Bile enhances glucose uptake, reduces permeability, and modulates effects of lectins, trypsin inhibitors and saponins on intestinal tissue.

Antinutritional factors (ANFs) can disrupt digestive and other intestinal functions. ANFs in soybean meal (SBM) are implicated in proliferative and inflammatory responses in the intestine of various (functionally) monogastric animals, including Atlantic salmon (Salmo salar L.). The goal of the current study was to investigate the effect of ex vivo exposure of mid and distal intestinal tissue of salmon to soybean saponins (SAP), lectin (LEC) and Kunitz' trypsin inhibitor (KTI), singly and in combination, on epithelial function, as assessed by measuring in vitro glucose uptake pathways along a glucose concentration gradient. As solubilization of SAP in the calcium-containing Ringer's solution was problematic but resolved with the addition of a physiological concentration of bile collected from the gall bladder of salmon, an evaluation of bile effects became an added element. Results indicated that bile increased baseline glucose absorption and possibly transport, and also had a protective effect on the epithelial barrier, at least partially due to taurocholate. Compared to controls, tissues exposed to LEC+bile, KTI+bile and LEC+KTI+bile exhibited increased glucose uptake at the higher glucose concentrations, apparently due to markedly increased tissue permeability. Addition of SAP, however, attenuated the response, possibly by binding bile components. SAP+bile, also in combination with LEC and/or KTI, as well as LEC, KTI and LEC+KTI without bile often reduced transcellular glucose uptake pathways, while maintaining low tissue permeability. SAP+LEC+KTI+bile, LEC and KTI caused the most marked reductions. The distal intestine was more affected, reflecting the restriction of in vivo SBM-induced inflammatory changes to this region.

Medium-chain and long-chain fatty acids have different postabsorptive fates in Atlantic salmon.

An increasingly larger proportion of the oils used in diets for farmed fish are plant derived and rapeseed oil is most commonly used. Despite high dietary lipid levels and a marked change in lipid composition, the transport and metabolic fate of absorbed fatty acids is not fully understood in teleost fish. The main purpose of this study was to trace the postabsorptive metabolic fate of 2 fatty acids of different chain length: oleic acid [(3)H-18:1(n-9)], constituting 70% of fatty acids in rapeseed oil, and the medium-chain decanoic acid [(14)C-10:0], which does not require carrier molecules for membrane passage. The fatty acids and their metabolites were traced in portal and peripheral blood, liver, heart, skeletal muscle, and visceral adipose tissue at time intervals from 3 to 48 h after feeding. The portal vein was the primary transport route for both 10:0 and 18:1(n-9) from the intestine to the liver the first 6 h after feed intake. From 12 to 48 h, the peripheral route became increasingly more important. The study also indicates a possible direct transport route of fatty acids from the intestine to the surrounding viscera. Our data demonstrate that whereas 18:1(n-9) is primarily deposited as TG in skeletal muscle and visceral adipose tissue, 10:0 is used by the heart and skeletal muscle as a source for rapid energy production.

Antibacterial peptides derived from caprine whey proteins, by digestion with human gastrointestinal juice.

Peptides in caprine whey were identified after in vitro digestion with human gastrointestinal enzymes in order to determine their antibacterial effect. The digestion was performed in two continuing steps using human gastric juice (pH 2·5) and human duodenal juice (pH 8) at 37°C. After digestion the hydrolysate was fractionated and 106 peptides were identified. From these results, twenty-two peptides, located in the protein molecules, were synthesised and antibacterial activity examined. Strong activity of the hydrolysates was detected against Escherichia coli K12, Bacillus cereus RT INF01 and Listeria monocytogenes, less activity against Staphylococcus aureus ATCC 25 923 and no effect on Lactobacillus rhamnosus GG. The pure peptides showed less antibacterial effect than the hydrolysates. When comparing the peptide sequences from human gastrointestinal enzymes with previously identified peptides from non-human enzymes, only two peptides, ß-lactoglobulin f(92-100) and ß-casein f(191-205) matched. No peptides corresponded to the antibacterial caprine lactoferricin f(14-42) or lactoferrampin C f(268-284). Human gastrointestinal enzymes seem to be more complex and have different cleavage points in their protein chains compared with purified non-human enzymes. Multiple sequence alignment of nineteen peptides showed proline-rich sequences, neighbouring leucines, resulting in a consensus sequence LTPVPELK. In such a way proline and leucine may restrict further proteolytic processing. The present study showed that human gastrointestinal enzymes generated different peptides from caprine whey compared with non-human enzymes and a stronger antibacterial effect of the hydrolysates than the pure peptides was shown. Antimicrobial activity against pathogens but not against probiotics indicate a possible host-protective activity of whey.

Human Gastrointestinal Juices Intended for Use in In Vitro Digestion Models.

The aim of this study was to characterise the individual human gastric and duodenal juices to be used in in vitro model digestion and to examine the storage stability of the enzymes. Gastroduodenal juices were aspirated, and individual variations in enzymatic activities as well as total volumes, pH, bile acids, protein and bilirubin concentrations were recorded. Individual pepsin activity in the gastric juice varied by a factor of 10, while individual total proteolytic activity in the duodenal juice varied by a factor of 5. The duodenal amylase activity varied from 0 to 52.6 U/ml, and the bile acid concentration varied from 0.9 to 4.5 mM. Pooled gastric and duodenal juices from 18 volunteers were characterised according to pepsin activity (26.7 U/ml), total proteolytic activity (14.8 U/ml), lipase activity (951.0 U/ml), amylase activity (26.8 U/ml) and bile acids (4.5 mM). Stability of the main enzymes in two frozen batches of either gastric or duodenal juice was studied for 6 months. Pepsin activity decreased rapidly and adjusting the pH of gastric juice to 4 did not protect the pepsin from degradation. Lipase activity remained stable for 4 months, however decreased rapidly thereafter even after the addition of protease inhibitors. Glycerol only marginally stabilised the survival of the enzymatic activities. These results of compositional variations in the individual gastrointestinal juices and the effect of storage conditions on enzyme activities are useful for the design of in vitro models enabling human digestive juices to simulate physiological digestion.

Different digestion of caprine whey proteins by human and porcine gastrointestinal enzymes.

The objective of the present study was twofold: first to compare the degradation patterns of caprine whey proteins digested with either human digestive juices (gastric or duodenal) or commercial porcine enzymes (pepsin or pancreatic enzymes) and second to observe the effect of gastric pH on digestion. An in vitro two-step assay was performed at 37 degrees C to simulate digestion in the stomach (pH 2, 4 or 6) and the duodenum (pH 8). The whey proteins were degraded more efficiently by porcine pepsin than by human gastric juice at all pH values. Irrespective of the enzyme source, gastric digestion at pH 2 followed by duodenal digestion resulted in the most efficient degradation. Lactoferrin, serum albumin and the Ig heavy chains were highly degraded with less than 6 % remaining after digestion. About 15, 56 and 50 % Ig light chains, beta-lactoglobulin (beta-LG) and alpha-lactalbumin remained intact, respectively, when digested with porcine enzymes compared with 25, 74 and 81 % with human digestive juices. For comparison, purified bovine beta-LG was digested and the peptide profiles obtained were compared with those of the caprine beta-LG in the digested whey. The bovine beta-LG seemed to be more extensively cleaved than the caprine beta-LG in the whey. Commercial enzymes appear to digest whey proteins more efficiently compared with human digestive juices when used at similar enzyme activities. This could lead to conflicting results when comparing human in vivo protein digestion with digestion using purified enzymes of non-human species. Consequently the use of human digestive juices might be preferred.

In vitro studies of the digestion of caprine whey proteins by human gastric and duodenal juice and the effects on selected microorganisms.

The in vitro digestion of caprine whey proteins was investigated by a two-step degradation assay, using human gastric juice (HGJ) at pH 2.5 and human duodenal juice (HDJ) at pH 7.5. Different protein and peptide profiles were observed after the first (HGJ) and second (HDJ) enzymatic degradation. The minor whey proteins serum albumin, lactoferrin and Ig were rapidly degraded by HGJ, while alpha-lactalbumin (alpha-LA) and beta-lactoglobulin (beta-LG) were more resistant and survived both 30 and 45 min of the enzymatic treatment. Further digestion with HDJ still showed intact beta-LG, and the main part of alpha-LA also remained unchanged. The protein degradation by HGJ and HDJ was also compared with treatment by commercial enzymes, by using pepsin at pH 2.5, and a mixture of trypsin and chymotrypsin at pH 7.5. The two methods resulted in different caprine protein and peptide profiles. The digests after treatment with HGJ and HDJ were screened for antibacterial effects on some selected microorganisms, Escherichia coli, Bacillus cereus, Lactobacillus rhamnosus GG and Streptococcus mutans. Active growing cells of E. coli were inhibited by the digestion products from caprine whey obtained after treatment with HGJ and HDJ. Cells of B. cereus were inhibited only by whey proteins obtained after reaction with HGJ, while the products after further degradation with HDJ demonstrated no significant effect. Screenings performed on cells of Lb. rhamnosus GG and S. mutans all showed no signs of inhibition.

Fish arsenic may influence human blood arsenic, selenium, and T4:T3 ratio.

The effects of an arsenic-rich fish diet and selenium (Se) supplementation on blood arsenic (As), Se, and thyroid hormones were studied in 32 women divided into four equal groups. Groups 1 and 4 received 400 microg Se-methionine daily, group 2 received 400 microg selenite daily, and group 3 received placebo tablets for 15 wk. In addition, groups 1-3 increased their fish intake, eating at least three fish dinners weekly. Mean blood Se concentrations (initially 1.68 +/- 0.24 micromol/L) increased twofold in the Se-methionine groups (p < 0.0001) and by 32% in the selenite group (p < 0.01). Group means of blood As concentrations increased by 63% (p < 0.01), 50% (p < 0.01), 106% (p < 0.01), and 29% (p < 0.05) in the four groups, respectively. Analyzed As intake from duplicate portions of consumed fish correlated with final blood As concentrations (r = 0.85, p < 0.001, n = 32). In the group not receiving Se, there was a positive correlation between final blood As concentrations and plasma T4:T3 ratio (r = 0.80, p < 0.02, n = 8). Initially, blood As concentrations correlated negatively with both T3 and T4 in plasma, but this correlation disappeared upon Se supplementation. The results demonstrate that increased intake of fish may influence blood As concentrations and that circulating thyroid hormones may be influenced by Se-As interactions.