1-Deoxy-D-xylulose: synthesis based on molybdate-catalyzed rearrangement of a branched-chain aldotetrose.

1-Deoxy-D-xylulose has been prepared in seven steps and approximately 21% overall yield from 2,3-O-isopropylidene-D-erythrono-1,4-lactone. The key reaction involves transformation of a branched-chain aldotetrose to the 1-deoxy-2-ketopentose catalyzed by molybdic acid. Other branched-chain aldotetroses containing bulkier substituents at C2 also engage in the conversion, suggesting routes to protected 2-ketoses and alpha-ketoacids/esters. This synthetic route mimics reactions of the non-mevalonate isoprenoid pathway in plants and bacteria. [reaction: see text]

Acyclic forms of [1-(13)C]aldohexoses in aqueous solution: quantitation by (13)C NMR and deuterium isotope effects on tautomeric equilibria.

High-resolution (13)C NMR spectra (150 MHz) have been obtained on the complete series of D-aldohexoses (D-allose 1, D-altrose 2, D-galactose 3, D-glucose 4, D-gulose 5, D-idose 6, D-mannose 7, D-talose 8) selectively labeled with (13)C at C1 in order to detect and quantify the percentages of acyclic forms, and to measure and/or confirm percentages of furanoses and pyranoses, in aqueous solution. Aldehyde and hydrate signals were detected for all aldohexoses, and percentages of these forms at 30 degrees C ranged from 0.006 to 0.7% (hydrate) and 0.0032 to 0.09% (aldehyde). Aldehyde percentages are largest for the altro, ido, and talo configurations, ranging from 0.01 to 0.09%; the ido configuration yielded the most hydrate (0.74%). Hydrate/aldehyde ratios vary with aldohexose configuration, ranging from 1.5 to 13, with gluco exhibiting the smallest ratio and gulo the largest. (2)H Equilibrium isotope effects (EIEs) on aldohexose anomerization were measured in D-galactose 3 and D-talose 8 selectively (13)C- and (2)H-labeled at C1 and H1. The (2)H isotope effect on (13)C chemical shift, and broadband (1)H- and (2)H-decoupling, were exploited to permit simultaneous observation and quantitation of the protonated and deuterated molecules in NMR samples containing equimolar mixtures of D-[1-(13)C]aldose and D-[1-(13)C; 1-(2)H]aldose. Small (2)H EIEs were observed for 8, but were undetectable for 3. These results suggest that configuration at C2 influences the magnitude of the (2)H isotope effect at H1 and/or that the observed effect cannot be reliably interpreted due to complications arising from the involvement of acyclic aldehyde forms as intermediates in the interconversion of cyclic forms. The observed (2)H isotope effects on aldohexose tautomeric equilibria provide new insights into the important question of whether (2)H substitutions can alter aldofuranose ring conformation, and lead to the identification of an optimal (2)H- and (13)C-substituted 2-deoxyribofuranose isotopomer on which to investigate this potential effect.

2-Deoxy-beta-D-erythro-pentofuranose: hydroxymethyl group conformation and substituent effects on molecular structure, ring geometry, and NMR spin-spin coupling constants from quantum chemical calculations.

The effect of hydroxymethyl conformation (gg, gt, and tg rotamers about the C4-C5 bond) on the conformational energies and structural parameters (bond lengths, bond angles, bond torsions) of the 10 envelope forms of the biologically relevant aldopentofuranose, 2-deoxy-beta-D-erythro-pentofuranose (2-deoxy-D-ribofuranose) 2, has been investigated by ab initio molecular orbital calculations at the HF/6-31G level of theory. C4-C5 bond rotation induces significant changes in the conformational energy profile of 2 (2gt and 2tg exhibit one global energy minimum, whereas 2gg exhibits two nearly equivalent energy minima), and structural changes, especially those in bond lengths, are consistent with predictions based on previously reported vicinal, 1,3- and 1,4-oxygen lone pair effects. HF/6-31G-optimized envelope geometries of 2gg were re-optimized using density functional theory (DFT, B3LYP/6-31G), and the resulting structures were used in DFT calculations of NMR spin-spin coupling constants involving 13C (i.e., J(CH) and J(CC) over one, two, and three bonds) in 2gg according to methods described previously. The computed J-couplings were compared to those reported previously in 2gt to assess the effect of C4-C5 bond rotation on scalar couplings within the furanose ring and hydroxymethyl side chain. The results confirm prior predictions of correlations between 2J(CH), 3J(CH), 2J(CC) and 3J(CC), and ring conformation, and verify the usefulness of a concerted application of these couplings (both their magnitudes and signs) in assigning preferred ring and C4-C5 bond conformations in aldopentofuranosyl rings. The new calculated J-couplings in 2gg have particular relevance to related J-couplings in DNA (and RNA indirectly), where the gg rotamer, rather than the gt rotamer, is observed in most native structures. The effects of two additional structural perturbations on 2 were also studied, namely, deoxygenation at C5 (yielding 2,5-dideoxy-beta-D-erythro-pentofuranose 4) and methyl glycosidation at O1 (yielding methyl 2-deoxy-beta-D-erythro-pentofuranoside 5) at the HF/6-31G level. The conformational energy profile of 4 resembles that found for 2gt, not 2gg, indicating that 4 is an inappropriate structural mimic of the furanose ring in DNA. Glycosidation failed to induce differential stabilization of ring conformations containing an axial C1-O1 bond (anomeric effect), contrary to experimental data. The latter discrepancy indicates that either the magnitude of this differential stabilization depends on ring configuration or that solvent effects, which are neglected in these calculations, play a role in promoting this stabilization.

13C NMR evidence of the failure of human erythrocytes to metabolize ascorbate and dehydroascorbate to lactate.

13C-NMR spectroscopy was used to record time courses of the metabolism of [1-(13)C]-L-ascorbic acid (AA) and [2-(13)C]-L-ascorbic acid and their dehydro-counterparts (DHAA) by human erythrocytes. Under a range of experimental conditions, but most notably in the absence of glucose in the incubation medium, no (13)C-NMR signal for lactate emerged during any of the 5 h time courses. The NMR resonances that did emerge over time were assigned to diketogulonic (DKG) acid and CO(2). Only very minor resonances from degradation products of DKG appeared from samples that contained physiologically high concentrations of DHAA. These results are in contrast with those in a recent report that lactate is derived from AA in human erythrocytes. However, an explanation for this possible artifact is given.

13C-labeled platinum(IV)-carbohydrate complexes: structure determination based on (1)H-(1)H, (13)C-(1)H, and (13)C-(13)C spin-spin coupling constants.

The reaction of D-mannose and D-allose with [PtMe(3)(Me(2)CO)(3)]BF(4) 1 in acetone affords complexes [PtMe(3)L]BF(4) 5 and 6 (5, L = alpha-D-mannofuranose; 6, L = beta-D-allofuranose). The coordination mode and conformation of the carbohydrate ligands in 5 and 6 in acetone-d(6) have been determined from an analysis of J(HH), J(CH), and J(CC) in complexes formed using site-specific (13)C-labeled D-mannose and D-allose. These coupling data are compared to those measured in (13)C-labeled complex [PtMe(3)L]BF(4) 2 (L = 1, 2-O-isopropylidene-alpha-D-glucofuranose) and 1, 2-O-isopropylidene-alpha-D-glucofuranose 3, whose solid-state structures are known, and in (13)C-labeled 1,2;5, 6-di-O-isopropylidene-alpha-D-glucofuranose 4. The preferred furanose ring conformations in 2 and 5 are very similar ((3)E/E(4) and E(4)/(o)E/E(1), respectively; eastern hemisphere of the pseudorotational itinerary), with platinum coordination involving O3, O5, and O6 of the saccharide. In contrast, the furanose ring of 6 prefers an (4)E/E(o)/(1)E geometry (western hemisphere of the pseudorotational itinerary) resulting from altered complexation involving O1, O5, and O6. Couplings within the exocyclic fragments of 2, 5, and 6 also support the existence of two different platinum coordination modes. In addition to establishing the structures and conformations of 2, 5, and 6 in solution, one-, two-, and three-bond J(CH) and J(CC) observed in these complexes provide new insights into the effect of structure and conformation on the magnitudes of these couplings in saccharides. Weak platinum(IV) complexation with the carbohydrate conformationally restricts the furanose and exocyclic fragment without introducing undesirable structural strain, thereby allowing more reliable correlations between structure and coupling magnitude.

Deuterium nuclear spin-lattice relaxation times and quadrupolar coupling constants in isotopically labeled saccharides.

(13)C and (2)H spin-lattice relaxation times have been determined by inversion recovery in a range of site-specific (13)C- and (2)H-labeled saccharides under identical solution conditions, and the data were used to calculate deuterium nuclear quadrupolar coupling constants ((2)H NQCC) at specific sites within cyclic and acyclic forms in solution. (13)C T(1) values ranged from approximately 0.6 to 8.2 s, and (2)H T(1) values ranged from approximately 79 to 450 ms, depending on molecular structure (0.4 M sugar in 5 mM EDTA (disodium salt) in (2)H(2)O-depleted H(2)O, pH 4. 8, 30 degrees C). In addition to providing new information on (13)C and (2)H relaxation behavior of saccharides in solution, the resulting (2)H1 NQCC values reveal a dependency on anomeric configuration within aldopyranose rings, whereas (2)H NQCC values at other ring sites appear less sensitive to configuration at C1. In contrast, (2)H NQCC values at both anomeric and nonanomeric sites within aldofuranose rings appear to be influenced by anomeric configuration. These experimental observations were confirmed by density functional theory (DFT) calculations of (2)H NQCC values in model aldopyranosyl and aldofuranosyl rings.

Catalytic mechanism and specificity for hydrolysis and transglycosylation reactions of cytosolic beta-glucosidase from guinea pig liver.

Cytosolic beta-glucosidase (CBG) from mammalian liver is known for its broad substrate specificity and has been implicated in the transformation of xenobiotic glycosides. CBG also catalyzes a variety of transglycosylation reactions, which have been been shown with other glycosylhydrolases to function in synthetic and genetic regulatory pathways. We investigated the catalytic mechanism, substrate specificity, and transglycosylation acceptor specificity of guinea pig liver CBG by several methods. These studies indicate that CBG employs a two-step catalytic mechanism with the formation of a covalent enzyme-sugar intermediate and that CBG will transfer sugar residues to primary hydroxyls and equatorial but not axial C-4 hydroxyls of aldopyranosyl sugars. Kinetic studies revealed that correction for transglycosylation reactions is necessary to derive correct kinetic parameters for CBG. Further analyses revealed that for aldopyranosyl substrates, the activation energy barrier is affected most by the presence of a C-6 carbon and by the configuration of the C-2 hydroxyl, whereas the binding energy is affected modestly by the configuration and substituents at C-2, C-4, and C-5. These data indicate that the transglycosylation activity of CBG derives from the formation of a covalently linked enzyme-sugar intermediate and that the specificity of CBG for transglycosylation reactions is different from its specificity for hydrolysis reactions.

13C NMR studies of vitamin C transport and its redox cycling in human erythrocytes.

13C NMR spectra of labeled [1-13C]- and [2-13C]ascorbic acid were seen to contain resonances arising from the intra- and extracellular populations in suspensions of human erythrocytes; i.e., they displayed the "split-peak" phenomenon. This new observation enabled the ready determination of the location, whether inside or outside cells, of the redox reactions in which the vitamin C was involved and to monitor the transport of the compounds into and out of the cells. Thus, the membrane permeability of ascorbic acid and the apparent Vmax and KM for the reduction of dehydroascorbic acid were determined in a noninvasive manner. In contrast to other work, evidence was found of a transporter of dehydroascorbic acid which is different from the glucose transporter. This transport system also appeared to be involved in the simultaneous reduction of dehydroascorbic acid on its passage into the cells. A second reduction process appeared to occur extracellularly, by the passage of reducing equivalents through the plasma membrane, as occurs with the reduction of ferricyanide. Evidence is presented that the processes of vitamin C recycling rely on different cellular sources of reducing equivalents. Whereas the transport and reduction via the membrane appeared to be dependent on glycolysis (NADH), the reduction of intracellular dehydroascorbic acid, formed in the process of transmembrane electron transfer or by transport from the outside of the cell, is currently thought to depend on NADPH.

13C-labeled oligodeoxyribonucleotides: a solution study of a CCAAT-containing sequence at the nuclear factor I recognition site of human adenovirus.

The solution behavior of the single-stranded CCAAT-containing octamer 1, d(AGCCAATA), that comprises part of the nuclear factor I (NF-I) recognition site at the origin of replication of human adenovirus has been studied by nmr spectroscopy at 500 and 600 MHz. Proton resonance assignments for 1 were aided by selective 13C enrichment at C1' of A1 or A5. High-resolution 13C-1H heteronuclear multiple-bond coherence spectra of the 13C-labeled oligomers permitted the selective detection of furanosyl ring protons within each labeled residue due to short- and long-range 13C-1H couplings to the enriched C1'. The resulting assignments provided firm starting points in the interpretation of double quantum filtered correlated spectra, yielding information supplemented by total correlated spectroscopy (TOCSY) and rotating frame nuclear Overhauser effect spectroscopic data to completely assign the 1H-nmr spectrum of 1 and extract 3JHH values for furanose conformational analysis. Several 13C-1H spin-coupling constants within the 13C-enriched A1 or A5 residues were measured from cross-peak shifts in TOCSY spectra, and their signs determined by inspection of the relative orientations of these shifts. 1H-1H and 13C-1H spin-couplings both indicate a preference (> 75%) for south (C2'-endo) conformations by the furanosyl rings of 1.

13C-labeled D-ribose: chemi-enzymic synthesis of various isotopomers.

Current interest in the use of heteronuclear multidimensional NMR methods to assess the structures, conformations and/or dynamics of oligonucleotides in solution has created an immediate need for nucleosides and their derivatives labeled in various ways with stable isotopes (13C, 2H, 15N and/or 17,18O). This short review focuses exclusively on chemienzymic methods to introduce one or more 13C labels into D-ribose, a precursor to ribo- and 2'-deoxyribonucleosides. It will be demonstrated that five convenient reactions, applied in specific sequences, provide access to 26 of the 32 13C-labeled isotopomers of D-ribose in acceptable yields. While not explicitly discussed herein, these same reactions, appropriately modified, can also be used to insert one or more 2H and/or 17,18O isotopes into this aldopentose.

19F and 13C NMR studies of polyol metabolism in freeze-tolerant pupae of Hyalophora cecropia.

Sorbitol biosynthesis and regulation in freeze tolerant pupae of Hyalophora cecropia have been investigated as a function of temperature by 19F and 13C nuclear magnetic resonance (NMR) spectroscopy using several 13C-labeled and/or fluorine-substituted carbohydrates. 3-Deoxy-3-fluoro-D-glucose (3DFG) was metabolized to 3-deoxy-3-fluoro-D-sorbitol (3DFS), 3-deoxy-3-fluoro-D-fructose (3DFF), and 3-deoxy-3-fluoro-D-gluconic acid (3DFGA), indicating that the enzymes required for sorbitol biosynthesis and metabolism are active in H. cecropia at warm (22 degrees C) and cold (4 and -10 degrees C) temperatures. Two additional metabolites were produced when pupae were injected with either 3DFG, 3DFS, 3DFF, or 3-deoxy-3-fluoro-D-mannose (3DFM). One of these was identified as 3-deoxy-3-fluoro-D-mannitol (3DFML) by 13C NMR using [1-13C]3DFM and [1-13C]3DFG as metabolic probes. H. cecropia pupae injected with D-glucose labeled with 13C at C-1, C-2, or C-3 and subsequently analyzed by 13C NMR clearly demonstrated the ability to generate sorbitol and fructose. In contrast, gas chromatography/mass spectrometric analysis of hemolymph failed to detect sorbitol in pupae reared under natural conditions (i.e. in the absence of injected enriched sugars). Thus, although H. cecropia pupae have the enzymic machinery to biosynthesize sorbitol, they do not appear to accumulate high steady-state concentrations of this polyol over the temperature range studied. The specificity of the enzymes involved in alditol biosynthesis in H. cecropia was examined by 13C NMR with a wide range of aldoses enriched with 13C at C-1. Pupae were capable of converting these sugars to their corresponding [1-13C]alditols, indicating that nonspecific dehydrogenase(s), in addition to aldose reductase, is(are) involved in polyol biosynthesis in H. cecropia pupae.

(13C)-substituted sucrose: 13C-1H and 13C-13C spin coupling constants to assess furanose ring and glycosidic bond conformations in aqueous solution.

Sucrose (beta-D-fructofuranosyl alpha-D-glucopyranoside, 1), methyl alpha-D-fructofuranoside (2), and methyl beta-D-fructofuranoside (3) have been prepared by chemical and/or enzymic methods with single sites of 13C-substitution at C-1, C-2, C-3, and C-6 of the fructofuranosyl ring. 1H (500 MHz) and 13C (75 and 125 MHz) NMR spectra of 1-3 have been obtained, yielding 1H-1H, 13C-1H, and 13C-13C spin coupling constants that were used to assess furanose ring and glycoside bond conformations in aqueous (2H2O) solution. Results show that the conformational mobility of the furanosyl ring in 3 is altered when incorporated into 1. Furthermore, 13C-13C and 13C-1H spin couplings across the glycosidic linkage suggest a psi torsion angle different from that observed in the crystal (phi appears similar). Interplay between the strength of the exoanomeric effect and hydrogen bonding in solution may be responsible, in part, for the apparent conformational flexibility of 1. In addition, spin couplings in 2 and 3 have been compared to those measured previously in alpha-D-threo-pentulofuranose (4) and beta-D-threo-pentulofuranose (5), respectively, as a means to study the effect of glycosidation and hydroxymethyl substitution on the solution conformation of the 2-ketofuranose ring. The conversion of 4 to 2 is accompanied by minimal conformational change, whereas a significant change accompanies the conversion of 5 to 3, showing that the effect of substitution on ring conformation depends highly on ring configuration before and after substitution.

Isotope-edited 1D and 2D n.m.r. spectroscopy of 13C-substituted carbohydrates.

Three isotope-edited n.m.r. methods have been applied to selectively 13C-substituted monosaccharides and nucleosides to simplify their spectra and/or measure 1H-1H, 13C-1H, or 13H-13C spin-couplings detected via the labeled site. 1D INADEQUATE spectra allowed the selective detection of the natural-abundance carbons that are spin-coupled to the labeled carbon, and adjustment of the mixing time permitted further discrimination between one-bond and longer-range 13C-13C coupling pathways. Geminal and vicinal 13C-1H coupling constants were determined from the analysis of 1H-1H COSY cross-peaks for those protons coupled to the labeled carbon. Long-range 13C-(HETCOR) and 1H-detected (HMBC) 13C-1H chemical-shift correlation spectra permitted the selective observation of those protons coupled to the labeled site, and JH,H values were measured from data projections. The implications of these methods for structural studies of more complex systems is briefly discussed.

Multiply 13C-substituted monosaccharides: synthesis of D-(1,5,6-13C3)glucose and D-(2,5,6-13C3)glucose.

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.

Ring-opening kinetics of the D-pentofuranuronic acids.

The ring-opening reactions of the furanose forms of the penturonic acids D-arabinuronic acid (1), D-lyxuronic acid (2), D-riburonic acid (3), and D-xyluronic acid (4) in aqueous solution have been studied as a function of temperature and solution pH by 13C saturation-transfer n.m.r. (s.t.-n.m.r.) spectroscopy using 1-13C-substituted compounds. Unidirectional rate constants of ring-opening (kopen) have been determined for the cyclic forms of 1-4 in their protonated (pH 1.5) and ionized (pH 4.5) forms, and have been compared to the k-values measured previously for structurally related furanose sugars. At 50 degrees and pH 1.5, kopen values decrease as follows: alpha-xyluronic acid (2.57 s-1) greater than alpha-riburonic (1.65 s-1) greater than beta-arabinuronic (1.52 s-1) greater than beta-xyluronic (1.09 s-1) greater than beta-riburonic (0.76 s-1) greater than beta-lyxuronic (0.55 s-1) greater than alpha-arabinuronic (0.46 s-1) greater than alpha-lyxuronic (0.40 s-1). At 50 degrees and pH 4.5, this order changes significantly (e.g., beta-arabinuronate is most reactive); in general kopen values for beta anomers appear to be enhanced relative to those for corresponding alpha anomers, suggesting the involvement of intramolecular catalysis in which the carboxylate anion assists in abstracting the hydroxyl proton from O-1. Activation energies of ring-opening, determined for the alpha and beta anomers of 1-4, were found to depend on ring configuration and solution pH.

Deoxygenated and alkylated furanoses: Thorpe-Ingold effects on tautomeric equilibria and rates of anomerization.

2-Deoxy-D-glycero-tetrose, 3-deoxy-DL-glycero-tetrose, 3-deoxy-3,3-di-C-methyl-DL-glycero-tetrose, 3-C-methyl-DL-erythrose, 3-C-methyl-DL-threose, 2-deoxy-5-O-methyl-D-erythro-pentose and 3-deoxy-5-O-methyl-D-erythro-pentose have been prepared, in some cases with 13C-substitution at the anomeric carbon, and characterized by 1H-(300 and 620 MHz) and 13C-n.m.r. (75 MHz) spectroscopy. The proportions of cyclic (alpha and beta furanoses) and acyclic (aldehyde and hydrate) forms were determined in aqueous (2H2O) solution, and ring-opening (kopen) and ring-closing (kclose) rate constants were measured by 1H and 13C saturation-transfer n.m.r. spectroscopy at p2H 5.0 (acetate buffer) and 60 degrees. The degree of furanose ring substitution was found to significantly affect both the thermodynamics and kinetics of furanose anomerization. Increased substitution enhances the proportion of cyclic forms in solution by stimulating furanose kclose. In contrast, furanose kopen was less affected by the degree of substitution; however, kinetic studies of 2-deoxyfuranose anomerization implicate furanose ring conformation as a potential determinant of kopen.

D-Penturonic acids: solution studies of stable-isotopically enriched compounds by 1H- and 13C-n.m.r. spectroscopy.

Methyl D-pentofuranosides were prepared by Fischer glycosidation of the aldopentoses D-arabinose, D-lyxose, D-ribose, D-xylose, and 2-deoxy-D-erythro-pentose, and oxidized with O2 over a platinum oxide catalyst to give the corresponding methyl D-pentofuranosiduronic acids. After purification by anion-exchange chromatography, these glycosides were hydrolyzed to give the corresponding D-penturonic acids [D-arabinuronic acid (1), D-lyxuronic acid (2), D-riburonic acid (3), D-xyluronic acid (4), and 2-deoxy-D-erythro-penturonic acid (5)] in 80% yield based on the starting pentofuranoside. 1-13C-Substituted D-aldopentoses were used to prepare D-(1-13C)penturonic acids. Aqueous solutions of the 1-13C-substituted penturonic acids, studied over a range of pH values by 13C-n.m.r. spectroscopy, were found to contain alpha- and beta-furanoses, acyclic aldehyde and hydrate, and/or hydrated 2,5-lactone. The ratio of D-riburonic acid anomers was most sensitive to solution pH (alpha/beta = 0.49 and 1.2 at pH 1.9 and 4.9, respectively). The values of the 1H and 13C chemical shifts, and 1H-1H. 13C-1H, and 13C-13C spin-coupling constants, were determined by 1H-(300, 500, and 620 MHz) and 13C-(75 MHz) n.m.r. spectroscopy with the aid of 2-D 13C-1H chemical shift correlation maps, 2-D 1H-1H COSY data, and 13C substitution, and were compared to those determined previously for structurally-related furanose rings. Isomerization of the penturonic acids at pH 5.0 and 50 degrees gave the corresponding 4-pentulosonic acids.

Synthesis of D-erythro-2-pentulose and D-threo-2-pentulose and analysis of the 13C- and 1H-n.m.r. spectra of the 1-13C- and 2-13C-substituted sugars.

The pentuloses D-erythro-2-pentulose (1) and D-threo-2-pentulose (2) and their 1-13C- and 2-13C-substituted derivatives were prepared by hydrogenating the corresponding isotopically normal and 13C-substituted D-pentos-2-uloses with a Pd-carbon catalyst. The threo isomer and its labeled derivatives were alternatively prepared from isotopically normal and 13C-substituted D-xyloses with immobilized D-xylose (D-glucose) isomerase (E.C.5.3.1.5). The equilibrium compositions of 1 and 2 (furanose anomers and acyclic keto forms) in 2H2O were determined from 13C-n.m.r. spectra (75 MHz) of the 2-13C-labeled derivatives. The conformational properties of the cyclic and acyclic forms in 2H2O were assessed with the use of 1H-1H, 13C-1H, and 13C-13C spin-coupling constants obtained from 1H-n.m.r. (620 MHz) and 13C-n.m.r. (75 MHz) spectra. Compared with the structurally related aldotetrofuranoses the 2-pentulofuranoses more strongly prefer conformations in which the anomeric hydroxyl group is oriented quasi-axially. The strongly dipolarized carbonyl group in the acyclic keto forms of 1 and 2 appears to stabilize chain conformations having O-1 and O-3 eclipsed with the carbonyl oxygen.

(1-13C)alditols: elimination of magnetic equivalence in 1H- and 13C-n.m.r. spectra of symmetric compounds through (13C)-substitution.

(1-13C)Glycerol, D-(1-13C)arabinitol, D-(1-13C)ribitol, D-(1-13C)xylitol, D-(1-13C)glucitol, D-(1-13C)mannitol, and D-(1-13C)talitol have been prepared by NaBH4 reduction of the corresponding (1-13C)aldoses. A comparison of the 1H- (300 and 620 MHz) and 13C (75 MHz) n.m.r. spectra of natural and (1-13C)-substituted dissymmetric alditols has permitted the unequivocal assignments of their hydroxymethyl proton and carbon signals and the measurement of several 13C-1H and 13C-13C spin-coupling constants. Similar spectra of (1-13C)-substituted symmetric alditols, however, are more difficult to interpret since they are composed of overlapping 13C-coupled and 13C-noncoupled subspectra. In some cases, 1H difference spectra and 1H-coupled 13C spectra may be used to extract the 13C-1H and 13C-13C spin couplings from the 13C-coupled component. These couplings have been examined in light of conformational models previously proposed, permitting a preliminary evaluation of standard 3JCH and 3JCC values for specific coupling pathways in these compounds.