High-pressure liquid chromatography of cobalamins and cobalamin analogs

Binder, Mitchell; Kolhouse, J F; Horne, K. C. Van; Allen, Robert H.

doi:10.1016/0003-2697(82)90003-3

Cited by 33 publications

(7 citation statements)

References 13 publications

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“…To determine which sections ofthe cbl molecule are important for in vivo activity, we synthesized 16 (16,25). The structural assignments of the monocarboxylic acids was followed as recommended by Anton et al (26).…”

Section: Introductionmentioning

confidence: 99%

“…Analogues with substitutions at the e-propionamide side chain of the C-ring were made by a modification of the methods of Armitage (27) (25).…”

Section: Introductionmentioning

confidence: 99%

“…A molar extinction coefficient of 30,800 M-' cm-' was used for each analogue except for CN-cbl [13-epi] for which a value of 20,600 MWcm- (16) was used. The final analogue solutions gave a single peak when they were analyzed for purity by HPLC utilizing separation methods previously described (25).…”

Section: Introductionmentioning

confidence: 99%

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Inhibition of cobalamin-dependent enzymes by cobalamin analogues in rats.

Stabler¹,

Brass²,

Marcell³

et al. 1991

J. Clin. Invest.

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Section: Introductionmentioning

confidence: 99%

“…Analogues with substitutions at the e-propionamide side chain of the C-ring were made by a modification of the methods of Armitage (27) (25).…”

Section: Introductionmentioning

confidence: 99%

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Inhibition of cobalamin-dependent enzymes by cobalamin analogues in rats.

Stabler¹,

Brass²,

Marcell³

et al. 1991

J. Clin. Invest.

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“…Collagenase (type II) used for the isolation of hepatocytes was from Worthington Biochemical Corp. (Freehold, NJ). HCCL was prepared by the reduction of the previously synthesized cyanocobalamin analogue (35,36). Octanoyl-and palmitoyl-CoA, and defatted BSA were purchased from Sigma Chemical Co. (St. Louis, MO).…”

Section: Introductionmentioning

confidence: 99%

Increased hepatic mitochondrial capacity in rats with hydroxy-cobalamin[c-lactam]-induced methylmalonic aciduria.

Krähenbühl¹,

Ray²,

Stabler³

et al. 1990

J. Clin. Invest.

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IntroductionTreatment of rats with the vitamin B12 analogue hydroxy-cobalaminic-lactaml (HCCL) impairs methylmalonyl-CoA mutase function and leads to methylmalonic aciduria due to intracellular accumulation of propionyl and methylmalonyl-CoA.Since accumulation of these acyl-CoAs disrupts normal cellular regulation, the present investigation characterized metabolism in hepatocytes and liver mitochondria from rats treated subcutaneously with HCCL or saline (control) by osmotic minipump. Consistent with decreased methylmalonyl-CoA mutase activity, 14C02 production from 1-"4C-propionate (1 mM) was decreased by 76% and 82% after 2-3 wk and 5-6 wk of HCCL treatment, respectively. In contrast, after 5-6 wk of HCCL treatment, "4CO2 production from 1-"4C-pyruvate (10 mM) and 1-'4C-palmitate (0. Vitamin B 12 (cobalamin) deficiency leads to decreased activity of the cobalamin-requiring enzymes methylmalonyl-CoA mutase and methionine synthetase (1). Decreased activity of methylmalonyl-CoA mutase results in accumulation of methylmalonyl-and propionyl-CoA, and methylmalonic aciduria (2-4). This defect is analogous to the metabolic defect in patients with hereditary methylmalonic acidurias. The vitamin B 12-deficient rat has provided a useful animal model for the human methylmalonic acidurias and allowed the investigation of hepatic metabolism in the presence of this specific, metabolic insult (4-6). Dietary vitamin B 12 deficiency is difficult to achieve, and the resulting metabolic defect shows a high variability between treated animals (7, 8). An alternative methodology to induce decreased activity of methylmalonyl-CoA mutase is treatment with hydroxy-cobalamin [c-lactam] (HCCL),' a synthetic vitamin B 12 analogue (4-6). Rats treated with HCCL develop decreased hepatic vitamin B 12 levels (4), decreased methylmalonyl-CoA mutase and methionine synthetase activity (6), and decreased propionate metabolism with an increased renal excretion of methylmalonic acid (4-6).Increased hepatocellular propionyl and methylmalonylCoA concentrations inhibit several metabolic activities, including gluconeogenesis (9, 10), urea synthesis (9, 11) fatty acid oxidation (11, 12) and pyruvate decarboxylation (13, 14). In contrast, in rats with chronic accumulation of propionyl and methylmalonyl-CoA induced by dietary vitamin B 12 deficiency, gluconeogenetic and ketogenetic responses to fasting were intact in vivo (4), suggesting compensatory mechanisms that mitigate the toxic effects of accumulated acyl-CoAs. One compensatory mechanism in rats with dietary and functional vitamin B 12 deficiency is increased hepatic total CoA concentration (4, 15).To further study hepatic metabolism under conditions of impaired methylmalonyl-CoA mutase activity, hepatic mitochondrial function in HCCL-and saline-treated rats was characterized. Mitochondrial function was quantified by measurement of fuel oxidation rates in isolated hepatocytes and mitochondria. In addition, activities of mitochondrial and extramitochondrial enzymes were determined in isolat...

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“…Compared with microbiological assays, HPLC combines high performance separation and identification and provides a relatively rapid method for determination of B 12 and its related compounds. Many HPLC systems have been developed for B 12 determination (Frenkel et al 1979;Binder et al 1982;Lambert et al 1992;Vinas et al 1996;Moreno and Salvado 2000;Wongyai 2000), but they are not sufficiently sensitive to directly detect B 12 at the low concentrations found in natural waters. More recently, C-18 solid phase extraction (SPE) techniques have been combined with HPLC-UV detection to concentrate and measure trace amounts of B 12 in seawater (Okbamichael and Sañudo-Wilhelmy 2004) and have improved our understanding of vitamin B 12 in some biogeochemical and ecological processes (Sañudo-Wilhelmy et al 2006;Gobler et al 2007;Taylor and Sullivan 2008;Panzeca et al 2008Panzeca et al , 2009).…”

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confidence: 99%

Analysis of vitamin B₁₂ in seawater and marine sediment porewater using ELISA

Zhu

Aller

Kaushik

2011

Limnology & Ocean Methods

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Vitamin B 12 (B 12 ) is a set of closely related organocobalt compounds required by phytoplankton. A highly sensitive and specific enzyme-linked immunosorbent assay (ELISA) was developed for the determination of B 12 in seawater and marine sediment porewater. An antibody directed against B 12 was coated on the surface of a 96-well microtiter plate, and horseradish peroxidase (HRP) was used as a labeling enzyme. In the indirect competitive immunoassay format, water samples or standards and a constant amount of HRP-labeled B 12 were added into the microtiter plate wells, HRP-labeled and free B 12 compete for binding to the plate-bound antibody. After immunoreaction, the immunochemically adsorbed HRP-B 12 conjugate was determined by measuring the absorbance produced in a solution containing substrate tetramethylbenzidine (TMB) and hydrogen peroxide. The calibration graph for B 12 was linear over the range of 0.4-100 ng/mL (0.3-74 nM; higher concentrations were not evaluated) with a detection limit of 0.2 ng/mL (3s). Coupled with C-18 column extraction-preconcentration, the method is readily applicable to seawater levels (~1-10 pM). No interferences from humic acids, total dissolved organic matter (DOM), and salinity were observed. ELISA determined B 12 in coastal seawater and surface tidal flat porewater (0-2 cm) ranged from 4.5-38 pM and 12-47 pM, respectively.

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High-pressure liquid chromatography of cobalamins and cobalamin analogs

Cited by 33 publications

References 13 publications

Inhibition of cobalamin-dependent enzymes by cobalamin analogues in rats.

Inhibition of cobalamin-dependent enzymes by cobalamin analogues in rats.

Increased hepatic mitochondrial capacity in rats with hydroxy-cobalamin[c-lactam]-induced methylmalonic aciduria.

Analysis of vitamin B₁₂ in seawater and marine sediment porewater using ELISA

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High-pressure liquid chromatography of cobalamins and cobalamin analogs

Cited by 33 publications

References 13 publications

Inhibition of cobalamin-dependent enzymes by cobalamin analogues in rats.

Inhibition of cobalamin-dependent enzymes by cobalamin analogues in rats.

Increased hepatic mitochondrial capacity in rats with hydroxy-cobalamin[c-lactam]-induced methylmalonic aciduria.

Analysis of vitamin B12 in seawater and marine sediment porewater using ELISA

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Analysis of vitamin B₁₂ in seawater and marine sediment porewater using ELISA