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PubMed1999

Induction of apoptosis by monosaccharide butyrate stable derivatives in chronic lymphocytic leukemia cells.

BACKGROUND AND OBJECTIVE: Different therapeutic approaches are needed to restore apoptotic mechanisms in CLL cells, as present ones are not successful. We assessed the apoptotic effects of stable butyrate derivatives on CLL lymphocytes: in these molecules a mannose molecule is bound as ester to one-five butyrate moieties, conferring pharmacological stability to the pro-drugs which are able to induce apoptosis in primary AML blasts. DESIGN AND METHODS: Peripheral blood samples obtained from 17 patients with typical B-CLL were cultured in the presence of 0.5-1mM D1 (O-n-butanoyl-2, 3-O-isopropylidene-a-D-mannofuranoside), F1 (1-O-n-butanoyl-2, 3-O-isopropylidene-D,L-xylitol) and G1 (1-O-n-butanoyl-D,L-xylitol) derivatives for 4 days and equimolar sodium butyrate as comparison. After culture, apoptosis was evaluated by cell morphology, cellular DNA content, pattern of DNA fragmentation, annexin V exposure on cell membrane, and cell cycle parameters. Bcl2, bax, and fas oncogene expression were also evaluated by the APAAP method. RESULTS: The addition to cell cultures of D1 or F1 or G1 butyrate monosaccharides as well as sodium butyrate 0.5 and 1 mM determined to different extents an increase in the percentage of apoptotic cells in all CLL samples, relatively to the method and butyrate molecule added in culture. Heterogeneity in CLL cell sensitivity to the three butyrates was observed. Up to 60-68% apoptotic bodies were present in treated cultures after exposure to D1 0.5-1 mM, 60-72% after F1 0.5-1 mM and 48-60% after G1 0.5-1 mM. Comparison of untreated versus treated cultures yielded important significance (p< 0.001). At DNA content analysis, analyzed by flow cytometry, apoptotic events were accounting for up to 70-77% of D1-treated and 68-74% of F1-treated CLL cells at 0.5 and 1 mM concentrations (p= 0. 0001, vs controls 0-39%), and for 72-81% of G1 (0.5-1 mM) treated cells (overall, p=0.005). Cell cycle parameters were not altered by addition of butyrates, but expression of Annexin V was greatly enhanced. In a limited number of CLL cases fas, bcl2/bax ratio was analyzed and found unmodified. INTERPRETATION AND CONCLUSIONS: Monosaccharide butyrate stable derivatives are potent inducers of primary CLL cell apoptosis, both in untreated and alkylating agent pre-treated cases. Our results suggest that the apoptotic pathways elicited by butyrate in CLL lymphocytes are direct, specific and most probably do not involve bcl2/bax. Pro-apoptotic agents like the stable monosaccharide butyrate derivatives here studied could bring more insights into CLL biology and resistance to apoptosis, and possibly originate alternative treatments for CLL.

Aged↗
PubMed1979

Relationship between changes in lactase activity and monosaccharide uptake in the small intestine of the rat during development.

The relationship between changes in intestinal lactase activity and monosaccharide uptake was studied in rats of different ages using the technique of intestinal everted sacs. In the postweaning period there is a sharp decrease in the rate of glucose and galactose uptake by the small intestinal mucosa. This change occurs simultaneously with a decrease in lactase activity in the tissue. The kinetic analysis showed a lower Vmax for monosaccharide uptake in the 50-days-old rats as compared with the fifteen-days-old animals, with no change in the apparent Kt. The addition of Tris+ (40 mM) to the incubation media was found to produce a decrease in Vmax for monosaccharide uptake only in the suckling rats but not in the 50-days-old animals; the K1 remained unchanged. Tris+, at the concentration used had previously been shown to completely inhibit lactase activity in homogenates of small intestinal mucosa. These observations lead us to postulate the lactase in the suckling rat might possess a translocating role for sugar, in addition to its hydrolytic function. The decrease in sugar uptake after weaning could be explained by the disappearance of this role.

Age Factors↗
PubMedDate not supplied

[Comparative study of the monosaccharide glycocalyx composition of Bacillus--Lactobacillus--Streptococcus bacteria group by biochemical methods and electron microscopy].

Results of comparison of the data on monosaccharide composition of glycocalix in group Bacillus--Lactobacillus--Streptococcus bacteria using biochemical and electronmicroscopic methods prove that the composition and quantity of monosaccharides varied depending on the genus and peculiarities of microorganisms strains (biological activity in respect of people and warm-blooded animals). At the same time, representatives of this group of microorganisms in the glycocalix had common monosaccharides: glucosamine, sialic acids, galactosamine, galactose, glucose, mannose, fucose, that is the phenotypical display of genetic relations.

Bacillus↗
PubMed1979

Suppression of cell-mediated immune reactions by monosaccharides.

Various monosaccharides have been shown capable of inhibiting lymphokine activity in in vitro assay systems. In this study, we demonstrate that L-fucose can inhibit the ability of lymphokine-containing supernatants to induce skin reactions or cause reductions in the macrophage content of peritoneal exudates. Moreover, L-fucose can inhibit the cutaneous delayed hyoersensitivity reaction and the peritoneal macrophage disappearance reaction (MDR) induced by antigen in actively immunized guinea pigs. The effect of L-rhamnose, another sugar with in vitro inhibitory activity, was investigated in the MDR, and was also shown to be inhibitory. L-arabinose, which has no in vitro effects on lymphokines, had no suppressive effect on any of the in vivo systems studied. No monosaccharide inhibition of Arthus reactions or nonspecific inflammation could be found in these studies. The results demonstrate that monosaccharides capable of inhibiting lymphokine activity in vitro are effective in suppressing in vivo manifestations of cellular immunity.

Animals↗
PubMed1990

[Study on the discrimination of bacteria by gas chromatographic profiles of cellular monosaccharides].

A procedure for obtaining gas chromatographic (GC) profiles of bacterial cellular monosaccharides was described. Some of the unknown component peaks in these profiles were identified. And, based on the complete linkage cluster analysis with the Euclidean distance coefficient, the interpretation of the resulting cellular monosaccharides of bacteria were performed by mini-computer. By means of this method, the discrimination of 5 species (24 strains) of aerobic endospore-forming bacteria. The results showed that there were defined differences between the profiles of cellular monosaccharides of B. anthracis and B. cereus. This procedure has provided a useful method for the classification and identification of microorganisms, for their physiological and biochemical studies, and for studies on their subcellular components.

Bacillus↗
PubMed1993

Monosaccharide and oligosaccharide analysis of proteins transferred to polyvinylidene fluoride membranes after sodium dodecyl sulfate-polyacrylamide gel electrophoresis.

We have developed an intermediate method toward the complete carbohydrate analysis of proteins, which should be universally applicable to all proteins and independent of sample matrix. Using only Coomassie Blue-stained proteins which have been electroblotted onto polyvinylidene fluoride membranes, we report a strategy for: (i) determining unequivocally whether a protein is glycosylated; (ii) obtaining a complete monosaccharide composition; (iii) oligosaccharide mapping which separates most forms according to size, charge and isomerity; and (iv) sequentially releasing and analyzing specific classes of oligosaccharides with endoglycosidases. The method was shown to be applicable to a variety of well characterized soluble glycoproteins and to the membrane-bound protein, the gastric H+, K(+)-ATPase. The monosaccharide composition of the H+,K(+)-ATPase revealed the absence of N-acetylneuraminic or N-glycolylneuraminic acids and a monosaccharide composition which indicated O-linked sugar chains. Oligomannosidic/hybrid and biantennary oligosaccharides were sequentially released and analyzed from one electroblotted band of recombinant tissue plasminogen activator using endo-beta-N-acetylglucosaminidase H and endo-beta-N-acetylglucosaminidase F2, respectively. Sialylated polylactosamine structures were identified and quantified by analyzing high performance liquid chromatography profiles of oligosaccharides first released by peptide-N4-(N-acetyl-beta-D-glucosaminyl)asparagine amidase and then treated with endo-beta-galactosidase, using a single, stained band of recombinant erythropoietin. This recombinant erythropoietin was found to contain eight times more tetrasialylated oligosaccharides than previously reported (Sasaki, H., Bothner, B., Dell, A., and Fukuda, M. (1987) J. Biol. Chem. 262, 12059-12076); 47% of released oligosaccharides were identified as polylactosamine structures.

Animals↗
PubMedDate not supplied

Micellar electrokinetic chromatography of monosaccharides derivatized with 1-phenyl-3-methyl-2-pyrazolin-5-one.

Mono- and disaccharides derivatized with 1-phenyl-3-methyl-2-pyrazolin-5-one (PMP) were separated by micellar electrokinetic chromatography (MEKC), on the basis of their ability to differentially partition between an electroendosmotically driven aqueous phase and sodium dodecyl sulfate micelles. The use of a Tris phosphate buffer, pH 7.5, containing 50 mM sodium dodecyl sulfate, provided good resolution of neutral and basic monosaccharides and disaccharides. Baseline resolution was accomplished for the monosaccharides most commonly found in glycoproteins. A mass detection limit of 20 femtomoles, at a signal-to-noise ratio of three, was achieved by monitoring UV absorbance at 245 nm. The applicability of the system described to the identification and quantitation of monosaccharides obtained from carbohydrate hydrolysates from glycoproteins was investigated.

Antipyrine↗
PubMed1996

[The terminal monosaccharide of antigenic determinant is involved in the binding of human sperm to specific monoclonal IgA antibody].

In order to know whether terminal monosaccharide of human sperm antigen is involved in stimulating antisperm immune response (especialy IgA production) in mucosal immune system, 33 IgA, 12 IgG and 35 IgM monoclonal antibodies (MAbs) all obtained by intragastrointestinal immunization were used in the present study. The molecular weight (MW) of sperm antigens reactive with MAbs were dectected by Western blotting. The MW range of sperm antigens to the above monclonal IgA, IgM and IgG class, except 12 MAbs not reactive with blotted sperm antigen on nitrocellulose strip, is 10-89 KDa, 11-75 KDa and 12-94 KDa respectively. The sperm antigens were separately blocked and digested with 5 lectins and 4 glycosidases. After that, the capacity of human sperm antigens bind with the above antibodies were determined by ELISA. One or more terminal monosaccharides of sperm antigens are involved with the reaction of most MAbs tested. The loss of terminal alpha-fucose, alpha-N-acetylgalactosamine, alpha-N-acetylglucosamine, mannose and beta-galactose but neuraminic acid in the antigenic determinant of human sperm were found to seriously affect the IgA binding with the corresponding sperm antigen. The binding of IgG with sperm antigen was obviously damaged by the removing of terminal alpha-fucose, alpha-N-acetylgalactosamine and alpha-mannose. The results suggest that the terminal monosaccharides of the sperm antigens play an important role in stimulation of antisperm IgA and other antibodies produced by using intragastrointestinal immunization.

Antibodies, Monoclonal↗
PubMed2004

Comparison of antigen constructs and carrier molecules for augmenting the immunogenicity of the monosaccharide epithelial cancer antigen Tn.

We have demonstrated previously that the optimal method for inducing an antibody response against defined cancer antigens is covalent conjugation of the antigen to keyhole limpet hemocyanin (KLH) and use of the potent saponin adjuvant QS-21. Single molecules of glycolipids (tetrasaccharides, pentasaccharides, or hexasaccharides) and MUC1 peptides (containing between one and five MUC1 tandem repeats) conjugated to KLH have proven sufficient for antibody recognition and vaccine construction. However, cancer specificity of monoclonal antibodies against the monosaccharide Tn and disaccharide sTn comes largely from recognition of clusters (c) of these molecules on the cell surface. Tn consists of a monosaccharide (GalNAc) O-linked to serine or threonine on epithelial cancer mucins which are uniquely rich in serines and threonines. We test here several Tn constructs: Tn monosaccharide, Tn(c) prepared on a triple threonine backbone, and Tn prepared on a partially or fully glycosylated MUC1 backbone. We determine that Tn(c) is more effective than Tn, and conjugation to KLH is more effective than conjugation to BSA or polystyrene beads for inducing ELISA reactivity against Tn, and FACS reactivity against Tn-positive tumor cells. Surprisingly, MUC1 glycosylated with Tn at three or five sites per 20 amino acid MUC1 tandem repeat and conjugated to KLH, induced the strongest antibody response against Tn and tumor cells expressing Tn, and had the additional advantage of inducing antibodies against MUC1.

Animals↗
PubMed1976

Serum glycoprotein-type sequence of monosaccharides in membrane glycoproteins of Semliki Forest virus.

Semliki Forest virus was grown in BHK-21 cells and labelled in vivo with radioactive monosaccharides. The virus was disrupted with sodium dodecyl sulphate and the polypeptides were hydrolyzed with pronase. A mixture of type A glycopeptides (for nomenclature, see Johnson and Clamp (1971) Biochem. J. 123, 739-745) of the membrane glycoproteins E1 and E3 was isolated by gel filtration and subjected to sequential degradation with exo-glycosidases. The reduction in the apparent molecular weight and the cleavage of radioactive monosaccharides were monitored with gel filtration. The results suggest that the type A oligosaccharides have similar average structures and contain at the non-reducing terminus 3.4 mol of alpha-D-sialic acid and 0.7 mol of alpha-L-focose, folloled by 3.1 mol of beta-D-galactose, 4.2 mol of N-acetyl-beta-D-glucosamine, 0.7-1.5 mol of alpha-D-mannose, 0.5 mol of beta-D-mannose and 0.6-2.2 mol of N-acetyl-beta-D-glucosamine attached to 1.0 mol of N-acetylglucosamine resistant to N-acetyl-beta-D-glucosaminidase. This innermost monosaccharide unit, therefore, appears to be attached to the peptide. The peptides attached to this N-acetyl-glucosamine had an apparent molecular weight of 720+/-100. We propose the following average structure, compatible with most of our data, for the type A glycopeptides of Semliki Forest virus:.

Animals↗
PubMed1999

Multicomponent quantification of diastereomeric hexosamine monosaccharides using ion trap tandem mass spectrometry.

A rapid means of stereochemical differentiation and quantification for the hexosamine monosaccharides was achieved using electrospray ionization quadrupole ion trap mass spectrometry. The hexosamine monosaccharides, glucosamine, galactosamine, and mannosamine, were derivatized with [Co(DAP)2Cl2]Cl, and the complex [Co(DAP)2(HexNH2)]Cl was generated. Subjecting this complex to collision-induced dissociation provided a unique product ion spectrum for each of the diastereomeric monosaccharide complexes, thus differentiating the stereoisomers. Furthermore, the stereoisomers were quantified. This was achieved by using the relative abundances of product ions from pure standards and using these values to determine the ratio of isomeric products in a mixture. The utility of this quantification method was demonstrated by successfully determining the composition of two- and three-component mixtures of the hexosamines.

Hexosamines↗
PubMed2002

Improved method for quantifying levoglucosan and related monosaccharide anhydrides in atmospheric aerosols and application to samples from urban and tropical locations.

An improved analytical method was developed and validated for the determination of the monosaccharide anhydrides levoglucosan, mannosan, and galactosan in atmospheric aerosol samples. The method uses an external recovery standard, extraction in dichloromethane, trimethylsilylation, addition of an internal standard (1-phenyl dodecane), and analysis by gas chromatography with flame ionization detection (GC-FID) and gas chromatography/mass spectrometry (GC/MS). As external recovery standard, we selected 1,2,3-trihydroxyhexane, which has a similar polarity as the monosaccharide anhydrides; furthermore, it was ensured that the trimethylsilylation step leads to complete derivatization into trimethylsilyl ethers. The reproducibility of the combined trimethylsilylation and analysis of levoglucosan was about 2% for standard solutions, whereas the precision of the entire method for the sum of all three monosaccharide anhydrides (MAs) in real aerosol filter samples was about 5%. The method was applied to aerosol samples from urban and tropical locations. The atmospheric concentration of the MAs in fine (<2.5 microm) aerosols at a primary forest site in Rondĵnia, Brazil, was on average 2.15 microg m(-3) during the dry season when intensive biomass burning occurs, which was almost 400 times higher than during the wet (nonburning) season. Urban total aerosols collected in Gent, Belgium, showed an average atmospheric concentration of MAs of 0.56 microg m(-3) for the winter season, which was a factor of 20 higher than for the summer season. The carbon in the MAs accounted on average for about 5.1% and 1.8% of the organic carbon in the Brazilian dry season and Gent winter aerosols, respectively. Levoglucosan was the major MA, with a relative abundance in the range of 76-93%.

Aerosols↗
PubMed1990

Cell mutants defective in synthesizing a heparan sulfate proteoglycan with regions of defined monosaccharide sequence.

We have demonstrated that mouse LTA cells synthesize cell-surface heparan sulfate proteoglycans (HSPGs) with regions of defined monosaccharide sequence that specifically interact with antithrombin (HSPGact). It remains unclear how HSPGact can be generated by a biosynthetic pathway with no simple template for directing the ordered assembly of monosaccharide units. To examine this issue, we treated LTA cells with ethyl methanesulfonate and then isolated seven stable mutants that synthesize only 8-27% of the wild-type HSPGact but produce normal amounts of other HSPGs. These mutants are recessive in nature and fall into at least two different complementation groups. The delineation of the molecular basis of these defects should help to elucidate the manner by which cells synthesize HSPGs with regions of defined monosaccharide sequence.

Animals↗
PubMed2004

NMR application probes a novel and ubiquitous family of enzymes that alter monosaccharide configuration.

By exploiting nuclear magnetic resonance (NMR) techniques along with novel applications of saturation difference analysis, we deciphered the functions of the previously uncharacterized products of three bacterial genes, rbsD, fucU, and yiiL, which are part of the ribose, fucose, and rhamnose operons of Escherichia coli, respectively. We show that RbsD catalyzes the pyran to furan conversion of ribose, whereas FucU and YiiL are involved in the catalysis of the anomeric conversion of their respective sugars. It was observed that the anomeric exchange of only ribofuranose, not ribopyranose, occurs spontaneously in solution rationalizing its evolutionary incorporation into the nucleic acid. The RbsD and FucU proteins share sequence homology and belong to the same protein family that is found from eubacteria to human, whereas the YiiL homologues exist in archaebacteria and lower eukaryotes. These enzymes, including the galactose mutarotase, exhibit a certain degree of cross-specificity to structurally analogous sugars thereby encompassing all existing monosaccharides in terms of their reactivities. The ubiquitous presence of enzymes involved in the anomeric changes of monosaccharides highlights an importance of these activities in various cellular processes requiring efficient monosaccharide utilization.

Carbohydrate Epimerases↗
PubMed1996

Maintenance of villus height and crypt depth, and enhancement of disaccharide digestion and monosaccharide absorption, in piglets fed on cows' whole milk after weaning.

The aims of the present study were (a) to maintain the structure and function of the small intestine of the piglet after weaning, and (b) to compare the capacity in vivo of sucking and weaned piglets to digest oral boluses of lactose and sucrose and absorb their monosaccharide products. Piglets were fed on cows' whole milk ad libitum every 2 h for 5 d after weaning. Physiological doses of lactose plus fructose (treatment LAC+FRU) and sucrose plus galactose (treatment SUC+GAL) were administered on day 27 of lactation and on the fifth day after weaning, after which time piglets were killed. Villus height and crypt depth were maintained (P > 0.05) by feeding cows' milk after weaning. The areas under the curves (AUC) for galactose and glucose, adjusted for live weight and plasma volume, increased (P < 0.05) after weaning. Despite the enhancement of gut function after weaning, the galactose index (GalI:AUC for galactose ingested as lactose divided by the AUC for the same dose of galactose ingested as the monosaccharide) and fructose index (FruI: AUC for fructose ingested as sucrose divided by the AUC for the same dose of fructose ingested as the monosaccharide), which are indices of digestive and absorptive efficiency, both decreased after weaning. This apparent anomaly may be reconciled by increased growth, and hence surface area, of the small intestine between weaning and slaughter such that 'total' digestion and absorption most probably increased despite apparent decreases in GalI and FruI. Positive correlations (P < 0.05) between villus height and GalI are consistent with the maximum activity of lactase occurring more apically along the villus. Significant linear relationships (P < 0.05) were recorded between villus height at the proximal jejunum and adjusted AUC for galactose and glucose following treatment LAC+FRU, and between villus height at the proximal jejunum and adjusted glucose AUC following treatment SUC+GAL. These relationships suggest that maximum digestion and absorption occurs at increasing distances along the crypt:villus axis in the weaned pig.

Animals↗
PubMed1998

Identification and characterization of human fructose or glucose taste variants with hypogeusia for one monosaccharide but not for the other.

Human psychophysical functions for sweetness are similar for sucrose and fructose, but different for glucose, and suggest different mechanisms for fructose and glucose. Drosophila behavioral and electrophysiological data are similar to the human data and indicate separate receptor cell mechanisms for the monosaccharides. Moreover, fructose 'nontasters' (NTs) and glucose NTs have been identified in two Drosophila species. Identification of human NTs would confirm separate mechanisms and could lead to identification of proteins in human sweet taste by molecular genetic techniques. To identify human NTs, we first obtained responses for sucrose, fructose and glucose from 20 subjects. They tasted seven concentrations of each sugar (2-128 mM), paired with water, and indicated the sweeter of each pair. Functions for recognition indices (RIs) (proportion of subjects recognizing the sugar as sweeter) were similar for sucrose and fructose and different for glucose; this result agrees with the previous studies and supports different mechanisms for the monosaccharides. At 128 mM, RIs for all three sugars were 1.0; this result is consistent with the monogeusia reported by Breslin et al. for concentrations higher than those tested here. Eleven rising-phase concentrations (10-35 mM fructose, 10-90 mM glucose) then were tested on 32 subjects. A statistically significant interaction indicated different regression slopes and supported different monosaccharide mechanisms. From these data, positive identification values (PIDs) (lowest concentration at which the sugar always was judged sweeter than the water) were determined for each subject. The fructose log(PID) and glucose log(PID) data were not well correlated; thus separate mechanisms were supported further. Next, NT traits were defined by log(PID)s > or = 2 SD above the mean for one sugar, while the PID for the other remained within 1 SD of the population mean log(PID). Ninety-two subjects were screened to identify 12 glucose NTs and four fructose NTs. Two glucose NTs and three average subjects were tested in six additional sessions. The NTs showed an experience-induced change: there was a statistically significant reduction of glucose PIDs, but not of fructose PIDs. No change occurred in PIDs of the average subjects for either sugar.

Animals↗
PubMed1979

Movements of monosaccharides between blood and tissues of vascularly perfused small intestine.

1. A method involving the analysis of pulse transients of the vascular concentrations of test sugar and extracellular marker has been used to study the movements of non-metabolized sugars between the cells and vascular fluid of the vascularly perfused small intestine of R. ridibunda. Reasons are given for supposing that the method does properly measure the net entry of sugars into a cellular compartment that includes at least the epithelium. 2. It is found that while the glucose analogues, 3-O-methyl-D-glucose (3MG) and 2-deoxy-D-glucose (2)DG) are able to enter a cellular compartment when they are added to the vascular bed, alpha-methyl-D-glucoside (alpha MG) is able to enter the compartment only at a very slow rate from the vascular bed. In contrast, 3MG and alaph MG are well absorbed from the lumen whereas the inward permeability of 2DG across the lumen face of the epithelium is very low and, unlike 3MG and alpha MG, is not influenced by the presence in the lumen of other transported sugars. 3. The presence of phlorizin in the intestinal lumen increases the flux of alpha MG and of 3MG in the direction vascular bed--bulk phase of lumen. Reasons are given for supposing that the movement of sugars from the vascular bed into the lumen may involve a cellular route but occurs, at least in part, through a paracellular, extracellular route. 4. The exit of monosaccharides that have been loaded previously into the epithelium either from the lumen or from the vascular bed has been investigated. 3MG washes out rapidly into the vascular bed and the exit is stimulated by the addition of D-glucose to the intestinal lumen and by the addition of 2DG, but not alpha MG, to the arterial infusate. In contrast, alpha MG, loaded into the epithelium from the lumen, washes out of the cells only slowly into the vascular bed, so that even with high rates of vascular perfusion alpha MG accumulates within the tissue. The sustained accumulation of alpha MG implies that not only is the permeability for exit into the blood restricted for this sugar, but also the permeability is low across the brush border in an outward direction, cell to lumen ('lobster pot effect'). The wash-out of 3MG into the vascular effluent is sufficently rapid that only in the absence of vascular perfusion is 3MG accumulated within the tissue. 5. The contrasting properties of the monosaccharide transport systems accessible from the intestinal lumen and the vascular bed respectively are discussed in relation to the problems of epithelial transport of monosaccharide.

Animals↗
PubMed1979

Studies on amino acid inhibition of monosaccharide exit from anuran small intestinal epithelium.

1. The effect of the addition of amino acids to the intestinal lumen upon the movement of the monosaccharide alpha-methyl-D-glucopyranoside (alpha MG) from the preloaded epithelium into the blood and into the lumen of the vascularly perfused frog small intestine has been studied. 2. The neutral hydrophobic amino acids tryptophan, leucine, phenylalanine, tyrosine, isoleucine, valine, norleucine and cycloleucine all rapidly inhibit the exit of alpha MG out of the epithelium into the vascular bed. They stimulate backflux of the sugar from the epithelium into the lumen to a very much smaller extent. 3. L-Leucine is a more effective inhibitor of alpha MG exit into the blood than is D-leucine. Near-maximal inhibition of alpha MG exit is seen with 10 mM-L-leucine in the intestinal lumen. 4. The addition of leucine (10 mM) to the lumen of the intestine preloaded with alpha MG approximately halves the rate constant for alpha MG washout into the blood from 12.6 +/- 1.7 (4) x 10(-3) to 5.5 +/- 1.4 (4) x 10(-3) min-1, without appreciably altering the pool of monosaccharide in the tissue. The inhibitory effect of L-leucine upon alpha MG exit into the blood is not abolished by the presence of phlorizin (5 x 10(-5) M) in the intestinal lumen. 5. The complex pattern of inhibition of alpha MG transfer from the lumen to the blood observed upon the addition of L-leucine to the lumen is consistent with the finding that the amino acid inhibits the exit of the monosaccharide out of the epithelium into the blood in addition to any inhibitory effect upon sugar entry across the brush border. 6. It is suggested that alpha MG may be a substrate for a proposed transport system for neutral hydrophobic amino acids which, it is suggested, is present in the basolateral membrane of the epithelial cell.

Amino Acids↗