Two diastereoisomeric 2,3:5,6-di-O-ethylidene-β-D-alloses and related derivatives

Direct condensation of β-D-allose with acetaldehyde in the presence of sulfuric acid formed two of eight possible 2,3:5,6-di-O-ethylidene-D-alloses in overall yields of 84–96%. Conditions of the reaction were varied to favor formation of either isomer. The presence of a furanose ring in both isomers was established by converting the diastereoisomers into 1,4-di-O-acetyl-D-allitol analogs. P.m.r. analysis of the reducing isomers, their 3-deuterio analogs, their 1-O-acetyl derivatives, and the 1,5,6-triacetate of a common hydrolysis product, 2,3-O-ethylidene-D-allose, established the anomeric configuration of D-allose as β-, and the C-2′ atom in the 2,3-O-ethylidene ring as R and as either R or S in the 5,6-O-ethylidene ring.     

The oxidation of terminal D-galactofuranose residues of a galactan and a glycoprotein by a D-galactose Oxidase preparation from Dactylium dendroides

A galactan, isolated from the unicellular organism Prototheca zopfii, and a glycoprotein from a hyphal cell-wall fraction of the fungus Pithomyces chartarum have been oxidised by a D-galactose oxidase preparation from Dactylium dendroides. The oxidised polymers were subsequently reduced with sodium borotritide. The site of oxidation was identified as C-6 of non-reducing D-galactofuranosyl residues in both polymers.     

Proton magnetic resonance spectra of D-gluco-oligosaccharides and D-glucans

The p.m.r. spectra of some D-gluco-oligosaccharides and D-glucans in deuterium oxide were studied with respect to the anomeric proton. In (1→2)-linked glucobioses, the effect of change in configuration of the hydroxyl group at C-1 on the chemical shifts of the glycosidic proton is noted. Equilibrium mixtures of (1→2)-linked glucobioses contained more α-anomer than did the other examples, despite the cis configuration of substituents at C-1 and C-2. Some D-glucans were investigated with regard to the degree of branching, although solubility was a limitation.     

Preparation of some acylated D-arabino-hexopyranosuloses and 4-deoxy-D-glycero-hex-3-enopyranosuloses

Oxidation of 1,3,4,6-tetra-O-benzoyl-α- and β-D-glucopyranose gave the tetra-O-benzoyl-α- and -β-D-arabino-hexopyranosuloses (3α and β), from which benzoic acid was readily eliminated to give the anomeric tri-O-benzoyl-4-deoxy-D-glycero-hex-3-enopyranosuloses (4α and β). The anomeric 1-O-acetyl-tri-O-benzoyl-D-arabino-hexopyranosuloses (7α and β) were obtained as very unstable syrups which readily lost benzoic acid. Treatment of tetra-O-benzoyl-2-O-benzyl-D-glucopyranose (1) with hydrogen bromide gave 3,4,6-tri-O-benzoyl-α-D-glucopyranosyl bromide (5) in one step.     

Metabolism of 14C-labelled D-glucose, D-fructose and allitol by Itea plants

The metabolism of D-[1-¹⁴C]glucose, D-[6-¹⁴C]glucose, D-[1-¹⁴C]fructose and D-[6-¹⁴C]fructose by leafy spurs of Itea plants results in rapid incorporation of label into allitol and D-allulose. The patterns of labelling found in the allitol and D-allulose are discussed, a direct interconversion from D-glucose and D-fructose being indicated. Allitol has been found to be an active metabolite in Itea plants.     

Proton and carbon-13 nuclear magnetic resonance studies on methyl (methyl d-galactosid)uronates and their per-O-acetyl derivatives

The ¹H- and ¹³C-n.m.r. spectra of the anomeric methyl (methyl d-galactosid)uronates, as well as the ¹H-n.m.r. spectra of their acetyl derivatives, were analyzed. The spectra of the unacetylated d-galactopyranosiduronates showed good correlation with those of the corresponding anomeric d-galactopyranuronic acids and their methyl esters, and with those of the anomeric methyl d-galactopyranosides. From the values of the chemical shifts and coupling constants, it was concluded that the anomeric methyl (methyl d-galactopyranosid)uronates and their corresponding peracetates are in the ⁴C₁(d) conformation. The chemical shifts in the ¹³C-n.m.r. spectra show good correlation with those of the methyl d-galactosides. The signals of the furanose derivatives appear at fields lower than those of the corresponding pyranose compounds.     

A non-hydrogen-bonding role for the 4-hydroxyl group of D-Galactose in its reaction with D-Galactose oxidase

Several 4-deoxy analogs of methyl β-D-galactopyranoside are oxidized by D-galactose oxidase. The rates associated with their various, axially attached 4-substituents follow the sequence OH>NH₂>F?>Cl> H; these differences are attributed mainly to variations in K_m. Other 4-deoxy analogs, namely, the 4-azido-4-deoxy, 4-bromo-4-deoxy-, 4-deoxy-4-iodo, and 4-thio derivatives were found to be inactive. These observations indicate that the axial 4-hydroxyl group of D-galactopyranose does not play a hydrogen-bonding role primarily, but constitutes a substituent of a size optimal for interaction with the enzyme.     

Synthesis of β-l-fucopyranosyl phosphate andl-fucofuranosyl phosphates by the MacDonald procedure

Fusion or β-l-fucopyranose tetraacetate with phosphoric acid for 1 min at 50° gives a 9:1 anomeric mixture of the α-and β-pyranosyl phosphates. Longer fusion times give the α-anomer exclusively. The l-fucofuranose tetraacetates were synthesized for the first time by acetolysis or methyl-2,3,5-tri-O-acetyl-β-l-fucofuranoside. Fusion or the furanose tetraacetates with phosphoric acid gave a mixture or the fucofuranosyl phosphates in which the β-anomer predominated (β/α = 2.4). Anomeric pairs in the fucofuranose series appear to be distinguishable by the chemical shift of the C-6 methyl protons, as already shown by Sinclair and Sleeter in the pyranose series.     

Reactions of 4,6-disulphonates of methyl d-glucopyranosides and methyl d-galactopyranosides with thio-nucleophiles

The reactions of some 4,6-disulphonates of methyl 2,3-di-O-acyl-(and di-O-methyl)-d-glucopyranosides and -galactopyranosides, with thiocyanate, thioacetate, and thiobenzoate anions, have been studied under a variety of conditions. In the glucoside series, somewhat similar reactivity is shown by isomers differing only in anomeric configuration, and there is no very great difference between the reactivities of a 2,3-dibenzoate and its 2,3-di-O-methyl analogue. In contrast to the situation in the β-d-galactoside series, the presence of O-benzoyl groups in an α-d-galactoside does not have an unfavourable effect on displacement at C-4. Two hexose derivatives containing the novel 4,6-epithio bridge are described.     

Equilibria of the anomeric and ring forms of ammonium 3-deoxy-d-manno-octulosonate in deuterium oxide

The tautomeric composition of a solution of ammonium 3-deoxy-d-manno-octulosonate (KDO, 1a) in D₂O at 28° was assessed by means of ¹³ C-F.t.-n.m.r. spectroscopy. The results revealed the presence of 6?0 and 11 % of the α and β anomers of the pyranose, and 20 and 9 % of the two furanoses, and suggested, but did not unequivocally prove, that the major furanose form is the α anomer. To facilitate interpretation of the spectral results for 1, ammonium 3,5-dideoxy-d-arabino(or ribo)-octulosonate (3a) was prepared by the reaction of 5-deoxy-d-erythro-pentose with sodium oxalacetate at pH 11. A chromatographically homogeneous, noncrystalline sample of 3 was obtained by lyophilization, and characterized as its (4-nitrophenyl)hydrazone (m.p. 162-163°). The ¹³C-n.m.r. spectrum of a solution of 3a in D₂O revealed it to be substantially all in the α-pyranose form. No signals were obtained for the possible 1,4-lactone of 3. As the 1,5-lactone and furanose forms are impossible for 3, it exhibited no signals analogous to those attributed to furanoid 1. On the basis of these results for 3, the two lactone forms of 1 were excluded from consideration, and the three pairs of ¹³C-n.m.r. signals observed at ≈45, 86, and 104 p.p.m. were assigned to the furanose forms of 1.     

Selenium derivatives of L-arabinose, D-ribose,and D-xylose

Attempted cyclization of 2,3,4-tri-O-methyl-5-seleno-L-arabinose dimethyl acetal in acidic solution gave the corresponding diselenide. Intramolecular attack by the selenobenzyl group at C-5 of 5-O-p-tolylsulfonyl-L-arabinose dibenzyl diseleno-acetal resulted in the formation of benzyl 1,5-diseleno-L-arabinopyranoside. Similarly, 2,3,5-tri-O-methyl-4-O-p-tolylsulfonyl-D-xylose dibenzyl diselenoacetal gave benzyl 2,3,5-tri-O-methyl-1,4-diseleno-L-arabinofuranoside, and 2,3,4-tri-O-acetyl-5-O-p-tolylsulfonyl-D-xylose (or ribose) dibenzyl diselenoacetal gave benzyl 2,3,4-tri-O-acetyl-1,5-diseleno-D-xylo- (or ribo-)pyranoside. The glycosylic benzylseleno group was removed from the pyranoside with mercuric acetate, but attempted deacetylation of the product led to decomposition and not to the expected 5-seleno-D-xylopyranose.     

Sex difference in L-glutamine D-fructose-6-phosphate aminotransferase activity of mouse submandibular gland

L-Glutamine D-fructose-6-phosphate aminotransferase (2-amino-2-deoxy-D-glucose-6-phosphate ketol-isomerase (amino transferring), EC 5.3.1.19) activities in the three main salivary glands of male and female mice were measured. It was found that the activity in the submandibular gland was about 10 times more in females than in males, whereas the activities in the sublingual and parotid glands of males and females were similar. The activity in the submandibular gland of female mice was not affected appreciably by ovariectomy but it decreased to the level in males on injection of testosterone. The activity in males was not affected appreciably by injection of progesterone or 17β-estradiol, but it increased to the level in females after castration. The increased acitivity in castrated male mice was decreased again to the normal level by testosterone injection. Thus, this sex difference is caused by androgen, not by female hormones. On the basis of in vivo experiments using actinomycin D, it was suggested that testosterone produced an “enzyme inhibitor”, which suppressed the enzyme activity in the submandibular glands of androgen-rich animals.     

Solvent effects in the reaction of 2,3,6-tri-O-methyl D-glucopyranose and phenyl isocyanate

The reaction of a 3:1 mixture of 2,3,6-tri-O-methyl-α- andβ-D-glucopyranose (1) with phenyl isocyanate, in acetone, benzene, dimethyl sulfoxide, 1,4-dioxane, pyridine, and tetrahydrofuran, showed the isomer ratio in the product mixture to be solvent-dependent. The ratio varied from 4.55β/α in benzene to 0.49 in Me₂SO. It is proposed that an activated complex formed between 1 and a 1-isocyanate complex provides for the simultaneous attack of a nucleophile on the anomeric hydroxyl proton, and of an electrophile on the ring-oxygen atom of 1, causing mutarotation. The rate of mutarotation of the activated complex is dependent on the degree of solvation of the anomeric hydroxyl group. Solvent association is highest in ME₂SO and lowest in benzene. The reaction rate is higher in benzene than 1,4-dioxane and is slowest in Me₂SO. The hydroxyl group at C-1 is ≈ 3 times as reactive as the one at C-4.     

A sensitive,specific radioisotope assay for l-glutamine-d-fructose-6-phosphate aminotransferase

A sensitive and specific radioassay for l-glutamine-d-fructose-6-phosphate aminotransferase (EC 5.3.1.19) activity is presented. Picomoles of product are measurable, and the assay can be applied to systems having limited quantities of available protein, particularly in extracts of either cell or organ cultures. The assay is at least 10,000 times more sensitive under K₁ concentrations of fructose 6-phosphate than the modified Elson-Morgan colorimetric assay and 20 times more sensitive under saturating conditions of fructose 6-phosphate. As little as 0.5 μg of cell-extract protein will yield measurable product. In contrast, 280 μg of crudeextract protein from colon is required with the modified Elson-Morgan colorimetric assay.     

6-Phosphofructo-2-kinase and D-fructose-2,6-bisphosphatase activities in mung bean seedlings

6-Phosphofructo-2-kinase (ATP: D-fructose-6-phosphate-2-phosphotransferase) and D-fructose-2,6-bisphosphatase activities have been found in extracts prepared from etiolated mung bean seedlings. The activity of 6-phosphofructo-2-kinase exhibits a sigmoidal shape in response to changes in concentrations of both substrates, D-fructose 6-phosphate and ATP (S_0.5 values of 1.8 and 1.2 mM, respectively). Inorganic orthophosphate (P_i) has a strong stimulating effect on the 2-kinase activity (A_0.5 at about 2 mM), moderately increasing the V_max and modifying the response into hyperbolic curves with K_m values of 0.4 and 0.2 mM for fructose 6-phosphate and ATP, respectively. 3-Phosphoglycerate (I_0.5 about 0.15 mM) partially inhibited the kinase activity by counteracting the P_i activation. In contrast, the activity of D-fructose-2,6-bisphosphatase (K_m 0.38 mM) is strongly inhibited by P_i (I_0.5 0.8 mM) lowering its affinity to fructose-2,6-P₂ (K_m 1.4 mM). 3-Phosphoglycerate activites the enzyme (A_0.5 at about 0.3 mM) without causing a significant change in its K_m for fructose-2,6-P₂. The activities of both of these enzymes in relationship to the metabolic role of D-fructose 2,6-bisphosphate in the germinating seed is discussed.     

Synthesis of D-ristosamine and its derivatives

A convenient preparative route involving eleven steps starting from D-glucose is described for the synthesis of D-ristosamine (15) hydrochloride. Methyl 2-deoxy-β-D-arabino-hexopyranoside, prepared from 3,4,6-tri-O-acetyl-1,5-anhydro-2-deoxy-D-arabino-hex- 1-enitol, was benzylidenated, and the product mesylated to give methyl 4,6-O-benzylidene-2-deoxy-3-O-methylsulfonyl-β-D-arabino-hexopyranoside. Azidolysis of this compound and subsequent opening of the 1,3-dioxane ring with N-bromosuccinimide gave methyl 3-azido-4-O-benzoyl-6-bromo-2,3,6-trideoxy-βD-ribo-hexopyranoside. Simultaneous reduction of the azido and bromo groups gave a mixture that was benzoylated to give methyl N,O-dibenzoyl-β-D-ristosaminide and then hydrolyzed to 15 hydrochloride (3-amino-2,3,6-trideoxy-D-ribo-hexopyranose hydrochloride).     

Chemical and physical studies on oligosaccharides. Isolation,configurations, and conformations of the per-O-acetyl-O-β-D-glucopyranosyl-(1→3)-D-arabinoses

The reaction of octa-O-acetylcellobiononitrile with sodium methoxide, and acetylation of the resulting mixture, afforded the anomeric peracetates of O-β-D-glucopyranosyl-(1→3)-D-arabinopyranose and O-β-D-glucopyranosyl-(1→3)-D-arabinofuranose, which were isolated by fractional recrystallization. Their structures, anomeric configurations, and conformations were studied by chemical, optical, and spectroscopic methods.     

D-Ribulose-1,5-bisphosphate carboxylase/oxygenase from chemolithotrophically grown Rhizobium japonicum

Ribulose-1,5-bisphosphate carboxylase/oxygenase has been purified from chemolithotrophically grown Rhizobium japonicum SR and ribulose-5-phosphate kinase activity has also been detected in extracts of such cells. Electrophoretically homogeneous ribulosebisphosphate carboxylase/oxygenase purified in the presence of PMSF showed two types of large subunits of 55 000 and 53 000 daltons and small subunits of 14 200 daltons. The heterogeneity of large subunits was not observed when the enzyme was prepared in the presence of PMSF and DIFP. Ribulose-1,5-bisphosphate carboxylase from R. japonicum was inhibited by antibodies to this enzyme and a single precipitin band from the antibody-enzyme interaction was observed on double diffusion plates. Antibodies to R. japonicum enzyme did not cross-react on immunodiffusion plates with the ribulosebisphosphate carboxylase/oxygenases from wheat, spinach, soybean and tobacco.     

Laser-raman spectra of d-glucose and sucrose in aqueous solution

Laser-Raman spectra of d-glucose in water at various concentrations were recorded below 1700 cm^?1. Assignments of the frequencies of d-glucose were proposed, based on earlier work on the vibrational spectra of this sugar, and determination by other techniques of the anomeric composition of aqueous solutions of d-glucose. The proportions of the anomers found from the ratio of the Raman intensities for the same models of vibrations were similar to those found by other techniques. Assignments of the frequencies for sucrose were proposed, in the light of previous results on d-glucose and d-fructose, and the effect, on their Raman spectra, of the condensation of these two monosaccharides was pointed out.