Homocysteine, vitamin B12, and serum and erythrocyte folate in peritoneal dialysis and hemodialysis patients.
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BACKGROUND: Plasma homocysteine (Hcy) is an independent risk factor for cardiovascular disease. High levels of plasma Hcy have been observed in end-stage renal disease patients. Few studies have compared peritoneal dialysis (PD) and hemodialysis (HD) patients and few data are available on erythrocyte folate (ery-F) levels in dialysis patients. OBJECTIVES: To evaluate plasma Hcy concentrations, vitamin B12 (B12), and folate status in dialysis patients; to analyze the possible causes of high Hcy levels; to follow up changes in folate and B12 concentrations after 6 months. DESIGN: A cross-sectional observational study. SETTING: Nephrology division and laboratory of hematology in a university and clinical research hospital. PATIENTS: The study included 82 patients treated with PD for 37 + 37 months and 70 patients treated with HD for 136 + 95 months. LABORATORY METHODS: Plasma Hcy was measured by the immunoenzymatic IMx Hcy FPIA method (Abbott Laboratories, Diagnostic Division, Abbott Park, IL, U.S.A.), serum folate (s-F) and ery-F by the Stratus folate fluorometric enzyme-linked assay, and B12 by the Stratus vitamin B12 fluorometric enzyme-linked assay (DADE-Behring, Newark, DE, U.S.A.). RESULTS: Ninety-six percent of PD and 97% of HD patients had Hcy levels above the cutoff (13.5 micromol/L). Homocysteine level was higher in HD than in PD patients, while the prevalence of hyperhomocysteinemia was similar with the two techniques. Erythrocyte folate was significantly higher in PD (1333 +/- 519 pmol/L) than in HD (1049 +/-511 pmol/L, p < 0.01). Statistically significant correlations were observed between Hcy and B12, s-F, ery-F, and dialysis duration. Multivariate analysis showed a strong correlation between s-F and Hcy. After 6 months there were no differences in Hcy, B12, s-F, and ery-F levels. CONCLUSIONS: Plasma Hcy levels were high in more than 95% of our dialysis patients, with no relation to the type of dialysis. Vitamin B12 and folate were normal in the majority of our patients. However, serum folate was the major determinant of Hcy levels. Such a relation between Hcy and folate suggests that levels of folate within the reference interval are inadequate for dialysis patients. (+info)
Interaction between ethanolamine ammonia-lyase and methylcobalamin. Half-site reactivity with an adenosylcobalamin-dependent enzyme.
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The adenosylcobalamin-dependent enzyme ethanolamine ammonia-lyase contains two active sites per molecule. The effects of methylcobalamin on the properties of this enzyme differ qualitatively depending on whether one or both of these sites is occupied by the cobamide. At 0.5 mol of methylcobalamin/mol of active sites, catalytic activity fell rapidly to approximately 30% of control levels, thereafter remaining constant for an hour. With the partially inhibited enzyme, Km values for ethanolamine and adenosylcobalamin were 5.5 muM and 1.6 muM, respectively, values that do not differ significantly from those of uninhibited enzyme. When the methylcobalamin per active site ratio was increased to 1, the decline in activity became progressive with time, eventually falling to levels much lower than seen at a cobamide per active site ratio of 0.5. Methylcobalamin also promotes the formation of a complex stable to gel filtration between ethanolamine and enzyme. Complex formation increased with increasing methylcobalamin per active site ratios up to a ratio of 0.7/1, at which point 0.5 mol of ethanol/mol of active sites was taken up. Ethanolamine uptake did not increase at higher methylcobalamin to active sites ratios. Methylcobalamin itself was taken up by enzyme, forming a complex containing 0.5 mol of methylcobalamin/mol of active sites that was stable to gel filtration. Measurement by the technique of Hummel and Dreyer ((1962) Biochim. Biophys. Acta 63, 530-532), however, showed one methylcobalamin binding site per active site. The formation of enzyme-ligand complexes stable to gel filtration was not affected by 5'-deoxyadenosine nor did 5'-deoxyadenosine by itself promote the formation of a stable complex between enzyme and ethanolamine. These observations were interpreted as evidence indicating half-site reactivity of ethanolamine ammonia-lyase with methylcobalamin. Comparison with previous results suggested that this half-site reactivity was an epiphenomenon not related to catalysis. (+info)
Dietary strategies for lowering homocysteine concentrations.
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BACKGROUND: Elevated plasma total homocysteine (tHcy) concentrations are associated with increased risk of vascular disease, and there is a strong inverse association between dietary and blood folate and blood tHcy concentrations. Increased folate consumption may lower the risk of tHcy-mediated cardiovascular disease. OBJECTIVES: The objective was to determine the most appropriate means of increasing dietary folate to reduce plasma tHcy. DESIGN: Sixty-five free-living subjects aged 36-71 y with tHcy concentrations >/=9 micromol/L participated in a randomized, controlled trial to compare 3 approaches for increasing dietary folate to approximately 600 microg/d: folic acid supplementation, consumption of folic acid-fortified breakfast cereals, and increased consumption of folate-rich foods. RESULTS: An intake of 437 microg folic acid/d from supplements resulted in a 27-nmol/L increase in serum folate and a 21% reduction in tHcy, relative to the change in a control group. In subjects who consumed folic acid-fortified breakfast cereal, folate intake increased by an average of 298 microg, serum folate increased by 21 nmol/L, and tHcy concentrations decreased by 24%. Increased intakes of folate-rich foods resulted in a 418-microg increase in dietary folate, a 7-nmol/L increase in serum folate, and a 9% reduction in tHcy concentrations. The decrease in tHcy was negatively correlated (r = -0.66) with the increase in serum folate. CONCLUSIONS: Daily consumption of folic acid-fortified breakfast cereals and the use of folic acid supplements appear to be the most effective means of reducing tHcy concentrations. The reduction in tHcy was significantly negatively correlated with the increase in serum folate, which may be a useful marker for measuring dietary change. (+info)
Anaerobic chlorophyll isocyclic ring formation in Rhodobacter capsulatus requires a cobalamin cofactor.
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The isocyclic ring of bacteriochlorophyll (BChl) is formed by the conversion of Mg-protoporphyrin monomethyl ester (MPE) to protochlorophyllide (PChlide). Similarities revealed by blast searches with the putative anaerobic MPE-cyclase BchE suggested to us that this protein also uses a cobalamin cofactor. We found that vitamin B(12) (B(12))-requiring mutants of the bluE and bluB genes of Rhodobacter capsulatus, grown without B(12), accumulated Mg-porphyrins. Laser desorption/ionization time-of-flight (LDI-TOF) MS and NMR spectroscopy identified them as MPE and its 3-vinyl-8-ethyl (mvMPE) derivative. An in vivo assay was devised for the cyclase converting MPE to PChlide. Cyclase activity in the B(12)-dependent mutants required B(12) but not protein synthesis. The following reaction mechanism is proposed for this MPE-cyclase reaction. Adenosylcobalamin forms the adenosyl radical, which leads to withdrawal of a hydrogen atom and formation of the benzylic-type 13(1)-radical of MPE. Withdrawal of an electron gives the 13(1)-cation of MPE. Hydroxyl ion attack on the cation gives 13(1)-hydroxy-MPE. Withdrawal of three hydrogen atoms leads successively to 13(1)-keto-MPE, its 13(2)-radical, and cyclization to PChlide. (+info)
The mechanism of action of ethanolamine ammonia-lyase, an adenosylcobalamin-dependent enzyme. The source of the third methyl hydrogen in the 5'-deoxyadenosine generated from the cofactor during catalysis.
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Ethanolamine ammonia-lyase is an adenosylcobalamin-dependent enzyme which catalyzes the conversion of ethanolamine and propanolamine to ammonia and the corresponding aldehydes. A mechanism has been proposed for this and other adenosylcobalamin-dependent reactions which involves cleavage of the carbon-cobalt bond of the cofactor followed by abstraction of a substrate hydrogen atom by the adenosyl fragment to form 5'-deoxyadenosine. In support of this proposal, a previous study demonstrated that the deamination of propanolamine by ethanolamine ammonia-lyase is accompanied by the reversible cleavage of the carbon-cobalt bond of the cofactor, with the production of 5'-deoxyadenosine (Babior, B.M., Carty, T.J., and Abeles, R.H. (1974) J. Biol. Chem. 249, 1689-1695). The present study is concerned with the origin of the third hydrogen atom on the methyl group of the 5'-deoxyadenosine produced in that reaction. The 5'-deoxyadenosine isolated from an incubation mixture initially containing enzyme, [5',5'-D2]adenosylcobalamin, and [1,1-D2]propanolamine was chemically degraded so that the 4' and 5' carbon atoms were, respectively, converted to the carbonyl and methyl carbons of acetaldehyde. Analysis of the p-nitrophenylhydrazone of the acetaldehyde by gas-liquid chromatography-mass spectroscopy revealed 3 deuterium atoms/molecule, indicating that two of the methyl hydrogens originated from adenosylcobalamin and the third was donated by substrate. This observation provides further support for the participation of 5'-deoxyadenosine in the mechanism of adenosylcobalamin-dependent reactions. (+info)
Gastrointestinal bleeding and iron absorption in the experimental blind loop syndrome.
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Rats with surgically created self-filling jejunal blind loops and the blind loop syndrome manifested gastrointestinal bleeding and hyperabsorption of iron. Although the mean hematocrit and serum iron levels of rats with self-filling blind loops became overtly anemic and manifested low-serum iron levels. It is suggested that the documented gastrointestinal bleeding in these rats with the experimental blind loop syndrome is another manifestation of damage to the intestinal epithelium in conditions of small intestinal bacterial overgrowth. (+info)
Folate and vitamin B12 status of women in Newfoundland at their first prenatal visit.
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BACKGROUND: Newfoundland has one of the highest rates of neural tube defects in North America. Given the association between low maternal folic acid levels and neural tube defects, a cross-sectional study was conducted to obtain base-line data on the folate and vitamin B12 status of a sample of women in Newfoundland who were pregnant. METHODS: Blood samples were collected between August 1996 and July 1997 from 1424 pregnant women in Newfoundland during the first prenatal visit (at approximately 16 weeks' gestation); this represented approximately 25% of the women in Newfoundland who were pregnant during this period. The samples were analysed for serum folate, vitamin B12, red blood cell folate and homocysteine. RESULTS: Median values for serum folate, red blood cell folate and serum vitamin B12 were 25 nmol/L, 650 nmol/L and 180 pmol/L, respectively. On the basis of the interpretive criteria used for red blood cell folate status, 157 (11.0%) of the 1424 women were deficient (< 340 nmol/L) and a further 180 (12.6%) were classified as indeterminate (340-420 nmol/L). Serum homocysteine levels, measured in subsets of the red blood cell folate status groups, supported the inadequate folate status. Serum vitamin B12 levels of 621 (43.6%) women were classified as deficient or marginal; however, the validity of the interpretive criteria for pregnant women is questionable. INTERPRETATION: A large proportion of pregnant women surveyed in Newfoundland in 1997 had low red blood cell folate levels. (+info)
Osmotic reflextion coefficients of capillary walls to low molecular weight hydrophilic solutes measured in single perfused capillaries of the frog mesentery.
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1. Individual capillaries of the transilluminated frog mesentery have been perfused with suspensions of human red cells in frog Ringer solution containing 1-0 g albumin 100 ml.-1. The outer surface of the mesentery has been washed with normal frog Ringer solution and with frog Ringer solutions made hypertonic by addition of one of the following solutes: sodium chloride (100 m-mole. 1.-1); urea (100 m-mole.1.-1); sucrose (20-50 m-mole. 1.-1); cyanocobalamin (8-5 m-mole. 1.-1). The temperature of the mesentery was between 14 and 16 degrees C in all experiments. 2. Wtih the mesentery superfused with normal Ringer, the filtration coefficient was determined from measurements of the rate of fluid filtration across the capillary wall, at a series of known capillary pressures (Michel, Mason, Curry & Tooke, 1974). Filtration coefficient varied from 0-69 X 10(-3) to 4-45 X 10(-3) mum. sec-1 .cm H2O-1 with an average value of 1-87 X 10(-3) mum. sec-1. cm H2O-1. 3. When the superfusate was made hypertonic by the addition of a test solute, the osmotic reflextion coefficient (sigma) of the capillary wall to test solute was calculated from the additional rate of filtration, the concentration of test solute in the superfusate and the filtration coefficient. Average values for sigma were: sodium chloride, 0-068 +/- 0-03 (three capillaries); urea, 0-071 +/- 0.015 (four capillaries); sucrose, 0-115 +/- 0-023 (seven capillaries); cyanocobalamin, 0-100 +/- 0-03 (three capillaries). 4. In further experiments, the osmotic reflextion coefficients to sodium chloride, urea and sucrose were determined in the same capillary. Five technically acceptable experiments were carried out. Although there were differences in the value of sigma between different capillaries, in any one capillary values of sigma were of the same magnitude and there appeared to be no significant trend with the molecular size of the test solute. 5. Our findings are inconsistent with the hypothesis that there is a single pathway for water and small hydrophilic molecules across the capillary wall. 6. Our results may be interpreted in terms of an exclusive channel for water in parallel with a channel shared by both water and small hydrophilic molecules. It is suggested that the exclusive water channel may be the membranes and cytoplasm of the endothelial cells and the shared channel may be located in the intercellular junctions. 7. Our data suggest the exclusive water channel represents about 10% of the total filtration coefficient in frog mesenteric capillaries. The shared channel shows relatively little restriction to the molecules investigated. Estimates of the volume flow throught the two channels are made for conditions where hydrostatic pressure differences and osmotic pressure differences are the driving forces. (+info)