Improvement and validation of the method to determine neutral detergent fiber in feed

To improve the performance of the analytical method for neutral detergent fiber in feed with heat‐stable α‐amylase treatment (aNDFom), the process of adding heat‐stable α‐amylase, as well as other analytical conditions, were examined. In this new process, the starch in the samples was removed by adding amylase to neutral detergent (ND) solution twice, just after the start of heating and immediately after refluxing. We also examined the effects of the use of sodium sulfite, and drying and ashing conditions for aNDFom analysis by this modified amylase addition method. A collaborative study to validate this new method was carried out with 15 laboratories. These laboratories analyzed two samples, alfalfa pellet and dairy mixed feed, with blind duplicates. Ten laboratories used a conventional apparatus and five used a Fibertec^® type apparatus. There were no significant differences in aNDFom values between these two refluxing apparatuses. The aNDFom values in alfalfa pellet and dairy mixed feed were 388 g/kg and 145 g/kg, the coefficients of variation for the repeatability and reproducibility (CV_r and CV_R) were 1.3% and 2.9%, and the HorRat values were 0.8 and 1.1, respectively. This new method was validated with 5.8% uncertainty (k = 2) from the collaborative study.     

Molecular characterisation of carbohydrate digestion and absorption in equine small intestine

Dietary carbohydrates, when digested and absorbed in the small intestine of the horse, provide a substantial fraction of metabolisable energy. However, if levels in diets exceed the capacity of the equine small intestine to digest and absorb them, they reach the hindgut, cause alterations in microbial populations and the metabolite products and predispose the horse to gastrointestinal diseases. We set out to determine, at the molecular level, the mechanisms, properties and the site of expression of carbohydrate digestive and absorptive functions of the equine small intestinal brush-border membrane. We have demonstrated that the disaccharidases sucrase, lactase and maltase are expressed diversely along the length of the intestine and D-glucose is transported across the equine intestinal brush-border membrane by a high affinity, low capacity, Na+/glucose cotransporter type 1 isoform (SGLT1). The highest rate of transport is in duodenum > jejunum > ileum. We have cloned and sequenced the cDNA encoding equine SGLT1 and alignment with SGLT1 of other species indicates 85-89% homology at the nucleotide and 84-87% identity at the amino acid levels. We have shown that there is a good correlation between levels of functional SGLT1 protein and SGLT1 mRNA abundance along the length of the small intestine. This indicates that the major site of glucose absorption in horses maintained on conventional grass-based diets is in the proximal intestine, and the expression of equine intestinal SGLT1 along the proximal to distal axis of the intestine is regulated at the level of mRNA abundance. The data presented in this paper are the first to provide information on the capacity of the equine intestine to digest and absorb soluble carbohydrates and has implications for a better feed management, pharmaceutical intervention and for dietary supplementation in horses following intestinal resection.     

Electrophysiological characterization of electrolyte and nutrient transport across the small intestine in horses

The aim of this study was to characterize the transport mechanisms of electrolytes and nutrients across the jejunum of nine healthy horses electrophysiologically. The stripped mucosa was mounted in Ussing chambers and tissue conductances (G_t) and short circuit currents (I_sc) were continuously monitored. After blocking the sodium and potassium channels with amiloride, tetraethylammonium chloride (TEA) and barium, chloride secretion was stimulated by carbachol and forskolin. Subsequently, chloride channels were inhibited by 4,4′‐diisothiocyanato‐stilbene‐2,2′‐disulfonic acid, 5‐nitro‐2‐(3‐phenylpropylamino)benzoic acid, CFTR_inh‐172, N‐(2‐naphtalenyl)‐(3.5‐dibromo‐2.4‐dihydroxyphenyl)methylene glycine hydrazide (GlyH‐101) and glibenclamide and their dose–response effect was investigated. The response to glucose, l ‐alanine and glycyl‐l ‐glutamine was determined at two different mucosal pH values (pH 7.4 and 5.4 respectively). Mean basal I_sc was ?0.47 ± 0.31 μEq/cm²h and mean G_t was 22.17 ± 1.78 mS/cm². Amiloride and TEA did not alter the baseline I_sc. Barium, carbachol and forskolin significantly increased I_sc. Irrespective of the dose, none of the chloride inhibitors changed I_sc. All nutrients induced a significant increase in I_sc with the increase being significantly higher at pH 7.4 than at pH 5.4. In conclusion, there is evidence that chloride secretion in horses may be different from respective transport mechanisms in other species. The glucose absorption is suggestive of a sodium‐dependent glucose cotransporter 1. However, a decrease in luminal pH did not stimulate current response to peptides as shown for other mammals.     

Nitrogen and amino acid absorption in the small intestine of growing pigs. 2. Apparent and true crude protein digestibility and amino acid absorption to the end of the small intestine

Studies of the apparent and true ileal digestibility of crude protein and the apparent and true amino acid (AA) absorption were carried out with 7 female pigs in the live weight range between 22 and 34 kg after ileorectostomy. Protein sources of varied quality with regard to the method of treatment (toasted) and untoasted soybean oilmeal) and to the AA composition (vital gluten or vital gluten + L-lysine X HCl) were used. Distinct differences regarding the apparent ileal digestibility of crude protein and the apparent absorption of amino acids were discovered between the protein sources, the cause of which is to be seen in the different amounts of endogenous N. The true ileal digestibility of crude protein and amino acid absorption were partly distinctly higher than the values of the apparent ileal digestibility of crude protein and AA absorption. On the whole one can say that the results of the investigation gained by means of ileorectostomy can be compared to the values gained from cannulated animals. Thus the experimental method used is suitable for studying the absorption process to the end of the small intestine of pigs.     

Intrauterine growth restriction alters nutrient metabolism in the intestine of porcine offspring

Background:Intrauterine growth restriction (IUGR) has negative impacts on the postnatal survival, growth and development of humans and animals, with not only on...     

Effect of dietary phytic acid on performance and nutrient uptake in the small intestine of piglets

An experiment was conducted with piglets to determine the effect of dietary phytic acid supplementation on performance, electrophysiological properties of jejunum mounted in Ussing chambers, sodium-dependent glucose transporter 1 (SGLT1) protein expression in jejunum, and plasma glucose and Na concentrations. Sixteen piglets with an average initial BW of 7.40 ± 0.36 kg were randomly assigned to 2 experimental diets with 8 piglets per diet. The diets were casein-cornstarch-based and were either unsupplemented or supplemented with 2% phytic acid (as Na phytate). The basal diet was formulated to meet the recommendation of NRC (1998) for energy, AA, minerals, and vitamins for piglets. The experiment lasted for 21 d, and at the end, BW gain and feed consumption were determined, and blood samples were collected for determination of plasma glucose and Na concentrations. The piglets were then euthanized to determine jejunal electrophysiological properties (transmural potential difference and short-circuit current) and SGLT1 protein expression. Phytic acid supplementation reduced ADG (P = 0.002), ADFI (P = 0.017), and G:F (P = 0.001) from 316.1 to 198.2 g, 437.4 to 360.3 g, and 0.721 to 0.539 g/g, respectively. Phytic acid supplementation also tended to reduce (P = 0.088) potential difference (-3.80 vs. -2.23 mV) and reduced (P = 0.023) short-circuit current from 8.07 to 0.1 μA/cm(2). However, phytic acid supplementation did not affect SGLT1 protein, and blood plasma glucose and Na concentrations. In conclusion, dietary phytic acid reduced growth performance and transmural short-circuit current in the jejunum of piglets. The reduced transmural short-circuit current in the jejunum by phytic acid implies reduced active Na transport in the jejunum by the phytic acid. Therefore, it seems that dietary phytic acid reduces growth performance of pigs partly through reduced capacity of the small intestine to absorb Na.     

Simultaneous isolation and characterization of brush border and basolateral membrane vesicles from bovine small intestine

Purified brush border and basolateral membranes were isolated from homogenized intestinal enterocytes of Holstein steers by divalent cation precipitation followed by differential and sucrose density gradient centrifugation. Alkaline phosphatase and Na/K adenosine triphosphatase served as marker enzymes for the brush border and basolateral membranes, respectively. The brush border and basolateral membrane fractions were enriched 5.1- and 10.1-fold, respectively, over the cellular homogenate. Electron micrographs, obtained with transmission electron microscopy, confirmed the vesicular nature of the membranes and revealed that basolateral membrane vesicles generally were smaller and more irregular in shape than brush border membrane vesicles. The vesicular nature of isolated membrane preparations was confirmed with osmotic activity experiments. Enrichment of brush border and basolateral membrane fractions compared to the initial homogenate and the vesicular configuration of both preparations indicate that the isolated brush border and basolateral membrane preparations were suitable models for evaluating nutrient transport properties of bovine small intestine. The number of transport experiments possible per animal using the membrane vesicle technique is many times more efficient than some other in vitro techniques (i.e., intestinal rings or everted sacs).     

Effect of dietary fibres on small intestine histomorphology and lipid metabolism in young broiler chickens

Two experiments were conducted to determine the influence of dietary fibres on small intestine histomorphology and lipid metabolism in broilers from 1 to 21 day of age. In experiment 1, diets containing insoluble [cellulose (CEL); 2% and 4%] or soluble [carboxymethyl cellulose (CMC); 2% and 4%] fibre were fed to broilers from day 1 to 21 post‐hatch and ileal tissue was collected at day 21 of age for histological evaluation. In experiment 2, broilers diet was supplemented with 0%, 1% or 2% insoluble fibre (Arbocel) during day 7 to 21 post‐hatch and plasma and liver lipid metabolism were evaluated at day 21. In experiment 1, inclusion of CMC reduced body weight gain (BWG) and feed intake (FI) and increased feed conversion ratio (FCR) compared with others. Intestinal histomorphology was unaffected by CEL, but CMC led to an increase in crypt depth (CD) and serosa thickness and a decrease in villus height (VH), villus width (VW), VH:CD ratio and villus surface area (VSA), rather than control and CEL groups. Treatment did not affect goblet cell type. Moreover, the CMC‐fed birds had greater total goblet cell count (GCC) as compared with others. In experiment 2, fibre inclusion was associated with increases in BWG from 7 to 14 day of age and an improvement in FCR, whereas FI was not influenced by treatments. Inclusion of fibre in the diet decreased the weight of the abdominal fat and cholesterol concentrations of liver and plasma. No significant effects on fatty acid composition of liver lipid were observed by fibre supplementation. These findings suggest dietary fibre affects performance, intestinal histomorphology and lipid metabolism in young chicks, which may directly affect poultry feeding strategies.     

Regional distribution and variation of gamma-globulin absorption from the small intestine of the neonatal calf

125I-labeled immunoglobulin (Ig)G1 in colostral whey was used to determine the region of maximum absorption of Ig from the small intestine of the neonatal calf and the variation in Ig absorption among calves at the intestinal level. In experiment 1, 5 segments (approx 5%, 35%, 60%, 80%, and 95% of the duodenocecal length) were formed in the small intestine of 9 colostrum-deprived calves shortly after birth. These segments were injected with colostral whey containing 125I-IgG1 4 hours after birth, and uptake, transfer, and absorption (defined as uptake plus transfer) were determined for each segment 2 hours later. Raw data were adjusted for the milligrams of IgG1 injected per gram of intestinal tissue to obtain the least squares mean (LSM) value. The LSM values for absorption of IgG1 from distal segments 3, 4, and 5 were significantly greater (P less than 0.05) than those values for proximal segments 1 and 2. The region of the maximum IgG1 absorption was the lower small intestine, 60% to 80% of the duodenocecal length. There was also an indication of independence between uptake and transfer in each of the segments. Significant differences (P less than 0.05) were present among calves in the LSM values for uptake and absorption, but not for transfer. In experiment 2, thoracic ducts of 8 newborn calves were cannulated 4 to 5 hours after birth. At 6 hours after birth, colostral whey with 125I-IgG1 was injected into an intestinal segment (approx 60% to 80% of the duodenocecal length).(ABSTRACT TRUNCATED AT 250 WORDS)     

The absorption of magnesium from the forestomachs, stomach and small intestine of sheep.

The passage of magnesium from the mouth to the proximal duodenum and from the proximal duodenum to the terminal ileum was measured continuously in two rams for 24 and 19 days. The results indicate a net disappearance of 25 per cent and 34 per cent of ingested magnesium from the forestomachs and stomach, and no set absorption from the small intestine.     

Physiological aspects of absorption and secretion in intestine

Two differently oriented approaches in intestinal physiology can be distinguished. One, mainly based on in vitro experiments, seeks explanations at the level of the epithelium itself. The other, mainly based on in vivo experiments, looks for explanations at the level of regulatory nervous and endocrine mechanisms and their interaction. These two approaches complement each other.     

Physiological aspects of absorption and secretion in intestine

Two differently oriented approaches in intestinal physiology can be distinguished. One, mainly based onin vitro experiments, seeks explanations at the level of the epithelium itself. The other, mainly based onin vivo experiments, looks for explanations at the level of regulatory nervous and endocrine mechanisms and their interaction. These two approaches complement each other.     

51Cr-EDTA absorption blood test: an easy method for assessing small intestinal permeability in dogs

The 51Cr-EDTA test is a valuable clinical tool for screening intestinal diseases in dogs. The test is performed by calculating the percentage of recovery from urine of a PO-ingested dose of 51Cr-EDTA after 6 or 24 hours. Careful urine collection is a practical limitation of this test in dogs, and our goal was to develop a simpler test that measures 51Cr-EDTA in blood. A 51Cr-EDTA absorption test was simultaneously performed on urine and serum 43 times in healthy Beagle Dogs. Timed blood samples were withdrawn, and urine was collected during a 6-hour period. Percentages of the ingested dose were then calculated in urine and serum. The mean +/- standard deviation (range) percentage in urine after 6 hours was 14.07 +/- 8.72% (3.81-34.18%), whereas results in serum from samples taken at 2, 3, 4, 5, and 6 hours were 0.49 +/- 0.45% (0.02-2.13%), 0.75 +/- 0.52% (0.03-1.89%), 0.82 +/- 0.57% (0.13-2.21%), 0.70 +/- 0.53% (0.12-1.99%), and 0.47 +/- 0.44% (0.11-1.79%), respectively. The results for blood specimens showed good concordance with those for urine, especially for the samples taken at 4 hours (r = 0.89). Moreover, the correlation between urine and blood was better when the sum of the percentages of the recovered analyte from various blood samples was compared with urine. The correlation coefficient when summing 4 blood samples was excellent (r = 0.97) and remained excellent when summing only 2 blood samples taken at 3 and 5 hours (r = 0.95) or at 3 and 4 hours (r = 0.94). We conclude that a serum 51Cr-EDTA test determined by summing successive blood samples provides an easier means of estimating small intestinal permeability in dogs and gives results comparable to those of the 6-hour urine test.     

Effect of mixing and sampling method on pelleted feed nutrient analysis and diet formulation validation

Samples of pelleted feed may be taken during manufacturing and analyzed for nutrient content in order to estimate if nutrients are in accordance with diet formulation. The objective of this study was to determine the impact of mix time, sample location, sample number, and sample blending technique on analyzed nutrients. Ingredients were mixed in a single screw vertical mixer for 30 s or 10 minutes. Samples were taken post extrusion from the pellet die as hot pellets or post cooling and conveyance as finished feed. Two or 10 samples were collected at equally spaced time intervals and pooled. Subsamples were obtained by blending pooled samples by hand or using a sample splitting device. Proximate and supplemental amino acid analysis was determined on each blended subsample and standardized for moisture content. Crude fat and free supplemental DL-methionine were affected by mix time (P < 0.05). The 10-minute mix time produced crude fat and free supplemental DL-methionine that had lower and higher values, respectively, compared to the 30-second mix time and agreed more with the calculated nutrient profile and diet formulation. Free supplemental DL-methionine was also affected by sampling location (P < 0.05). Samples obtained at the pellet die were higher and agreed more with diet formulation compared to finished feed samples. Analysis of free supplemental DL-methionine from samples obtained at the pellet die may best indicate pelleted diet homogeneity and validate the diet formulation.     

Goblet cell mucin distribution in the small intestine of newborn goat kids fed lyophilized bovine colostrum

The number of goblet cells containing neutral and acidic mucins, including sulphomucins and sialomucins, was investigated in the small intestine of goat kids fed with lyophilized bovine colostrum in the period of passive immunity acquisition. At 0, 7 and 14 h of life, 15 male newborns received 5% of body weight of lyophilized bovine colostrum (LBC) and 14 male newborns received goat colostrum (GC), both with 55 mg/mL of IgG. Three additional animals were sampled at birth, without colostrum intake. Duodenum, jejunum and ileum samples were collected at 18, 36 and 96 h of life. Histological stains, periodic acid-Schiff, 1% alcian blue pH 2.5 and 1% alcian blue pH 1.0 were used to identify neutral and acidic mucins and acidic sulphated mucins, respectively. The number of goblet cells containing neutral and acidic mucins, including sulphomucins and sialomucins, does not differ in the duodenum (P>0.05). In the jejunum, LBC showed a higher number of goblet cells containing sialomucins compared to GC (P<0.05). The highest number of goblet cells containing acidic and neutral mucins and total number of goblet cells were observed at 96 h (P<0.05). In this segment, vacuoles of colostrum were present at 18 and 36 h mainly in the upper region of the villi, while the goblet cells were located at the bottom. At 96 h, vacuoles of colostrum were not detected, only goblet cells distributed throughout the villi. In the ileum, the number of goblet cells containing sulphomucins was higher (P<0.05) at 96 h than at 18 h. The LBC group showed higher (P<0.05) number of goblet cells containing sulphomucins at 96 h and total number of goblet cells at 36 and 96 h than the 0-h group. The present work revealed that the greater the absorption of colostrum in the goat kids' jejunum epithelium, the smaller the number of goblet cells. Considering this segment, feeding newborns with heterologous colostrum caused alteration in the number of goblet cells containing sialomucin. This condition suggested a reaction of the intestinal epithelium with increasing secretion due to the presence of non-recognized substances from the lyophilized bovine colostrum.     

The validation of an enzyme immunoassay for the detection of antibodies to bovine leukosis virus

Validation of an enzyme immuno-assay for detection of antibodies against bovine leucosis virus is described in this paper. Internal standardisation of the test was done by means of a negative control serum. With absolute extinction of the negative control serum between 100 and 200 mE, a serum sample is rated positive, if its extinction is 1.5 times above the control. The methodological sensitivity of the enzyme immuno-assay described has proved to be four times as high as that of the immunodiffusion test. The results recorded at five diagnostic laboratories suggested a sensitivity of the test of 97.6 percent (92.1 to 100 percent) and a specificity of 98.1 percent (94.4 to 100 percent). The high efficiency of the test can be confirmed by immunoblotting.     

Effects of weaning and enterotoxigenic Escherichia coli on net absorption in the small intestine of pigs

The purpose of this study was to measure the net absorption of fluid, sodium, potassium and chloride in the small intestine of weaned pigs and of their unweaned littermates and to correlate these values with villus height and crypt depth. Five pairs of segments of the small intestine were prepared in each of 80 pigs; the cranial segment of each pair was injected with an enterotoxigenic strain of Escherichia coli and the caudal segment with a control solution. Net absorption was measured on the day of weaning and four, seven, 11 and 14 days after weaning. In unweaned pigs the net absorption of fluid, potassium and chloride did not vary with time. In weaned pigs the net absorption of fluid in the control segments was significantly less on days 4, 7 and 14 after weaning and of sodium and chloride on days 4 and 7 than in unweaned littermates. In infected segments of weaned pigs the net absorption of fluid was significantly less than in unweaned littermates on day 11 and 14, of sodium and potassium on day 11 and of chloride on days 4 and 11 after weaning. Net absorption was negatively correlated with villus height but only in the infected segments of weaned pigs; no other correlations were found. It was concluded that after weaning the net absorption of fluid and electrolytes in the small intestine of pigs is temporarily decreased, a condition that may initiate diarrhoea.