High Field 2D Research Articles

Alkaloids from millipedes: a re-evaluation of defensive exudates from Polyzonium germanicum

Millipedes are known to produce various sets of chemical compounds in exocrine defensive glands to protect themselves against predators and microorganisms. Here, we reanalyzed the gland secretion of Polyzonium germanicum, a millipede of the order Polyzoniida, by using a combination of analytical techniques such as GC-MS, LC-HRMS and high field 1D and 2D NMR spectroscopy. Previously only one compound (polyzonimine, 1) had been described, but our approach allowed us to add six compounds to the defensive chemistry of this species. Besides polyzonimine (1), we found nitropolyzonamine (2) and five new compounds: 3 (2,3-dimethyl-7’-nitro-2’,3’,5’,6’,7’,7a’-hexahydrospiro[cyclopent-2-ene-1,1’-pyrrolizine]), 4 (2,3-dimethyl-7’-nitro-2’,3’,5’,6’,7’,7a’-hexahydrospiro[cyclopentane-1,1’-pyrrolizin]-2-ene), 5 ((1Z)-8,9-dimethyl-1-(nitromethylidene)-2-azaspiro[4.4]non-8-en-7-one), and not fully identified compounds A, B. For compounds 3–5 we were able to determine the molecular constitution, for two of them (4, 5) we were able to give relative configurations. Overall, the combination of advanced analytical techniques applied herein allowed detailed insights into the defensive chemistry of P. germanicum with a low number of individuals needed for analysis and without prior compound isolation.

Advanced solid state 1D/2D NMR investigation of the B2O3-Zn(PO3)2 glasses

In order to understand how the B2O3 insertion alters the structure and properties of the widely used zinc metaphosphate glass composition, advanced 1D/2D solid state MAS-NMR techniques were applied in this contribution to B2O3-Zn(PO3)2 glasses. High field (18.8 T) 1D NMR experiments allowed for a detailed characterization of the borate local order but did not provide much information about the phosphate speciation. We show how a set of 2D correlation NMR techniques could be applied to obtain additional data. First, 11B 2D DQ-SQ NMR spectra were edited to monitor the formation of the borate network through [x]BO[x]B linkages. In a second time, 11B(31P) D-HMQC technique was applied to characterize the formation of [x]BOP bonds and to provide additional information used to decompose the 1D 31P MAS-NMR experiments. This first 1D/2D NMR analysis of the B2O3-Zn(PO3)2 line was finally used to construct a structural model which shed light upon the glass transition temperature non-linear evolution.

Karmitoxin: An Amine-Containing Polyhydroxy-Polyene Toxin from the Marine Dinoflagellate Karlodinium armiger.

Marine algae from the genus Karlodinium are known to be involved in fish-killing events worldwide. Here we report for the first time the chemistry and bioactivity of a natural product from the newly described mixotrophic dinoflagellate Karlodinium armiger. Our work describes the isolation and structural characterization of a new polyhydroxy-polyene named karmitoxin. The structure elucidation work was facilitated by use of 13C enrichment and high-field 2D NMR spectroscopy, where 1H-13C long-range correlations turned out to be very informative. Karmitoxin is structurally related to amphidinols and karlotoxins; however it differs by containing the longest carbon-carbon backbone discovered for this class of compounds, as well as a primary amino group. Karmitoxin showed potent nanomolar cytotoxic activity in an RTgill-W1 cell assay as well as rapid immobilization and eventual mortality of the copepod Acartia tonsa, a natural grazer of K.armiger.

Cellulose Structural Polymorphism in Plant Primary Cell Walls Investigated by High-Field 2D Solid-State NMR Spectroscopy and Density Functional Theory Calculations.

The native cellulose of bacterial, algal, and animal origins has been well studied structurally using X-ray and neutron diffraction and solid-state NMR spectroscopy, and is known to consist of varying proportions of two allomorphs, Iα and Iβ, which differ in hydrogen bonding, chain packing, and local conformation. In comparison, cellulose structure in plant primary cell walls is much less understood because plant cellulose has lower crystallinity and extensive interactions with matrix polysaccharides. Here we have combined two-dimensional magic-angle-spinning (MAS) solid-state nuclear magnetic resonance (solid-state NMR) spectroscopy at high magnetic fields with density functional theory (DFT) calculations to obtain detailed information about the structural polymorphism and spatial distributions of plant primary-wall cellulose. 2D (13)C-(13)C correlation spectra of uniformly (13)C-labeled cell walls of several model plants resolved seven sets of cellulose chemical shifts. Among these, five sets (denoted a-e) belong to cellulose in the interior of the microfibril while two sets (f and g) can be assigned to surface cellulose. Importantly, most of the interior cellulose (13)C chemical shifts differ significantly from the (13)C chemical shifts of the Iα and Iβ allomorphs, indicating that plant primary-wall cellulose has different conformations, packing, and hydrogen bonding from celluloses of other organisms. 2D (13)C-(13)C correlation experiments with long mixing times and with water polarization transfer revealed the spatial distributions and matrix-polysaccharide interactions of these cellulose structures. Celluloses f and g are well mixed chains on the microfibril surface, celluloses a and b are interior chains that are in molecular contact with the surface chains, while cellulose c resides in the core of the microfibril, outside spin diffusion contact with the surface. Interestingly, cellulose d, whose chemical shifts differ most significantly from those of bacterial, algal, and animal cellulose, interacts with hemicellulose, is poorly hydrated, and is targeted by the protein expansin during wall loosening. To obtain information about the C6 hydroxymethyl conformation of these plant celluloses, we carried out DFT calculations of (13)C chemical shifts, using the Iα and Iβ crystal structures as templates and varying the C5-C6 torsion angle. Comparison with the experimental chemical shifts suggests that all interior cellulose favor the tg conformation, but cellulose d also has a similar propensity to adopt the gt conformation. These results indicate that cellulose in plant primary cell walls, due to their interactions with matrix polysaccharides, and has polymorphic structures that are not a simple superposition of the Iα and Iβ allomorphs, thus distinguishing them from bacterial and animal celluloses.

23Na and 1H NMR Relaxometry of Shale at High Magnetic Field

Formation evaluation of unconventional reservoirs is challenging due to the coexistence of different phases such as kerogen, bitumen, movable and bound light hydrocarbon and water. Current low-frequency (0.05 T) nuclear magnetic resonance (NMR) laboratory and logging methods are incapable of quantitatively separating the different phases. We demonstrate the utility of high-field (9 T) NMR 2D T1-T2 measurements for separating hydrocarbon and the clay-interacting aqueous phases in shale based on the difference in the frequency dependence of the spin-lattice relaxation time. Furthermore, we demonstrate 23Na NMR as a promising complementary technique to conventional 1H NMR for shale fluid typing, taking advantage of the fact that sodium ions are only present in the aqueous phase. We validate high-field (9 T) 23Na-1H NMR relaxometry for assessing brine-filled porosity and brine salinity in various porous materials, including porous glass, conventional rocks, clays, and shale, and apply it for differentiating hydrocarbon versus aqueous components and also the clay-associated versus free water in Eagle Ford shale cores. This work lays the groundwork for developing future downhole 23Na-1H NMR logging techniques.

Probing Dynamics and Interaction of Maltenes with Asphaltene Aggregates in Crude Oils by Multiscale NMR

Asphaltenes cause many problems in crude oil transportation, refinery, and production. A clear understanding of asphaltene behavior in crude oils is thus crucial for improving crude oil production. Here, we used noninvasive multiscale NMR techniques such as low field NMR 2D D–T2, T1–T2, nuclear magnetic relaxation dispersion (NMRD) techniques as well as high field two-dimensional (2D) NMR DOSY for probing the dynamics and interaction of maltenes with asphaltene aggregates in crude oil. We prepared different asphaltene concentrations ranging between 0 wt % (pure maltenes) and 9 wt % (native crude oil) by using a dilution procedure that maintains the asphaltene aggregates structure initially present in native crude oil. We showed by multiscale NMR techniques that changing asphaltene concentration induces a drastic modification of hydrocarbon dynamics. In native crude oil, we observe anomalous relation between the diffusion coefficient (D) and the transverse relaxation time (T2) that we can reproduce theoretically by a quasi one-dimensional (1D) diffusion of hydrocarbons in between slowly rotating asphaltene macroaggregates. The relation D α T2 usually observed in crude oils without asphaltenes progressively appeared when the asphaltene concentration decreased. Our 2D D–T2 results at 2.5 and 23 MHz show that the diffusion coefficient varies by a factor 4, while the T2 changes by more than an order of magnitude when the asphaltene concentrations vary. These observations prove that the hydrocarbon dynamics changes drastically at nanoscale and vary much less at large length scales (μm). Moreover, the 2D D–T2 also shows two different dynamics for short and long chain hydrocarbons in crude oil. This is confirmed by high field 2D NMR DOSY. We found also that the ratio T1/T2 in 2D T1–T2 distribution was progressively reduced when the asphaltene concentration decreases with an upward bent away in native crude oil at short T2 that we succeeded to interpret with the quasi 1D hydrocarbon dynamics. We observed that the NMRD profiles are completely different for crude oil with and without asphaltenes. In native crude oil, NMRD techniques clearly evidence a 2D translational diffusion of short hydrocarbon chains at proximity of the asphaltene nanoaggregates. In crude oil without asphaltene, the NMRD profile is explained in terms of a rotational reorientation of hydrocarbon chains. Most of our proposed multiscale NMR techniques could be made down-hole and then can provide an invaluable tool for a clear understanding of asphaltene behavior in crude oil.

Cytotoxity of two new coumarin derivatives isolated from Launaea mucronata

The chloroform fraction of methanol (MeOH) extract of the aerial parts of Launaea mucronata was in vitro investigated for cytotoxicity against HCT116, HepG2 and MCF-7 cell lines, and resulted with IC50 = 20.0, 18.6 and 14.30 μg/mL, respectively. The chloroform fraction of the MeOH extract was subjected to further fractionation, which led to the isolation of two new coumarin compounds (6-isobutyl coumarin and 6-isobutyl-7-methyl- coumarin). The structures of the new compounds were elucidated by high field 1D and 2D NMR and ESI-MS spectroscopies. Low polar fractions revealed the isolation of other known triterpene compounds which were identified according to its spectral data and comparison with the literature. New coumarin compounds show high cytotoxicity against MCF-7, HCT116 and HepG2 cell lines.

Diterpenos y otros constituyentes de <i>Croton hirtus</i> (Euphorbiaceae)

The roots of Croton hirtus (Euphorbiaceae) collected in Paraiso, Cartago (Costa Rica) afforded, in addition to the few known metabolites, 20 new compounds: the bis-nor dolabradane 1, the dolabradanes 2 and 3, the kauranes 4-7, the cyclopropakauranes 8 and 9, the hirtusanes 10-15, the germacradiene esters 16-18 and the C-25 compound 21, presumably formed by a Diels-Alder reaction between compound 18 and a monoterpene. All structures were elucidated using high field 1D and 2D NMR techniques. MS fragmentation patterns are here reported. The absolute configurations of 4 and 9 were elucidated by using circular dichroism. The separation was performed with modern chromatographic technics.

Demonstration of a common indole-based aromatic core in natural and synthetic eumelanins by solid-state NMR.

Despite the essential functions of melanin pigments in diverse organisms and their roles in inspiring designed nanomaterials for electron transport and drug delivery, the structural frameworks of the natural materials and their biomimetic analogs remain poorly understood. To overcome the investigative challenges posed by these insoluble heterogeneous pigments, we have used l-tyrosine or dopamine enriched with stable (13)C and (15)N isotopes to label eumelanins metabolically in cell-free and Cryptococcus neoformans cell systems and to define their molecular structures and supramolecular architectures. Using high-field two-dimensional solid-state nuclear magnetic resonance (NMR), our study directly evaluates the assumption of structural commonality between synthetic melanin models and the corresponding natural pigments, demonstrating a common indole-based aromatic core in the products from contrasting synthetic protocols for the first time.

Antiplasmodial Alkaloids from the Bark of Cryptocarya nigra (Lauraceae)

A dichloromethane extract of the stem bark of Cryptocarya nigra showed strong in vitro inhibition of Plasmodium falciparum growth, with an IC50 value of 2.82 μg/mL. The phytochemical study of this extract has led to the isolation and characterization of four known alkaloids: (+)-N-methylisococlaurine (1), atherosperminine (2), 2-hydroxyathersperminine (3), and noratherosperminine (4). Structural elucidation of all alkaloids was accomplished by means of high field 1D- and 2D-NMR, IR, UV and LCMS spectral data. The isolated extract constituents (+)-N-methylisococlaurine (1), atherosperminine (2) and 2-hydroxy-atherosperminine (3) showed strong antiplasmodial activity, with IC50 values of 5.40, 5.80 and 0.75 μM, respectively. In addition, (+)-N-methylisocolaurine (1) and atherosperminine (2) showed high antioxidant activity in a DPPH assay with IC50 values of 29.56 ug/mL and 54.53 ug/mL respectively. Compounds 1 and 2 also both showed high antioxidant activity in the FRAP assay, with percentages of 78.54 and 70.66 respectively and in the metal chelating assay, with IC50 values of 50.08 ug/mL and 42.87 ug/mL, respectively.

Probing Structure and Dynamics of Bulk and Confined Crude Oils by Multiscale NMR Spectroscopy, Diffusometry, and Relaxometry

We propose using a set of noninvasive multiscale NMR techniques for probing the structure and dynamics of bulk and confined crude oils with and without asphaltene. High-field 1D (1)H and (13)C NMR spectroscopies evidence the proton species and the amount of asphaltene and give an average chain length for the hydrocarbon aliphatic chains. Two-dimensional (1)H diffusion-ordered NMR spectroscopy (DOSY) spectra allow us to identify two populations of hydrocarbons characterized by two distributions of translational diffusion coefficients in the presence of asphaltene and a single one without asphaltene. A detailed analysis of the distributions of longitudinal, T1, relaxation times measured at different magnetic fields is proposed in terms of highly skewed bimodal (or monomodal) log-normal distributions, confirming the two environments in the presence of asphaltene and a single one without asphaltene. We show that these distributions are similar to the gas and gel permeation chromatography distributions, thus showing a connection of the hydrocarbon dynamics with their chain lengths. The remarkable observed features of the nuclear magnetic relaxation dispersion (NMRD) profiles of <1/T1> for bulk and confined crude oils with and without asphaltene are interpreted with an original relaxation model of intermittent surface dynamics of proton species at the proximity of asphaltene nanoaggregates and bulk dynamics in between clusters of these nanoaggregates. This allows us to probe the 2D translational diffusion correlation time and the time of residence of hydrocarbons in the proximity of the asphaltene nanoaggregates. Provided that the diffusion of the hydrocarbons close to the asphaltene nanoaggregates is three times smaller than the bulk diffusion, as the DOSY experiments show, this time of residence gives an average radius of exploration for the 2D hydrocarbon diffusion, r2D ≈ 3.9 nm, of the same order of magnitude as the aggregate sizes found by J. Eyssautier with SAXS and SANS in asphaltene solutions and by O. C. Mullins with the observation of gravitational gradients of asphaltenes in oilfield reservoirs.

Probing multi-dynamics of crude oils by high field 2D NMR DOSY and relaxometry

We propose using a set of non-invasive multiscale NMR techniques for probing the structure and dynamics of bulk and confined crude oils with and without asphaltene. High field NMR spectroscopy identifies the proton species. 2D NMR DOSY spectra for bulk crude oils in presence or absence of asphaltene favor an interpretation of two populations of hydrocarbon aliphatic chains in two environments. In the presence of asphaltene, an analysis of the distributions of longitudinal relaxation time, T1, in terms of a bimodal log-normal distribution, confirms the two environments clearly evidenced by varying the Larmor frequency. The remarkable features of the nuclear magnetic relaxation dispersion (NMRD) of 〈1/T1〉 for bulk and confined crude oils with and without asphaltene have been interpreted with an original model of intermittent surface dynamics of proton species at proximity of the surface of asphaltene nanoaggregates and bulk dynamics in between the clusters of these nanoaggregates.

Bioassay-Guided Isolation and Identification of Cytotoxic Compounds from Gymnosperma glutinosum Leaves

Bioassay-guided fractionation of hexane extracts of Gymnosperma glutinosum (Asteraceae) leaves, collected in North Mexico, afforded the known compounds hentriacontane (1) and (+)-13S,14R,15-trihydroxy-ent-labd-7-ene (2), as well as the new ent-labdane diterpene (−)-13S,14R,15-trihydroxy-7-oxo-ent-labd-8(9)-ene (3). In addition, D-glycero-D-galactoheptitol (4) was isolated from the methanolic extract of this plant. Their structures were established on the basis of high-field 1D- and 2D NMR methods supported by HR-MS data. The cytotoxic activity was determined by using the in vitro L5178Y-R lymphoma murine model. Hentriacontane (1) and the new ent-labdane 3 showed weak cytotoxicity, whereas the ent-labdane 2 showed significant (p < 0.05) and concentration dependent cytotoxicity (up to 78%) against L5178Y-R cells at concentrations ranging from 7.8 to 250 µg/mL.

Cytotoxicity and acetylcholinesterase inhibitory activity of an isolated crinine alkaloid from Boophane disticha (Amaryllidaceae)

Ethnopharmacological relevanceBoophane disticha of the family Amaryllidaceae is used traditionally in southern Africa in the treatment of several neurological disorders. Aim of the studyAlthough acetylcholinesterase (AChE) inhibitory activity has been reported for this plant, the aim of the study was to identify and characterise the compound responsible for this activity using bioassay guided fractionation. Toxicity of the isolated compound was also assessed. Materials and methodsBioassay guided isolation of the active compound from the methanol extract was carried out using column chromatography, TLC and preparative thin layer chromatography. Structural elucidation was carried out using high field 1D and 2D NMR and mass spectroscopy. AChE inhibitory activity was determined using the Ellman’s colorimetric method. Cytotoxicity assessment was determined in human neuroblastoma (SH-SY5Y) cells using the MTT and neutral red uptake assays. ResultsThe data obtained from the integration of the 1H spectra confirmed the compound to be a 3:1 mixture of two epimers, with epimer A, 6α-hydroxycrinamine as the major epimer. The IC50 value for AChE inhibitory activity of the compound was 445μM. The compound was observed to be cytotoxic in both the MTT and neutral red assays with IC50 values of 54.5 and 61.7μM, respectively. ConclusionThe present study describes for the first time, the isolation of 6-hydroxycrinamine, a crinine alkaloid, from the methanol extract of the bulbs of B. disticha. Although this compound possessed AChE inhibitory activity, it was found to be toxic to the neuroblastoma cells. Quantitative structure–activity relationship studies could be carried out to modify the structure in order to make it less toxic and improve its activity.

Anti-inflammatory effects of Terminalia phanerophlebia (Combretaceae) and identification of the active constituent principles

The ‘Lebombo cluster leaf’ Terminalia phanerophlebia (Combretaceae) is used in traditional medicine in South Africa for a host of ailments, including aches and pain, wounds and infections, gynaecological conditions, inflammation, mental illness such as epilepsy, and sexually-transmitted diseases. Based on the anti-inflammatory activity (92.4% inhibition of the cyclooxygenase enzyme COX-2) initially observed for a crude extract, stems of T. phanerophlebia were here examined for the underlying phytochemical principles. Three cholestane triterpenoids β-sitosterol, β-sitostenone and stigmast-4-ene-3,6-dione were isolated from the ethanol extract by silica gel column chromatography via gradient elution with hexane/ethyl acetate mixtures. Structure elucidation was facilitated by use of physical and spectroscopic methods, including high field 2D NMR and mass spectroscopic techniques. The crude extract as well as β-sitosterol was seen to be selective inhibitors of the cyclooxygenase enzyme COX-2. This work is of chemosystematic relevance as it represents the first isolation of β-sitostenone and stigmast-4-ene-3,6-dione from any species of Terminalia.

Antineoplastic Agents. 587. Isolation and Structure of 3-Epipancratistatin from Narcissus cv. Ice Follies

Bioassay-guided (cancer cell line) separation of an extract prepared from Narcissus cv. Ice Follies (from The Netherlands) led to the isolation of a new Amaryllidaceae isocarbostiryl, 3-epipancratistatin (1b), as well as narciclasine (2). This Narcissus cultivar was found to be a good source of narciclasine. The structure of 1b was established by high-resolution mass and high-field 2D NMR spectroscopic analyses. Against a panel of murine and human cancer cell lines, 3-epipancratistatin (1b) led to cell growth inhibition (GI(50) 2.2-0.69 μg/mL) some 100× less than that found for pancratistatin (1a) and narciclasine (2), thereby revealing an important configurational requirement in 1a for strong cancer cell growth inhibition.

Antibacterial activity of crinane alkaloids from Boophone disticha (Amaryllidaceae)

Ethnopharmacological relevanceBoophone disticha (Amaryllidaceae) is one of the most common bulbous plants used for medicinal purposes by the indigenous people of southern Africa. Its use as a narcotic substance by the Khoi/San tribes has been known for several centuries, while the Sotho, Xhosa and Zulu people are known to use the plant to treat a host of ailments, including inflammation, wounds, gynaecological conditions and psychosis. Aim of the studyMuch of the pharmacological work on the plant, such as affinity to the serotonin transporter, has been based on its reputed usage for narcotic purposes. However, its widespread use to treat wounds and infections has not been linked to a specific chemical entity. In this regard, Boophone disticha was here examined for its phytochemical composition which could shed light on the use of the plant for such purposes. Materials and methodsThe known crinane alkaloids buphanidrine and distichamine were isolated via column chromatography of the ethanolic extract of bulbs of Boophone disticha. Structural details of the compounds were determined by high field 2D NMR and mass spectroscopic techniques. Microbial activity against selected Gram-positive and Gram-negative bacteria was ascertained according to the micro-dilution assay. ResultsBoth buphanidrine and distichamine were uncovered as novel, broad spectrum moderately active, antibacterial agents with the best MIC value detected at 0.063mg/ml for Staphylococcus aureus, Escherichia coli and Klebsiella pneumoniae. MIC values for Bacillus subtilis were two-fold less than that observed for the other three bacteria, suggesting that the extract and pure compounds were selective in their interaction with the bacterial pathogens. ConclusionPhytochemical investigation of Boophone disticha has led to the identification of two known crinanes, buphanidrine and distichamine. Based on the reputed traditional use of the plant for wounds and infections, both compounds were screened for antibacterial activity which revealed them to be novel, broad spectrum antibacterial agents with the best MIC value set at 0.063mg/ml. Their close structural similarity may have bearing on their similar activity profiles.

Polyhydroxypregnane Glycosides from the Roots of Cynanchum otophyllum

AbstractSix new polyhydroxypregnane glycosides, namely cynotophyllosides A–F (1–6), together with five known steroids, were isolated from the roots of Cynanchum otophyllum. Their structures were elucidated by extensive spectroscopic methods (especially 2D‐NMR techniques) and acid‐catalyzed hydrolysis. Furthermore, high field 1D‐ and 2D‐NMR experiments were employed to elucidate the structure of 8 previously deduced only on the basis of LC/MS data.

Isolation of narciprimine from Cyrtanthus contractus (Amaryllidaceae) and evaluation of its acetylcholinesterase inhibitory activity

Ethnopharmacological relevancePlants of the family Amaryllidaceae are used widely in traditional medicine in South Africa. Several of these, including representatives of the genus Cyrtanthus find use in the treatment of mental illness and age-related dementia. Aim of the studyBased on the distribution of central nervous system-activating alkaloidal constituents within the genus Cyrtanthus, Cyrtanthus contractus was here explored for such compounds which could interact with acetylcholinesterase (AChE), of significance in the progression of neurodegeneration associated with Alzheimer's disease. Materials and methodsThe known phenanthridone alkaloid narciprimine was isolated via column chromatography of the ethanolic extract of bulbs of Cyrtanthus contractus. The structure of the compound was determined by high field 2D NMR and mass spectroscopic techniques. The classical method of Ellman et al. was used in the determination of AChE inhibitory activity. ResultsThe isolation of narciprimine from Cyrtanthus contractus is a landmark find since it has previously only been identified in Zephyranthes, Narcissus and Lycoris, genera endemic to the Americas, Europe and Asia, respectively. Narciprimine exhibited micromolar inhibitory activity (IC50 78.9) against the enzyme acetylcholinesterase. ConclusionThis work represents the first isolation of narciprimine from an African Amaryllidaceae species, which may be of chemotaxonomic significance. The AChE inhibitory activity of narciprimine, taken together with activities of other structurally similar inhibitors within the series affords further insight to the structural details of the lycorine alkaloid acetylcholinesterase inhibitory pharmacophore.

Isolation and characterization of a new flavone diglucoside from Salix denticulata

A new flavone diglucoside named 7,3′-dihydroxy-4′-methoxyflavone-5-O-β-D-glucopyranosyl (6″ → 1‴)-β-D-glucopyranoside (1), along with four known flavonoids, luteolin (2), isoquercetin (3), catechin (4), and diosmetin (5), has been isolated and characterized from Salix denticulata. The structure of the new flavone diglucoside was characterized by means of high field 1D and 2D NMR and MS spectral analysis.