Heteronuclear Correlation NMR Experiments Research Articles

Two-dimensional 13C– 1H heteronuclear correlation NMR spectroscopic studies for the inclusion complex of cyclomaltoheptaose (β-cyclodextrin) with a new Helicobacter pylori eradicating agent (TG44) in the amorphous state

The interaction of a newly developed Helicobacter pylori eradicating agent (TG44, 4-methylbenzyl-4′-[ trans-4-(guanidinomethyl)cyclohexylcarbonyloxy]-biphenyl-4-carboxlylate monohydrochloride) with cyclomaltoheptaose (β-cyclodextrin, β-CyD) in the solid state was studied by high-speed frequency-switched Lee-Goldburg (FSLG) 13C– 1H heteronuclear correlation (HETCOR) NMR experiments. The TG44/β-CyD solid complex in a 1:1 stoichiometry was prepared by the grinding method. Powder X-ray diffractometry confirmed that the complex is in an amorphous state. The solid-state 13C signals of TG44 and β-CyD were significantly broadened by the complexation. As the temperature increased, the 13C signals of the aromatic moieties of TG44 were insignificantly influenced, whereas those of the cyclohexyl moiety became sharper. The T 1 ρ H values of the aromatic moieties of TG44 were almost the same as those of the β-CyD carbons, whereas those of other TG44 carbons gave much smaller values. The 13C– 1H HETCOR spectra gave the intermolecular correlation peaks between the aromatic carbons of TG44 and the β-CyD protons or between the biphenyl protons of TG44 and the β-CyD carbons, when measured using longer contact times (500 and 1500 μs). On the basis of these solid NMR spectroscopic data together with aqueous NMR data, we assume that β-CyD includes predominantly the biphenyl moiety of TG44 in the solid state. 13C– 1H HETCOR spectroscopy is particularly useful for the determination of inclusion modes of the complexes that occurring in an amorphous form.

SPAM-MQ-HETCOR: an improved method for heteronuclear correlation spectroscopy between quadrupolar and spin-1/2 nuclei in solid-state NMR

The recently introduced concept of soft pulse added mixing (SPAM) is used in two-dimensional heteronuclear correlation (HETCOR) NMR experiments between half-integer quadrupolar and spin-1/2 nuclei. The experiments employ multiple quantum magic angle spinning (MQMAS) to remove the second order quadrupolar broadening and cross polarization (CP) or refocused INEPT for magnetization transfer. By using previously unexploited coherence pathways, the efficiency of SPAM-MQ-HETCOR NMR is increased by a factor of almost two without additional optimization. The sensitivity gain is demonstrated on a test sample, AlPO(4)-14, using CP and INEPT to correlate (27)Al and (31)P nuclei. SPAM-3Q-HETCOR is then applied to generate (27)Al-(31)P spectra of the devitrified 41Na(2)O-20.5Al(2)O(3)-38.5P(2)O(5) glass and the silicoaluminophosphate ECR-40. Finally, the method allowed the acquisition of the first high resolution solid-state correlation spectra between (27)Al and (29)Si.

Activity of lupane triterpenoids from Maytenus species as inhibitors of nitric oxide and prostaglandin E 2

In the present study, we report that three new lupane triterpenes ( 1– 3), in addition to 16 known ones ( 4– 19), were isolated from the root bark of Maytenus cuzcoina and the leaves of Maytenus chiapensis. Their structures were elucidated by spectral analysis, including homonuclear and heteronuclear correlation NMR experiments (COSY, ROESY, HSQC, and HMBC). The natural compounds and derivatives 6a, 6b, 9a, and 9b have been tested for potential anti-inflammatory activity, and several compounds including 3- epicalenduladiol ( 2), 11α-hydroxy-glochidone ( 3), rigidenol ( 6), acetoxy-rigidenol ( 6a), 11α-acetoxy-30-chloro-3-oxo-lup-20(29)-ene ( 6b), betulin ( 9), 28-acetoxy-betulin ( 9a), epibetulin ( 12), epibetulinic acid ( 13), and betulonic acid ( 16) exhibited potent inhibitory effects on NO and prostaglandin E 2 production in mouse macrophages (RAW 264.7) stimulated with bacterial endotoxin. The structure–activity relationship is discussed in detail.

Toward direct determination of conformations of protein building units from multidimensional NMR experiments VI. Chemical shift analysis of his to gain 3D structure and protonation state information

NMR--chemical shift structure correlations were investigated by using GIAO-RB3LYP/6-311++G(2d,2p) formalism. Geometries and chemical shifts (CSI values) of 103 different conformers of N'-formyl-L-histidinamide were determined including both neutral and charged protonation forms. Correlations between amino acid torsional angle values and chemical shifts were investigated for the first time for an aromatic and polar amino acid residue whose side chain may carry different charges. Linear correlation coefficients of a significant level were determined between chemical shifts and dihedral angles for CSI[(1)H(alpha)]/phi, CSI[(13)C(alpha)]/phi, and CSI[(13)C(alpha)]/psi. Protonation of the imidazole ring induces the upfield shift of CSI[(13)C(alpha)] for positively charged histidines and an opposite effect for the negative residue. We investigated the correspondence of theoretical and experimental (13)C(alpha), (13)C(beta), and (1)H(alpha) chemical shifts and the nine basic conformational building units characteristic for proteins. These three chemical shift values allow the identification of conformational building units at 80% accuracy. These results enable the prediction of additional regular secondary structural elements (e.g., polyProlineII, inverse gamma-turns) and loops beyond the assignment of chemical shifts to alpha-helices and beta-pleated sheets. Moreover, the location of the His residue can be further specified in a beta-sheet. It is possible to determine whether the appropriate residue is located at the middle or in a first/last beta-strand within a beta-sheet based on calculated CSI values. Thus, the attractive idea of establishing local residue specific backbone folding parameters in peptides and proteins by employing chemical shift information (e.g., (1)H(alpha) and (13)C(alpha)) obtained from selected heteronuclear correlation NMR experiments (e.g., 2D-HSQC) is reinforced.

Lupane Triterpenoids from Maytenus Species

Five new lupane triterpenes (1-5), in addition to 24 known ones, were isolated from Maytenus cuzcoina and M. chiapensis. Their structures were elucidated on the basis of spectroscopic analysis, including homonuclear and heteronuclear correlation NMR (COSY, ROESY, HSQC, and HMBC) experiments. The compounds were assayed for antimicrobial and cytotoxic activities, with 3-epi-betulinic acid and 28,30-dihydroxy-3-oxolup-20(29)-ene showing moderate cytotoxicity.

Cheiloclines A—I. First Examples of Octacyclic Sesquiterpene‐Triterpene Hetero‐Diels—Alder Adducts.

AbstractFor Abstract see ChemInform Abstract in Full Text.

Organic Coprecipitates with Calcite: NMR Spectroscopic Evidence

Dissolved organic ligands are well known to interact strongly with the calcite surface, altering precipitation and dissolution rates, crystal morphology, and possibly the ability of calcite to sequester metal contaminants. We show, using NMR spectroscopic techniques, that some of the citrate molecules present in a solution of precipitating calcite are incorporated structurally into the calcite crystal. Calcite grown by a seeded constant-addition method contains approximately 1 wt % coprecipitated citrate and yields 13C{1H} cross-polarization magic-angle spinning NMR spectra that contain broad peaks for citrate plus a signal from carbonate. Results from 13C{1H} heteronuclear correlation NMR experiments show that citrate is located in close spatial proximity to carbonate groups. In addition, calcite/citrate coprecipitates contain about 0.4 wt % excess water, which is not present as fluid inclusions, and some of which occurs as rigid structural water. These results suggestthat water and hydrogen-bonding interactions play a role in the interface between included organic molecules and the calcite host. Additional NMR data obtained for calcite coprecipitates of aspartic and glutamic acids suggest they are also incorporated structurally but at concentrations much lower than for citrate, whereas no evidence was found for phthalate incorporation.

Triterpenoids from Hippocratea excelsa. The Crystal Structure of 29-Hydroxytaraxerol

The root bark of Hippocratea excelsa afforded a new derivative of β -amyrin, which was identified as its ferulate, together with components new in this species. They were identified as the rare 29-hydroxytaraxerol, 29-hydroxyglutinol, 29-hydroxyfriedelin and the sterol 6β -hydroxystigmast-4- en-3-one. The known triterpene quinone methides pristimerin and tingenone characteristics of this genus, β -sitosterol, trans-polyisoprene, squalene, β -amyrin, and the alditol galacticol characteristic of the Celastraceae were also isolated. The structures were established on the basis of spectral analysis, including homo- and heteronuclear correlation NMR experiments (COSY, DEPT, HMQC and HMBC) and by comparison with data reported in the literature. The structure of 29-hydroxytaraxerol was confirmed by X-ray diffraction. The antimicrobial and antifungal activities of the compounds were studied, but no significant activity was found.

New phenolic triterpenes from Maytenus blepharodes. Semisynthesis of 6-deoxoblepharodol from pristimerin

Four new phenolic triterpenes with a 24- nor-D:A-friedoleane skeleton, isoblepharodol, 7-oxoblepharodol, blepharotriol and 6-deoxoblepharodol, were isolated from Maytenus blepharodes. Their structures were elucidated on the basis of spectroscopic analysis, including homo and heteronuclear correlation NMR experiments (COSY, ROESY, HSQC, and HMBC). The semisynthesis of 6-deoxoblepharodol and its epimer at C-8 was achieved by catalytic reduction of pristimerin, a quinone-methide triterpene present in the plant. The biosynthetic formation of the phenolic triterpenes isolated from this species is also discussed. The compounds were assayed for antimicrobial and cytotoxic activities.

Cheiloclines A–I. First examples of octacyclic sesquiterpene-triterpene hetero-Diels–Alder adducts

Cheiloclines A–I ( 1– 9 ) have been isolated from the root bark of Cheiloclinium hippocratioides (Celastraceae). They represent the first examples of hetero-Diels–Alder adducts between a nor-triterpenequinone and a sesquiterpene. Their structures were elucidated on the basis of spectral analysis, including homonuclear and heteronuclear correlation NMR experiments (COSY, ROESY, HSQC and HMBC). These compounds were tested for antitumoral, antibacterial and aldose reductase inhibitory activities.

Measuring 1H–1H and 1H–13C RDCs in methyl groups: example of pulse sequences with numerically optimized coherence transfer schemes

The optimization of coherence-transfer pulse-sequence elements (CTEs) is the most challenging step in the construction of heteronuclear correlation NMR experiments achieving sensitivity close to its theoretical maximum (in the absence of relaxation) in the shortest possible experimental time and featuring active suppression of undesired signals. As reported in the present article, this complex optimization problem in a space of high dimensionality turns out to be numerically tractable. Based on the application of molecular dynamics in the space of pulse-sequence variables, a general method is proposed for constructing optimized CTEs capable of transferring an arbitrary (generally non-Hermitian) spin operator encoding the chemical shift of heteronuclear spins to an arbitrary spin operator suitable for signal detection. The CTEs constructed in this way are evaluated against benchmarks provided by the theoretical unitary bound for coherence transfer and the minimal required transfer time (when available). This approach is used to design a set of NMR experiments enabling direct and selective observation of individual 1H-transitions in 13C-labeled methyl spin systems close to optimal sensitivity and using a minimal number of spectra. As an illustrative application of the method, optimized CTEs are used to quantitatively measure 1H–1H and 1H–13C residual dipolar couplings (RDCs) in a 17kDa protein weakly aligned by means of Pf1 phages.

Aspergillus fumigatus CY018, an endophytic fungus in Cynodon dactylon as a versatile producer of new and bioactive metabolites

Aspergillus fumigatus CY018 was recognized as an endophytic fungus for the first time in the leaf of Cynodon dactylon. By bioassay-guided fractionation, the EtOAc extract of a solid-matrix steady culture of this fungus afforded two new metabolites, named asperfumoid ( 1) and asperfumin ( 2), together with six known bioactive compounds including monomethylsulochrin, fumigaclavine C, fumitremorgin C, physcion, helvolic acid and 5α,8α-epidioxy-ergosta-6,22-diene-3β-ol as well as other four known compounds ergosta-4,22-diene-3β-ol, ergosterol, cyclo(Ala-Leu) and cyclo(Ala-Ile). Through detailed spectroscopic analyses including HRESI-MS, homo- and hetero-nuclear correlation NMR experiments (HMQC, COSY, NOESY and HMBC), the structures of asperfumoid and asperfumin were established to be spiro-(3-hydroxyl-2,6-dimethoxyl-2,5-diene-4-cyclohexone-(1,3′)-5′-methoxyl-7′-methyl-(1′H, 2′H, 4′H)-quinoline-2′,4′-dione) and 5-hydroxyl-2-(6-hydroxyl-2-methoxyl-4-methylbenzoyl)-3,6-dimethoxyl-benzoic methyl ester, respectively. All of the 12 isolates were subjected to in vitro bioactive assays against three human pathogenic fungi Candida albicans, Tricophyton rubrum and Aspergillus niger. As a result, asperfumoid, fumigaclavine C, fumitremorgin C, physcion and helvolic acid were shown to inhibit C. albicans with MICs of 75.0, 31.5, 62.5, 125.0 and 31.5 μg/mL, respectively.

Five New Alkaloids from the Leaves of Remijia peruviana

Three new indolylquinuclidine-type alkaloids, remijinine (1), epiremijinine (2), and 5-acetyl-apocinchonamine (3), and two new cinchonine-derived alkaloids, N-acetyl-deoxycinchonicinol (4) and N-acetyl-cinchonicinol (5), as well as the known alkaloids quinamine, conquinamine, cinchonine, and quinidine were isolated from the leaves of Remijia peruviana. The structures of the new alkaloids were elucidated on the basis of spectroscopic analysis, including homonuclear and heteronuclear correlation NMR experiments (COSY, ROESY, HMQC, and HMBC). The relative configuration at C-7 for remijinine (1) and, in consequence, for epiremijinine (2) was established by X-ray crystal structure analysis of the former.

19F and 13C NMR Signal Assignment and Analysis in a Perfluorinated Ionomer (Nafion) by Two-Dimensional Solid-State NMR

The 19F and 13C NMR resonances of the perfluorinated ionomer, Nafion, are assigned to their corresponding chemical groups using two-dimensional (2D) 13C−19F heteronuclear correlation and 19F-exchange NMR experiments under 28 or 30 kHz magic-angle spinning, combined with peak area and relaxation time information. On the basis of these new experimental data, we revise the assignment of more than half of the resolved 19F NMR peaks. In particular, the backbone CF group is shown to resonate at −138 ppm, the side-group CF at −144 ppm, and the SCF2 group, which can be selected by a T2 filter, at −117 ppm. The OCF2 groups resonate slightly downfield from the CF3 fluorines. Deconvolution of the 19F and 13C spectra based on cross sections from the 2D spectra provides the peak widths and positions of all side group and several backbone sites. The inhomogeneous broadening observed in both 13C and 19F NMR spectra for the sites near the backbone CF group reveals static disorder near the branch point, which contrasts with the high conformational order of the rest of the backbone and the mobility of the ends of the side group.

An evaluation of detergents for NMR structural studies of membrane proteins.

Structural information on membrane proteins lags far behind that on soluble proteins, in large part due to difficulties producing homogeneous, stable, structurally relevant samples in a membrane-like environment. In this study 25 membrane mimetics were screened using 2D (1)H-(15)N heteronuclear single quantum correlation NMR experiments to establish sample homogeneity and predict fitness for structure determination. A single detergent, 1-palmitoyl-2-hydroxy-sn-glycero-3-[phospho-RAC-(1-glycerol)] (LPPG), yielded high quality NMR spectra with sample lifetimes greater than one month for the five proteins tested - R. sphaeroides LH1 alpha and beta subunits, E. coli and B. pseudofirmus OF4 ATP synthase c subunits, and S. aureus small multidrug resistance transporter - with 1, 2, or 4 membrane spanning alpha-helices, respectively. Site-specific spin labeling established interhelical distances in the drug transporter and genetically fused dimers of c subunits in LPPG consistent with in vivo distances. Optical spectroscopy showed that LH1 beta subunits form native-like complexes with bacteriochlorophyll a in LPPG. All the protein/micelle complexes were estimated to exceed 100 kDaltons by translational diffusion measurements. However, analysis of (15)N transverse, longitudinal and (15)N[(1)H] nuclear Overhauser effect relaxation measurements yielded overall rotational correlation times of 8 to 12 nsec, similar to a 15-20 kDalton protein tumbling isotropically in solution, and consistent with the high quality NMR data observed.

Two new 24-isopropenyl-lanostanoids from Tillandsia brachycaulos.

The leaves of Tillandsia brachycaulos afforded two novel tetracyclic triterpenoids identified as (24S)-24-isopropenyl-29-nor-5alpha-lanosta-7-en-3beta-ol (1) and (24S)-24-isopropenyl-29-nor-5alpha-lanosta-7-en-3-one (2), in addition to the known isopimaric acid (3) and chlorogenic acid (4). Their structures were elucidated on the basis of spectral analysis, including homo- and heteronuclear correlation NMR experiments (COSY, ROESY, HMQC and HMBC) and by comparison with data in the literature. The antimicrobial and antifungal activities were studied. The compounds did not show significant activity.

Facile determination of the protecting group location of Nim-protected histidine derivatives by 1H-15N heteronuclear correlation NMR.

The positioning of the imidazole protecting group of several histidine derivatives was determined by means of (1)H-(15)N heteronuclear multiple-bond correlation NMR experiments. The cross-peak originated from the three-bond correlation between the histidine side-chain H(beta) and the imidazole N(pi) was used for the identification of the N(pi) signal in the (15)N spectrum. Therefore, based on the fact that the signal of the substituted imidazole nitrogen appears always at lower chemical shift (delta) than the unsubstituted one, the position of the blocking group could easily be inferred. The obtained data confirmed previous findings that were accomplished with other less generally applicable spectroscopic or crystallographic techniques.

Antileishmanial activities of dihydrochalcones from piper elongatum and synthetic related compounds. Structural requirements for activity

Two dihydrochalcones ( 1 and 2) were isolated from Piper elongatum Vahl by activity-guided fractionation against extracellular promastigotes of Leishmania braziliensis in vitro. Their structures were elucidated by spectral analysis, including homonuclear and heteronuclear correlation NMR experiments. Derivatives 3– 7 and 20 synthetic related compounds ( 8– 27) were also assayed to establish the structural requirements for antileishmanial activity. Compounds 1– 11 that proved to be more active that ketoconazol, used as positive control, were further assayed against promastigotes of Leishmania tropica and Leishmania infantum. Compounds 7 and 11, with a C 6–C 3–C 6 system, proved to be the most promising compounds, with IC 50 values of 2.98 and 3.65 μg/mL, respectively, and exhibited no toxic effect on macrophages (around 90% viability). Correlation between the molecular structures and antileishmanial activity is discussed in detail.

Absolute configuration of sesquiterpenes from Crossopetalum tonduzii and their inhibitory effects on Epstein-Barr virus early antigen activation in Raji cells.

Two new sesquiterpenoids (1 and 2) with a dihydro-beta-agarofuran skeleton were isolated from Crossopetalum tonduzii. Their structures were elucidated on the basis of spectral analysis, including homonuclear and heteronuclear correlation NMR experiments (COSY, ROESY, HSQC, and HMBC). Their absolute configurations were determined by CD studies on 3, the benzoylated derivative of 1. Chemical correlations have allowed the absolute configurations of 4 and 5, two previously known dihydro-beta-agarofuran analogues, to be reported for the first time. Compounds 1, 2, and 5 showed strong antitumor-promoting effects on Epstein-Barr virus early antigen (EBV-EA) activation.

A study of conformational stability of polypeptide blends by solid state two-dimensional 13C– 1H heteronuclear correlation NMR spectroscopy

The intermolecular hydrogen-bond interactions in polyglycine (PG)/poly( l-valine) (PLV) blend with a ratio of 1/1 have been studied through high-speed frequency-switched Lee–Goldburg (FSLG) 13C– 1H heteronuclear correlation (HETCOR) NMR experiments, where the PG/PLV blend sample is prepared by adding trifluoroacetic acid (TFA) solution of their polypeptide mixture containing a 2.0 wt/wt% amount of sulfuric acid (H 2SO 4) to alkaline water. The spectral assignment of the polypeptide blend is made by using multiple proton cross-peaks, which appeared in the HETCOR spectra. Intermolecular correlation peaks between PG(β-sheet) and PLV(β-sheet) appear in the HETCOR spectra with long contact time. From these experimental results, it has been clarified that intermolecular hydrogen-bond interactions ( C O⋯ H– N) between the PG(β-sheet) and PLV(β-sheet) are formed and thus the PG/PLV blend with high miscibility is formed.