1.Multiply 13C-substituted monosaccharides: synthesis of D-(1,5,6-13C3)glucose and D-(2,5,6-13C3)glucose.
Wu J1, Serianni AS, Bondo PB. Carbohydr Res. 1992 Mar 30;226(2):261-9.
D-(1,5,6-13C3)Glucose (7) has been synthesized by a six-step chemical method. D-(1,2-13C2)Mannose (1) was converted to methyl D-(1,2-13C2)mannopyranosides (2), and 2 was oxidized with Pt-C and O2 to give methyl D-(1,2-13C2)mannopyranuronides (3). After purification by anion-exchange chromatography, 3 was hydrolyzed to give D-(1,2-13C2)mannuronic acid (4), and 4 was converted to D-(5,6-13C2)mannonic acid (5) with NaBH4. Ruff degradation of 5 gave D-(4,5-13C2)arabinose (6), and 6 was converted to D-(1,5,6-13C3)glucose (7) and D-(1,5,6-13C3)mannose (8) by cyanohydrin reduction. D-(2,5,6-13C3)Glucose (9) was prepared from 8 by molybdate-catalyzed epimerization.
2.Theoretical aspects of 13C metabolic flux analysis with sole quantification of carbon dioxide labeling.
Yang TH1, Heinzle E, Wittmann C. Comput Biol Chem. 2005 Apr;29(2):121-33.
The potential of using sole respirometric CO2 labeling measurement for 13C metabolic flux analysis was investigated by metabolic simulations. For this purpose a model was created, considering all CO2 forming and consuming reactions in the central catabolic and anabolic pathways. To facilitate the interpretation of the simulation results, the underlying metabolic network was parameterized by physiologically meaningful flux parameters such as flux partitioning ratios at metabolic branch points and reaction reversibilities. For real case flux scenarios of the industrial amino acid producer Corynebacterium glutamicum and different commercially available (13)C-labeled tracer substrates, observability and output sensitivity towards key flux parameters was investigated. Metabolic net fluxes in the central metabolism, involving, e.g. glycolysis, pentose phosphate pathway, tricarboxylic acid cycle, anaplerotic carboxylation, and glyoxylate pathway were found to be determinable by the respirometric approach using a combination of [1-13C] and [6-13C] glucose in two parallel studies.
3.Measurement of pentose phosphate-pathway activity in a single incubation with [1,6-13C2,6,6-2H2]glucose.
Ross BD1, Kingsley PB, Ben-Yoseph O. Biochem J. 1994 Aug 15;302 ( Pt 1):31-8.
The isotopically substituted molecule D-[1,6-13C2,6,6-2H2]glucose is introduced for measuring the relative activities of the pentose phosphate pathway (PPP) and glycolysis in a single incubation. PPP activity in cultured cells was determined by gas chromatography/mass spectrometric analysis of lactate produced by cells incubated with [1,6-13C2,6,6-2H2]glucose. Two other isotopes, [1,5,6-13C3]glucose and [6-13C,1,6,6-2H3]glucose, were not satisfactory for measurements of this activity. This method has four advantages over the traditional one in which 14CO2 production from [1-14C]glucose and [6-14C]glucose is compared: (1) repeated measurements can be made on a single set of cells, (2) only a single incubation is required, (3) extensive CO2 production by Krebs-cycle activity does not interfere with the measurements and (4) it is not necessary to measure the amount of glucose consumed in order to calculate relative activities of the PPP and glycolysis.