Structural determination and anticholinesterase assay of C-glycosidic ellagitannins from Lawsonia inermis leaves: A study supported by DFT calculations and molecular docking.

An ellagitannin monomer, lythracin M (1), and a dimer, lythracin D (2), along with eight known monomers (3-10) were isolated from Lawsonia inermis (Lythraceae) leaves. Lythracin M (1) is a C-glycosidic ellagitannin with a flavogallonyl dilactone moiety that participates in the creation of a γ-lactone ring with the anomeric carbon of the glucose core. Lythracin D (2) was determined as an atropisomer of the reported lythcarin D. These newly discovered structures (1 and 2) were determined by intensive spectroscopic experiments and by comparing DFT-calculated 1H1H coupling, 1H NMR chemical shifts, and ECD data with experimental values. The anti-acetylcholinesterase assay of the compounds 1-10 revealed that the C-1 ellagitannin epimers [casuarinin (7; IC50 = 34 ± 2 nM) and stachyurin (8; IC50 = 56 ± 3 nM)], and the new dimer (2; IC50 = 61 ± 4 nM) possess enzyme inhibitory effects comparable to the reference drug (donepezil, IC50 = 44 ± 3 nM). Molecular docking of compounds 1-10 with AChE identified the free galloyl moiety as an important pharmacophore in the anticholinesterase activity of tannins.

Design, Characterization, and Antimicrobial Evaluation of Copper Nanoparticles Utilizing Tamarixinin a Ellagitannin from Galls of Tamarix aphylla.

The application of plant extracts or plant-derived compounds in the green synthesis of metal nanoparticles (NPs) was researched. Determining the exact metabolite implicated in the formation of NPs would necessitate comprehensive investigations. Copper nanoparticles (CuNPs) are gaining a lot of attention because of their unique properties and effectiveness against a wide range of bacteria and fungi, as well as their potential for usage in catalytic, optical, electrical, and microelectronics applications. In the course of this study, we aimed to formulate CuNPs utilizing pure tamarixinin A (TA) ellagitannin isolated from Tamarix aphylla galls. The main particle size of the formed CuNPs was 44 ± 1.7 nm with zeta potential equal to -23.7 mV, which emphasize the stability of the CuNPs. The X-ray diffraction spectroscopy showed a typical centered cubic crystalline structure phase of copper. Scanning electron microscopy images were found to be relatively spherical and homogeneous in shape. The antimicrobial properties of TA, as well as its mediated CuNPs, have been evaluated through well diffusion assays against four bacterial, Bacillus subtilis NCTC 10400, Staphylococcus aureus ATCC 25923, Escherichia coli ATCC 25922, and Pseudomonas aeruginosa ATCC 27853, and two fungal, Candida albicans and Aspergillus flavus, strains. The distinctive antimicrobial activities were noted against the fungal strains and the Gram-negative bacterial strains P. aeruginosa ATCC 27853, and E. coli ATCC 25922. In conclusion, CuNPs mediated by TA can be applied for combating a wide range of bacterial and fungal species especially C. albicans, Asp. flavus, and P. aeruginosa in a variety of fields.

Ellagitannins and simple phenolics from the halophytic plant Tamarix nilotica.

Three new [nilotinins M8‒M10 (1‒3)] and two known [tamarixinin A (4) and gemin D (5)] ellagitannins and seven simple phenolics [gallic acid (6), methyl gallate (7), 3,4-di-O-methylgallic acid (8), ellagic acid (9), 3-O-methylellagic acid (10), methyl ferulate 3-O-sulphate (11), and 7,4'-di-O-methylkaempferol (12)] were isolated from the halophytic plant Tamarix nilotica (Ehrenb.) Bunge (Tamaricaceae). Their structures were determined based on intensive spectroscopic studies and comparisons with reported data. Compounds 4, and 6-8 were evaluated for their cytotoxicity against lung adenocarcinoma cell line (A549) and anti-leishmanial activity against Leishmania major. Compounds 4, 6 and 7 showed promising cytotoxic properties against A549 (IC50 29 ± 2.3, 10.5 ± 0.7, and 20.7 ± 1.9 µg/mL), while compounds 4 and 7 showed higher growth-inhibitory effects against L. major promastigotes (IC50 40.5 ± 2.7 and 38.4 ± 2.5 µg/mL), as compared with the standards doxorubicin (IC50 0.42 µg/mL) and miltefosine (IC50 9.43 µg/mL), respectively.

Two new C-glycosidic ellagitannins and accompanying tannins from Lawsonia inermis leaves and their cytotoxic effects.

Investigation on tannins having antitumor properties led to the isolation of two new C-glycosidic ellagitannins (1 and 2) along with seven known ellagitannins (3-9) and a related polyphenolic constituent (10) from Lawsonia inermis leaves. Our intensive HRESIMS, 1D and 2D NMR, and ECD spectroscopic studies of new tannins have shown that one (1) has a monomer structure of C-glycosidic tannin, and the other (2) has a dimeric structure of 2,3-O-hexahydroxydiphenoyl glucopyranose and a C-glycosidic tannin. Among the known compounds, one (3) is a C-glycosidic tannin that was isolated first of all from nature, five were C-glycosidic tannins, vescalagin (4), 1-O-methylvescalagin (5), castalagin (6), stachyurin (7), and casuarinin (8), and one was an O-glycosidic ellagitannin, tellimagrandin II (9). The remaining phenolic constituent from the leaves was identified as valoneic acid dilactone (10). The ellagitannins 1, and 3-9 demonstrated noticeable cytotoxicity on human oral squamous cell carcinoma cell lines (HSC-2, HSC-4, and Ca9-22), and lower effects on human oral normal cells (HGF, HPC, and HPLF). Tellimagrandin II (9) had the highest tumor-specific cytotoxicity, and also cleaved poly (ADP-ribose) polymerase 1 in HSC-2 cells. These findings showed that L. inermis ellagitannins may be a candidate for the production of anti-oral cancer materials.

Ethyl acetate extract of Ceiba pentandra (L.) Gaertn. reduces methotrexate-induced renal damage in rats via antioxidant, anti-inflammatory, and antiapoptotic actions.

Methotrexate (MTX) is a chemotherapeutic agent and an immunosuppressant used to treat cancer and autoimmune diseases. However, its use is limited by its multi-organ toxicity, including nephrotoxicity, which is related to MTX-driven oxidative stress. Silencing oxidative stressors is therefore an important strategy in minimizing MTX adverse effects.Medicinal plants rich in phenolic compounds are probable candidates to overcome these oxidants. Herein, C. pentandra ethyl acetate extract showed powerful in vitro radical-scavenging potential (IC50 = 0.0716) comparable to those of the standard natural (ascorbic acid, IC50 = 0.045) and synthetic (BHA, IC50 = 0.056) antioxidants. The effect of C. pentandra ethyl acetate extract against MTX-induced nephrotoxicity in rats was evaluated by administering the extract (400 mg/kg/day) or the standard antioxidant silymarin (100 mg/kg/day) orally for 5 days before and 5 days after a single MTX injection (20 mg/kg, i.p.).C. pentandra showed slight superiorities over silymarin in restoring the MTX-impaired renal functions, with approximately twofold decreases in overall kidney function tests. C. pentandra also improved renal antioxidant capacity and reduced the MTX-induced oxidative stress. Moreover, C. pentandra inhibited MTX-initiated apoptotic and inflammatory cascades, and attenuated MTX-induced histopathological changes in renal tissue architecture.Phytochemical investigation of the extract led to the purification of the phenolics quercitrin (1), cinchonains 1a (2) and 1b (3), cis-clovamide (4), trans-clovamide (5), and glochidioboside (6); a structurally similar with many of the reported antioxidant and nephroprotective agents. In conclusion, these data demonstrate that C. pentandra exhibits nephroprotective effect against MTX-induced kidney damage via its antioxidant, antiapoptotic and anti-inflammatory mechanisms. TAXONOMY: Functional Disorder, Traditional Medicine, Herbal Medicine.

High-performance liquid chromatographic profile and 1H quantitative nuclear magnetic resonance analyses for quality control of a Xinjiang licorice extract.

Various pharmacological properties of Xinjiang licorice flavonoids have been reported recently. We have investigated constituents corresponding to distinct peaks on the high-performance liquid chromatography (HPLC) profile of a flavonoid-rich extract from licorice, and identified 13 flavonoids, including licochalcone A (1), licochalcone B (3), glabrone (4), and echinatin (5), by isolating them and then performing high-resolution electrospray ionization mass spectrometry and 1H nuclear magnetic resonance (NMR) spectral analyses. We then applied the 1H quantitative NMR (qNMR) method for analysis of major flavonoids, 1 and 3-5 in the extract. The 1H qNMR results were supported by 13C NMR analysis. The results demonstrated the utility of the combination of HPLC profiling and qNMR analyses for quality control of Xinjiang licorice. Additionally, we observed a moderate inhibitory effect of the most abundant constituent, licochalcone A (1), on acetylcholine esterase activity, suggesting utility as a seed for drug development.

1H Quantitative NMR analyses of ß-asarone and related compounds for quality control of Acorus rhizome herbal drugs in terms of the effects of their constituents on in vitro acetylcholine esterase activity.

We used quantitative nuclear magnetic resonance analyses to measure the contents of major constituents of Acorus rhizome materials used as herbal drugs. The inhibitory effects of crude n-hexane extracts and their individual constituents on in vitro acetylcholine esterase activity were evaluated. The crude extracts had unexpectedly weak inhibitory effects (46-64% inhibition at 1.0 mg/mL), despite the high content (46-64%) of ß-asarone, which independently had a potent effect (IC50 2.9 µM [0.61 µg/mL]). Further investigation revealed participation of eudesmin A, a lignan constituent, in the suppression of the inhibitory effect of ß-asarone.

Simultaneous Quantification of Ellagitannins and Related Polyphenols in Geranium thunbergii Using Quantitative NMR.

Compared to commonly employed liquid chromatography-based methods, quantitative nuclear magnetic resonance (qNMR) is a recently developed method for accurate quantification of natural compounds in extracts. The simultaneous quantification of ellagitannins and the related polyphenols of Geranium thunbergii were studied using qNMR after a short-term and long-term decoction. The qNMR fingerprint for quantifying ellagitannin was presented in this work. Geraniin was observed in the short-term decoction as a major component while corilagin was the major component of the long-term decoction. An aqueous acetone extract of G. thunbergii after long-term decoction was extracted with diethyl ether, ethyl acetate, and n-butanol. Corilagin was found as a major constituent in the ethyl acetate and n-butanol extracts. Furthermore, the contents of these polyphenols in G. thunbergii from six locations in Japan and three locations in China were quantified. The contents of geraniin and corilagin in G. thunbergii from Japan were higher than those from China. Our finding raised the possibility that qNMR can be effectively employed as a simple, accurate, and efficient method for quantification of ellagitannins in medicinal plants.

Antifungal and Ichthyotoxic Sesquiterpenoids from Santalum album Heartwood.

In our continuing study on a survey of biologically active natural products from heartwood of Santalum album (Southwest Indian origin), we newly found potent fish toxic activity of an n-hexane soluble extract upon primary screening using killifish (medaka) and characterized α-santalol and ß-santalol as the active components. The toxicity (median tolerance limit (TLm) after 24 h at 1.9 ppm) of α-santalol was comparable with that of a positive control, inulavosin (TLm after 24 h at 1.3 ppm). These fish toxic compounds including inulavosin were also found to show a significant antifungal effect against a dermatophytic fungus, Trichophyton rubrum. Based on a similarity of the morphological change of the immobilized Trichophyton hyphae in scanning electron micrographs between treatments with α-santalol and griseofulvin (used as the positive control), inhibitory effect of α-santalol on mitosis (the antifungal mechanism proposed for griseofulvin) was assessed using sea urchin embryos. As a result, α-santalol was revealed to be a potent antimitotic agent induced by interference with microtubule assembly. These data suggested that α-santalol or sandalwood oil would be promising to further practically investigate as therapeutic agent for cancers as well as fungal skin infections.

Ellagitannins of Davidia involucrata. I. Structure of Davicratinic Acid A and Effects of Davidia Tannins on Drug-Resistant Bacteria and Human Oral Squamous Cell Carcinomas.

We isolated a new ellagitannin, davicratinic acid A (5), together with four known ellagitannins, davidiin (1), granatin A (2), pedunculagin (3), and 3-O-galloylgranatin A (4), from an aqueous acetone extract of dried Davidia involucrata leaves. The known ellagitannins were identified based on spectroscopic data. The structure of davicratinic acid A (5), a monomeric ellagitannin possessing a unique, skew-boat glucopyranose core, was established based on spectroscopic data. Additionally, we examined the effects of several tannins with good yields from this plant on drug-resistant bacteria and human oral squamous cell carcinomas, and found that davidiin (1) exhibited the most potent antibacterial and antitumor properties among the tannins examined.

Ellagitannins, gallotannins, and gallo-ellagitannins from the galls of Tamarix aphylla.

Chromatographic separation of an aqueous acetone extract of the galls from Tamarix aphylla using gels resulted in isolation of an ellagitannin, phyllagallin M1 (13), a gallo-ellagitannin, phyllagallin D1 (14), and four gallotannins, phyllagallin M2 (15) and phyllagallins D2-D4 (16-18), in addition to four known ellagitannins and three phenolics of lower molecular weight structurally related to hydrolyzable tannins. The structures of the six new tannins were elucidated based on spectroscopic and chemical data. Among the phenolics, flavogallonic acid dilactone (8), which is presumed to be biogenetically produced by C-C oxidative coupling of an ellagic acid unit with a galloyl residue, shows an exceptional oxidative pattern of gallic acid residues in plants of the family Tamaricaceae. Although the ellagitannin tamarixellagic acid (4) was reported to be a constituent of the galls of T. aphylla, such compounds with anomalous location of the DHDG moiety at O-3 on the glucopyranose core have not been observed among the tannins of tamaricaceous plants.

Action mechanism of 6, 6'-dihydroxythiobinupharidine from Nuphar japonicum, which showed anti-MRSA and anti-VRE activities.

BACKGROUND: Multidrug-resistant bacteria, such as methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin resistant enterococci (VRE), cause serious infections at clinical sites, for which the development of new drugs is necessary. We screened candidates for new antibiotics and investigated its action mechanism. METHODS: An antimicrobial compound was isolated from an extract of Nuphar japonicum. Its chemical structure was determined by NMR, MS, and optical rotation. We measured its minimum inhibitory concentration (MIC) using the microdilution method. The effects of the compound on DNA gyrase and DNA topoisomerase IV were investigated with DNA supercoiling, decatenation, and cleavage assay. RESULTS: We isolated and identified 6,6'-dihydroxythiobinupharidine as the antimicrobial compound. The MIC of this compound was 1-4 µg/mL against various MRSA and VRE strains. We also demonstrated that this compound inhibited DNA topoisomerase IV (IC50 was 10-15 µM), but not DNA gyrase in S. aureus, both of which are known to be the targets of quinolone antibiotics and necessary for DNA replication. However, this compound only exhibited slight cross-resistance to norfloxacin-resistant S. aureus, which indicated that DTBN might inhibit other targets besides topoisomerase IV. These results suggest that 6,6'-dihydroxythiobinupharidine may be a potent candidate or seed for novel antibacterial agents. CONCLUSIONS: DTBN from N. japonicum showed anti-MRSA and anti-VRE activities. DTBN might be involved in the inhibition of DNA topoisomerase IV. GENERAL SIGNIFICANCE: DTBN might be useful as a seed compound. The information on the inhibition mechanism of DTBN will be useful for the modification of DTBN towards developing novel anti-MRSA and anti-VRE drug.

Structures of new phenolics isolated from licorice, and the effectiveness of licorice phenolics on vancomycin-resistant Enterococci.

Licorice, which is the underground part of Glycyrrhiza species, has been used widely in Asian and Western countries as a traditional medicine and as a food additive. Our continuous investigation on the constituents of roots and stolons of Glycyrrhiza uralensis led to the isolation of two new phenolics, in addition to 14 known compounds. Structural studies including spectroscopic and simple chemical derivatizations revealed that both of the new compounds had 2-aryl-3-methylbenzofuran structures. An examination of the effectiveness of licorice phenolics obtained in this study on vancomycin-resistant strains Enterococcus faecium FN-1 and Enterococcus faecalis NCTC12201 revealed that licoricidin showed the most potent antibacterial effects against both of E. faecalis and E. faecium with a minimum inhibitory concentration (MIC) of 1.9 × 10-5 M. 8-(γ,γ-Dimethylallyl)-wighteone, isoangustone A, 3'-(γ,γ-dimethylallyl)-kievitone, glyasperin C, and one of the new 3-methyl-2-phenylbenzofuran named neoglycybenzofuran also showed potent anti-vancomycin-resistant Enterococci effects (MIC 1.9 × 10-5-4.5 × 10-5 M for E. faecium and E. faecalis). The HPLC condition for simultaneous detection of the phenolics in the extract was investigated to assess the quality control of the natural antibacterial resource, and quantitative estimation of several major phenolics in the extract with the established HPLC condition was also performed. The results showed individual contents of 0.08%-0.57% w/w of EtOAc extract for the major phenolics in the materials examined.

Structures of two new flavonoids and effects of licorice phenolics on vancomycin-resistant Enterococcus species.

Since our previous study revealed that several licorice phenolics have antibacterial effects on methicillin-resistant Staphylococcus aureus (MRSA), and suppressive effects on the oxacillin resistance of MRSA, we further investigated effectiveness of licorice constituents on vancomycin-resistant Enterococcus (VRE) bacteria, and purified 32 phenolic compounds. Two flavonoids among them were characterized structurally, and identified their structures as demethylglycyrol (31) and 5,7-di-O-methylluteone (32), respectively. Examination of antibacterial effects of licorice phenolics showed that 3-arylcoumarins such as licoarylcoumarin (9) and glycycoumarin (26), and 2-arylcoumarones such as gancaonin I (17), have moderate to potent antibacterial effects on the VRE strains used in this study.

Polyphenolic constituents of Cynomorium songaricum Rupr. and antibacterial effect of polymeric proanthocyanidin on methicillin-resistant Staphylococcus aureus.

Oligomeric and polymeric flavan-3-ols were obtained by chromatographic fractionation of extracts from Cynomorium songaricum Rupr. The structure of the polymeric constituent, cynomoriitannin, was characterized using spectral and chemical data. Results from acid-catalyzed degradation indicated that cynomoriitannin is a polymeric proanthocyanidin predominantly composed of epicatechin, together with low proportions of epicatechin-3-O-gallate and catechin as extension units. The terminal unit was chiefly composed of catechin, with an admixture of epicatechin. Size exclusion chromatographic analysis demonstrated a mean polymerization degree of 14. Two new phloroglucinol adducts (cynomoriitannin-phloroglucinol adducts A and B) obtained by acid-catalyzed degradation of cynomoriitannin in the presence of phloroglucinol were characterized using spectral analyses. Six oligomeric flavan-3-ols were also identified as follows: procyanidin B3, catechin-(6'-8)-catechin, catechin-(6'-6)-catechin, epicatechin-(4ß-8)- epicatechin-(4ß-8)-catechin, epicatechin-(4ß-6)-epicatechin-(4ß-8)-catechin, and arecatannin A1, respectively. These flavan-3-ols were isolated from C. songaricum. This is the first time that this procedure has been described. The antibacterial activity of the fractions and constituents was tested against methicillin-resistant Staphylococcus aureus (MRSA). The crude acetone-water (7:3) extract had moderate activity against MRSA. Cynomoriitannin was the most effective of the plant constituents against MRSA.

In vivo anti-inflammatory and antioxidant properties of ellagitannin metabolite urolithin A.

Urolithin A is a major metabolite produced by rats and humans after consumption of pomegranate juice or pure ellagitannin geraniin. In this study, we investigated the anti-inflammatory effect of urolithin A on carrageenan-induced paw edema in mice. The volume of paw edema was reduced at 1h after oral administration of urolithin A. In addition, plasma in treated mice exhibited significant oxygen radical antioxidant capacity (ORAC) scores with high plasma levels of the unconjugated form at 1h after oral administration of urolithin A. These results indicate strong associations among plasma urolithin A levels, the plasma ORAC scores, and anti-inflammatory effects and may help explain a mechanism by which ellagitannins confer protection against inflammatory diseases.

In vitro photochemical and phototoxicological characterization of major constituents in St. John's Wort (Hypericum perforatum) extracts.

Extracts from St. John's Wort (SJW: Hypericum perforatum) have been used for the treatment of mild-to-moderate depression. In spite of the high therapeutic potential, orally administered SJW sometimes causes phototoxic skin responses. As such, the present study aimed to clarify the phototoxic mechanisms and to identify the major phototoxins of SJW extract. Photobiochemical properties of SJW extract and 19 known constituents were characterized with focus on generation of reactive oxygen species (ROS), lipid peroxidation, and DNA photocleavage, which are indicative of photosensitive, photoirritant, and photogenotoxic potentials, respectively. ROS assay revealed the photoreactivity of SJW extract and some SJW ingredients as evidenced by type I and/or II photochemical reactions under light exposure. Not all the ROS-generating constituents caused photosensitized peroxidation of linoleic acid and photodynamic cleavage of plasmid DNA, and only hypericin, pseudohypericin, and hyperforin exhibited in vitro photoirritant potential. Concomitant UV exposure of quercitrin, an SJW component with potent UV/Vis absorption, with hyperforin resulted in significant attenuation of photodynamic generation of singlet oxygen from hyperforin, but not with hypericin. In conclusion, our results suggested that hypericin, pseudohypericin, and hyperforin might be responsible for the in vitro phototoxic effects of SJW extract.

Effects of mace and nutmeg on human cytochrome P450 3A4 and 2C9 activity.

Pharmacokinetic or pharmacodynamic interactions between herbal medicines or food constituents and drugs have been studied as crucial factors determining therapeutic efficacy and outcome. Most of these interactions are attributed to inhibition or induction of activity of cytochrome P450 (CYP) metabolic enzymes. Inhibition or induction of CYP enzymes by beverages, including grapefruit, pomegranate, or cranberry juice, has been well documented. Because spices are a common daily dietary component, other studies have reported inhibition of CYP activity by spices or their constituents/derivatives. However, a systematic evaluation of various spices has not been performed. In this study, we investigated effects of 55 spices on CYP3A4 and CYP2C9 activity. Cinnamon, black or white pepper, ginger, mace, and nutmeg significantly inhibited CYP3A4 or CYP2C9 activity. Furthermore, bioassay-guided fractionation of mace (Myristica fragrans) led to isolation and structural characterization of a new furan derivative (1) along with other 16 known compounds, including an acylphenol, neolignans, and phenylpropanoids. Among these isolates, (1S,2R)-1-acetoxy-2-(4-allyl-2,6-dimethoxyphenoxy)-1-(3,4-dimethoxyphenyl)propane (9) exhibited the most potent CYP2C9 inhibitory activity with an IC50 value comparable to that of sulfaphenazole, a CYP2C9 inhibitor. Compound 9 competitively inhibited CYP2C9-mediated 4'-hydroxylation of diclofenac. The inhibitory constant (K(i)) of 9 was determined to be 0.037 µM. Compound 9 was found to be 14-fold more potent than was sulfaphenazole.

[Antifungal activity of essential oils and their constituents against Candida spp. and their effects on activity of amphotericin B].

Candidiasis is a common opportunistic fungal infection that responds well to amphotericin B (AMPH) treatment. However, AMPH often causes adverse effects such as kidney injury and hypokalemia. Because some essential oils have been reported to have antifungal effects, we investigated the antifungal activity of various essential oils and their major constituents against Candida spp. Most essential oils examined in this study showed antifungal activity, and several enhanced the antifungal effect of AMPH. Clove oil in particular, and its major constituent eugenol, had potent effects. These findings suggest that combining certain essential oils or their constituents with AMPH may be useful for suppressing the adverse effects of AMPH treatment.