Impact of α-amylase supplementation on energy balance and performance of high-yielding dairy cows on moderate starch feeding.

In dairy cows, exogenous α-amylase is suggested to improve starch utilization and positively affect performance and health traits linked to energy balance and fertility. In a 1-year feeding experiment, 421 cows were orally supplemented with α-amylase (treatment: 12.5 g/kg dry matter (DM) addition rate to a concentrated feed) or non-supplemented (control) on the basis of an ad libitum total mixed ration (TMR). Every cow was allocated to a high- (≥32 kg milk/day) or late-lactation group (<32 kg milk/day), in which the TMR starch content was 220 ± 20.8 g/kg DM and 183 ± 24.8 g/kg DM, respectively. The energetic effect of α-amylase supplementation seemed to be exclusively related to the high-lactation stage (5-100 days in milk) in primiparous cows, where the daily milk yield was 32 ± 0.49 versus 31 ± 0.50 kg per cow in the treatment versus control group (P < 0.05). The pluriparous cows did not benefit from the supplementation that way. In neither primiparous nor pluriparous cows, was the milk composition, the fat-to-protein ratio, the somatic cell score, the backfat thickness, serum total bilirubin, ß-hydroxybutyrate and the fertility found to be systematically affected by α-amylase supplementation.

Effect of alginate and inulin on intestinal microbial ecology of weanling pigs reared under different husbandry conditions.

The effects of inulin and alginate on intestinal microbial ecophysiology were investigated in piglets fed a diet (C) with 0.1% alginate (C+A) or 1.5% inulin (C+I) from weaning at day 28. The experiment was performed at an experimental farm (EF) and a commercial farm (CF). Digesta was collected from the ileum, caecum and colon of four piglets from each group on days 29, 30, 33 and 39. The metabolite concentrations changed with age. Colonic and caecal metabolites were affected by prebiotic treatment. Changes in microbiota composition were assessed by cultivation and denaturing gradient gel electrophoresis (DGGE) of 16S rRNA gene fragments. Enterococci increased in C+A at EF and decreased in C+I at both farms. Lactobacilli decreased in all segments in the experimental groups on days 30 and 33. Yeasts in C+I were five times lower at CF than at EF on day 39. The richness and diversity of DGGE profiles increased in the experimental groups. The evenness of colon digesta-derived DGGE profiles was higher in the experimental groups than in C and this situation was reversed in the distal small intestine. Multivariate redundancy analysis confirmed the recorded effects. In summary, both prebiotics affected the intestinal microbiota, and the changes were more pronounced at the CF.

Susceptibility of phaseolin (Phaseolus vulgaris) subunits to trypsinolysis and influence of dietary level of raw phaseolin on protein digestion in the small intestine of rats.

The aim of the present work was (a) to investigate trypsinolysis of denatured purified T phaseolin (Phaseolus vulgaris) subunits by MS and (b) to test the effect of raw T phaseolin inclusion level in diets fed chronically to rats on digestion in the small intestine. The diets contained casein as the sole protein source, or casein substituted with 33, 67 and 100 % of purified T phaseolin. Rats were fed for 10 d and then euthanised. Digesta and tissues from the first and second halves of the small intestine were prepared for electrophoresis, immunoblotting and densitometry. alpha-Phaseolin subunit for the T phaseolin was more resistant to trypsinolysis than beta-phaseolin subunit. Nearly intact phaseolin subunits (molecular weight, MW 44-54 kDa) and partially digested phaseolin fragments (MW 17-19 and 20-24 kDa) were identified in small intestinal digesta. The concentration of intact phaseolin and of most undigested phaseolin fragments in digesta increased in the second half of the small intestine with increasing phaseolin intake (P < 0.05-0.01). The concentration of phaseolin fragments of a MW of 21-22.5 and 23-24.5 kDa in the mucosa increased linearly (P = 0.016-0.084) when the level of the T phaseolin was increased in the diet. In conclusion, the present work provides evidence that denatured T phaseolin subunits display different trypsinolysis patterns in vitro. Moreover, a high intake of raw T phaseolin impacts digestion in the small intestine of rats.

Effect of barley and oat cultivars with different carbohydrate compositions on the intestinal bacterial communities in weaned piglets.

This experiment was aimed at comparing the intestinal microbial community composition in pigs fed hulled common barley supplemented with isolated barley mixed-linked beta-glucan, four hulless barley varieties and breeding lines with mixed-linked beta-glucan contents ranging from 41 to 84 g kg(-1) and different amylose/amylopectin ratios as well as two oat varieties. Seventy-two weaned piglets were allocated to one of nine diets composed of 81.5% cereal, 6% whey, 9% soy protein isolate and 3.5% minerals. After 15 days, pigs were sacrificed and ileum and colon contents were collected for quantitative real-time PCR (qPCR) and denaturing gradient gel electrophoresis to evaluate microbial communities. Shifts in intestinal microbial communities were observed with the hulless barley cultivars with a normal to high beta-glucan content and from normal starch toward either high-amylopectin or high-amylose starch. These hulless barleys had the lowest (P<0.05) microbial diversity, whereas oats had intermediate diversity compared with low-beta-glucan hulless cultivars and hulled varieties. Furthermore, hulless varieties favoured xylan- and beta-glucan-degrading bacteria whereas mixed-linked beta-glucan-supplemented hulled barley favoured lactobacilli. Numbers of lactobacilli decreased in the ileum of pigs fed hulless/high mixed-linked beta-glucan barley-based diets. Thus, cultivar differences in both the form and the quantity of carbohydrates affect gut microbiota in pigs, which provides information for future feeding strategies.

Biological diversity of yeasts in the gastrointestinal tract of weaned piglets kept under different farm conditions.

The study was conducted to determine yeasts present in the gastrointestinal tract (GIT) of piglets kept under experimental farm (EF) and commercial farm (CF) conditions. Ninety five German Landrace full- and half-sibling piglets were sacrificed at 39 days of age. Sixty eight piglets were weaned at 28th day of life, when they were offered one diet ad libitum. Twenty seven piglets remained unweaned by their dams. None of the piglets received any creep feed before weaning. Digesta samples were collected from 1/3 distal small intestine (SI), caecum and proximal colon. One hundred seventy three colonies of isolated yeasts were characterized by sequence analysis of the PCR-amplified D1/D2 domain of the 26S rRNA gene with following alignment of the recovered sequences to GenBank entries. From the 17 phylotypes found, isolates most closely related to Galactomyces geotrichum, Kazachstania slooffiae and Candida catenulata dominated in the GIT of CF piglets. Kazachstania slooffiae and Candida glabrata dominated in GIT of EF piglets. Sørenson and Morisita-Horn similarity indices between farms were low (0.44 and 0.54 respectively) and the Simpson diversity index was higher for EF (7.58) than for CF (4.34). The study brings new data on yeasts composition in the pig GIT and shows differences in yeasts biodiversity between farms operated at different hygiene conditions.

Effect of thymol on microbial diversity in the porcine jejunum.

Thirty two weaned pigs (24 d-old) were fed a diet without or with 1% (w/w) thymol. Pigs from each dietary treatment remained unchallenged or were challenged with Salmonella enterica serovar typhimurium. Jejunal content was collected and molecular microbial diversity was investigated using 16S rRNA gene polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE). Shannon index analysis showed no differences in diversity but Pearson coefficient analysis showed clear clustering of profiles, which delineated thymol fed and control groups irrespective of Salmonella challenge. Moreover, absence of bands corresponding to Actinobacillus minor (98-99% identity) was observed in thymol groups. A band corresponding to Citrobacter freundii (98% identity) was observed in almost all pigs from thymol groups, and only in 4 samples from pigs fed no thymol in the diet. Supplementation of pig diet with thymol caused clear changes in small intestine microbacterial community. Surprisingly, Salmonella infection caused no major perturbations to the community structure.

Susceptibility of phaseolin to in vitro proteolysis is highly variable across common bean varieties (Phaseolus vulgaris).

A study was conducted to investigate the amino acid (AA) composition and the susceptibility to in vitro proteolysis (pepsin, 120 min and pancreatin, 240 min) of a collection of purified phaseolins ( n = 43) in unheated or heat-treated form. The AA composition of phaseolin varied little across bean varieties. At 360 min of in vitro proteolysis, the degree of hydrolysis varied from 11 to 27% for unheated and from 57 to 96% for heated phaseolins ( P < 0.001). Heat treatment markedly increased the susceptibility of phaseolin to proteolysis ( P < 0.001). The AA scores (AAS) and the protein digestibility corrected for AAS indicated S-containing AA as the limiting AA (39 +/- 3 and 30 +/- 5%, respectively). In conclusion, susceptibility to proteolysis of heat-treated phaseolin rather than its AA composition affects the nutritional value of phaseolin estimated in vitro. Therefore, it should be the criterion of choice in breeding programs aimed at improving the nutritional value of common beans for humans.

Phaseolin type and heat treatment influence the biochemistry of protein digestion in the rat intestine.

The study aimed to investigate the in vivo digestion of Phaseolus vulgaris phaseolin types differing in their subunit pattern composition. Diets contained either casein as the sole source of protein or a mixture (1:1) of casein and pure Sanilac (S), Tendergreen (T) or Inca (I) phaseolin either unheated or heated. Rats were fed for 11 d with the experimental diets. Their ileal content and mucosa were collected and prepared for electrophoresis, Western blotting, densitometry and MS. Differences in digestion among native phaseolin types were observed for intact phaseolin at molecular weights (MW) of 47-50.5 kDa and for an undigested fragment at MW of 19-21.5 kDa in ileal digesta. In both cases, the concentration of these protein bands was lower for I phaseolin than for S or T phaseolin (P < 0.05). In the mucosa, the concentration of a protein band at MW of 20.5-21.5 kDa was lower for S phaseolin as compared to T or I phaseolin (P < 0.001). The presence of phaseolin subunits and their fragments was confirmed by Western blotting. MS analysis revealed the presence of undigested alpha and beta subunit fragments from phaseolin and endogenous proteins (anionic trypsin I and pancreatic alpha-amylase) in ileal digesta. Thermal treatment improved digestion (P < 0.01), acting on both dietary and endogenous protein components. In conclusion, this study provides evidence for differences in intestinal digestion among phaseolin types, S phaseolin being more resistant and I phaseolin more susceptible. These differences were affected by the origin of the phaseolin subunit precursor. Heat treatment enhanced phaseolin digestion.

First-pass metabolism limits the intestinal absorption of enteral alpha-ketoglutarate in young pigs.

Our results in a previous study indicated that the portal absorption of intragastrically fed alpha-ketoglutarate (AKG) was limited in young pigs. Our aim was to quantify the net portal absorption, first-pass metabolism, and whole-body flux of enterally infused AKG. In study 1, we quantified the net portal nutrient absorption in young pigs (n = 9) given an intraduodenal infusion of milk replacer [10 mL/(kg . h)] and either saline (control) or 930 micromol/(kg . h) AKG for 4 h. In study 2, we quantified the luminal disappearance of a duodenal AKG bolus in young pigs (n = 7). In study 3, we quantified the whole-body kinetics of (13)C-AKG metabolism when infused either enterally (n = 9) or intravenously (n = 9) in young pigs. In study 1, when compared with the control group, enteral AKG infusion increased (P < 0.01) the arterial (13.8 +/- 1.7 vs. 27.4 +/- 3.6 micromol/L) and portal (22.0 +/- 1.4 vs. 64.6 +/- 5.9 micromol/L) AKG concentrations and the net portal absorption of AKG [19.7 +/- 2.8 vs. 95.2 +/- 12.0 micromol/(kg . h)]. The mean fractional portal appearance of enterally infused AKG was 10.23 +/- 1.3%. In study 2, the luminal disappearance of AKG was 663 micromol/(kg . h), representing 63% of the intraduodenal dose. In study 3, the whole-body (13)C-AKG flux [4685 +/- 666 vs. 801 +/- 67 micromol/(kg . h)] was higher (P < 0.05) when given enterally than intravenously, but (13)CO(2) recovery was not different (37.3 +/- 1.0 vs. 36.2 +/- 0.7%dose). The first-pass splanchnic (13)C-AKG utilization was approximately 80%, of which 30% was oxidized to (13)CO(2). We conclude that the intestinal absorption of AKG is limited in young pigs largely due to substantial first-pass gastrointestinal metabolism.

Effect of partial dehulling of two- and six-row barley varieties on precaecal digestibility of amino acids in pigs.

Five barrows (German Landrace; initial BW 58 kg, final BW 80 kg) fitted with an ileo-rectal anastomosis were used to determine the effect of partial dehulling and addition of barley hulls of two- and six-row barley varieties on the precaecal digestibility (pD) of CP and amino acids. The following diets were provided according to a standardized diet formulation and tested in seven consecutive periods (repeated group-period design): two-row barley (TRB) + casein (C), dehulled TRB + C, TRB + C + 10% hulls, six-row barley (SRB) + C, dehulled SRB + C, SRB + C + 1% hulls, and wheat starch + C. The diets were supplied at daily rates of 79-86 g DMI x kg BW(-0.75) in barley containing diets and at 49 g DMI x kg BW(-0.75) in the casein diet. The digestibility of amino acids in barley varieties was determined by the difference method (casein as basal diet) using quantitative digesta collection. In both varieties of barley the pD of CP and amino acids did not differ. The pD of CP was unchanged in regard to the treatments in both barley varieties. Due to dehulling in TRB the pD was improved significantly for most indispensable amino acids and in SRB for Met and Cys. Addition of 10% hulls to TRB led to equivalent pD of Arg, His, Leu, Tyr, and Trp compared to TRB, but the pD of Lys, Phe, Thr and Val was significantly decreased below the levels of TRB. Addition of even 1% hulls to SRB impaired the pD of Lys below the level in SRB. In conclusion, addition of barley hulls to pig diets impairs amino acids absorption in the small intestine. The pD values, measured under standardized experimental conditions (without a correction using basal endogenous amino acids), are similar to the values of true digestibility published by NRC (1998).

Influence of the Phaseolus vulgaris phaseolin level of incorporation, type and thermal treatment on gut characteristics in rats.

Phaseolus vulgaris phaseolin has been shown to stimulate faecal losses of endogenous N in rats. Experiments with purified phaseolin were carried out in rats to test the hypothesis that these losses reflect intestinal disorders. Phaseolin composition varies depending on its constitutive subunits. Therefore, three phaseolin types (S, T, I) were tested. Phaseolin T was incorporated in varying levels (0, 33, 67 or 100 % of the dietary protein) as raw material in experiment 1. In experiment 2, the three phaseolin types were incorporated at 50 %, with or without previous thermic treatment. Raw casein was the basal protein source and was also heated in experiment 2. Faecal digestibility of phaseolin and gut integrity were evaluated in both experiments. The incorporation level or type of phaseolin had little effect on gross anatomy of gut segments but these factors influenced the weight and pH of fresh contents of the stomach and caecum (P<0.05). Raw phaseolin T incorporated at various levels led to an enlargement of duodenal villi together with a tendency for increased crypt depth in the jejunum (P=0.06). Activities of both alkaline phosphatase in the duodenum and aminopeptidase N in the ileum decreased (P<0.05) after thermal treatment of casein while they increased (P<0.05) for heat-treated phaseolin S and T, respectively. In conclusion, raw phaseolin had no effect on the tissue weight of gut segments and induced limited alterations in the small intestine. Differences due to phaseolin level or type were limited too.

Intraduodenal infusion of alpha-ketoglutarate decreases whole body energy expenditure in growing pigs.

BACKGROUND AND AIMS: alpha-Ketoglutarate (AKG) has been suggested to play a particular role as an oxidative fuel for the gut, and thus may have a sparing function for fuels such as glutamate and aspartate. Using the pig model we aimed to quantify how the route of administration (intravenous, i.v.; intragastric, i.g.; intraduodenal, i.d.) affects AKG utilization, whole body energy expenditure (EE) and nutrient oxidation. METHODS: Pigs (15 kg) were supplied with a complete nutrient solution (NS) via catheters. To explore the metabolic effects of AKG, 1.0 g AKG kgBW(-1)d(-1) was infused simultaneously with the NS using either the i.d., i.v. or i.g. route. [1-(13)C]AKG (15 mg kgBW(-1)) was infused i.d., i.v. or i.g., respectively, for 3h. AKG utilization (AKG UTIL) was estimated as AKG UTIL=100-(13)C recovery (% of (13)C dose). (13)C recovery was calculated from the (13)C enrichment in breath CO(2) and the whole-body CO(2) production. RESULTS: AKG infusion and NS via the i.d. route resulted in a reduced AKG UTIL (40.1+/-6.7) as compared to the i.v. route (62.9+/-2.4, P<0.001) and i.g. route (62.3+/-1.6, P<0.001). The total EE was lower with the i.d. route of AKG and NS (745+/-68 kJkgBW(-0.62)d(-1)) as compared to the i.v. route (965+/-54 kJkgBW(-0.62)d(-1), P<0.005) and i.g. route (918+/-43 kJkgBW(-0.62)d(-1), P<0.005). Carbohydrate oxidation was increased with the i.d. route (38.2g+/-3.4 kgBW(-0.62)d(-1)) as compared to the i.v. route (27.8+/-2.9 gkgBW(-0.62)d(-1), P<0.08) and i.g. route (23.9+/-8.5 gkgBW(-0.62)d(-1), P<0.05). Fat oxidation was decreased (2.1+/-1.9 gkgBW(-0.62)d(-1); P<0.001) with the i.d. route as compared to the i.v. route (11.5+/-1.4 gkgBW(-0.62)d(-1), P<0.001) and i.g. route (11.9+/-3.1 gkgBW(-0.62)d(-1), P<0.001). CONCLUSIONS: The i.d. infusion of AKG in combination with the NS affected the whole body EE and nutrient oxidation, in comparison to that obtained with the i.v. and i.g. routes. It was concluded that the i.d. administration of AKG markedly controlled the nutrient partitioning in the oxidation processes. Finally, in contrary to the observations with glutamine or glutamate, a considerable percentage of the AKG infusion was retained in the body irrespective of the route of administration.