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Transport of 3H-labeled nucleosides across the bovine intestinal brush border membrane (BBM) was characterized with BBM vesicles (BBMV) isolated from mid-jejunum of cows because large amounts of nucleic acids are digested in the small intestine of ruminants. Two Na+-dependent electrogenic nucleoside transporters with overlapping substrate specificity were shown to be present in the jejunal BBM, one for pyrimidine nucleosides and one for purine nucleosides. As indicated by inhibitory studies, thymidine seemed to be a specific substrate for the pyrimidine nucleoside transporter, while this applied to guanosine and deoxyguanosine for the purine nucleoside transporter. Uridine and adenosine appear to have an affinity to both transporters. This also applies to deoxyadenosine and deoxyuridine. Nucleobases (uracil, hypoxanthine) did not affect transport of nucleosides. The kinetic constants (Km and Vmax) for Na-dependent thymidine and guanosine transport were 29 and 24 micromol/L and 78 and 51 pmol (mg protein)(-1) s(-1), respectively. These values are much higher than those reported for monogastric species.

To detect whether pentoses and hexoses occurring in rumen bacteria or in hemicellulose ingested with feed and partly released in the small intestine have an affinity for the Na(+)-dependent glucose transporter of the bovine intestinal brush border membrane (BBM), we investigated whether these monosaccharides inhibit Na(+)-dependent transport of 14C-labelled D-xylose across the BBM using brush border membrane vesicles (BBMV) isolated from the mid-jejunum of cows. We used D-xylose as the transport substrate, because it has a low affinity for the Na(+)-dependent glucose transporter and thus its uptake into BBMV is more efficiently competitively inhibited by other sugars than that of D-glucose. D-Ribose, D-mannose and L-rhamnose occurring in rumen bacteria significantly inhibited Na(+)-dependent uptake of D-xylose into BBMV, but their inhibitory effect was less than that of D-glucose, D-xylose and phlorizin. This also applied to L-arabinose (and D-arabinose), which is, like D-xylose and D-galactose, a constituent of hemicellulose, and to 2-deoxy-D-glucose. Of all monosaccharides tested, only D-fructose did not affect Na(+)-dependent D-xylose transport. It is concluded that some pentoses and hexoses occurring in rumen bacteria (D-ribose, D-mannose and L-rhamnose) or hemicellulose (L-arabinose and D-xylose) have a low affinity for the Na(+)-dependent glucose transporter of the bovine BBM and may therefore be absorbed from the jejunum when released in the small intestine.

Due to their ubiquitous occurrence in the plant kingdom, plant phenolics, including monomeric cinnamic acids, are ingested by man and animals in variable amounts with their natural diets. Recently, Na(+)-dependent saturable transport of cinnamic acid across the brush-border membrane of rat jejunum has been described. It was the aim of the present study to characterize this mechanism in more detail. We therefore determined the transport kinetics of mucosal uptake of radioactively labelled cinnamic acid under various conditions using a short-term mucosal uptake technique. In addition, the transfer of cinnamic acid across the jejunal wall was investigated using everted intestinal sacs. Investigations of the kinetics of cinnamic acid uptake by the mid-jejunal mucosa revealed the involvement of two transport components, a diffusive Na(+)-independent mechanism and a saturable Na(+)-dependent mechanism. The results obtained with everted sacs provided further evidence of the existence of an active Na+ gradient-driven transport of cinnamic acid across the intestinal epithelium. In the presence of Na+, a significant accumulation of cinnamate occurred inside the serosal compartment and this was strongly inhibited by serosal ouabain. A decrease in the extracellular pH stimulated mucosal cinnamate uptake by increasing the apparent affinity (1/km). This may be attributable to the involvement of a transmembrane H+ gradient in Na(+)-dependent cinnamate transport because the protonophore FCCP caused a significant reduction of cinnamate uptake only in the presence of Na+. The kinetics of cinnamate transport in the absence or presence of a surplus of either unlabelled cinnamate or unlabelled butyrate indicates a reduction in the apparent affinity of the Na(+)-dependent mechanism involved in cinnamate uptake. These results may be explained by a modification of the mechanism by the intracellular pH. Additionally, competitive inhibition of cinnamate uptake by substances structurally related to cinnamic acid may also be involved.

Phenolic acids are present in all plant-derived foods and in most diets. Indirect evidence indicates substantial absorption of phenolic monomers from the gastrointestinal tract. However, the mechanisms involved in the absorptive process are unknown. The present study investigates mucosal uptake of radioactively labeled cinnamic acid as a model substance for monomeric cinnamic acid derivatives (e.g., cinnamic, ferulic or caffeic acid) in the rat jejunum using an in vitro mucosal uptake technique. The results indicate the existence of a Na(+)-dependent saturable transport mechanism for uptake of cinnamic acid across the jejunal brush border membrane. The observed Na+ dependence of jejunal cinnamate uptake seems not to be related to the activity of the Na+,H+ exchanger. Lowering the pH of the incubation medium resulted in a pronounced increase in mucosal cinnamate uptake that can be only partially explained by an increase in nonionic diffusion of cinnamic acid. Furthermore, jejunal uptake of cinnamate seems to be influenced by intracellular HCO3- and/or pH, since the addition of methazolamide to a HCO3(-)- and CO2-free incubation medium significantly inhibited mucosal cinnamate uptake, whereas methazolamide was without an effect in the presence of HCO3- and CO2 in the incubation medium. Unlabeled cinnamic and ferulic acid as well as short-chain fatty acids (acetic, propionic and butyric acid) significantly inhibited Na(+)-dependent uptake of radioactivity labeled cinnamic acid.(ABSTRACT TRUNCATED AT 250 WORDS)

1. Transport of citrate and fumarate across the pig intestinal brush-border membrane (BBM) was investigated using isolated BBM vesicles. 2. Citrate and fumarate uptake was stimulated by an inwardly directed Na+ gradient consistent with Na+/citrate (fumarate) co-transport. Cis-inhibition and trans-stimulation experiments strongly suggest the existence of a common transport site for tri- and dicarboxylates. 3. The protonated forms of citrate (citrate-1, citrate-2) seem to be much better transported than citrate-3, indicated by the strong stimulation of citrate uptake at an extravesicular pH of 5.6 compared to pH 7.8. 4. Uptake of tri- and dicarboxylates seems to be potential-sensitive since citrate and in particular fumarate transport was enhanced by an inside negative potential difference. 5. Kinetics of succinate transport revealed a single carrier-mediated component with apparent kinetic constants of 0.43 nmol/mg protein-3 s (Vmax) and 0.14 mmol/l (Km).

Transport of tri- and dicarboxylic acids across the intestinal brush border membrane was investigated using citrate and fumarate as transport substrates. The experiments were performed with brush border membrane vesicles isolated from calf proximal jejunum. Citrate and fumarate uptake by the brush border membrane vesicles occurred by a common Na(+)-dependent transport mechanism that appears to be specific for tri- and dicarboxylates. The protonated forms of citrate (citrate-(1) and citrate-(2] seem to be much better transported than the trivalent form, as indicated by the strong stimulation of citrate uptake at an extravesicular pH of 5.6 compared to that at pH 7.8. Furthermore, citrate transport across the intestinal brush border membrane appears to be mediated by an electroneutral process.

In ruminants, intestinal digesta can contain considerable amounts of lactic acid derived from ingestion of lactic acid-containing feed and from production of lactic acid during ruminal digestion of readily fermentable carbohydrates. The aim of the present study was to investigate L-lactate transport across the bovine intestinal brush border membrane. The experiments were performed using isolated brush border membrane vesicles from the midjejunum of heifers. The results demonstrate the existence of Na+-stimulated L-lactate uptake by the brush border membrane vesicles. Acetate, propionate, and butyrate strongly inhibited Na+-dependent L-lactate transport. Acetate caused a 58% inhibition, whereas propionate and butyrate completely inhibited Na+-dependent uptake. Kinetic evaluation of L-lactate uptake in the presence or absence of extravesicular butyrate suggests a competitive inhibition by butyrate. Among the phenolic acids tested in this study only trans-cinnamic acid caused a significant reduction of L-lactate uptake, whereas cumaric acid, ferulic acid, and gallic acid only slightly reduced L-lactate transport. Thus, the L-lactate transporter appears to have some affinity for transcinnamic acid.

The objectives of the present study were to find out whether selenate and sulphate are transported by a common Na+-dependent transport mechanism across the intestinal brush-border membrane and whether selenate is also transported by the anion-exchange system transporting sulphate across the brush-border membrane. Results from cis-inhibition as well as from trans-stimulation experiments show that selenate and sulphate are transported across the brush-border membrane by both Na+ co-transport and anion exchange. The influence of various dicarboxylates (oxalate, oxaloacetate, glutamate) and of the anion-exchange inhibitor 4,4'-diisothiocyanostilbene-2,2'-disulphonate (DIDS) on selenate and sulphate transport across the brush-border membrane was also investigated. Under Na+-gradient conditions selenate uptake across the brush-border membrane was reduced by oxalate and oxaloacetate but not by L-glutamate or L-leucine. Additionally sulphate uptake under this condition was significantly inhibited by oxalate (about 50%) and DIDS (25%). Sulphate transport by anion exchange was inhibited about 80% by both oxalate and DIDS. These results suggest that under Na+-gradient conditions sulphate (selenate) uptake in part occurs via anion exchange. Furthermore, oxalate seems to inhibit both Na+-sulphate (selenate) co-transport and sulphate (selenate) transport by anion exchange.

The properties of hypoxanthine transport were investigated in purified brush border membrane vesicles isolated from calf proximal and distal jejunum. Hypoxanthine uptake in the vesicles was stimulated by a transmembrane Na+ gradient and an inside negative potential resulting in a transient accumulation of intravesicular hypoxanthine, especially in the proximal jejunum. Na+-dependent hypoxanthine uptake at this site seemed to occur by two saturable transport systems, a high affinity (Km=0.33 μmol/l) and a low affinity (Km=165 μmol/l) transporter. Guanine, hypoxanthine, thymine and uracil inhibited intravesicular hypoxanthine uptake, whereas adenine and the nucleosides inosine and thymidine were without effect. These findings represent the first demonstration of active Na+ gradient-dependent nucleobase transport in intestinal brush border membrane vesicles. [ABSTRACT FROM AUTHOR]