Pablo Hueso

Universidad de Salamanca, Helmantica, Castille and León, Spain

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Publications (40)88.23 Total impact

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    ABSTRACT: The content, distribution of individual species, and the fatty acid composition of phospholipids (PL) from ewe's and goat's milk were analyzed. The binding of enterotoxigenic and uropathogenic Escherichia coli strains to PL and the inhibition of bacterial hemagglutination by PL were addressed using high performance thin-layer chromatography-overlay assays and microtiter plates, respectively. Ovine and caprine milk contained more PL than bovine milk but less than human milk. The profile of individual PL was similar, including sphingomyelin, phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine and phosphatidylinositol in both ovine and caprine milk. Regarding the fatty acid composition, a high content of long-chain fatty acids (more than C16) and unsaturated fatty acids, with C18:1 as the most abundant was found in ovine and caprine milk PL. Ovine milk has longer and less saturated fatty acids while caprine milk has shorter and more saturated ones. Neither the adhesion of any bacterial strains assayed to the individual PL from ovine or caprine milk nor the inhibition of bacterial hemagglutination by PL were observed. These are important constituents of the milk fat globule membrane, but it seems that they do not play a role in the defence of new-borns against bacteria if the results obtained are taken into account.
    No preview · Article · Jun 2013 · Grasas y Aceites
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    ABSTRACT: Sialic acid and gangliosides content and profiles were analyzed in infant formulae, whey protein concentrates and human milk. In infant formulae, N-acetylneuraminic (Neu5Ac) and N-glycolylneuraminic (Neu5Gc) acid ranged from 147.6–199.7 to 3.3–8.3 mg L−1, in whey protein concentrate from 1.6–2.4 g 100 g−1 to 2.8–20.2 mg 100 g−1 and in human milk from 299.9 to 2.1 mg L−1. Gangliosides ranged from 0.25 to 2.29 mg lipid-bound sialic acid L−1 in reconstituted infant formula, the main gangliosides being GD3 (73.3–55.5%), GM3 (3.5–36.6%) and GT3 (1.4–14.1%) and O-acetyl-GD3 (5.3–18.9%). In whey protein concentrates, 0.7–55.6 mg lipid-bound sialic acid 100 g−1 were found with the profile 82.1–86.9% GD3 and 1.8–2.5% GM3. Human milk gave 2 mg lipid-bound sialic acid L−1, with the profile 97.4% GM3 and 2.5% GD3. Intakes of sialic acid and gangliosides are lower with infant formulae versus human milk, especially during the first stages of life.
    No preview · Article · Nov 2011 · International Dairy Journal
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    ABSTRACT: The effects of simulated gastrointestinal digestion upon sialic acid and gangliosides in infant and follow-on formulas and human milk, as well as their bioaccessibility, have been evaluated. The gastric stage is the step that causes a greater decrease in sialic acid and ganglioside contents. The intestinal stage only decreases the total and individual contents of gangliosides. After gastrointestinal digestion, neither sialic acid nor gangliosides were found in the nonbioaccessible fraction. The highest bioaccessibility (100 × content in soluble fraction after gastrointestinal digestion/total content) of sialic acid is found in human milk (87%), followed by infant formula (77%) and follow-on formula (16%). In the case of gangliosides, the highest bioaccessibility is present in the follow-on formula (51%), followed by human milk (29%) and infant formula (5%).
    No preview · Article · May 2011 · Journal of Agricultural and Food Chemistry
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    ABSTRACT: Milk glycosphingolipids (GSL) have been reported to participate in the newborn's defense against pathogens. Taking this into account, in this study we determined the neutral GSL content of ovine milk, including its fatty acid profile. Its role in bacterial adhesion was also addressed by immunodetection of separate GSL in a high-performance thin-layer chromatography overlay assay. Ovine milk has a neutral GSL pattern similar to human milk and includes lactosylceramide (LacCer; 45.7%), monohexosylceramide (glucosylceramide and galactosylceramide, 31.2%), globotriaosylceramide (Gb3; 19.1%), and globotetraosylceramide (Gb4; 3.5%). Globotriaosylceramide and Gb4 are present in human but not bovine milk. Neutral GSL contained C23:0 and C24:0 as the most abundant fatty acids, a finding consistent with its high content of very long chain fatty acids (longer than C20). Most fatty acids were saturated and had a low content of polyunsaturated fatty acids. Bovine enterotoxigenic Escherichia coli strains bound strongly to LacCer and showed a weak binding to monohexosylceramide. The K99 strain also bound strongly to Gb3, and F41 to Gb4. Lactosylceramide, monohexosylceramide, and Gb3 were also observed to bind to human uropathogenic E. coli strains. The results reported here show the ability of neutral GSL in ovine milk to bind to E. coli strains. These compounds could be used as an alternative and available source to supplement infant or bovine formulas with a view to preventing bacterial infections.
    No preview · Article · Jan 2010 · Journal of Dairy Science
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    ABSTRACT: Sialoglycoconjugate contents of several milk replacers (MR) currently used in Spain have been determined. The ingestion of these compounds by calves fed bovine milk or MR is also discussed. Total sialic acids and glycoprotein-, oligosaccharide-, casein-, and lipid-bound sialic acids and free sialic acid were determined. High sialic acid contents in all the fractions studied, including total sialic acids, were found. N-acetylneuraminic acid was found to be the major sialic acid in MR. However, MR-D and H had a high N-glycolylneuraminic acid (NeuGc) content in all fractions (15-40%), while E and F only had a high NeuGc content in the glycoprotein fraction. Five different sialyloligosaccharides -3'-sialyllactose, 6'-sialyllactose, 3'-sialyllactosamine, 6'-sialyllactosamine, and disialyllactose, which are the most abundant oligosaccharides in bovine milk- were detected in the MR. Samples B to I contained high amounts of these oligosaccharides.The content of individual gangliosides was very similar to that of bovine milk, with GD3 as the major ganglioside. Although MR are not formulated as regards their sialic acid content, high amounts of sialic acid-containing glycoconjugates with high levels of NeuGc, a sialic acid critical in the adhesion of some Escherichia coli strains to calf intestinal epithelium were detected.The ingestion of MR analyzed in this work could protect new-born calves from several enteric pathogens.
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    ABSTRACT: The phospholipid (PL) content was determined comparatively in the milk fat globule membrane (MFGM) and whole milk including their fatty acid profiles. The possible role of milk PLs in defence against pathogens was also addressed. The MFGM and whole milk showed a similar distribution of PL species; however, the fatty acid contents of the PL species were different. Total PL from MFGM showed a decrease in C18:0 content in parallel with an increase in C18:1 and C18:2 and very long-chain fatty acid (more than C20) content. No significant differences in the fatty acid content of phosphatidylcholine and sphingomyelin from either source were found. However, the phosphatidylethanolamine from MFGM had more C18:1 and C18:2 and less C14:0 and C16:0 than that from whole milk. A similar but less pronounced result was found for phosphatidylserine/phosphatidylinositol. Enterotoxigenic Escherichia coli strains failed to bind to PL, which had been previously separated by high-performance thin-layer chromatography.
    No preview · Article · May 2009 · International Dairy Journal
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    ABSTRACT: Several components of milk fat globule membranes (MFGMs) have been reported to display beneficial health properties and some of them have been implicated in the defense of newborns against pathogens. These observations prompted us to determine the glycosphingolipid content of MFGMs and their interaction with pathogens. A comparative study with whole milk components was also carried out. Milk fat globules and MFGMs were isolated from milk. Gangliosides and neutral glycosphingolipids were obtained from MFGMs and whole milk and their fatty acid contents were determined by gas chromatography-mass spectrometry (GC-MS). MFGMs and whole milk showed similar ganglioside and neutral glycosphingolipid contents, with whole milk having more GM3 and glucosylceramide and less GD3, O-acetyl GD3, O-acetyl GT3, and lactosylceramide. The fatty acid content of gangliosides from both sources showed a similar composition. However, the neutral glycosphingolipid fatty acid content seemed to be quite different. Whole milk had fewer very-long-chain fatty acids (18.1% vs. 46.4% in MFGMs) and more medium-chain and unsaturated C18:1 and C18:2 fatty acids. Milk fat globules, MFGMs, lactosylceramide, and gangliosides GM3 and GD3 were observed to bind enterotoxigenic Escherichia coli strains. Furthermore, bacterial hemagglutination was inhibited by MFGMs and glycosphingolipids.
    No preview · Article · Nov 2008 · Biological Chemistry
  • Pablo Hueso · Enrique Villar
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    ABSTRACT: Without Abstract
    No preview · Article · Mar 2006 · Glycoconjugate Journal
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    ABSTRACT: The protective effect of human milk against infection is well known. Several non-immunologic components, including complex carbohydrates, have been described. The present study was undertaken to determine the sialic acid distribution in different milk fractions (complex carbohydrates). Milk samples from 12 Spanish women at three different lactational stages (colostrum, transitional milk and mature milk) were analyzed. Total and glycoprotein-bound, oligosaccharide-bound, casein-bound, and lipid-bound sialic acids were determined. Sialic acids from human milk are mainly bound to oligosaccharides and only a small amount is present bound to glycoproteins or in the free form. All the fractions analyzed showed a similar trend: sialic acids decrease rapidly along lactation. Casein-bound sialic acid does not follow this trend. We detected the presence of an O-acetylated species of N-acetylneuraminic acid. In human milk from Spanish women we observed slightly different values than those previously reported. This could be a result of population differences but nutritional or methodological aspects can not be discarded.
    No preview · Article · Dec 2004 · Journal of Pediatric Gastroenterology and Nutrition
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    ABSTRACT: The objectives of this work were to study the FA composition of milk gangliosides, as well as to gain further insight into the characterization of human milk gangliosides. The potential capacity of human milk gangliosides to adhere to human enterotoxigenic Escherichia coli (ETEC-strains) was also studied. Human milk gangliosides were isolated and identified by high-performance TLC or immunoassay. The latter also was used to assay bacterial adhesion. The FA composition of gangliosides was studied by GC. The presence of O-acetyl GD3 (Neu5,9Ac2alpha2-8 NeuAcalpha2-3Galbeta1-4GlcCer) and trace amounts of GM1 [Galgamma1]3-3GalNAcgamma1,-3(Neualpha2-3)Galbeta1-4GlcCerl in human milk was confirmed. Medium-chain FA were almost absent in colostrum, whereas in the subsequent stages they rose to 20%. The levels of long-chain FA decreased after colostrum. With respect to the degree of saturation, gangliosides from colostrum were richer in monounsaturated FA than gangliosides synthesized during the rest of the lactation period, opposite to the pattern for PUFA. A human-ETEC colonization factor antigen II-expressing strain showed binding capacity to human milk GM3 (NeuAcalpha2-3Gal[1-4GlcCer). New data on human milk gangliosides have been gathered. A thorough knowledge of their composition is needed since they may have important biological implications in regard to newborns' defense against infection.
    No preview · Article · Mar 2004 · Lipids
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    ABSTRACT: Diarrhea caused by enterotoxigenic Escherichia coli (ETEC) is the main infectious disease of newborn calves. The first step of infection involves bacterial attachment to the intestinal mucosa. This adhesion is mediated by fimbriae that recognize some glycoconjugates on the host cell surface, in particular, several gangliosides. Because milk also contains gangliosides, these have been suggested to serve as ligands for bacterial fimbriae and thus prevent the bacterial attachment to mucosa. The most relevant ETEC strains in calves, including those with K99 and F41 fimbriae, were assayed to determine whether they are able to bind gangliosides isolated from several stages of bovine lactation. Both GM3 and GD3, the main gangliosides of milk, were recognized by ETEC strains, although the different fimbriae showed diverse levels of affinity. Unexpectedly, the adhesion to colostral gangliosides was considerably weaker than that to gangliosides from the other stages of lactation. Because the carbohydrate moiety did not change and because differences in the percentages of unsaturated FA and sphingosine between colostrum and other stages were observed, we conclude that the differences in adhesion could be due to a different composition of the ganglioside ceramide.
    No preview · Article · Aug 2003 · Lipids
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    ABSTRACT: Several lines of research support a role for human milk oligosaccharides in the defense of breast-fed infants against pathogens. Some ofthese oligosaccharides contain at least one moiety of sialic acid and are, thus, termed sialyloligosaccharides. These constitute a significant component (>1 g/L) of human milk. It is well established that milk composition varies among species, and previous reports have indicated that one ofthe differences between human and bovine milk is precisely their contents of sialyloligosaccharides. Because most infant formulas are manufactured with bovine milk components, it follows that formula-fed and breast-fed infants ingest dissimilar quantities of these carbohydrate structures. To ascertain these differences and their impact along lactation, the contents of oligosaccharide-bound sialic acids and major sialyloligosaccharides in samples of human and bovine milk (obtained at different lactation stages) were determined. In addition, infant formulas were assayed for their sialyloligosaccharide contents. Seven sialyloligosaccharides were identified in human milk; namely, 3'-sialyl-3-fucosyllactose and sialyllacto-N-tetraoses (a and b+c), the predominant structures at all lactation stages. Five sialyloligosaccharides were identified in bovine milk, of which 6'-sialyllactosamine and 3'-sialyllactose were the most abundant. In addition, sialyloligosaccharides in human and bovine milk decreased along lactation, and infant formulas did not contain significant amounts of sialyloligosaccharides. The results point to the general conclusion that regarding both qualitative and quantitative aspects, milk from humans and cows and infant formulas have different oligosaccharide contents. In this sense, bottle-fed infants are subject to reduced sialyloligosaccharide intake as compared to breast-fed infants.
    Preview · Article · Jan 2003 · Journal of Dairy Science
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    ABSTRACT: Milk oligosaccharides can act as soluble receptors that block bacterial adhesion to the different epithelia. Colonization factor antigens (CFA)/I- and CFA/II-expressing enterotoxigenic Escherichia coli (ETEC) strains constitute one of the main causes of diarrhea in infants. Here, the inhibition of hemagglutination mediated by these strains by milk oligosaccharides was tested. Human milk oligosaccharides showed a strong inhibitory capacity, which decreased when the oligosaccharides were desialylated. Because milk oligosaccharides also are present in the urine of neonates receiving mothers' milk, their ability to bind two uropathogenic Escherichia coli (UPEC) strains was also examined. UPEC strains expressing P (Pap) and P-like (Prs) fimbriae are responsible for infections of the urinary tract such as pyelonephritis and cystitis. The hemagglutination mediated by these strains was inhibited by human milk oligosaccharides. The sialylated fraction was partially responsible for this inhibition in the case of the UPEC expressing the P-like fimbria because differences were found after desialylation. Although bovine milk oligosaccharides were less efficient at inhibiting the hemagglutination of ETEC strains, they were still quite good inhibitors of UPEC strains.
    No preview · Article · Nov 2002 · Journal of Nutrition
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    ABSTRACT: Milk oligosaccharides have been proposed to play an important role in newborn defense, blocking bacterial adhesion to the intestinal mucosa and preventing infections. Some studies have been performed on human milk oligosaccharides. Here we checked whether bovine milk oligosaccharides would achieve the same protective action against the most common calf enteric pathogens. Seven enterotoxigenic Escherichia coli strains, isolated from diarrheic calves, were selected. All strains managed to agglutinate horse erythrocytes, and we therefore used the inhibition of hemagglutination in the presence of oligosaccharides as an indicator of the union between oligosaccharide and bacterial adhesins. Oligosaccharides from different stages of bovine lactation and standard oligosaccharides were assayed. Midlactation milk, in particular that corresponding to the transition period, proved to be the most efficient at inhibiting hemagglutination. The standard oligosaccharides used pointed to the preference of several strains (K99-, F41-, and F17-fimbriated) for alpha2,6-linked sialic acid. By contrast, B23 fimbriae exhibited higher affinity for alpha2,3-sialylated isomers and B64 seemed to require N-acetylglucosamine for binding. Our results suggest a general trend for milk oligosaccharides. Probably they participate in the protection of newborn mammals from pathogens.
    No preview · Article · Feb 2002 · Glycoconjugate Journal
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    ABSTRACT: The sialoglycoconjugate content of human milk has been extensively studied. However, little attention has been paid to the changes occurring in these compounds in bovine milk during lactation. Since sialoglycoconjugates are very abundant in milk from the early stages of lactation, they have been suggested to be important for the nutrition of the newborn during the first months of life. The distribution of sialoglycoconjugates (expressed as glycoconjugate-bound sialic acid) from four different stages of lactation (colostrum, transitional, mature, and late-lactation milks) was investigated in four Spanish-Brown cows. All the fractions studied (total sialic acids, glycoproteins, oligosaccharides, casein, and gangliosides) showed a similar trend. We found the highest values in the colostrum, these decreasing in transitional and mature milks and increasing again in late-lactation milk. We also found a selective change in the relative contents of glycoprotein- and oligosaccharide-bound sialic acids. In mature milk, the latter increased up to 80% (59% in colostrum) and the former decreased to 3.9% (35.3% in colostrum). It would appear that the decrease in oligosaccharide-bound sialic acid is compensated by the increase in glycoprotein-bound sialic acid. From these results, it is deduced that newborn infants or calves fed with infant formulas or milk replacers, respectively, should be supplemented with sialoglycoconjugates to approximate the composition of human and cow milk as far as is practicable.
    No preview · Article · Jun 2001 · Journal of Dairy Science
  • M J Martín · Samuel Martín-Sosa · Pablo Hueso
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    ABSTRACT: The stage of lactation is one of the most important factors that influence milk composition. Changes in fatty acids from triacylglycerols and phospholipids have already been reported. In this study, we looked for a lactational change in the ganglioside lipid moiety since ganglioside contents and patterns vary strongly with stage of lactation. Individual gangliosides from four stages were isolated, methanolized to cleave the bonds between individual constituents, and derivatized for gas-liquid chromatography and gas chromatography/mass spectrometry analyses. Ceramide components, both fatty acids (as methyl esters derivatives) and long-chain bases, were identified and quantified. The results pointed to a marked change in ceramide from colostrum to milk that was characterized by a dramatic decrease in saturated and the longest-chain fatty acids as well as an increase in 18:1 and 18:2. The major long-chain base along lactation was a recently described structure, 3-ethoxy-15:0 sphinganine. Other new long-chain base structures appeared in these gangliosides. All these changes suggest differences in the fluidity of the fat globule membrane, reflecting physiological variations in cows with respect to milk production.
    No preview · Article · Apr 2001 · Lipids
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    ABSTRACT: In a previous work (Zanetta et al. Glycobiology 9, 255-266 (1999)), it was reported that all constituents of gangliosides could be obtained as heptafluorobutyrate derivatives after methanolysis in a single gas chromatography analysis. This report demonstrates that gas chromatography coupled with mass spectrometry in the electron impact mode allows identification and quantification of long-chain bases and fatty acids without interference from monosaccharides. On the basis of ions specific for families and for individual compounds, sphingosines, sphinganines, and phytosphingosines (including ramified, unsaturated, hydroxylated, and etherified compounds) can be identified. Fatty acid methyl esters, including linear, ramified, unsaturated, and hydroxylated species, are identified and quantified in the same way. Possible extensions of this method to the fatty moiety of other lipids (alkylacylglycerol and dimethyl acetal) are discussed.
    No preview · Article · Oct 2000 · Analytical Biochemistry
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    ABSTRACT: The effects of chronic ethanol or sucrose administration to rats on acetylcholinesterase from brain and liver were investigated. Membrane-bound and soluble acetylcholinesterase activities were determined in fractions prepared by centrifugation. The thermal stability and the effects of temperature and different types of alcohols on acetylcholinesterase activity were also studied. Membrane-bound acetylcholinesterase activity increased (p < 0.01) in the liver after chronic ethanol administration, whereas no differences among groups in the encephalic areas, except in the brain stem soluble form, were found. Membrane-bound acetylcholinesterase from the ethanol- and sucrose-treated groups was more stable at the different temperatures assayed between 10 and 50°C than that corresponding to the control group. Non-linear Arrhenius plots were obtained with preparations of membrane-bound acetylcholinesterase from rat liver, with discontinuities at 30°C (control or sucrose groups) or 34-35°C (alcohol group). Assays made with membrane-bound or soluble enzyme from brain showed linear Arrhenius plots in all groups studied. The inhibitory effects of increasing concentrations of ethanol, n-propanol and n-butanol on acetylcholinesterase preparations from forebrain, cerebellum, brain stem and liver of the three experimental groups (control, sucrose-fed and ethanol-fed) were very similar. However, n-butanol displayed a biphasic action on particulate or soluble preparations of rat forebrain. n-butanol inhibited (competitive inhibition) at higher concentrations (250-500 mM), while at lower concentrations (10-25 mM), the alcohol inhibited at low substrate concentrations but activated at high substrate concentration. These results suggest that the liver is more affected by ethanol than the brain. Moreover, the lipid composition of membranes is probably modified by ethanol or sucrose ingestion and this would affect membrane fluidity and consequently the behaviour of acetylcholinesterase.
    No preview · Article · Apr 2000 · Neurochemical Research
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    ABSTRACT: The chemical behaviour of cytidine 5′-monophospho-N-acetylneuraminic acid (CMP–Neu5Ac) under different conditions was studied. We found that the stability of this sugar nucleotide in water and cacodylate buffer was high and similar in both systems, although it was more stable in ethanol/water. Hydrolysis rates increased with temperature. 2-Deoxy-2,3-dehydro-N-acetylneuraminic acid (NeuAc2en) was formed in the first few minutes of the treatments. CMP–Neu5Ac was more stable in alkaline (pH 8 and 10) than in acidic (pH 3 and 5) conditions; its stability was very similar in neutral (water and ethanol/water) and alkaline conditions. The yield of Neu5Ac2en increased with pH and temperature. Additionally, CMP–Neu5Ac dissolved in the solvent system n-butanol/n-propanol/0.1 N HCl (pH 1) was hydrolyzed very fast (98–99% after 1 min). Moreover, this sugar nucleotide dissolved in water or cacodylate buffer (pH 5.8) and when subjected to successive freezing and thawing cycles did not show any increase in the hydrolysis rate. Finally, no decomposition of N-acetylneuraminic acid (Neu5Ac) or Neu5Ac2en was observed under the above-mentioned conditions (pH, temperature, freeze–thaw cycles).
    No preview · Article · Jul 1999 · Comparative biochemistry and physiology. B, Comparative biochemistry
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    ABSTRACT: The activity of UDP-N-acetylglucosamine 2-epimerase was determined in the liver of rats and guinea-pigs of different ages. The activity of this enzyme in rats was low at birth, increased to a maximum value on day 15, and fell gradually until day 30. Thereafter, it increased up to the 60th day. The activity profile of the enzyme from guinea-pig liver was very similar. However, guinea-pig activity was 2-5 times lower than in rats. Both rats and guinea-pigs displayed similar liver sialic acid contents which increased from birth to 2 months of age. Rats also showed a N-glycolylneuraminic acid content that decreased from birth to 2 months. From these results we can inferred that postnatal UDP-N-acetylglucosamine 2-epimerase activity seems to be correlated with age and the developmental states of rats and guinea-pigs.
    No preview · Article · Oct 1997 · Comparative Biochemistry and Physiology Part B Biochemistry and Molecular Biology

Publication Stats

677 Citations
88.23 Total Impact Points


  • 1987-2013
    • Universidad de Salamanca
      • • Department of Biochemistry and Molecular Biology
      • • Faculty of Biology
      Helmantica, Castille and León, Spain
  • 1988-1992
    • Universidad de León
      • • Facultad de Veterinaria
      • • Department of Biochemistry and Molecular Biology
      León, Castile and Leon, Spain