13C nmr studies of aurothioglucose: Ligand exchange and redox disproportionation reactions

¹³C nmr studies of gold thioglucose, AuSTg, and solutions containing added β-1-D-thioglucose, TgSH, have been conducted at PD 7.4 and interpreted in terms of complexation and ligand exchange reactions that are consistent with the known preference of gold(I) for linear two-coordinate structures. The upper limit of the half-life for ligand exchange between 0.25 M Au(STg)₂^? and TgSH at pD 7.4 is 2.2 msec. The ¹³C nmr spectra of various thioglucose derivatives have been assigned. A novel oxidation reduction reaction was discovered that leads to the formation of metallic gold and a product tentatively identified as the sulfinic acid derivative of thioglucose. The presence of sulfinic acid in AuSTg was indicated by the infrared absorption at 1050 cm^?1. The same product was formed by slow hydrolysis of thioglucose disulfide. A mechanism for the formation of the sulfinic acid derivative from AuSTg is proposed.     

Metabolism of progesterone and testosterone by a Bacillus sp.

Microbial transformations by a Bacillus sp. were employed as a means of preparing potentially important derivatives of progesterone and testosterone. Each microbial metabolite was subjected to structure elucidation employing ¹H and ¹³C nmr, mass spectral and cd analysis. Hplc was used for the determination of the percentages of the metabolites formed. The progesterone metabolites were characterised as 14-hydroxy-4-pregnene-3,20-dione (II), 14-hydroxy-5 α -pregnane-3,6,20-trione (III)., 11 α — hydroxy-5 α — pregnane-3, 6,20-trione (IV) and 11 α, 14-dihydroxy-4-pregnene-3,20-dione (V). The testosterone analogs were identified as 4-androstene-3,17-dione (VII), 17 β-hydroxy-5 α -androstene-3,6-dione (VIII), 14-hydroxy-4-androstene-3,17-dione (IX) and 14, 17 β-dihydroxy-4-androsten -3-one (X)₁. The availability of the metabolites enabled complete elucidation of their ¹³C nmr spectra.     

The structure of the antitumor complex cis-(diammino) (1,1-cyclobutanedicarboxylato)-Pt(II): X ray and nmr studies

X-ray crystallography shows that Pt(NH₃)₂(CBDCA) is a square-planar complex with the dicarboxylate chelate ring in the boat conformation and a planar cyclobutane ring. ¹H and ¹³C nmr studies suggest that rapid chelate ring flipping occurs in solution. The value of ¹⁹⁵Pt nmr combined with ¹⁵N labeling as an informative new method of studying carboxylate coordination is illustrated. nmr results are also reported for the analogous ethylmalonate complex.     

Comparison of the effects of calcium and magnesium on the conformation of tetra cycline in Me2SO solution

The effects of anhydrous Ca(NO₃)₂ on the ¹³C nuclear magnetic resonance (nmr) and circular dichroism spectra of tetracycline in Me₂SO-d₆ solution have been investigated in order to make a comparison with the results of a previous study in which the effects of Mg(NO₃)₂ were determined under the same conditions. The results of experiments described in this article provide strong evidence that Ca²⁺ and Mg²⁺ bind tetracycline at the same sites in Me₂SO and that both ions induce the same change in molecular conformation of tetracycline upon binding. The Ca²⁺ complex, in contrast to the Mg²⁺ complex, has a lifetime that is short on the nmr time scale.     

Properties of metal-ion coordinated imidazoles: nmr and C-2 H Exchange in Co(III) Complexes

The ¹H and ¹³C nmr spectra of Co(NH₃)₅ImH³⁺ and the ¹H nmr spectra of αCotrien(ImH)₂³⁺ and βCotrien(ImH)₂³⁺ are reported. The pK_a values determined from the dependence of the chemical shift on pH are 10.0, 9.6, and 10.1, respectively. The range of the chemical shift between the acid and base forms is unusually small in the ¹H nmr, 0.5–0.7 ppm for the C-2 H and about 0.25 ppm for the C-4 H and C-5 H. In the ¹³C nmr, C-2 and C-4 have large shifts to low field and C-5 a small shift to high field on deprotonation. The C-2 proton is not exchanged with solvent ²H under acidic or basic conditions, in marked contrast to the corresponding proton in both imidazole and 1-methylimidazole. These spectroscopic and chemical properties should be useful for the direct identification of metal-ion coordinated histidines in proteins.     

Investigation of the binding of inorganic complexes to blue copper proteins by 1H nmr spectroscopy,I. The interaction between the [Cr(phen)3]3+ and [Cr(CN)6]3− ions and the copper(I) form of parsley plastocyanin

The effects of the addition of [Cr(phen)₃](ClO₄)₃ and K₃[Cr(CN)₆] on the ¹H nmr spectrum of the copper(I) form of parsley plastocyanin are described. It is concluded that the ions [Cr(phen)₃]³⁺ and [Cr(CN)₆]^3? bind to different parts of the protein.     

1H-1H and 13C-13C vicinal coupling constants and amino acid side chain conformation in peptides

The vicinal coupling constants ¹³C′-¹³C^γ were measured in aspartic acid and phenylalanine (85 % ¹³C enrichment) as free amino acids and in the peptides Asp-Pro and Gly-Pro-Phe. These coupling constants used in connection with those between the α -and the β-protons provide the unambiguous assignment of rotamers I and II in the Asp and Phe side chains. The method is generally applicable to other amino acids and residues even in large peptides. A possible set of J_g^c,c and J_t^c,c values is proposed for the use of carbon 13-carbon 13 vicinal coupling constants in the side chain conformational studies of amino acid residues with a free carboxyl group.     

Complexes of binucleating ligands. XVII. Some 3-atom N,C bridged palladium(II) complexes

The acetate-bridged complex, LPd₂(CH₃CO₂), in which L^3? is a binucleating ligand, reacts with 2-vinylpyridine in the presence of methanol or ethanol to generate the 3 atom N,C bridged complexes LPd₂(2-C₅H₄N·CH·CH₂OR) (R = Me or Et) whose ¹H and ¹³C nmr spectra indicate the presence in solution of two slowly interconverting forms at room temperature. The ¹H and ¹³C nmr spectra of two closely related pairs of 3 atom N,C bridged complexes of the form LPd₂(2-C₅H₄N·CH·X) and LPd₂ (HN = C(CH₃)·CH·X) (where X = COCH₃ or COOCH₃) show that the complexes with pyridine-containing bridges exist in solution at room temperature in two distinguishable forms whilst the corresponding imine-bridged complexes behave as single species. The existence of two forms of the complexes with pyridine-containing 3 atom N,C bridges, the natures of which are discussed in this paper, appears to be a consequence of steric interaction between the pyridine α hydrogen atom and the closely adjacent oxygen donor of L.     

Determination of the mode of interaction of Mn2+ and Cu2+ with cis-inositol by 13C NMR spectroscopy

Natural abundance ¹³C nuclear magnetic resonance spectroscopy (¹³C NMR) was used to study the mode of binding of Mn²⁺ and Cu²⁺ to the cyclitol, cis-inositol. Resonance linewidths and the electron nuclear relaxation rates [(T₁^e)^?1 values] were used to establish that a unique binding site exists for these metal-ions on this cyclitol involving only the three axial hydroxyl groups. This work may aid in the development of new organometallic complexes used as paramagnetic relaxation agents in magnetic resonance imaging research.     

An NMR study of the loss of carbon-20 in the biosynthesis of gibberellin A3 by Gibberella fujikuroi

Resuspension cultures of Gibberella fujikuroi, strain GF-1a, were shown to metabolise potassium [3′-¹³C] mevalonate to ¹³C-enriched C₁₉-gibberellins, plus ¹³CO₂ (derived from the loss of carbon-20). The formation of [¹³C]-gibberellins could be observed in vivo using ¹³C NMR; however that of ¹³CO₂ could not. In contrast, removal of the mycelium and concentration of the filtrate at pH 12 enabled the ¹³CO₂ produced to be observed using ¹³C NMR. During incubations of H¹⁴CO₂Na with this fungus, complete conversion to other radioactive products was observed, and the significance of these results in the light of previous work is discussed.     

A 31P nuclear magnetic resonance and fluorescence study of the interaction of an anti-arthritic gold phosphine drug with albumin. A bioinorganic approach

¹H decoupled ³¹P nmr spectra were recorded for a series of gold complexes of formulae PEt₃AuL and [PEt₃AuL′]⁺ClO₄^? where L and L′ are ligands containing biologically relevant donor atoms. This series of a model compounds provide a ³¹P nmr scale for the interaction of the ‘PEt₃Au’ moiety with proteins. The reactions of albumin and SH blocked albumin with PEt₃AuCl were monitored by ³¹P nmr spectroscopy. Comparison of the observed chemical shifts to those of the model compounds revealed preferential binding of gold to S occurs. Fluorescence studies of the gold-protein interactions imply that a protein conformational charge occurs on binding of gold. The implications of these studies on the mechanism of action of anti-arthritic gold drugs is discussed.     

Different forms of the cobalt-bleomycin A2 complex

Chromatographic analysis of Co-bleomycin on CM Sephadex C25 or silica shows that known components like bleomycin A₂, B₂, and A₁ can be separated into two forms. We observed an equilibrium between both forms and saw that the addition of certain salts influences the transition of one form into the other. A reaction scheme is suggested in which a different occupation of the sixth binding site of cobalt is correlated with the different forms observed. These different complexes may be an intramolecular adduct of the carbamoyl moiety of bleomycin, an external carboxylate adduct or an oxygen or hydroxyl adduct.     

Preparations and properties of tripodal and linear tetradentate N,S-Donor ligands and their complexes containing the MoO22+core

New linear and tripodal tetradentate ligands, LH₂, are reported and their syntheses are described. The new linear ligands L = HSCH₂CH₂SCH₂CH₂NRCH₂CR₂SH, R = H, CH₃) and the new tripodal ligands N(CH₂CH₂SH)₂CH₂Z, Z = CH₂NH₂, CH₂N(CH₃)₂, CH₂N(C₂H₅)₂, CH₂SCH₃ and CO₂^- were synthesized. The known linear ligands HSCH₂CH₂NCH₃(CH₂)_nNCH₃CH₂CH₂SH (n = 2, 3) and HSCR₂CH₂NHCH₂CH₂NHCH₂CR₂SH (R = H, CH₃) were also utilized. These ligands react with MoO₂(acac)₂ in CH₃OH to yield MoO₂L complexes in high yield. Infra-red and ¹H nmr spectra provide evidence to supplement X-ray crystallographic results reported elsewhere for selected numbers of the series. Octahedral structures with cis MoO₂²⁺ groupings are assigned. Solution ¹H nmr studies are consistent with a trans placement of the two thiolate donors in agreement with the X-ray studies.     

Native gibberellins and the metabolism of [14C]gibberellin A53 and of [17-13C, 17-3H2]gibberellin A20 in tassels of Zea mays

The native hormones from tassels of maize (Zea mays) were re-investigated. The previous identification by GC/SIM of GA₁, GA₈ and GA₂₉ in normal tassels was confirmed by full GC/MS scans at the correct Kovats retention indices. In tassels of dwarf-1 mutants, GA₄₄,^?GA₁₉, GA₁₇, GA₂₀ and the 16,17-dihydro, 7β,16α,17-trihydroxy derivative of ent-kaurenoic acid were identified by GC/MS. Gibberellin A₁ was not found in the mutant tassels. [¹⁴C]Gibberellin A₅₃ was fed to tassels of the dwarf-5 mutant. In the ethyl acetate-soluble acidic fraction from the feeds, [¹⁴C]GA₄₄ was identified by GC/MS; [¹⁴C]GA₁₉ and [¹⁴C]GA₂₉ were identified by GC/SIM. The GA₂₉ is probably a metabolite of the feeds because the dwarf-5 mutant is known to control the step copalyl pyrophosphate to ent-kaurene in the maize GA-biosynthetic pathway and because GA₂₉ was not identified in a control experiment. The n-butanol fractions obtained from the feeds were shown, by GC/MS, to contain [¹⁴C]GA₅₃ after hydrolysis, suggesting that conjugated [¹⁴C]GA₅₃ is a major metabolite from GA₅₃ feeds. [17-¹³C, 17-³H₂]Gibberellin A₂₀ was fed to normal, dwarf-1 and dwarf-5 tassels. In each case, analysis of the purified ethyl acetate-soluble acidic extracts by GC/MS led to the identification of [¹³C]GA₂₉ and unmetabolized [¹³C]GA₂₀ in which no ¹³C-isotope dilution was observed.     

Photosynthesis and the influence of CO2-enrichment on δ13C values in a C3 halophyte

Abstract Shifts in ?¹³C of the graminaceous C₃ halophyte Puccinellia nuttalliana (Schultes) Hitch. can be induced by salinization. To investigate this phenomenon, three approaches were taken: assay of carboxylases, CO₂-enrichment studies, and gas exchange analysis. Although ribulose-1,5-bisphosphate carboxylase activity decreased with salinity, phosphoenolpyruvate carboxylase activity did not increase and its levels were not atypical of C₃ plants. When plants were grown at four NaCl concentrations under atmospheres of 310 and 1300 cm³ m^?3 CO₂, the CO₂-enrichment enhanced the effects of salinity on ?¹³C. This is consistent with a biophysical explanation for salt-induced shifts in ?¹³C, whereby there is a steepening of the CO₂ diffusion gradient into the leaf. Gas exchange analysis indicated that intercellular CO₂ concentrations were depressed in the leaves of salt-affected plants. This resulted from a greatly decreased stomatal conductance coupled with only small effects on intrinsic photosynthetic capacity. Water-use efficiency was enhanced.     

Carbon-13 NMR studies of C2H5N13C binding to various hemoproteins

The addition of an excess of C₂H₅N¹³C to myoglobin and human adult and fetal hemoglobins, gives three characteristic NMR spectra with new ¹³C resonances respectively at δ = ?10,56 ppm, δ = ?7,03 and ?7,95 ppm and δ = ?6,28 and ?7,95 ppm (CH₃CO₂Na as external standard). These signals correspond to the C₂H₅N¹³C bound to the Fe(II) of the different heme units, according to CO exchange experiments. Characteristic resonances can be assigned to C₂H₅N¹³C bound to α, β and γ subunits. C₂H₅N¹³C appears as a more sensitive probe than ¹³CO for hemoprotein NMR studies.     

Hysteretic kinetic behavior of beef liver pyruvate carboxylase.

Natural abundance ¹³C nuclear magnetic resonance (nmr) spectra have been obtained for samples of a variety of native collagens by use of cross-polarization (CP) techniques which permit high resolution natural abundance ¹³C nmr spectra of solids to be obtained with high sensitivity. The CP ¹³C nmr spectra of lyophilized skin and tendon collagens consisted of two broad resonance envelopes spanning a five kHz range. Hydrated tendon collagen gave rise to a CP spectrum very similar to that obtained for the lyophilized sample, indicating that it retains its solid-like properties. In contrast, hydrated skin collagen became denatured under the conditions of the CP experiment and subsequently gave rise to a conventional high-resolution Fourier transform (FT) nmr spectrum. The CP ¹³C nmr spectrum of ivory was similar to those of lyophilized skin and tendon collagens, demonstrating the solid-like character of the collagen in dentine, whereas the CP spectrum of bovine nasal cartilage reflected the presence of highly mobile proteoglycan components in addition to relatively rigid collagen molecules. In the case of ivory, the resolution of the CP spectrum was enhanced by “magic angle” spinning to a degree approaching that of conventional FT ¹³C nmr spectra of denatured collagen in solution. Because of its ability to probe the dynamic properties of solid-like biological molecules, CP ¹³C nmr spectroscopy should be a valuable investigative tool for future studies.     

NMR studies on UO22+ complexes with pyridoxal

The interaction of pyridoxal with dioxouranium(VI) acetate was studied by ¹H and ¹³C NMR measurements in D₂O and CD₃OD.The results indicate that the preferred bonding site is the C-3O^? donor, and the major species obtained under the experimental conditions used is the equimolar complex.     

Concerning the biosynthesis of prostaglandin I2

Two diastereoisomers, 5R,6R-5-hydroxy-6(9α)-oxido-11α,15S-dihydroxyprost-13-enoic acid (7) and 5S,6S-5-hydroxy-6(9α)-oxido-11α,15S-dihydroxyprost-13-enoic acid (10) were synthesized for evaluation as possible biosynthetic intermediates in the enzymatic transformation of PGH₂ or PGG₂ into PGI₂. The synthetic sequence entails the stereospecific reduction of the 9-keto function in PGE₂ methyl ester after protecting the C-11 and C-15 hydroxyls as tbutyldimethylsilyl ethers. The resulting PGF_2α derivative was epoxidized exclusively at the C-5 (6) double bond to yield a mixture of epoxides, which underwent facile rearrangement with SiO₂ to yield the 5S,6S and 5R,6R-5-hydroxy-6(9α)-oxido cyclic ethers. It was found that dog aortic microsomes were unable to transform radioactive 9β-5S,6S[³H] or 9β-5R,6R[³H]-5-hydroxy-6(9α)-oxido cyclic ethers into PGI₂. Also, when either diastereoisomer was included in the incubation mixture, neither isomer diluted the conversion of [1-¹⁴C]arachidonic acid into [1-¹⁴C]PGI₂.     

Acquisition and within-plant allocation of 13C and 15N in CO2-enriched Quercus robur plants

We assessed the effects of doubling atmospheric CO₂ concentration, [CO₂], on C and N allocation within pedunculate oak plants (Quercus robur L.) grown in containers under optimal water supply. A short-term dual ¹³CO₂ and ¹⁵NO₃^? labelling experiment was carried out when the plants had formed their third growing flush. The 22-week exposure to 700 μl l^?1 [CO₂] stimulated plant growth and biomass accumulation (+53% as compared with the 350 μl l^?1 [CO₂] treatment) but decreased the root/shoot biomass ratio (-23%) and specific leaf area (-18%). Moreover, there was an increase in net CO₂ assimilation rate (+37% on a leaf dry weight basis; +71% on a leaf area basis), and a decrease in both above- and below-ground CO₂ respiration rates (-32 and -26%, respectively, on a dry mass basis) under elevated [CO₂]. ¹³C acquisition, expressed on a plant mass basis or on a plant leaf area basis, was also markedly stimulated under elevated [CO₂] both after the 12-h ¹³CO₂ pulse phase and after the 60-h chase phase. Plant N content was increased under elevated CO₂ (+36%), but not enough to compensate for the increase in plant C content (+53%). Thus, the plant C/N ratio was increased (+13%) and plant N concentration was decreased (-11%). There was no effect of elevated [CO₂] on fine root-specific ¹⁵N uptake (amount of recently assimilated ¹⁵N per unit fine root dry mass), suggesting that modifications of plant N pools were merely linked to root size and not to root function. N concentration was decreased in the leaves of the first and second growing flushes and in the coarse roots, whereas it was unaffected by [CO₂] in the stem and in the actively growing organs (fine roots and leaves of the third growth flush). Furthermore, leaf N content per unit area was unaffected by [CO₂]. These results are consistent with the short-term optimization of N distribution within the plants with respect to growth and photosynthesis. Such an optimization might be achieved at the expense of the N pools in storage compartments (coarse roots, leaves of the first and second growth flushes). After the 60-h ¹³C chase phase, leaves of the first and second growth flushes were almost completely depleted in recent ¹³C under ambient [CO₂], whereas these leaves retained important amounts of recently assimilated ¹³C (carbohydrate reserves?) under elevated [CO₂].