2D NMR Spectroscopy Research Articles

Multi-Function of a New Bioactive Secondary Metabolite Derived from Endophytic Fungus Colletotrichum acutatum of Angelica sinensis.

In the current study we assessed a new crystallized compound, 5-(1-hydroxybutyl)-4-methoxy-3-methyl-2H-pyran-2-one (C-HMMP), from the endophytic fungus Colletotrichum acutatum residing in the medicinal plant Angelica sinensis for its in vitro antimicrobial, antibiofilm, antioxidant, antimalarial, and anti-proliferative properties. The promising compound was identified as C-HMMP through antimicrobial-guided fraction. The structure of C-HMMP was unambiguously confirmed by 2D NMR and HIRS spectroscopic analysis. Antimicrobial property testing of C-HMMP showed it to be effective against a variety of pathogenic bacteria and fungi with MICs ranging from 3.9 to 31.25 μg/ml. The compound displayed excellent antibiofilm activity against C. albicans, S. aureus, and K. pneumonia. Furthermore, the antimalarial and radical scavenging activities of C-HMMP were clearly dosede-pendent, with IC50 values of 0.15 and 131.2 μg/ml. The anti-proliferative activity of C-HMMP against the HepG-2, HeLa, and MCF-7 cell lines in vitro was investigated by MTT assay, revealing notable anti-proliferative activity with IC50 values of 114.1, 90, and 133.6 μg/ml, respectively. Moreover, C-HMMP successfully targets topoisomerase I and demonstrated beneficial anti-mutagenicity in the Ames test against the reactive carcinogenic mutagen, 2-aminofluorene (2-AF). Finally, the compound inhibited the activity of α-glucosidase and α-amylase with IC50 values of 144.7 and 118.6 μg/ml, respectively. To the best of our knowledge, the identified compound C-HMMP was obtained for the first time from C. acutatum of A. sinensis, and this study demonstrated that C-HMMP has relevant biological significance and could provide better therapeutic targets against disease.

Anonazepine, a new alkaloid from the leaves of Annona muricata (Annonaceae).

From the CHCl3-soluble extract of Annona muricata L. (Annonaceae) leaves, one new 3-benzazepine-type alkaloid, anonazepine (1), and four known aporphine-type alkaloids, (+)-laurotetanine (2), (+)-norglaucine (3), (-)-xylopine (4), and lanuginosine (5), were isolated. Except for (-)-xylopine (4), these remaining known alkaloids were first reported in A.muricata. The structures of the isolated alkaloids were established by 1D and 2D NMR spectroscopy and MS, as well as comparison with literature data. Thenew3-benzazepine-type alkaloid existed in an inseparable mixture of two equilibrium conformers. Its absolute configuration was determined based on comparingtheirexperimental and calculated ECD data. Theanti-inflammatory activity of the isolated alkaloids was investigated, but none of the alkaloids showed a significant result.

Discovery of New Triterpenoids Extracted from Camellia oleifera Seed Cake and the Molecular Mechanism Underlying Their Antitumor Activity.

Theasaponin derivatives, which are reported to exert antitumor activity, have been widely reported to exist in edible plants, including in the seed cake of Camellia oleifera (C.), which is extensively grown in south of China. The purpose of this study was to isolate new theasaponin derivatives from C. seed cake and explore their potential antitumor activity and their underlying molecular mechanism. In the present study, we first isolated and identified four theasaponin derivatives (compounds 1, 2, 3, and 4) from the total aglycone extract of the seed cake of Camellia oleifera by utilizing a combination of pre-acid-hydrolysis treatment and activity-guided isolation. Among them, compound 1 (C1) and compound 4 (C4) are newly discovered theasaponins that have not been reported before. The structures of these two new compounds were characterized based on comprehensive 1D and 2D NMR spectroscopy and high-resolution mass spectrometry, as well as data reported in the literature. Secondly, the cytotoxicity and antitumor property of the above four purified compounds were evaluated in selected typical tumor cell lines, Huh-7, HepG2, Hela, A549, and SGC7901, and the results showed that the ED50 value of C4 ranges from 1.5 to 11.3 µM, which is comparable to that of cisplatinum (CDDP) in these five cell lines, indicating that C4 has the most powerful antitumor activity among them. Finally, a preliminary mechanistic investigation was performed to uncover the molecular mechanism underlying the antitumor property of C4, and the results suggested that C4 may trigger apoptosis through the Bcl-2/Caspase-3 and JAK2/STAT3 pathways, and stimulate cell proliferation via the NF-κB/iNOS/COX-2 pathway. Moreover, it was surprising to find that C4 can inhibit the Nrf2/HO-1 pathway, which indicates that C4 has the potency to overcome the resistance to cancer drugs. Therefore, C1 and C4 are two newly identified theasaponin derivatives with antitumor activity from the seed cake of Camellia oleifera, and C4 is a promising antitumor candidate not only for its powerful antitumor activity, but also for its ability to function as an Nrf2 inhibitor to enhance the anticancer drug sensitivity.

One-Pot Solvent-Involved Synthesis of 5-O-Substituted 5H-Chromeno[2,3-b]pyridines.

Chromeno[2,3-b]pyridines are substances demanded in medicinal and material chemistry. PASE (pot, atom, and step economy) and in particular one-pot approaches are key green chemistry techniques that are applied for the synthesis of heterocyclic compounds. In this case, the PASE approach was extended with 'component economy', as solvent was used also as reactant (solvent-involved reaction). This approach was adopted for the one-pot synthesis of previously unknown O-substituted 5-alkoxy-5H-chromeno[2,3-b]pyridines via two-step transformation, namely the reaction of salicylaldehydes and malononitrile dimer, with the subsequent addition of alcohol. The mechanistic studies revealed the possibility of concurrent reaction. The studies aided in optimizing the reaction conditions for the best yields (77-93%). Thus, the one-pot reaction proceeds efficient and quickly, and the work-up procedure (only simple filtering) is very convenient. The structure of synthesized chromeno[2,3-b]pyridines was confirmed by 2D NMR spectroscopy.

Hemiactinomycin, an undescribed intermediate of actinomycin biosynthesis from an actinomycetes strain Streptomyces antibioticus H41-55

Hemiactinomycin (1), an intermediate derivative of actinomycin biosynthesis, together with three known actinomycins (2-4) , were isolated from the ethanolic extract of Streptomyces antibioticus H41-55 fermentation mycelium by using various column chromatography. The structure of the derivative was established by extensive spectroscopic analysis, including HRESIMS, 1D, and 2D NMR spectroscopy. In addition, the anti-inflammatory activities of all the isolates were tested. The derivative (1) showed inhibiting NO release activities in LPS-induced RAW 264.7 macrophages with the IC50 values of 15.41 ± 0.66 μM.

Synthesis and Studies of Fused Benzo-Benzisapphyrins.

A series of rare examples of fused benzo-benzisapphyrins were synthesized readily by (3 + 2) condensation of benzodipyrrole-derived diol and para-benzitripyrrane in the presence of 0.5 equiv of TFA in CH2Cl2 under inert atmosphere conditions accompanied by DDQ oxidation in open air. The crude compounds were separated by basic alumina column chromatography and afforded pure fused benzo-benzisapphyrins in 20-22% yields. The fused sapphyrins were characterized in detail by high-resolution mass spectrometry (HRMS) and one-dimensional (1D) and two-dimensional (2D) NMR spectroscopy. The 1H NMR spectra recorded at both 298 and at 233 K clearly exhibited the presence of a strong diatropic ring current in benzo-benzisapphyrins, and the macrocycles are of aromatic nature. The DFT-optimized structure of benzo-benzisapphyrin revealed that the macrocycle was planar to a great extent due to the rigid structure of the dibenzopyrrole moiety, and the NICS(0) value of -11.2 ppm supports the aromatic nature of macrocycles. The absorption spectra of benzo-benzisapphyrins showed three weak Q bands approximately in the region of 650-900 nm and a strong Soret band at 480 nm, along with a shoulder band at ∼510 nm. The diprotonated derivative generated by the addition of excess TFA to the benzo-benzisapphyrin macrocycle exhibited bathochromically shifted absorption bands compared to the free base macrocycle.

Unambiguous Identification of Glucose-Induced Glycation in mAbs and other Proteins by NMR Spectroscopy

ObjectiveGlycation is a non-enzymatic and spontaneous post-translational modification (PTM) generated by the reaction between reducing sugars and primary amine groups within proteins. Because glycation can alter the properties of proteins, it is a critical quality attribute of therapeutic monoclonal antibodies (mAbs) and should therefore be carefully monitored. The most abundant product of glycation is formed by glucose and lysine side chains resulting in fructoselysine after Amadori rearrangement. In proteomics, which routinely uses a combination of chromatography and mass spectrometry to analyze PTMs, there is no straight-forward way to distinguish between glycation products of a reducing monosaccharide and an additional hexose within a glycan, since both lead to a mass difference of 162 Da.MethodsTo verify that the observed mass change is indeed a glycation product, we developed an approach based on 2D NMR spectroscopy spectroscopy and full-length protein samples denatured using high concentrations of deuterated urea.ResultsThe dominating β-pyranose form of the Amadori product shows a characteristic chemical shift correlation pattern in 1H-13C HSQC spectra suited to identify glucose-induced glycation. The same pattern was observed in spectra of a variety of artificially glycated proteins, including two mAbs, as well as natural proteins.ConclusionBased on this unique correlation pattern, 2D NMR spectroscopy can be used to unambiguously identify glucose-induced glycation in any protein of interest. We provide a robust method that is orthogonal to MS-based methods and can also be used for cross-validation.

New Benzofuran Oligomers from the Roots of Eupatorium heterophyllum Collected in China.

The chemical constituents of two root samples of Eupatorium heterophyllum DC. collected in Yunnan Province, China, were investigated. Five new oligomeric benzofurans (1-5), nine new benzofuran/dihydrobenzofuran derivatives, and a new thymol analog were isolated, and their structures were determined using extensive spectroscopic techniques, such as 1D and 2D NMR spectroscopy and DFT calculations of the CD spectra. Most of the new compounds, including oligomeric benzofurans (1-5), were obtained from only one of the root samples. Furthermore, this is the first example that produces oligomeric benzofurans in this plant. These results imply that diversification of secondary metabolites in E. heterophyllum is ongoing. Plausible biosynthetic pathways for 1-5 are also proposed.

New steroids from the pericarps of Datura metel L

Three new steroids (1–3) and 13 reported analogs (4–16) were extracted from Datura metel L. pericarps. Structure analysis of these extracted compounds was performed by 1 D-NMR and 2 D-NMR spectroscopy, and their spectra were compared with those of similar compounds previously described in the literature. The extracted steroids (1–3) and known compounds (4–16) were evaluated for anti-inflammatory activity against LPS-activated RAW 264.7 cells. Compounds 5, 7, 9, 12 and 15 showed potential anti-inflammatory activity with IC50 less than 35 μM, while compounds 3 and 11 showed weak anti-inflammatory activity with IC50 less than 100 μM.

Cannabinoid-like meroterpenoids from Peperomia incana

Ten previously undescribed metabolites were isolated from Peperomia incana (Haw.) A. Dietr. (Piperaceae), among which four contained a chromene moiety, two were identified as meroterpene lactones, and four were cannabinoid-like compounds. While the chemical structures of the compounds were assigned based on HRESIMS and 1D and 2D-NMR spectra analyses, the relative and absolute configurations were assigned from NOE correlations and a combination of ECD data and X-ray single crystal analyses, respectively. In a cytotoxic assay against a panel of seven human cancer cell lines (A549, MDA-MB-231, HeLa, DU 145, 5637, Hep G2, and MIA PaCa-2, which represent non-small cell lung cancer, as well as breast, cervical, prostate, bladder, liver, and pancreas carcinomas, respectively) most of the isolated compounds showed promising cytotoxic activities. The incanachromenes B, and incanabinoids A and C exhibited the highest cytotoxicity toward all tested cancer cell lines with IC50 values in the range of 5.0–10.0 μM, whereas incanolides A, B, and incanabinoid B showed the lowest cytotoxic activity. In addition, incanachromene C and incanabinoid C produced a significant antibacterial effect toward planktonic cells and biofilms of multidrug-resistant Staphylococcus aureus strains.

PAMAM-Calix-Dendrimers: Second Generation Synthesis, Fluorescent Properties and Catecholamines Binding.

A convenient method for the synthesis of the second generation of PAMAM dendrimers based on a p-tert-butylthiacalix[4]arene core in cone, partial cone and 1,3-alternate conformations was developed. Unusual fluorescence of the obtained PAMAM-calix-dendrimers has been found and explained. The binding ability of the synthesized dendrimers toward catecholamines (dopamine, L-adrenaline and L-noradrenaline) was shown by UV-Vis, fluorescence, 1D and 2D NMR spectroscopy and the binding constants (logKa 3.85-4.74) calculated. As was shown, the PAMAM-calix-dendrimers bind catecholamines by the internal cavities. All the studied hormones were most efficiently bound by the dendrimers bearing a macrocyclic core in 1,3-alternate conformation. The size of the formed supramolecular systems of dendrimer/catecholamine was established by the DLS method. A decrease in hemolytic activity of the PAMAM-calix-dendrimers with an increase in the generation number of a dendrimer was shown for the dendrimers with a core in 1,3-alternate conformation. The prospects for the use of the synthesized dendrimers with the macrocyclic core as drug delivery agents were discussed.

Polyacetylenic caffeoyl amides from Ammodaucus leucotrichus

Six undescribed polyacetylenic caffeoyl amides, five known flavones and three known lignans were obtained from the fruits of the North African traditional medicinal plant Ammodaucus leucotrichus Coss. & Durieu (Apiaceae). Isolation was achieved by a combination of chromatographic methods, and structures were established by extensive 1D and 2D NMR spectroscopy, mass spectrometry, electronic circular dichroism, and by GC-MS analysis of sugar derivatives. Polyacetylenic caffeoyl amides are reported for the first time as specialized metabolites.

Bioactive Compounds from Endophytic Bacteria Bacillus subtilis Strain EP1 with Their Antibacterial Activities.

Endophytic bacteria boost host plant defense and growth by producing vital compounds. In the current study, a bacterial strain was isolated from the Boswellia sacra plant and identified as Bacillus subtilis strain EP1 (accession number: MT256301) through 16S RNA gene sequencing. From the identified bacteria, four compounds-1 (4-(4-cinnamoyloxy)phenyl)butanoic acid), 2 (cyclo-(L-Pro-D-Tyr)), 3 (cyclo-(L-Val-L-Phe)), and 4 (cyclo-(L-Pro-L-Val))-were isolated and characterized by 1D and 2D NMR and mass spectroscopy. Moreover, antibacterial activity and beta-lactam-producing gene inhibition (δ-(l-α-aminoadipyl)-l-cysteinyl-d-valine synthetase (ACVS) and aminoadipate aminotransferase (AADAT)) assays were performed. Significant antibacterial activity was observed against the human pathogenic bacterial strains (E. coli) by compound 4 with a 13 ± 0.7 mm zone of inhibition (ZOI), followed by compound 1 having an 11 ± 0.7 mm ZOI. In contrast, the least antibacterial activity among the tested samples was offered by compound 2 with a 10 ± 0.9 mm ZOI compared to the standard (26 ± 1.2 mm). Similarly, the molecular analysis of beta-lactam inhibition determined that compounds 3 and 4 inhibited the two genes (2- to 4-fold) in the beta-lactam biosynthesis (ACVS and AADAT) pathway. From these results, it can be concluded that future research on these compounds could lead to the inhibition of antibiotic-resistant pathogenic bacterial strains.

Ruthenium Complexes of Rigid, Dianionic, Tetradentate N‐Donor Ligands and their Potential as Catalysts for Water Oxidation

AbstractTwo mononuclear ruthenium(II) complexes based on dianionic {N4} ligands and with axial pyridines have been prepared and characterized crystallographically (1) or by 2D NMR spectroscopy using residual dipolar couplings (2). The {N4} ligands provide a constrained equatorial coordination with one large N−Ru−N angle, and additional non‐coordinating N atoms in case of 2. Their redox properties have been investigated (spectro)electrochemically, and their potential to serve as water oxidation catalysts has been probed using cerium ammonium nitrate (CAN) at pH 1.0. Complex 1 undergoes rapid degradation, likely via ligand oxidation, whereas 2 is more rugged and exhibits 80 % efficiency in the CeIV‐driven water oxidation, with a high initial turnover frequency (TOFi) of 3.07×10−2 s−1 (at 100 equiv. CAN). The initial rate of O2 evolution exhibits 1st order dependence on catalyst concentration, suggesting a water nucleophilic attack mechanism. Repeated addition of CAN and control experiments show that high ionic strength conditions (both NO3− and CeIII) significantly decrease the TOF.

Diverse undescribed compounds from the rhizome of Zingiber officinale Rosc. And their anti-inflammatory activity

Seven undescribed compounds, including a sesquiterpenoid derivative, a γ-lactone, four gingerols, and a dihydroferulic acid lactate were isolated from the rhizome of Zingiber officinale, and named gingerterpenoid G, gingerlactone A, zingibergingerols A−D and L-dihydroferulic acid lactate, respectively. Zingibergingerols (±)-B and (±)-C were two pairs of enantiomers. The structures of all compounds were determined by 1D-NMR, 2D-NMR and mass spectrometry. The absolute configurations were determined by comparison of the experimental and theoretically calculated ECD curves or X-ray single-crystal diffraction. Bioassay showed that gingerterpenoid G, gingerlactone A, and zingibergingerols A and B exhibited significant anti-inflammatory activity in the model of LPS-induced RAW 264.7 cells.

Green multicomponent approach to novel 5‐[(2H‐Pyran‐3‐yl)(aryl)methyl]‐1,3‐dimethylpyrimidines

AbstractThe multicomponent green methodologies are an ideal instrument in organic synthesis. Environmentally benign, facile, and efficient multicomponent synthesis of [(2H‐pyran‐3‐yl)(aryl)methyl]‐1,3‐dimethylpyrimidines by the reaction of aromatic aldehydes, N,N′‐dimethylbarbituric acid and 4‐hydroxy‐6‐methyl‐2H‐pyran‐2‐one employing simple and eco‐friendly catalyst and solvent—sodium acetate and ethanol is reported. The reaction proceeds at reflux conditions with good and excellent yields (65%–97%). The isolation of final compounds does not require complex purification procedures. All novel synthesized compounds are characterized by 1H and 13C NMR spectroscopy, FT‐IR and mass spectrometry, and elemental analysis. The [(2H‐pyran‐3‐yl)(aryl)methyl]‐1,3‐dimethylpyrimidine structure was proven using 2D NMR spectroscopy methods.

Antimicrobial phenolic metabolites from the aerial parts of Orthosiphon aristatus

Two new compounds oraristatin A (1) and oraristatinoside A (2), one new natural compound (2 S)-methyl-6,7-dihydroxytropate (3), and 11 known phenolic metabolites were isolated from the aerial parts of Orthosiphon aristatus. Their chemical structures were identified by 1D- and 2D NMR and HRESIMS spectroscopic analyses. The absolute configurations of 1 and 3 were determined by TD-DFT ECD spectroscopic analyses. Of the isolates, compound 2 weakly inhibited both Gram-positive and -negative bacteria (MIC = 150–300 μM), while 6 and 7 suppressed the growth of three Gram-positive bacteria and a yeast (MIC = 75–150 μM). This is the first report of the isolation of 6 − 9, 12, and 14 from the genus Orthosiphon and the antimicrobial effects of compounds 3, 7, 9, 12, and 14.

Neuroprotective effects of phenolic glycosides from Populus tomentiglandulosa roots in HT22 mouse hippocampal neuronal cells

The present work isolated a total of three novel phenolic glycosides (1 − 3), and 12 known compounds (4 − 15) from methanol extract of the roots of Populus tomentiglandulosa. The chemical structures of these compounds were determined by spectroscopic analyses (1D and 2D NMR and mass spectrometry). The compounds (5, 7, and 11) having the 1‑hydroxy-6-oxo-2-cyclohexenecarboxylate (HCH) moiety showed strong neuroprotective effects in glutamate-induced HT22 hippocampal cells. However, among these compounds, tremulacin (compound 5; TRC) was evaluated for its ability to prevent glutamate-induced apoptosis. TRC reduced the glutamate-induced apoptosis by downregulating of cleaved poly (ADP-ribose) polymerase (PARP), caspase-3, Bax, and phosphorylated mitogen-activated protein kinase (MAPK) pathways while upregulating Bcl-2 and Bid. Also, TRC triggered the expression of hemeoxygenase-1 (HO-1) and nuclear erythroid-2 related factor 2 (Nrf2) in a dose-dependent manner. Furthermore, TRC treatment attenuated intracellular ROS generation and MMP loss. Molecular docking results also demonstrated that TRC interacted with Kelch-like ECH-associated protein 1 (Keap1). Overall, this study revealed that TRC might be a promising treatment option for neurological diseases such as Alzheimer's and Parkinson's.

Nimonol from Chisocheton macrophyllus (Meliaceae) Seeds and Their Cytotoxic Activity against P-388 Leukaemia Cells

The Chisocheton genus belongs to the Meliaceae family which produces various structures and activities of compounds, such as antimalarial, antimicrobial, antitumor, anti-inflammatory, and cytotoxic. This plant has 53 species that are spread in tropical and sub-tropical forests, including Indonesia. Chisocheton plants have been known as plants that produce limonoids, namely triterpenoid compounds that have been modified to lose four terminal carbons (tetranortriterpenoids). One of the species whose phytochemical reports are still few and interesting for research on limonoid content is Chisocheton macrophyllus. Chisocheton macrophyllus is a tall plant that grows in the rainforest in the northern part of the island of Sulawesi, Indonesia, has the regional name ma aa, gula, pasak lingga, gending, ta suea, bekak, or pithraj tree. This paper will describe a limonoid compound, namely nimonol which has been isolated from Chisocheton macrophyllus. Nimonol is known to have the molecular formula C28H36O5 from a group of havanensin. The structure was determined by spectroscopic methods UV, IR, 1D-NMR (1H-NMR, 13C-NMR, and DEPT), 2D-NMR (1H-1H COSY, HMQC, and HMBC), and mass spectroscopy.

Crystal structure and biological activity of tetra-tosyl derivative of quercetin

At present time one of the great interest is represented on the biologic activities ofquercetin a polyphenol relating to the class of flavonoids, natural products generally known for theirbeneficial actions on health. With the aim of obtaining new derivatives of quercetin which wouldpossess a potential of showing valuable biological activity, 5-hydroxy-3,3’,4’,7-tetratosyl flavone wassynthesized by base catalyzed etherification of the starting material with 4-toluenesulfonyl chloride(TsCl). The chemical structure of the derived molecule was established by X-ray crystallographicanalysis. The 1 H and 13 C NMR spectra of this compound were interpreted using 2D NMR spectroscopy. The new synthesized compound was tested for cytotoxic and antiradical activities and itsother possible pharmacological properties were predicted using PASS software.