effect on the diet selected. At higher levels, total food intake was reduced and the reduction was sustained entirely by the high-salt ration. The intake of the low-salt ration remained the same or increased, so that the selected diet then contained an increased proportion of the low-salt ration. The effects of adding NaCl to the drinking water and of water restriction were similar.

3. It was concluded that sheep with a choice of high- and low-salt rations can partly avoid the stress of saline drinking-water or restricted water supply by changing the proportion of each ration eaten.

1. The rates of production, or entry rates, of acetic, propionic and butyric acids in the rumen of grazing sheep have been measured by a radioisotope technique previously used on penned animals.

2. Regression equations were derived relating entry rates to ruminal concentrations. The equations from penned and grazing sheep did not differ significantly.

3. The standard errors of entry rates predicted for a single sheep from equations based on all results were calculated to be ± 0·06, 0·18 and 0·15 m-mole/min for acetic, propionic and butyric acids respectively.

4. Interconversions between acetic and butyric acids in the rumen were similar in extent to those found in penned sheep. These values were used to calculate from entry rates the net amounts of the acids becoming available to the animals.

5. An equation was derived that permits the amount of energy supplied to an animal by the acids to be predicted from concentrations with a standard error of ± 0·17 kcal/min.

6. The applicability of the prediction equations is discussed.

1. The 51Cr complex of ethylenediamine tetra-acetic acid was used as a soluble marker substance in sheep fed restricted amounts (700 g daily) of roughage diets. It was injected into the rumen at various stages of the feeding cycle and rumen volumes and flow rates were calculated.

2. The average resting rumen volume for sixteen sheep (average body-weight 39 kg) was 3.9 l.; the average resting dilution rate was 0.077 h-1 so that the net inflow rate averaged 0.29 l./h; the average rate of absorption of water from the resting rumen into the blood was 0.05 l./h.

3. There were marked differences in all these values, both within sheep on different days (coefficient of variation nearly 20%) and between sheep (coefficient of variation more than 25%). The resting salivary inflow rate was also found to fluctuate considerably.

4. During feeding, the salivary inflow rate increased; there was usually both an expansion of rumen volume and an increased rate of outflow. There was no evidence of diffusion of water from the blood into the rumen.

5. Following eating, return to resting conditions was slow.

6. When the sheep drank, there was an expansion of the rumen volume and usually an increased outflow rate. The rate of absorption of water from the rumen increased slightly, but only for a short time. Occasionally after drinking it was found that rumen marker concentrations were slightly higher than expected probably because a small part of the imbibed water bypassed the rumen down the reticular groove into the omasum.

7. Most of the water drunk in a day was consumed in a few minutes. Compared with this rapid rate of drinking, the rate of absorption of water from the rumen was always a very slow process; this finding is in disagreement with recent views expressed in the literature. However, when considered over the whole day, the amount of water absorbed from the rumen may have been approximately equal to the amount of water drunk.

8. During periods of both feeding and drinking, the mean outflow rate was about 0.7 l./h in several series of experiments.

9. When sheep were held without food or water past their normal time for eating and drinking, the net inflow rate into the rumen soon started to decrease, but there was no immediate effect on rumen volume.

10. The present work reveals the potential usefulness of the soluble marker technique in defining the changing patterns of fluid movement in the rumen throughout the day.

1. Lambs fed on skim milk containing either maize oil or cod-liver oil or both were observed for signs of muscular dystrophy, changes in serum enzymes and increases in the susceptibility of red cells to haemolysis by peroxide (peroxide haemolysis).

2. Four lambs fed on the milk containing cod-liver oil, and not receiving α-tocopheryl acetate injections, developed acute muscular dystrophy, but no abnormal changes occurred in peroxide haemolysis, packed cell volume or haemoglobin concentration in the blood. Marked increases occurred in the aspartate transaminase, alanine transaminase and glutamate dehydrogenase, and in lactate dehydrogenase isoenzyme activities of the serum, presumably due to leakage from the degenerating tissues. One lamb given weekly injections of 100 mg α-tocopheryl acetate remained clinically normal and maintained normal serum enzyme levels. Treatment of dystrophic animals with 200 mg α-tocopheryl acetate by injection produced prompt clinical recovery and an exponential decline in the raised serum enzyme activities.

3. In the five lambs fed on the skim-milk containing maize oil, muscular dystrophy did not occur. Small but significant increases occurred, however, in serum aspartate transaminase and glutamate dehydrogenase activities. In all five animals peroxide haemolysis was greatly increased, but in only two did packed cell volume and haemoglobin values fall slightly below the lower limit of normal (mean − 2 × SD). Injections of 100–200 mg α-tocopheryl acetate neither prevented nor cured the abnormally high levels of serum enzymes and peroxide haemolysis, although a change in the diet was followed by a gradual return to normal. In one lamb given massive doses of α-tocopherol by mouth, peroxide haemolysis was not reduced. Large amounts of α-tocopherol added in vitro to erythrocytes from the lamb did, however, prevent peroxide haemolysis.

4. Peroxide haemolysis did not increase in any of three lambs fed on skim milk containing both maize oil and cod-liver oil, although two of them, which were given much larger amounts of cod-liver oil than the third, developed muscular dystrophy with elevated serum enzyme levels.

5. Peroxide haemolysis in lambs is evidently not a measure of vitamin E deficiency. It is suggested that increased peroxide haemolysis in lambs, like encephalomalacia in chicks, depends on the fatty acid composition of the unsaturated lipids in the diet.

1. Glucose entry rates into the blood and propionate production rates in the rumen have been measured in sheep given rations containing varying proportions of starch and roughage (lucerne).

2. Glucose entry rates and propionate production rates were similar for all rations studied.

3. The proportion of the glucose entry rate arising from propionate produced in the rumen was highest on the ration containing the greatest quantity of lucerne and decreased as the proportion of starch in the ration increased. Rate of conversion of propionate into glucose was estimated and was found to decrease as the amount of starch in the ration increased.

4. Concentrations of total volatile fatty acids (VFA) in the ruminal fluid were lowest in the ration with the greatest proportion of starch, implying lower VFA production rates with the starch rations although intakes of digestible energy were approximately the same. The mean concentrations and rates of production of propionate in ruminal fluid were similar for all rations.

5. The low VFA concentrations and the reduced conversion of propionate into glucose on the high rations, despite similar propionate production rates and glucose entry rates, may have been due to starch escaping ruminal fermentation. It is suggested that this glucose absorption may have reduced gluconeogenesis from propionate.

1. Fifty-one pregnant fine-wool Merino ewes were slaughtered at intervals during pregnancy. The gravid uteri were dissected and were separated into uterus, membrane and fluid, and foetus fractions, which were analysed separately for water, nitrogen, ash, fat, calcium, phosphorus, sodium and potassium content.

2. Uteri from twenty-one similar non-pregnant ewes were also analysed for these constituents.

3. A number of relationships between the composition of the gravid uterus minus the composition of the uterus taken from non-pregnant sheep, and time from conception were calculated.

4. The rate of nutrient deposition was calculated from these relationships and estimates were made of the nutrients utilized for pregnancy.

1. A laboratory-prepared groundnut flour defatted at room temperature (DGF) was subjected to dry heat or to pressure steaming under varying conditions of time and temperature, and its amino acid composition and nutritive value, the latter assessed by a chick growth test—the gross protein value (GPV) test, were compared with those of some commercial groundnut meals. Trypsin inhibitor activity, available lysine value (ALV) and ‘arachin’ and ‘conarachin’ content and, in some instances, GPV were estimated in the heated samples.

2. The amino acid composition of the DGF and of commercial meals of high, medium and low GPV did not differ markedly, and the GPV of the DGF fell within the range of the three commercial samples.

3. Both dry and moist heat under specified conditions lowered ALV in the DGF and in the ‘arachin’ fractions, but had little effect on the ALV of the ‘conarachin’ fraction.

4. Moist, but not dry, heat rapidly removed trypsin inhibitor activity, and dry, but not moist, heat lowered GPV.

5. Neither ‘conarachin’ content nor trypsin inhibitor activity correlated with GPV in a range of commercial groundnut meals.

6. Dry heat (125. for 5 h) lowered nutritive value and ‘conarachin’ content but did not reduce the amount of total nitrogen soluble in sodium chloride solution.

7. No trypsin-inhibiting activity was found in the testa (skins) but these did exhibit growth-depressant properties for chicks. This property was removed by mild moist heat treatment.

8. ‘Arachin’ isolated from a commercial groundnut meal was valueless as a protein supplement for a cereal ration for chicks; ‘conarachin’ by itself, and mixed with arachin (1:3) was equivalent in GPV to the parent meal.

9. A factor other than those considered here, and possibly unassociated with processing, is primarily responsible for the differences in growth-promoting qualities of the commercial groundnut meals used in this work.

1. The effects of heat treatment on the proteins of the groundnut were studied by examining the electrophoretic patterns of the proteins extracted from samples of a laboratory-prepared groundnut flour after various heat treatments.

2. Marked changes in the electrophoretic pattern of the conarachin fraction were found and these changes correlated with changes in nutritive value.

3. Marked differences were found between the electrophoretic patterns of the conarachin fractions extracted from twenty commercial groundnut meals, and these differences could be correlated with nutritive value.

4. It is suggested that an examination of the electrophoretic pattern of the conarachin fraction could provide a useful and rapid means of determining the nutritive value of a groundnut meal.

1. The skeletal response of vitamin D-deficient chicks to dietary ascorbic acid was tested.

2. The epiphyseal cartilaginous plate width, which was increased by the vitamin D deficiency, was further increased when ascorbic acid was added and the effect appeared sooner.

3. Absence of vitamin D was associated with a reduction in the relative amount of cancellous to compact bone ash in the tibia. Dietary ascorbic acid intensified this effect to a significant degree.

4. Plasma ascorbic acid levels in rachitic chicks were approximately 25% less than in controls. This observation may explain the skeletal response to added vitamin C and suggests that avian bone tissue has a particularly high requirement for this vitamin.

1. The purpose of this paper is to propose the measurement of scalp hair diameter as a useful index of protein synthesis in the management of cases of malnutrition.

2. Observations on scalp hair from ten control infants showed that each hair tapered very gradually towards its free end.

3. Observations on scalp hair from twenty-six infants with kwashiorkor showed that each hair narrowed towards the follicle; the effect was statistically significant.

4. After I month's treatment for kwashiorkor, scalp hair thickened towards the follicle and the effect was statistically significant.

5. Serial measurements of the greatest diameter at 0·5 mm intervals along the hair shaft had a main trend which was related to nutritional history.

6. The period of time over which narrowing of the hair shaft took place was calculated from the rate of hair growth and the serial diameter measurements: this time was always longer than the history of kwashiorkor.

1. The effects, on the secretion in milk of fat and its component fatty acids, of supplementing a basal diet low in fat with red palm oil at three levels are reported.

2. The secretion of total fat and of the fatty acids contained in the oil supplement was positively correlated with the dietary intakes, except for linoleic acid which was hydrogenated in the rumen before its absorption and secretion in milk.

3. Evidence for the synthesis de novo of palmitic acid and for the possible depression of intramammary de novo synthesis by high levels of dietary fat is discussed.

1. The total body calcium (Expt 1) of litter-mate male rats given a diet adequate in phosphorus and high (0·74%) or low (0·13%) in Ca from the age of 3 weeks was determined after 21, 48 and 60 weeks on the diet. In Expt 2, the ash content of ten groups of bones from 10-week-old rats given these same diets for 7 weeks was studied to ascertain whether all groups were equally affected by the difference in dietary Ca level. Four groups of bones, i.e. skull and mandibles, vertebras, the shafts and the ends of long bones, were consequently chosen for examination in Expt 3 where their growth and composition were studied in rats given the Ca diets for 24, 48 or 60 weeks from the age of 3 weeks.

2. Total body Ca (Expt 1), expressed as g Ca or as a percentage of the net body-weight or of net dry, fat-free carcass weight, was always significantly higher in rats given the higher level of Ca. The dietary effect was greatest after 21 weeks.

3. The higher level of dietary Ca led to a highly significant increase in the weight of the dry, fat-free bone and in the percentage ash content of all ten bone groups after 7 weeks (Expt 2). The greater treatment differences in ash content occurred with the less well mineralized bones.

4. In Expt 3, irrespective of diet, the four groups of bones developed at different rates. In rats given the diets for 24 weeks or longer, the vertebras and the shafts of long bones showed the greatest proportional increase in weight of dry, fat-free bone and ash relative to the corresponding values for 3-week-old rats; the greatest change in percentage ash content occurred in the ends of the long bones.

5. Increasing the level of Ca in the diet increased the weight of the dry, fat-free bone and its ash content. The effect of diet decreased with increasing time on the diets, in general persisting most strongly in the skull and mandibles and declining most rapidly in the ends of the long bones. The percentage Ca and P content of the bone ash was only slightly affected by diet and, except for the Ca in the ash of vertebras, significant differences were only found in the composition of the ash of bones from 27-week-old rats.

1. Total lipids were extracted from digesta obtained from the rumen, abomasum and upper small intestine (jejunum) of each of four slaughtered sheep. The lipids were fractionated into unesterified fatty acids, neutral lipids and phospholipids and the proportional contribution of each fraction to the total fatty acids was determined.

2. The contribution made by phospholipids to the total fatty acids in the digesta showed a marked increase in the samples from the small intestine compared with those from the rumen and abomasum. This increase was apparently due to the presence of biliary phospholipids.

3. Total lipids and conjugated bile acids were extracted from sheep bile, the lipids were fractionated and their fatty-acid composition was determined. Phospholipids predominated and these consisted mainly of phosphatidylcholine, together with some lysophosphatidylcholine.

4. Both phospholipids contained significant amounts of unsaturated C18 components which could account, at least in part, for the previously reported increament to the proportion of these acids in the digesta when it enters the upper jejunum.

5. The overall fatty acid compositions of the two biliary phospholipids were very similar and, in common with other naturally occurring phosphatidylcholines, the fatty acids present in position 2 of the phosphatidylcholine of bile were found to consist almost entirely of unsaturated components.

6. Total lipids and conjugated bile acids were extracted from samples of digesta obtained from three sheep with cannulas in different positions in the jejunum. Analysis of the lipids indicated that biliary phospholipids, in particular phosphatidylcholine, underwent progressive hydrolysis in the intestinal lumen.

7. The distribution of conjugated bile acids, unesterified fatty acids and phospholipids between the solid (particulate) and liquid (micellar) phases of the intestinal digesta was determined. These chyme constituents were, for the most part, associated with the particulate matter and thus, at any given time, it appears that only a small fraction of the total fatty acids is available for absorption in micellar form. It is suggested that the micellar solubilization of fatty acids may be facilitated by the presence of lysophosphatidylcholine.