First-pass uptake and oxidation of glucose by the splanchnic tissue in young goats fed soy protein-based milk diets with or without amino acid supplementation: glucose metabolism in goat kids after soy feeding.

The study was designed to examine whether feeding soy protein isolate as partial replacement of casein (CN) affects glucose metabolism in young goats and whether effects may be ameliorated by supplementation of those AA known to be lower concentrated in soy than in CN. Goat kids (d 20 of age) were fed comparable milk protein diets, in which 50% of the crude protein was either CN (control, CON), soy protein isolate (SPI), or soy protein isolate supplemented with AA (SPIA) for 43 d (n=8 per group). On d 62 of age, a single bolus dose of d-[(13)C6]glucose (10mg/kg of BW) was given with the morning diet, and simultaneously, a single bolus dose of d-[6,6-(2)H2]glucose (5mg/kg of BW) was injected into a jugular vein. Blood samples were collected between -30 and +420 min relative to the tracer administration to measure the (13)C and (2)H enrichments of plasma glucose and the (13)C enrichment of blood CO2. Glucose first-pass uptake by the splanchnic tissues was calculated from the rate of appearance of differentially labeled glucose tracer in plasma. Glucose oxidation was calculated from (13)C enrichment in blood CO2. In addition, plasma concentrations of triglycerides, nonesterified fatty acids, glucose, insulin, and glucagon were measured. On d 63 of age, kids were killed and jejunal mucosa and liver samples were collected to measure lactase mRNA levels and lactase and maltase activities in the jejunum and activities of pyruvate carboxylase and phosphoenolpyruvate carboxykinase (PEPCK) in the liver. Basal plasma glucose concentration tended to be higher in the CON than the SPIA group, whereas basal insulin was higher in the CON group than the SPI and SPIA groups, and glucagon was higher in the CON than the SPIA group. Plasma glucose and insulin concentrations increased during the first hour after feeding, whereas plasma glucagon increased immediately after feeding and after 1h of feeding. First-pass uptake and glucose oxidation were not affected by diet. Maltase activities in proximal and mid jejunum and lactase activities in mid jejunum were lower in the CON than in the SPIA group. Activities of PEPCK were higher in the SPIA than in the SPI group. In conclusion, feeding milk diets with soy protein isolate seems to affect glucose status in kids, but has no effect on first-pass uptake and oxidation of glucose. The highest activities of lactase and maltase were observed after supplementation with AA. Higher PEPCK activities in the liver may point at elevated gluconeogenic activities after AA supplementation in soy-fed kids.

Glucose transporters and enzymes related to glucose synthesis in small intestinal mucosa of mid-lactation dairy cows fed 2 levels of starch.

Diets containing corn starch may improve glucose supply by providing significant amounts of intestinal starch and increasing intestinal glucose absorption in dairy cows. Glucose absorption in the small intestine requires specific glucose transporters; that is, sodium-dependent glucose co-transporter-1 (SGLT1) and facilitated glucose transporter (GLUT2), which are usually downregulated in the small intestine of functional ruminants but are upregulated when luminal glucose is available. We tested the hypothesis that mRNA and protein expression of intestinal glucose transporters and mRNA expression of enzymes related to gluconeogenesis are affected by variable starch supply. Dairy cows (n=9/group) were fed for 4 wk total mixed rations (TMR) containing either high (HS) or low (LS) starch levels in the diet. Feed intake and milk yield were measured daily. After slaughter, tissue samples of the small intestinal mucosa (mid-duodenum and mid-jejunum) were taken for determination of mRNA concentrations of SGLT1 and GLUT2 as well as pyruvate carboxylase, cytosolic phosphoenolpyruvate carboxykinase, and glucose-6-phosphatase by real-time reverse transcription PCR relative to a housekeeping gene. Protein expression of GLUT2 in crude mucosal membranes and of SGLT1 and GLUT2 in brush-border membrane vesicles was quantified by sodium dodecyl sulfate-PAGE and immunoblot. A mixed model was used to examine feeding and time-related changes on feed intake and milk yield and to test feeding and gut site effects on gene or protein expression of glucose transporters and enzymes in the intestinal mucosa. Dry matter intake, but not energy intake, was higher in cows fed HS compared with LS. Abundance of SGLT1 mRNA tended to be higher in duodenal than in jejunal mucosa, and mRNA abundances of pyruvate carboxylase tended to be higher in jejunal than in duodenal mucosa. In brush-border membrane vesicles, SGLT1 and GLUT2 protein expression could be demonstrated. No diet-dependent differences were found concerning mRNA and protein contents of glucose transporter or mRNA level of gluconeogenic enzymes. In conclusion, our investigations on glucose transporters and gluconeogenic enzymes in the small intestinal mucosa of dairy cows did not show significant diet regulation when TMR with different amounts of intestinal starch were fed. Therefore, predicted intestinal glucose absorption after enhanced starch feeding is probably not supported by changes of intestinal glucose transporters in dairy cows.

Energy expenditure, urea kinetics, and body weight gain within a segregating resource family population.

Beef and dairy cattle represent divergent metabolic types that disseminate nutrients into either meat or milk and differ in nutrient accretion. To investigate nutrient flow and turnover in an animal model combining beef and dairy cattle, a crossbred experiment has been started. An F(2) resource population was generated from Charolais (beef breed) sires and German Holstein (dairy breed) cows as P(0) founders by consistent use of embryo transfer to establish the F(1) and F(2) generations, which accordingly comprised half- and full-sib offspring. In 64 bulls of 5F(2) families, dry matter intake and growth performance were measured monthly, and carcass composition was determined after slaughtering at 18 mo of age. Energy expenditure and urea kinetics were investigated via stable isotope tracer techniques using an intravenous single bolus dose of sodium [(13)C]bicarbonate [2.5 µmol/kg of body weight (BW), 99 atom% (13)C] at 8 and 18 mo of age and of [(15)N]urea (0.28 mg/kg of BW, 99 atom% (15)N) at 8 mo of age, respectively. Insulin responses were measured via glucose tolerances tests at the age of 8 mo. The results revealed significant differences between families for growth performance, energy expenditure, and urea kinetics. In summary, low energy expenditure was associated with high average body mass gain and high insulin response. A greater urea loss was associated with reduced muscle protein in carcass. In addition, corresponding half-sib and full-sib sisters from bulls with highest growth rate indicated highest milk production. In conclusion, we have demonstrated that differences in energy expenditure and urea kinetics result in differences in average daily gain and carcass traits and vice versa in F(2) crossbred bulls with common beef and dairy genetic backgrounds.

Morphology, proliferation, and ribonucleic acid and fractional protein syntheses in the small intestinal mucosa of young goats fed soy protein-based diets with or without amino acid supplementation.

The study was designed to examine whether feeding soy protein isolate as partial replacement of casein (CN) affects jejunal protein synthesis and whether effects may be ameliorated by supplementation of those AA known to be at lower concentrations in soy protein isolate than in CN. Goat kids (14 d) were fed comparable milk protein diets, in which 50% of the crude protein was CN (CAS), soy protein isolate (SPI), or soy protein isolate supplemented with AA (SPIA) for 43 d (n=8 per group). On d 42, plasma concentrations of protein, urea, and AA were measured before and after morning feeding. In the morning of d 43, [15N]RNA from yeast [13 mg/kg of body weight (BW)] was given with the diet to measure the reutilization of dietary RNA precursors for mucosal RNA biosynthesis. Four hours later, an oral dose of l-[1-(13)C]leucine (180 mg/kg of BW) was administered and blood samples were collected between -15 and +45 min relative to tracer administration for analysis of plasma 13C alpha-ketoisocaproic acid and 13C recovery in blood CO2. Kids were killed 60 min after the tracer application, and jejunal tissue was collected to determine mucosal morphology, cell proliferation, enzyme activities, RNA synthesis, and fractional protein synthesis rate. Plasma protein concentrations were higher in CAS than in SPI and SPIA. Plasma concentrations of Thr were higher in CAS than in SPI and SPIA, and those of Met were lower in SPI than in CAS and SPIA. In mid-jejunum, villus circumferences were higher in CAS than in SPI and SPIA, and villus height and villus height:crypt depth ratio were higher in CAS than in SPI. In mid-jejunum, mucosal protein concentrations were higher in CAS than in SPI and SPIA and mucosal activities of aminopeptidase N tended to be higher in CAS than in SPI, whereas activities of dipeptidyl peptidase IV tended to be lower in SPI than in SPIA. Activities of 5' nucleotidase and xanthine oxidase were lower in CAS than in SPI. The 13C recovery in blood CO2 tended to be higher in SPI than in CAS. In mid-jejunum, 15N enrichment of RNA tended to be higher in CAS than in SPI, and 13C enrichment of protein-bound Leu was higher in SPI than in CAS. In mid-jejunum, the fractional protein synthesis rate tended to be higher in SPI than in CAS. Our results revealed changes in intestinal growth after soy protein feeding that were associated with effects on intestinal RNA and protein synthesis but that were not ameliorated by AA supplementation.

Alterations in the jejunum of young goats caused by feeding soy protein-based diets.

This study was designed to investigate whether soy protein or soy protein supplemented with indispensable amino acids (AA) change the protein expression pattern and utilization of pre-cursors for RNA biosynthesis in jejunal mucosa in relation to casein and whether these changes affect mucosal cell growth. Kids were fed comparable diets based on cow;s milk, of which 50% of crude protein were replaced by either casein (CAS), soy protein (SP) or soy protein supplemented with indispensible AA (SPA) for 34 days (n = 4/group). Jejunal tissue was collected 5 h after adding a single dose of (15)N-RNA to the diet, in order to determine morphology, protein repertoire by two-dimensional gel electrophoresis and matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry, and RNA biosynthesis by isotope ratio-mass spectrometry. In mid-jejunum, morphological alterations induced by partial replacement of casein with soy protein were accompanied by changes in mucosal proteins related to generation of the cytoskeleton and in pathways for mucosal RNA biosynthesis, resulting in a smaller re-utilization of dietary RNA pre-cursors and in an increased activity of enzymes involved in nucleic acid breakdown. Soy protein supplemented with indispensible aminoacids tended to revise mucosal growth retardation with no impact on salvage of dietary RNA pre-cursors for mucosal RNA biosynthesis, but changes in cytoskeleton generation. Feeding soy protein with supplementation of indispensible AA does not ameliorate soy protein effects on mucosal morphology and RNA metabolism in the jejunum in a significant manner.

Effect of a soy protein-based diet on ribonucleic acid metabolism in the small intestinal mucosa of goat kids.

This study was designed to investigate the effect of soy protein inclusion in milk replacer diets for goat kids on protein, RNA, and DNA contents in small intestinal mucosa, on the importance of RNA biosynthesis from dietary RNA precursors for mucosal RNA synthesis, and on the activities of enzymes involved in nucleotide degradation in small intestinal mucosa. Diets were based on cow's milk. In the control group, 35% of the milk protein was replaced by casein (CN) protein, and in the soy group (SPAA), the same amount of milk protein was replaced by soy protein supplemented with essential AA known to be at lower concentrations in soy than in CN (Thr, Val, Ile, Leu, His, Lys, Met). Diets were isonitrogenous and isoenergetic. At 47 d of age, goats were harvested and samples of proximal, middle, and distal jejunal mucosa were collected 5 h after feeding 15N-labeled RNA from yeast (13 mg/kg of body weight). Growth and feed conversion did not differ between the control and SPAA kids. Mucosal protein concentrations were lower in the SPAA than the control kids. Concentrations of RNA and DNA did not differ between feeding groups, but in all kids mucosal RNA concentrations were higher in proximal than in middle and distal jejunum. Protein:RNA ratios were higher in the control than the SPAA kids and were lowest in proximal jejunum. Activities of alkaline phosphatase in enterocytes were higher in proximal than in middle and distal jejunum. Activities of mucosal xanthine oxidase were highest in distal jejunum and were higher in the SPAA than the control kids, especially in the middle and distal sites. The 15N-enrichment of mucosal RNA was higher in the control than the SPAA kids, especially in distal jejunum, and was lowest in distal jejunum. In contrast, 15N-enrichment of urea in plasma tended to be higher and Gly concentration in plasma was lower in the SPAA than the control kids. Data indicate that protein content and the protein:RNA ratio were lower in jejunal mucosa of goat kids fed milk replacer with partial replacement of CN protein by soy protein. These findings were accompanied by a lower level of reutilization of preformed dietary RNA precursors for RNA biosynthesis in jejunal mucosa and a higher activity of xanthine oxidase. Thus, feeding soy protein instead of CN protein reduced the incorporation of preformed dietary RNA precursors for RNA biosynthesis in the mucosa and activated key enzymes involved in nucleic acid breakdown.

[Utilization of glucose and long-chain fatty acids in lactating dairy cows fed a fat-enriched diet]. / Verwertung von Glucose und langkettigen Fettsäuren durch die laktierende Milchkuh bei Fütterung einer fettangereicherten Diät.

The fate of carbon from long-chain fatty acids and glucose in dairy cows which were fed with protected fat was studied using stable isotope technique. The experiment was carried out on two groups of dairy cows (n=16 in each group) during the first 15 weeks of the lactation period. The cows were fed isoenergetic and isoproteinogenous diets based on corn silage. About 1.8 kg of tapioca starch in the diet of the starch group was substituted by about 0.7 kg of rumen protected fat (Ca salts of palm oil and soybean oil) in the diet of the fat group. The carbon atoms of dietary fat were naturally depleted in 13C as compared to carbon atoms of starch. Daily milk performance and lactose output were significantly (P < 0.05) higher among the cows fed with fat diet. In comparison to the starch group, the enrichment of milk fat with 13C was significantly lower, while that of breath CO2 was significantly higher in the fat group (P < 0.05). This means the fatty acids were incorporated into milk fat in preference to metabolic oxidation. Further studies showed that blood glucose is oxidized to a lower extent and is used for the synthesis of lactose to a higher proportion if the cows were fed with the fat diet. The glucose entry rate into the body glucose pool was not different between the diets. In conclusion, the dietary fatty acids perform a glucose sparing effect and improve the glucose supply for the mammary gland.

Lactation Biology Symposium: role of colostrum and colostrum components on glucose metabolism in neonatal calves.

In neonatal calves, nutrient intake shifts from continuous glucose supply via the placenta to discontinuous colostrum and milk intake with lactose and fat as main energy sources. Calves are often born hypoglycemic and have to establish endogenous glucose production (eGP) and gluconeogenesis, because lactose intake by colostrum and milk does not meet glucose demands. Besides establishing a passive immunity, colostrum intake stimulates maturation and function of the neonatal gastrointestinal tract (GIT). Nutrients and nonnutritive factors, such as hormones and growth factors, which are present in high amounts in colostrum of first milking after parturition, affect intestinal growth and function and enhance the absorptive capacity of the GIT. Likely as a consequence of that, colostrum feeding improves the glucose status in neonatal calves by increasing glucose absorption, which results in elevated postprandial plasma glucose concentrations. Hepatic glycogen concentrations rise much greater when colostrum instead of a milk-based colostrum replacer (formula with same nutrient composition as colostrum but almost no biologically active substances, such as hormones and growth factors) is fed. In contrast, first-pass glucose uptake in the splanchnic tissue tended to be greater in calves fed formula. The greater plasma glucose rise and improved energy status in neonatal calves after colostrum intake lead to greater insulin secretion and accelerated stimulation of anabolic processes indicated by enhanced maturation of the postnatal somatotropic axis in neonatal calves. Hormones involved in stimulation of eGP, such as glucagon and cortisol, depend on neonatal diet, but their effects on eGP stimulation seem to be impaired. Although colostrum feeding affects systemic insulin, IGF-I, and leptin concentrations, evidence for systemic action of colostral insulin, IGF-I, and leptin in neonatal calves is weak. Studies so far indicate no absorption of insulin, IGF-I, and leptin from colostrum in neonatal calves, unlike in rodents where systemic effects of colostral leptin are demonstrated. Therefore, glucose availability in neonatal calves is promoted by perinatal maturation of eGP and colostrum intake. There may be long-lasting effects of an improved colostrum supply and glucose status on postnatal growth and development, and colostrum supply may contribute to neonatal programming of performance (milk and growth) in later life, but data proving this concept are missing.

Energy metabolism in the newborn farm animal with emphasis on the calf: endocrine changes and responses to milk-born and systemic hormones.

Neonatal mammals need adaption to changes in nutrient supply because energy intake shifts from continuous parenteral supply of nutrients (mainly glucose, lactate, and amino acids) via the placenta to discontinuous colostrum and milk intake with lactose and fat as main energy sources. Besides ingested lactose, endogenous glucose production is essential in the neonate to assure sufficient glucose availability. Fetal endogenous glucose production is low, but endocrine changes (especially the prenatal rise of glucocorticoid production) promote maturation of metabolic pathways that enable marked glycogen synthesis before and enhanced gluconeogenesis after birth to establish an adequate glucose status during postnatal maturation. In preterm born farm animals gluconeogenic activity is low, mainly because of a low glucocorticoid and thyroid status. In full-term neonates, endogenous glucose production increases with age. Colostral bioactive components (such as growth factors, hormones, bioactive peptides, and cytokines) do not have a direct effect on endogenous glucose production. However, colostrum feeding stimulates intestinal growth and development, an effect at least in part mediated by bioactive substances. Increased nutrient and glucose absorption thus allows increased glucose supply and hepatic glycogen storage, which improves the glucose status. The improved energetic status of colostrum-fed neonates is reflected by an accelerated maturation of the somatotropic axis, leading especially to enhanced production of IGF-I in the neonate. Secretion and production of hormones involved in the regulation of glucose and fat metabolism in neonates depend on the developmental stage and the response to feeding. In addition, many such hormones have actions in the neonate that differ from adult animals. Endocrine action to support endogenous energy supply in neonates is probably not fully established, and therefore, needs postnatal maturation. Therefore, our knowledge on energy metabolism in the neonate needs to be extended to better understand the function and the failure and to assess endocrine responses during the neonatal period.

Ruminal degradability of 15N labelled ribonucleic acid in grass.

The ruminal degradation of RNA in rye grass (Lolium perenne) was studied using the bag method. A non-lactating cow (BW 550 kg) fitted with a rumen cannula was used and fed twice daily at maintenance level with a chopped grass hay-based ration containing 30% ground barley. Rye grass, labelled during growth by fertilization with 15N2-urea (9.5 atom% 15N, 20 g N/m2), was cut at seven stages of growth and maturity and freeze-dried. RNA-N represented 6 to 17% of total N. Labelled grass samples (milled to 5.0 mm screen, 5.0+/-0.1 g DM) were incubated in polyester bags (100 x 200 mm, pore size 50 microm) in the rumen for periods of 1, 3, 6, 9, 12, 24, and 48 h. Data of N and RNA disappearances from the bags were fitted to an exponential equation to estimate parameters of degradation. The effective degradability of RNA in the rumen averaged 90+/-4%, for N it was 11% units lower (P < 0.001). Degradability of RNA was correlated to that of N (R2 = 0.92). Degradability of RNA (R2 = 0.96) and N (R2 = 0.93) decreased with increasing fibre content of grass. Increasing the fibre content by 1% diminished the degradability of RNA and N by 1.1% units and 2.4% units, respectively (P < 0.001). Assuming a microbial protein synthesis in the rumen of 150 g/kg DOM, a N: RNA ratio of 1:1.35 in rumen microbes and a rumen outflow rate of 0.06 h(-1), a model calculation indicates that about 9 to 19% of duodenal RNA are of dietary origin in animals fed grass. This should be taken into account for the calculation of microbial N on the basis of RNA as marker.

[Evaluation of different markers for the determination of microbial nitrogen flow into the duodenum of dairy cows]. / Bewertung verschiedener Marker für die Messung der mikrobiellen Stickstoffpassage am Duodenum der Milchkuh.

2,6-Diaminopimelic acid (DAPA), ribonucleic acid (RNA), 15N, D-alanine (D-ALA) and the amino acid profiles (AAP) were compared as microbial markers for determination of the microbial protein synthesis in the rumen. Three dairy cows (Schwarzbuntes Milchrind, LW 602 kg), each fitted with a rumen cannula and a re-entrant cannula in the proximal duodenum, were offered four isoenergetic and isonitrogenous diets (mean daily intake 15.0 +/- 0.45 kg DM; forage: concentrate = 50:50) in a periodic experiment. The diets contained soyabean extracted meal, meat and bone meal, pea meal and dried clover as major sources of protein. On the 4th day after administration of 9 g 15N-labelled urea (95 atom-% 15N-excess) per day, samples of rumen fluid and duodenal digesta were obtained 3 h after feeding. The bacteria were isolated by differential centrifugation. Bacteria harvested from the rumen had significantly higher 15N enrichment and D-ALA: N ratio than 'duodenal' bacteria. However, DAPA: N ratio was higher in 'duodenal' bacteria compared to rumen bacteria. There were no differences in RNA: N ratio between rumen and 'duodenal' bacteria. The source of the bacteria in the digestive tract has an influence on the ratio of microbial N: total N, especially when 15N, AAP, DAPA and D-ALA but not RNA were used as markers. The most reproducible method was D-ALA (C.V. 4.7 for rumen and 6.8 for 'duodenal' bacteria) followed by 15N (10.8 resp. 4.8) and RNA (9.7 resp. 8.2). The results obtained with 15N and D-ALA agreed closely at the same source of bacteria. The RNA method reached the level of these markers (15N, D-ALA) when the bacteria were isolated from the duodenum. It is concluded that D-ALA (bacteria isolated from rumen and duodenum) and also 15N (bacteria isolated from duodenum) were the best markers for estimation of the microbial protein synthesis.

Effect of the pH value when treating concentrate protein with formaldehyde on protein protection in the rumen.

Nine samples of soybean extracted meal were treated by fumigation in plastic bag with 0.5 g formaldehyde/100 g crude protein, the pH ranging from 2.1 to 10.7. The studies showed that with rising pH (x) the proportion of totally bound (y1, % of formaldehyde amount used) and irreversibly bound (y2) formaldehyde increased (y1 = 77.7 + 1.56x, y2 = 28.2 + 2.04x), whilst the reversible proportion remained constant (45.4% of formaldehyde amount used). The lysine detectable (g/16 g N) after HCl hydrolysis was reduced due to pH raising (y = 101 - 0.67x). Solubility and fermentability of the soybean protein in the rumen were found to rise though. Consequently, the formaldehyde content of the protein was positively correlated with the content of soluble N and fermentable N, respectively, and negatively correlated with the lysine content. These experimental results allow to conclude that the desired protein protection to be attained by treating soybean extracted meal with formaldehyde, is greatly influenced by the respective pH value. To reach maximum protection of the protein against microbial degradation in the rumen (N degradation after 12 hours incubation less than or equal to 20% of total N), the pH value should be below 5. The contents of totally, reversibly or irreversibly bound formaldehyde does not allow any conclusion regarding the protein protection attained. The apparently increased binding rate of formaldehyde is presumably due to the fact that here the reaction stops at the stage of methylol formation (molar proportion 1:1). Under the conditions of acid protein hydrolysis for lysine determination, the released formaldehyde obviously reacts irreversibly with the E-NH2 group of lysine.

[The use of ribonucleic acid as a marker for measuring microbial protein yield in the rumen. 1. Chemical determination of ribonucleic acid]. / Untersuchungen zur Nutzung von Ribonukleinsäure als Marker für die Messung des mikrobiellen Proteinertrages im Pansen. 1. Mitteilung. Chemische Bestimmung von Ribonukleinsäure.

The determination of ribonucleic acid (RNA) in the duodenal digesta and the rumen microbes was carried out colorimetrically with orcinol after extraction with hot NaCl solution, precipitation with phosphotungstic acid and alkaline hydrolysis. 96 +/- 1% of the RNA supplemented to the digesta content was recovered (y +/- s, n = 5). The methodic error (repetition accuracy) was 1.1%. The relation of RNA-N:N (mg/g) in the bacteria mass amounted to 93.8 +/- 3.99 (y +/- s(-y), n = 48) and to 16.1 +/- 0.8 (y +/- s(-y), n = 51) mg RNA/g DM in the duodenal content.

[The use of ribonucleic acids as markers for the measuring of microbial protein yield in the rumen. 2. The effect of sample treatment, the time of sampling and the composition of the ration on the RNA:N ratio in rumen microbes]. / Untersuchungen zur Nutzung von Ribonukleinsäure als Marker für die Messung des mikrobiellen Proteinertrages im Pansen. 2. Mitteilung. Einfluss der Probenbehandlung, des Probenahmezeitpunktes und der Rationszusammensetzung auf das RNS: N-Verhältnis in den Pansenmikroben.

In two experiments the influence of the treatment of samples, the sampling time and the composition of the rations on the RNA: N ratio in the rumen microbes was checked. Experiment I proved that freezing (-21 degrees C), thawing and freeze-drying of isolated bacteria and protozoa from the rumen fluid and from the duodenal content did not result in a change of the RNA content and the RNA-N: total N relation. If, however, the rumen fluid is stored deep-frozen before the isolation of the bacteria the N content in the DM of the bacteria decreases by 17% and that of RNA by 30%. This results in a change of the RNA: N relation of 16%. In conclusion, the bacteria are to be isolated immediately after rumen fluid sampling. Isolated bacteria can be stored deep-frozen before RNA determination and then freeze-dried. Experiment II showed that the RNA content of the rumen protozoa varies according to the period after feeding. The RNA: N relation was 0.50, 0.92, 0.70 and 0.58 on average 0, 3, 6 and 8 h after feeding, in which the 3rd hour after feeding can obviously be considered the time of increased microbial activity. The conclusion from this variation is that more than one isolation of microbes must be carried out in the course of the day in order to achieve representative samples. These statements apply to easily and not easily fermentable protein as N source in the feed. It could also be proved that no essential variation is to be expected in the RNA: N relation in the microbes isolated from the rumen fluid in the range of 8-21% crude protein in the DM of the ration (roughage: concentrate = 55: 45). On average the rumen microbes contained 1.7 g RNA-N/16 g N, essential differences between bacteria and protozoa could not be ascertained. From the slight variation of the RNA-N: N relation in the isolated bacteria from various cows one can conclude that there is no need to isolate the microbes of each individual animal.

Investigations on the influence of duodenal histidine infusion on nitrogen and amino acid turnover of growing German Holstein bulls.

The effect of a continuous duodenal infusion of L-histidine (His) (8 g/d) on the retention of nitrogen was investigated in two experiments (I, II), each of which was carried out using two young bulls. In Exps. I and II, the animals (150-250 kg BW) were fitted with a re-entrant cannula in the proximal duodenum and were fed diets containing 125 g CP/kg DM and 11.5 MJ ME/kg DM. A third experiment (III) using two young bulls (140-200 kg BW) fitted with a simple T-cannula was carried out infusing 6 g L-His. The animals were fed a low protein diet (94 g CP/kg DM and 11 MJ ME/kg DM). The study was done to find out whether or not L-His is the first limiting amino acid (AA) for growing ruminants. N retention was 28 and 31, 38 and 38, 22 and 24 g/d without L-His infusion and with L-His infusion for Exps. I, II and III, respectively. Both in the experiments with a standard protein supply (I, II) and in the experiment with reduced protein supply (III), no significant differences were found between periods with and without infusion of L-His. The utilisation of duodenal NAN varied between 39% and 50% and was also not significantly influenced by the duodenal infusion of L-His. No significant effect was observed on the flow of AA into the duodenum. The faecal excretion of AA was also not significantly influenced by the infusion of L-His. The utilisation of individual amino acids as calculated by the ratio of retained AA to intestinal apparently digested AA, did not differ significantly following the duodenal infusion of L-His. As expected, the utilisation of His decreased. Of the different essential AA, L-His was the most utilised (80%) followed by Arg (72%), Met (60%), Leu (45%) and Lys (44%), during periods without supplementation of L-His. It is concluded that the intestinal supply of L-His from the basal diet was sufficient for the potential growth level of animals under these experimental conditions. In all AA present at the proximal duodenum, L-His could have at first a limiting effect on the performance of growing young bulls with high body gain. Arg and Met, but not Lys, could be second or co-limiting AA.

[The effect of starch sources barley, maize and potatoes and their ration portions on the nutrient digestibility and energy utilization in ruminants. 4. Nitrogen metabolism in the rumen]. / Einfluss der Stärkeherkünfte Gerste, Mais und Kartoffeln und ihrer Rationsanteile auf die Nährstoffverdaulichkeit und die Energieverwertung bei Wiederkäuern. 4. Mitteilung--Stickstoffumsatz im Pansen.

In 9 experimental periods on four adult bulls (LW 550 kg) fitted with re-entrant cannulae in the proximal duodenum isoenergetic rations were used on feeding level 1.7 with ground barley, ground maize or fresh potatoes as starch sources. The net energy parts of these concentrates in the ration amounted to 50, 25 and 10%. 50 to 80% of the ration DM consisted of dried grass and about 10% of sugar beet pulp. The dried grass supplied on an average 87, 79 and 62% of the feed crude protein. The intake of DM was 7.74 +/- 0.42 (mean +/- SD) kg/d. The energetic efficiency of microbial N synthesis in the rumen (g N/kg organic matter true fermented in the rumen, TFOM) was averaged 16.4 with a range of 10.6 to 21.4. The microbial efficiency achieved a maximum when the ratio of nitrogen-free extract to crude fibre in the diet was 1.7 and 2.1 with barley, 1.8 with potatoes and 2.1 and 3.3 with corn as starch source. Changes in the microbial efficiency were positively correlated with the rate of passage of non-microbial organic matter from the rumen (g/d) and with the duodenal flow rate (kg digesta/kg DM intake). The relation to the rate of carbohydrate fermentation in the rumen (in %) and to the amount of TFOM (g/d) was negative. The duodenal flow of microbial N and non-ammonia N (g/d) correlated negatively with the organic matter apparently fermented in the rumen (AFOM) and positively with the non AFOM. The amino acid (AA) profile of the duodenal protein was affected by the starch source. It was concluded that the metabolism of nitrogen in the forestomachs of cattle is affected by the source of starch and the ratio of forage to concentrate. There exists a relationship between both factors. The net synthesis of microbial protein in the rumen is not only the result of substrate fermentation. The passage of non-AFOM from the rumen significantly affects the energetic efficiency of microbial nitrogen synthesis and the duodenal supply of AA.

Effects of protozoa on methane production in rumen and hindgut of calves around time of weaning.

Effects of the presence or absence of ciliate protozoa on methanogenesis in the rumen and hindgut were investigated in young calves during a 7-week period. Ten Holstein calves, aged 7 days, were divided in two groups (n = 5) and fed an increasing amount of a commercial milk replacer and small amounts of a calves starter. One group was inoculated with ciliate fauna on two occasions, week 5 and 6, while the second remained ciliate-free. The absence of protozoa in the rumen decreased rumen empty weight (-23%, P < 0.01), and rumen pool size of N (-36%, P < 0.01) and crude fat (-37%, P < 0.05). Rumen bacteria of non-faunated calves contained a higher proportion of total amino acid-N per 16 g N (+3%, P < 0.01) and D-alanine-N per 16 g N (+13%, P < 0.05) compared to faunated calves. Further results contain a reference for a higher bacterial mass in the ciliate-free rumen with an increased number of bacteria adherent to rumen mucosa. The CH4 production in the rumen increased exponentially with the increase in protozoa population size (R2 = 0.68). In presence of 46 x 10(4) protozoa per ml rumen fluid, the in vitro CH4 production of rumen fluid per mol total VFA was about 34% higher in faunated than in non-faunated calves (P < 0.001). Hydrogen (2H) recovery of rumen fermentation was positively correlated (R2 = 0.55) to the CH4 production rate. Methanogens were attached on rumen mucosa. Methanogenesis, induced by rumen mucosa attached bacteria, was stimulated by ruminal protozoa. In the absence of protozoa in the rumen, the acetate-propionate ratio and butyrate proportion of VFA were reduced. In vivo, in the absence of protozoa not only the whole animal CH4 production (-30%, P < 0.05) but also the digestibility of carbohydrates (-4%, P < 0.05) was reduced. Thereby no difference was observed in the intake of ME per kg DM between the groups. In conclusion, the methanogenesis in the rumen, but not in hindgut, is associated with the development of the ruminal protozoa population. The level of methanogenesis (mol/mol VFA) in the hindgut amounts to 20% of the ruminal methanogenesis.

Passage of ribonucleic acid along the intestine of sheep.

Sheep (Flemish female x Texel male, 55 kg BW), fitted with a PVC cannula in the dorsal rumen and single T-shaped PVC cannulas in the proximal duodenum, distal duodenum, mid-jejunum and terminal ileum were fed hay or hay-concentrate diets at various levels of nitrogen and cell walls (NDF) (22 to 32 g N/d; 150 to 699 g NDF/d). Co-EDTA and Cr-NDF were used as markers to measure the flow rate of digesta. Ribonucleic acid (RNA) intestinal digesta and in rumen bacteria was determined with orcinol after extraction with sodium chloride, precipitation with tungstophosphoric acid and alkaline hydrolysis. The RNA:total N ratio in bacteria, harvested from the rumen, amounted to 0.70 (CV 4.4%). The apparent digestibility of RNA in different sections of the intestine was higher than of total N. About 6% of RNA entering the duodenum disappeared between the proximal and distal duodenum. At jejunum, the net disappearance of RNA amounted to 68% of the quantity which entered the proximal duodenum. A higher result of 71% was obtained at the ileum. Total net disappearance of RNA between the proximal duodenum and rectum averaged 75%. Sixteen percent of RNA leaving the ileum was apparently digested in the large intestine. The true digestibility of RNA between the proximal duodenum and the terminal ileum, as estimated by multiple regression analysis, amounted to 78%. Of the amount of RNA entering the ileum, 24% was of endogenous origin. At ileum, the RNA passage was positively related to the ileal flow of NDF (R2 = 0.67) and N (R2 = 0.94). The passage of RNA increased by 3 mg RNA per g ileal indigestible NDF. Ileal endogenous N consisting of approximately 2% of endogenous RNA-N. In conclusion, the digestion capacity in the first part of the small intestine is high. Rising flows of indigestible cell walls and nitrogen increase the loss of ileal RNA. Further, using RNA as a microbial marker to assess the amount of microbial protein entering the duodenum of ruminants, digesta samples should be collected immediately post pylorus at the proximal duodenum, in order to avoid underestimation of the microbial protein synthesis in the rumen.

Studies on the evaluation of feed protein for ruminants. 1. Passage of nitrogen fractions to the duodenum of dairy cows fed different protein and carbohydrate sources.

The duodenal passages of non-ammonia nitrogen (NAN), amino acid N (AA-N) and microbial N (MN) were measured in seven duodenal fistulated dairy cows (6130 kg FCM/305 d) during lactation receiving 30 different rations similar to those used in dairy practice. The rations consisted of roughage (protein-rich silages, hay) and concentrate mixture (corn, barley, oat) in a ratio of 60:40 on a DM basis or roughage/concentrate mixture/fodder beets in the proportions 60:20:20 or 60:0:40 respectively. N supplements consisted of soya bean meal and peas meal (untreated or treated with formaldehyde in each case), rapeseed meal, fish meal and urea. DM intake varied between 9.6 and 19.1 kg/d, the crude fibre content between 166 and 270 g/kg DM, the crude protein (CP) content between 111 and 184 g/kg DM, the AA content (N basis) of the CP between 51 and 82%, the unfermentable CP content of the CP between 23 and 49% and the organic matter digestibility between 70 and 79%. The duodenal passage of NAN was 24.2 +/- 2.7 g/kg DM intake, 34.9 +/- 4.1 g/kg apparently digestible organic matter (in total tract, DOM), 43.3 +/- 5.5 g/kg apparently digestible carbohydrates or 4.2 +/- 0.5 g/MJ net energy fat, (y +/- s, n = 90). The value measured for AA-N was 16.8 +/- 2.9 g/kg DM intake, and that for RNA-labelled microbial CP was 146 +/- 26 g/kg DOM. NAN passage (g/kg DM intake) correlated more closely with the duodenal digesta flow rate (DFR) (kg digesta/kg DM intake) as an intrinsic animal factor (r = 0.78) and with the rumen content dilution rate (RDR) (passage of microbial-free organic matter at duodenum/kg BW0.75.h) as a ration dependent factor (r = 0.64) than with the UDP fraction (% of ration CP) (r = 0.50) or other ration parameters. It was concluded that the specific effect of protein concentrates in mixed rations on duodenal NAN yields in lactating dairy cows is lower than hitherto assumed. The duodenal NAN yield of a single feedstuff or ration (g/kg DM) in cows is dependent on animal and feeding factors. The DFR could represents a suitable target for breeding activities.

Influence of dietary concentrate to forage ratio on the development of rumen mucosa in calves.

Effects of structural and non-structural carbohydrates on the development of rumen fermentation and ruminal mucosa in calves were examined during the weaning period. Barley/soybean meal (SBM) group was fed a concentrate starting from 2 weeks of age, whereas alfalfa group received a mixture of concentrate and alfalfa hay in which the proportion of the latter was gradually increased from 20% to 70% between weeks 2 and 9 of age. The total volatile fatty acid concentration in rumen fluid of calves increased with age, but at 9 weeks there were no significant differences between the two diets (barley/SBM group 153 mmol/l, alfalfa group 150 mmol/l). Rumen papillae at 9 weeks of age, as compared to 6 weeks of age, were longer and fewer in number per square centimetre mucosa, with larger cut surface. This resulted in a higher surface of papillae per square centimetre mucosa at 9 weeks (barley/SBM group 286 mm2/cm2, alfalfa group 245 mm2/cm2) than at 6 weeks of age (barley/SBM group 217 mm2/cm2, alfalfa group 198 mm2/cm2). At 9 weeks of age, the pH (barley/SBM 5.0, alfalfa 5.7), the acetate to propionate ratio (barley/SBM 2.2, alfalfa 3.2) as well as the length of the papillae in the ventral ruminal sac (barley/SBM 1.96 mm, alfalfa 2.37 mm) were increased in the alfalfa group when compared to the barley/SBM group (P < 0.1). In the former group, the proportion of butyrate revealed significantly increased values at 4 and 6 weeks of age. In animals of the barley/SBM group at 9 weeks of age, characteristic protrusions with proliferated thick epithelium occurred on the papillae and increased the surface for absorption. On the epithelium (Stratum corneum) desquamating cells with parakeratosis could be observed. In the alfalfa group the papillae of the ventral ruminal sac were longer, without protrusions. The morphotypes of the adhering rumen microflora differed between the groups. It can be concluded that feeding greater amounts of non-structural carbohydrates increases the surface for absorption of the rumen epithelium in calves. The absence of hyperkeratosis and rumenitis in the barley/SBM group indicated that there is no reason to limit high starch diets in the early weaning period of calves.