189663a0Nature1894765196102256636630028-0836196110.1038/189663a0ukNatureNatureNATUREnatureNature is a weekly international journal publishing the finest peer-reviewed research in all fields of science and technology on the basis of its originality, importance, interdisciplinary interest, timeliness, accessibility, elegance and surprising conclusions. Nature also provides rapid, authoritative, insightful and arresting news and interpretation of topical and coming trends affecting science, scientists and the wider public./nature/journal/v189/n4765issueJournal homeArchiveCurrent issueAdvance online publicationPrivacy policySubscribeNature Publishing GroupCurrent issue189663a0Chemical Oxidation of Uridine Diphosphate-Glucose to Uridine Diphosphate-Glucuronic Acid
AU  - JACOBSON, BERNARD
AU  - DAVIDSON, EUGENE A.Department of Biochemistry and Center for the Study of Aging, Duke University, Durham, North Carolina.THE importance of uronic acids in carbohydrate metabolism has only recently begun to be elucidated. The metabolism of inositol and ascorbate is known to involve uronic acid intermediates1,2, and nucleotide-bound glucuronic acid has been implicated in hyaluronic acid synthesis3.Uridine diphosphate (UDP)-glucuronic acid has been prepared chemically by Khorana and may also be produced enzymatically by the action of UDP-glucose dehydrogenase4. Both these preparations are relatively time-consuming and require the separation of the product from several contaminating reactants and side products.
Marsh has reported the oxidation of glucose-1-phosphate to glucuronic acid-1-phosphate5, and this report describes a similar oxidation of TJDP-glucose.
Powdered platinum catalyst was prepared in aqueous solution from Adam's platinous oxide catalyst by hydrogenation at atmospheric pressure. 100 mgm. of reduced catalyst were suspended in 10 c.c. of water and introduced into a narrow vessel similar to that described by Marsh. The cylindrical vessel and sintered glass disk were attached to a U-shaped tube and immersed in a constant tempera ture bath; oxygen was bubbled through the solution at a rate adequate to keep the heavy catalyst particles in suspension. The UDP-glucose solution (63 jjimoles) was then added, and oxygenation was allowed to proceed for 10^ hr. at a temperature of 55 , the pH being maintained between 7 and 8 by addition of 1 per cent potassium carbonate. This contrasted with the oxidation of glucose-1-phosphate at 45 , at which temperature no UDP-glucuronic acid was produced. At the end of the reaction the catalyst was removed by filtration, washed, and the filtrate and washings assayed for uronic acid content6.
In order to show that the uronic acid produced was still nucleotide-linked, an aliquot was spotted for paper electrophoresis in 0-1 M citrate-phosphate buffer, pH. 2. The electrophoresis was carried out at 800 V. for 2 hr., and the two ultra-violet absorbing spots were eluted from the paper with water; one of them gave a positive carbazole test, uridine spectrum, and showed a 1: 1 ratio of uronic acid to uridine based on a molar extinction coefficient for uridine of 10,000, the other material being unreacted UDP-glucose. The UDP-glucuronic acid could also be assayed by enzymatic transfer to o-aminophenol as described by Dutton and Storey, utilizing a fraction of rat liver homogenate sedimenting between 500gr and 37,000gr as the enzyme system7.
Use of 5 per cent palladium on charcoal as catalyst and oxygenation of the reaction mixture for 7 hr. resulted in no increase in the amount of uronic acid present, indicating that no oxidation had occurred.
Table 1. VARIATION OF YIELD WITH TIME
Time Yield[ast] (hr.) (per cent)
5-0 12
5-5 24
10-5 39
[ast] Based on recovered UDP-glucose.
On the basis of recovered UDP-glucose, the best yield under the conditions studied has been 39 per cent. It is felt that the ease of isolation and relatively short reaction time required make the reaction feasible for the production of UDP-glucuronic acid.Charalampos, , F. C., and Lyras, , C., J. Biol. Chem., 228, 1 (1957).PubMedHorowitz, , H., and King, , C., J. Biol. Chem., 205, 815 (1953).PubMedISIChemPortGlaser, , L., and Brown, , D., Proc. U.S. Nat. Acad. Sci., 41, 5 (1955).Strominger, , J., Kalckar, , H. M., Axelrod, , J., and Maxwell, , E. S., J. Amer. Chem. Soc., 76, 6411 (1954).ArticleISIChemPortMarsh, , C. A., J. Chem. Soc., 1578 (1952).Dische, , Z., J. Biol. Chem., 167, 189 (1947).ISIChemPortDutton, , G. J., and Storey, , I. D. E., Biochem. J., 57, 275 (1954).PubMedISIChemPort