Natural Deep Eutectic Solvents for the Extraction of Spilanthol from Acmella oleracea (L.) R.K.Jansen.

With a growing focus on green chemistry, the extraction of natural products with natural deep eutectic solvents (NADES), which are eutectic mixtures of hydrogen bond donors and acceptors, has become an ever-expanding field of research. However, the use of NADES for the extraction of spilanthol from Acmella oleracea (L.) R.K.Jansen has not yet been investigated. Therefore, in this study, 20 choline chloride-based NADES, and for comparison, ethanol, were used as green extraction agents for spilanthol from Acmella oleracea flower heads. The effects of time, water addition, and temperature on NADES extractions were investigated and analysed by HPLC-DAD quantification. Additionally, UHPLC-DAD-ESI-MSn results for dichloromethane extracts, as well as the isolation of spilanthol and other main constituents as reference compounds, are reported. The best green extraction results were achieved by choline chloride (ChCl) with 1,2-propanediol (P, 1:2 molar ratio, +20% water) at 244.58 µg/mL, comparable to yields with ethanol (245.93 µg/mL). Methylurea (MeU, 1:2, +20% water) also showed promising results as a hydrogen bond donor in combination with choline chloride (208.12 µg/mL). In further experiments with NADES ChCl/P (1:2) and ChCl/MeU (1:2), extraction time had the least effect on spilanthol extraction with NADES, while yield decreased with water addition over 20% and increased with extraction temperature up to 80 °C. NADES are promising extraction agents for the extraction of spilanthol, and these findings could lead to applicable extracts for medicinal purposes, due to their non-toxic constituents.

Diverse diterpenoids and a triterpenoid from Euphorbia spinidens Bornm. ex Prokh.

Four previously unreported diterpenoids including three ent-atisanes (1-3) and one ent-abietane (4), along with one known linear triterpenoid (5) and five known diterpenoids including four myrsinanes (6-9), and one abietane (10) have been isolated from the roots of Euphorbia spinidens Bornm. ex Prokh. The structures were determined on the basis of extensive spectroscopic analyses including HR-ESI-MS, 1D and 2D NMR and comparison of the data with those reported in the literature. Antimicrobial potential of isolated compounds were also evaluated. Guionianol B (10) showed good antibacterial activity against Staphylococcus aureus and Bacillus subtilis with MIC value of 6.25 µg/mL.

Structures and bioactivities of monomeric and dimeric carvotacetones from Sphaeranthus africanus.

Four previously undescribed carvotacetones including one monomeric (1) and three dimeric (8, 9, 10) derivatives, together with six known compounds were isolated from the n-hexane extract of the aerial parts of Sphaeranthus africanus L. The structures of the previously undescribed compounds were elucidated as 3-angeloyloxy-5-isobutanoyloxy-7-hydroxycarvotacetone (1), 7,7'-oxybis{3-angeloyloxy-5-[(2R*,3R*)-2,3-dihydroxy-2-methylbutanoyloxy]carvotacetone} (8), (2″S*,3″R*)-7-{3-angeloyloxy-5-[(2R*,3R*)-2,3-dihydroxy-2-methylbuta-noyloxy]carvotaceton-7-yloxy}-3-angeloyloxy-5-(2,3-dihydroxy-2-methylbutanoyloxy)carvo-tacetone (9), and 7,7'-oxybis{3-angeloyloxy-5-[(2S*,3R*)-2,3-dihydroxy-2-methylbutanoyl-oxy]carvotacetone} (10). The three dimeric derivatives (8-10) showed potent anti-proliferative activity against human cancer cell lines (CCRF-CEM, MDA-MB-231, U-251, HCT-116) with IC50 values ranging from 0.2 to 2.0 µM. Caspases 3 and 7 were found to be activated by all compounds, indicating apoptosis induction activity. Monomers exhibited a specific inhibition of NO production in BV2 and RAW 264.7 cells with IC50 values ranging from 4.2 to 6.8 µM which were 2-3.5-fold lower than IC50 values causing cytotoxicity. In addition, the carvotacetones reduced NF-κB1 (p105) mRNA expression at concentrations of 10 and 2.5 µM. Altogether, the results indicate that carvotacetones may be interesting lead structures for the development of anti-cancer and anti-inflammatory drugs.

Cycloartane Saponins from Astragalus glycyphyllos and Their In Vitro Neuroprotective, Antioxidant, and hMAO-B-Inhibiting Effects.

Astragalus glycyphyllos (Fabaceae) is used in the traditional medicine of many countries against hepatic and cardiac disorders. The plant contains mainly flavonoids and saponins. From a defatted methanol extract from its overground parts, a new triterpenoid saponin, 3-O-[α-L-rhamnopyranosyl-(1→2)]-ß-D-xylopyranosyl]-24-O-α-L-arabinopyranosyl-3ß,6α,16ß,24(R),25-pentahydroxy-20R-cycloartane, together with the rare saponin astrachrysoside A, were isolated using various chromatography methods. The compounds were identified via extensive high resolution electrospray ionisation mass spectrometry (HRESIMS) and NMR analyses. Both saponins were examined for their possible antioxidant and neuroprotective activity in three different in vitro models. Rat brain synaptosomes, mitochondria, and microsomes were isolated via centrifugation using Percoll gradient. They were treated with the compounds in three different concentrations alone, and in combination with 6-hydroxydopamine or tert-butyl hydroperoxide as toxic agents. It was found that the compounds had statistically significant dose-dependent in vitro protective activity on the sub-cellular fractions. The compounds exhibited a weak inhibitory effect on the enzyme activity of human recombinant monoamine oxidase type B (hMAO-B), compared to selegiline.

Discrimination of Zicao Samples Based on DNA Barcoding and HPTLC Fingerprints, and Identification of (22E)-Ergosta-4,6,8(14),22-tetraen-3-one As a Marker Compound.

The unambiguous identification of plant material is a prerequisite of rational phytotherapy. Misidentification can even cause serious health problems, as in the case of the Chinese medicinal herb Zicao. Commercial material labelled "Zicao" may be derived from the roots of Arnebia euchroma (ruan zicao), Lithospermum erythrorhizon (ying zicao), or Onosma paniculata (dian zicao). All of these roots contain shikonin derivatives as main bioactive constituents, but ying zicao and dian zicao contain also hepatotoxic pyrrolizidine alkaloids in high amounts. Therefore, the use of A. euchroma with a very low pyrrolizidine alkaloid content is desirable. Confusions of the species occur quite often, indicating an urgent need for an unambiguous identification method. Discrimination of 23 zicao samples has been achieved by analyses of the nuclear internal transcribed spacer ITS2 and trnL-F intergenic spacer of the chloroplast DNA. Data were analyzed using Bioedit, ClustalX, Mega 11 and BLAST. Results indicate that ITS2 barcoding can accurately distinguish Arnebia euchroma from their adulterants. Subsequently, an HPTLC method has been developed allowing a chemical discrimination of the most widely used species. (22E)-Ergosta-4,6,8(14),22-tetraen-3-one has been identified as characteristic marker compound, allowing an unambiguous discrimination of A. euchroma and L. erythrorhizon.

Characterization of Constituents with Potential Anti-Inflammatory Activity in Chinese Lonicera Species by UHPLC-HRMS Based Metabolite Profiling.

This study centered on detecting potentially anti-inflammatory active constituents in ethanolic extracts of Chinese Lonicera species by taking an UHPLC-HRMS-based metabolite profiling approach. Extracts from eight different Lonicera species were subjected to both UHPLC-HRMS analysis and to pharmacological testing in three different cellular inflammation-related assays. Compounds exhibiting high correlations in orthogonal projections to latent structures discriminant analysis (OPLS-DA) of pharmacological and MS data served as potentially activity-related candidates. Of these candidates, 65 were tentatively or unambiguously annotated. 7-Hydroxy-5,3',4',5'-tetramethoxyflavone and three bioflavonoids, as well as three C32- and one C34-acetylated polyhydroxy fatty acid, were isolated from Lonicera hypoglauca leaves for the first time, and their structures were fully or partially elucidated. Of the potentially active candidate compounds, 15 were subsequently subjected to pharmacological testing. Their activities could be experimentally verified in part, emphasizing the relevance of Lonicera species as a source of anti-inflammatory active constituents. However, some compounds also impaired the cell viability. Overall, the approach was found useful to narrow down the number of potentially bioactive constituents in the complex extracts investigated. In the future, the application of more refined concepts, such as extract prefractionation combined with bio-chemometrics, may help to further enhance the reliability of candidate selection.

Targeting fish spoilers Pseudomonas and Shewanella with oregano and nettle extracts.

To control Pseudomonas and Shewanella as important psychrotrophic spoilage bacteria in fish meat, we used ethanolic extracts of oregano (Origanum vulgare subsp. vulgare) and nettle (Urtica dioica), with phytochemical characterisation of the extracts and their bioactive compounds. Liquid chromatography coupled with photodiode array detection and electrospray ionisation-mass spectrometry was used for qualitative compositional determination of the extracts. Four main compounds were identified in the oregano extract, with rosmarinic acid the most abundant, followed by three glycosylated phenolics, one of which is reported for the first time in O. vulgare: 4'-O-ß-d-glucopyranosyl-3',4'-dihydroxybenzyl-4-hydroxybenzoate. Six main compounds were identified in the nettle extract, as caffeoylmalic acid and five flavonoid glycosides. These oregano and nettle ethanolic extracts showed in-vitro antimicrobial activities against selected Pseudomonas and Shewanella strains in broth and fish meat homogenate when evaluated at two inoculum concentrations. The antimicrobial activities were more pronounced for the nettle extract at the lower inoculum concentration, and for both the Shewanella strains. Growth inhibition in the fish meat homogenate was evaluated at 3.13 mg/mL and 1.56 mg/mL at 5 °C. Again, the nettle extract showed greater antimicrobial activity, which was seen as the lowest maximum growth rate, followed by the oregano extract, which was inhibitory only at 3.13 mg/mL. Finally, the extracts were applied to fish meat that was then stored at 5 °C for 9 days. Evaluation here was for the counts of the mesophilic, psychrotrophic, Pseudomonas and H2S producers. These confirmed the better antimicrobial effects of the nettle extract, especially against the H2S-producing bacteria, which included Shewanella. Both of the extracts were rich in glycosides of flavonoids and phenolic acids. The enzymatic activities of the Pseudomonas and Shewanella spoilage bacteria and their actions on the phenolic glycosides from natural sources will be further investigated.

Polyacetylenes from Oplopanax horridus and Panax ginseng: Relationship between Structure and PPARγ Activation.

Oplopanax horridus and Panax ginseng are members of the plant family Araliaceae, which is rich in structurally diverse polyacetylenes. In this work, we isolated and determined structures of 23 aliphatic C17 and C18 polyacetylenes, of which five are new compounds. Polyacetylenes have a suitable scaffold for binding to PPARγ, a ligand-activated transcription factor involved in metabolic regulation. Using a reporter gene assay, their potential was investigated to activate PPARγ. The majority of the polyacetylenes showed at least some PPARγ activity, among which oplopantriol B 18-acetate (1) and oplopantriol B (2) were the most potent partial PPARγ activators. By employing in silico molecular docking and comparing the activities of structural analogues, features are described that are involved in PPARγ activation, as well as in cytotoxicity. It was found that the type of C-1 to C-2 bond, the polarity of the terminal alkyl chain, and the backbone flexibility can impact bioactivity of polyacetylenes, while diol structures with a C-1 to C-2 double bond showed enhanced cytotoxicity. Since PPARγ activators have antidiabetic and anti-inflammatory properties, the present results may help explain some of the beneficial effects observed in the traditional use of O. horridus extracts. Additionally, they might guide the polyacetylene-based design of future PPARγ partial agonists.

Phenolic compounds of Iris adriatica and their antimycobacterial effects.

Little is known about the pharmacological activities of Iris adriatica (Iridaceae), a plant endemic to Dalmatia (Croatia). We therefore performed a bioassay-guided fractionation including high-performance counter current chromatography (HPCCC) and antibacterial tests using Mycobacterium smegmatis mc2 155. One obtained fraction was found to be antimycobacterially active with a MIC of 64 mg L-1. Furthermore, fractions were tested for resistance modulatory effects using ethidium bromide as substrate. We were able to identify the pure isoflavonic compounds irigenin and irilone and a fraction containing mainly benzophenone 2,4,6-trihydroxy-4-methoxy-benzophenone, responsible for the resistance-modulatory activity of this plant.

Anti-inflammatory and antiproliferative compounds from Sphaeranthus africanus.

BACKGROUND: Sphaeranthus africanus has been used in traditional Vietnamese medicine to treat sore throat, and to relieve pain and swelling. However, the anti-inflammatory activity of this plant had not yet been investigated. Previously, we isolated five carvotacetones (1-5) from this plant that displayed cytotoxicity against several cancer cell lines. PURPOSE: The objective of this study was to isolate further constituents from S. africanus and to investigate the anti-inflammatory activity of all constituents. Furthermore, the anti-proliferative activity of the newly isolated compounds was evaluated. STUDY DESIGN AND METHODS: Compounds were isolated from the upper parts of S. africanus by chromatographic methods. Structures were determined using spectroscopic techniques, like NMR and MS. All nine compounds isolated from S. africanus were evaluated for inhibitory activity against COX-1 and COX-2 isoenzymes in-vitro, COX-2 mRNA expression and influence on NO production. The anti-proliferative activities of newly isolated compounds (6-9) were evaluated by XTT viability assay with four cancer cell lines, namely CCRF-CEM, MDA-MB-231, HCT-116, and U-251 cells. RESULTS: Two diastereomeric carvotacetones (3-angeloyloxy-5-[2″S,3″R-dihydroxy-2″-methyl-butanoyloxy]-7-hydroxycarvotacetone (6) and 3-angeloyloxy-5-[2″R,3″R-dihydroxy-2″-methyl-butanoyloxy]-7-hydroxycarvotacetone (7), asperglaucide (8) and chrysoplenol D (9) were isolated from S. africanus. COX-1 and COX-2 assays of compounds 1-9 revealed that compounds 1 and 2 possess potent and selective COX-2 inhibitory activity with IC50 values of 3.6 and 0.5 µM, respectively. COX-2 gene expression assay showed that some carvotacetones exhibited inhibitory effects on COX-2 gene expression in THP-1 macrophages. Compound 4 is the most active compound inhibiting the synthesis of COX-2 by 55% at 2.06 µM. In the iNOS assay, all seven carvotacetones inhibited NO production in BV2 and RAW cell lines with IC50 values ranging from 0.2 to 2.9 µM. Compound 4 showed potent inhibitory activity with IC50 values of 0.2 µM in both BV2 and RAW cell lines. Molecular docking studies revealed the binding orientations of 1 and 2 in the active sites of COX-2. XTT assay of the newly isolated compounds revealed that the two isomeric carvotacetones (6-7) exhibited considerable anti-proliferative activity against four cancer cell lines (CCRF-CEM, MDA-MB-231, HCT-116, U-251) with IC50 values ranging from 1.23 to 8 µM. CONCLUSION: For the first-time, the diastereomeric carvotacetones (6-7) were isolated as separate compounds, and their anti-proliferative activity was determined. Selective COX-2 inhibitory, COX-2 mRNA expression and NO production inhibitory activities by some of the major constituents of S. africanus supports the traditional medical application of this plant for the treatment of inflammation-related disorders.

Periplocin, the most anti-proliferative constituent of Periploca sepium, specifically kills liposarcoma cells by death receptor mediated apoptosis.

BACKGROUND: During a screening of Chinese plants traditionally used for the treatment of cancer and related diseases, extracts of the root bark of Periploca sepium Bunge showed strong cytotoxic activity. PURPOSE: Isolate and identify cytotoxic compounds from P. sepium and investigate the effects and mechanism of action on different cancer cell lines. METHODS: Extracts obtained with solvents of different polarities of the root bark of P. sepium were tested for their anti-proliferative effects. The most active extract was subjected to activity-guided fractionation using different chromatographic methods. The most active compound was further investigated on sarcoma cell lines regarding its effects concerning apoptosis, DNA damage and death receptor expression. RESULTS: We isolated the cardiac glycosides periplocin, glucosyl divostroside, periplogenin, periplocymarin and periplocoside M with periplocin exhibiting the lowest IC50 value against leukemia and liposarcoma cells. Liposarcomas are rare tumors within the heterogeneous group of soft tissue sarcomas and respond poorly to conventional treatments. Periplocin led to growth inhibition and apoptosis induction by changing the expression of death receptors and inducing DNA double strand breaks in SW-872 cells. CONCLUSION: Periplocin displays a promising mechanism of action in sarcoma cells because altering the death receptor expression is an interesting target in sarcoma treatment especially to overcome TRAIL resistance.

Bupleurum chinense Roots: a Bioactivity-Guided Approach toward Saponin-Type NF-κB Inhibitors.

The roots of Bupleurum chinense have a long history in traditional medicine to treat infectious diseases and inflammatory disorders. Two major compounds, saikosaponins A and D, were reported to exert potent anti-inflammatory activity by inhibiting NF-κB. In the present study, we isolated new saikosaponin analogues from the roots of B. chinese interfering with NF-κB activity in vitro. The methanol-soluble fraction of the dichloromethane extract of Radix Bupleuri was subjected to activity-guided isolation yielding 18 compounds, including triterpenoids and polyacetylenes. Their structures were determined by spectroscopic methods as saikogenin D (1), prosaikogenin D (2), saikosaponins B2 (3), W (4), B1 (5), Y (6), D (7), A (8), E (9), B4 (10), B3 (11), and T (12), saikodiyne A (13), D (14), E (15) and F (16), falcarindiol (17), and 1-linoleoyl-sn-glycero-3-phosphorylcholine (18). Among them, 4, 15, and 16 are new compounds, whereas 6, previously described as a semi-synthetic compound, is isolated from a natural source for the first time, and 13-17 are the first reports of polyacetylenes from this plant. Nine saponins/triterpenoids were tested for inhibition of NF-κB signaling in a cell-based NF-κB-dependent luciferase reporter gene model in vitro. Five of them (1, 2, 4, 6, and 8) showed strong (> 50%, at 30 µM) NF-κB inhibition, but also varying degrees of cytotoxicity, with compounds 1 and 4 (showing no significant cytotoxicity) presenting IC50 values of 14.0 µM and 14.1 µM in the cell-based assay, respectively.

Efficient identification of flavones, flavanones and their glycosides in routine analysis via off-line combination of sensitive NMR and HPLC experiments.

We present a standardized, straightforward and efficient approach applicable in routine analysis of flavonoids combining sensitive NMR and HPLC experiments. The determination of the relative configuration of sugar moieties usually requires the acquisition of 13C NMR shift values. We use a combination of HPLC and sensitive NMR experiments (1D-proton, 2D-HSQC) for the unique identification of known flavones, flavanones, flavonols and their glycosides. Owing to their broad range of polarity, we developed HPLC and UHPLC methods (H2O/MeOH/MeCN/HCOOH) which we applied and validated by analyzing 46 common flavones and flavanones and exemplified for four plant extracts. A searchable data base is provided with full data comprising complete proton and carbon resonance assignments, expansions of HSQC-spectra, HPLC parameters (retention time, relative retention factor), UV/Vis and mass spectral data of all compounds, which enables a rapid identification and routine analysis of flavones and flavanones from plant extracts and other products in nutrition and food chemistry.

Two Novel Spirostene Glycosides from Selaginella chrysocaulos and their Chemotaxonomic Significance.

During an investigation of the Indian heterosporous fern Selaginella chrysocaulos, two novel C28 spirostene monosides, chrysocauloside A and B were identified. Chrysocauloside A (1ß,3ß-dihydroxy-20S,22R-spirost-5-ene-1-yl ß-D-glucopyranoside) and chrysocauloside B (1ß,3ß-dihydroxy-20S,22R-spirost-5-ene-1-yl ß-D-galactopyranoside) are O-glycosylated at C-1 and each bear a methyl group at C-24 and C-25. Spectroscopic (NMR) and MS data for the new compounds are given and the taxonomic significance of the occurrence of spirostenes in the group of Lycophyta is discussed.

Aldose reductase inhibition of a saponin-rich fraction and new furostanol saponin derivatives from Balanites aegyptiaca.

BACKGROUND: Balanites aegyptiaca Del. (Zygophyllaceae) fruits are used to treat hyperglycemia in Egyptian folk medicine and are sold by herbalists in the Egyptian open market for this purpose. Nevertheless, the fruits have not yet been incorporated into pharmaceutical dosage forms. The identity of the bioactive compounds and their possible mechanisms of action were not well understood until now. PURPOSE: Aldose reductase inhibitors are considered vital therapeutic and preventive agents to address complications caused by hyperglycemia. The present study was carried out to identify the primary compounds responsible for the aldose reductase inhibitory activity of Balanites aegyptiaca fruits. STUDY DESIGN: The 70% ethanolic extract of Balanites aegyptiaca fruit mesocarp and its fractions were screened for inhibition of the aldose reductase enzyme. Bio-guided fractionation of the active butanol fraction was performed and the primary compounds present in the saponin-rich fraction (D), which were responsible for the inhibitory activity, were characterized. HPLC chromatographic profiles were established for the different fractions, using the isolated compounds as biomarkers. METHODS: Aldose reductase inhibition was tested in vitro on rat liver homogenate. The butanol fraction of the 70% ethanolic extract was fractionated using vacuum liquid chromatography (VLC, RP-18 column). The most active sub-fraction D, which was eluted with 75% methanol, was subjected to preparative HPLC to isolate the bioactive compounds. RESULTS: The butanol fraction displayed inhibitory activity against the aldose reductase enzyme (IC50 = 55.0 ± 6 µg/ml). Sub-fraction D exhibited the highest inhibitory activity (IC50 = 12.8 ± 1 µg/ml). Five new steroidal saponin derivatives were isolated from this fraction. The isolated compounds were identified as compound 1a/b, a 7:3 mixture of the 25R:25S epimers of 26-O-ß-D-glucopyranosyl-furost-5-ene-3,22,26-triol 3-O-[α-L-rhamnopyranosyl-(1→3)- ß-D-glucopyranosyl-(1→2)]- α-L-rhamnopyranosyl-(1→4)-ß-D-glucopyranoside; compound 2, 26-O-ß-D-glucopyranosyl-(25R)-furost-5-ene-3,22,26-triol 3-O-[ ß-D-glucopyranosyl-(1→2)]- α-L-rhamnopyranosyl-(1→4)-ß-D-glucopyranoside; compound 3, 26-O-ß-D-glucopyranosyl-(25R)-furost-5,20-diene-3,26-diol 3-O-[α-L-rhamnopyranosyl-(1→3)- ß-D-glucopyranosyl-(1→2)]- α-L-rhamnopyranosyl-(1→4)-ß-D-glucopyranoside; compound 4, 26-O-ß-D-glucopyranosyl-(25R)-furost-5,20-diene-3,26-diol 3-O-[ ß-D-glucopyranosyl-(1→2)]- α-L-rhamnopyranosyl-(1→4)-ß-D-glucopyranoside; and compound 5, which is the 25S epimer of compound 4, by using various spectroscopic methods [MS,1D and 2D NMR (HSQC, HMBC, DQF-COSY, HSQC-TOCSY)]. Compounds 1a/b, 2, 3, 4, 5 exhibited highly significant aldose reductase inhibitory activities (IC50 values were 1.9 ± 0.2, 1.3 ± 0.5, 5.6 ± 0.2, 5.1 ± 0.4, 5.1 ± 0.6 µM, respectively) as compared to the activity of the reference standard quercetin (IC50 = 6.6 ± 0.3 µM). CONCLUSION: The aldose reductase inhibitory activity of Balanites fruits is due to the steroidal saponins present. HPLC chromatographic profiles of the crude butanol fraction and its 4 sub-fractions showed that the most highly bioactive fraction D contained the highest amount of steroidal saponins (75%) as compared to the 21% present in the original butanol fraction. The isolated furostanol saponins proved to be highly active in an in vitro assay.

In vitro growth inhibition by Hypericum extracts and isolated pure compounds of Paenibacillus larvae, a lethal disease affecting honeybees worldwide.

The in vitro inhibitory potential of 50 extracts from various species of the flowering plant genus Hypericum was investigated using the Kirby-Bauer disk diffusion susceptibility test against Paenibacillus larvae, a spore-forming, Gram-positive bacterial pathogen that causes American foulbrood (AFB), a lethal disease affecting honeybee brood worldwide. Of the tested extracts, 14 were identified as highly active against P. larvae as compared to the activity of the positive control, indicating the presence of highly potent antibacterial compounds in the extracts. Examination of these extracts using TLC and HPLC/MS analyses revealed the presence of acylphloroglucinol and filicinic-acid derivatives. Six pure compounds isolated from these extracts, viz., hyperforin (1), uliginosin B (2), uliginosin A (3), 7-epiclusianone (4), albaspidin AA (5), and drummondin E (6), displayed strong antibacterial activity against the vegetative form of P. larvae (MIC ranging from 0.168-220 µM). Incubation of P. larvae spores with the lipophilic extract of Hypericum perforatum and its main acylphloroglucinol constituent 1 led to the observation of significantly fewer colony forming units as compared to the negative control, indicating that the acylphloroglucinol scaffold represents an interesting lead structure for the development of new AFB control agents.

Inhibition of NO production by Grindelia argentina and isolation of three new cytotoxic saponins.

A bioassay-guided phytochemical analysis of the ethanolic extract of Grindelia argentina Deble & Oliveira-Deble (Asteraceae) allowed the isolation of a known flavone, hispidulin, and three new oleanane-type saponins, 3-O-ß-D-xylopyranosyl-(1→3)-ß-D-glucopyranosyl-2ß,3ß,16α,23-tetrahydroxyolean-12-en-28-oic acid 28-O-ß-D-xylopyranosyl-(1→2)-ß-D-apiofuranosyl-(1→3)-ß-D-xylopyranosyl-(1→3)-α-L-rhamnopyranosyl-(1→2)-α-L-arabinopyranosyl ester (2), 3-O-ß-D-glucopyranosyl-2ß,3ß,23-trihydroxyolean-12-en-28-oic acid 28-O-ß-D-xylopyranosyl-(1→2)-ß-D-apiofuranosyl-(1→3)-ß-D-xylopyranosyl-(1→3)-α-L-rhamnopyranosyl-(1→2)-α-L-arabinopyranosyl ester, (3) and 3-O-ß-D-xylopyranosyl-(1→3)-ß-D-glucopyranosyl-2ß,3ß,23-trihydroxyolean-12-en-28-oic acid 28-O-ß-D-xylopyranosyl-(1→2)-ß-D-apiofuranosyl-(1→3)-ß-D-xylopyranosyl-(1→3)-α-L-rhamnopyranosyl-(1→2)-α-L-arabinopyranosyl ester (4), named grindeliosides A-C, respectively. Their structures were determined by extensive 1D- and 2D-NMR experiments along with mass spectrometry and chemical evidence. The isolated compounds were evaluated for their inhibitory activities against LPS/IFN-γ-induced NO production in RAW 264.7 macrophages and for their cytotoxic activities against the human leukemic cell line CCRF-CEM and MRC-5 lung fibroblasts. Hispidulin markedly reduced LPS/IFN-γ-induced NO production (IC50 51.4 µM), while grindeliosides A-C were found to be cytotoxic, with grindelioside C being the most active against both CCRF-CEM (IC50 4.2±0.1 µM) and MRC-5 (IC50 4.5±0.1 µM) cell lines.

Polyacetylenes from Radix et Rhizoma Notopterygii incisi with an inhibitory effect on nitric oxide production in vitro.

Notopterygium roots (Qiang Huo) have been used in traditional Chinese medicine for treating colds, inflammatory diseases like rheumatoid arthritis, and as an analgesic. The anti-inflammatory activity of the roots of Notopterygium incisum has been evaluated by testing the inhibitory activity on nitric oxide production by inducible nitric oxide synthase. The apparent authenticity of the sample was checked by DNA sequence comparison. Using activity-guided isolation, different compounds were isolated and structurally characterized by means of NMR and mass spectroscopy. Eight polyacetylenes could be identified and were tested on their inhibitory activity on nitric oxide production in RAW 264.7 mouse macrophages using the Griess assay. Different 3-hydroxy allyl polyacetylenes exhibited significant activity (IC50: 8-acetoxyfalcarinol, 20.1 µM; falcarindiol, 9.2 µM; 9-epoxyfalcarindiol, 8.8 µM; and crithmumdiol, 23.6 µM).

Accelerated sample preparation and formation of astragaloside IV in Astragali Radix.

Abstract Context: Astragali Radix (Huangqi; Astragalus mongholicus BUNGE, Fabaceae) is used in herbal medicinal products as well as in many food supplements. In traditional Chinese medicine, the roots are used for its Qi tonifying, immunostimulant, cardioprotective, hepatoprotective and hypoglycemic effects. Objective: Astragaloside IV (AGS-IV), a cycloartane-type triterpene glycoside is used as a marker compound for the quality control of Astragali Radix in various pharmacopoeias. Materials and methods: In this study, we analyzed the content of AGS-IV and other astragalosides in various commercial samples of Huangqi by reversed-phase HPLC using evaporative light scattering detection. Results: The analyses revealed that AGS-IV is formed during sample preparation from acylated astragalosides like astragaloside I and astragaloside II, when using the assay method of the European Pharmacopoeia. Discussion and conclusion: For consistent assay results, the extraction methods of the pharmacopoeias should be re-evaluated and optimized. Alternatively, the hydrolysis by ammonia could be omitted and the genuine compounds like astragaloside I, II and malonyl-AGS-I could be considered for the quality control of Astragali Radix.

Polyacetylenes from Notopterygium incisum--new selective partial agonists of peroxisome proliferator-activated receptor-gamma.

Peroxisome proliferator-activated receptor gamma (PPARγ) is a key regulator of glucose and lipid metabolism and therefore an important pharmacological target to combat metabolic diseases. Since the currently used full PPARγ agonists display serious side effects, identification of novel ligands, particularly partial agonists, is highly relevant. Searching for new active compounds, we investigated extracts of the underground parts of Notopterygium incisum, a medicinal plant used in traditional Chinese medicine, and observed significant PPARγ activation using a PPARγ-driven luciferase reporter model. Activity-guided fractionation of the dichloromethane extract led to the isolation of six polyacetylenes, which displayed properties of selective partial PPARγ agonists in the luciferase reporter model. Since PPARγ activation by this class of compounds has so far not been reported, we have chosen the prototypical polyacetylene falcarindiol for further investigation. The effect of falcarindiol (10 µM) in the luciferase reporter model was blocked upon co-treatment with the PPARγ antagonist T0070907 (1 µM). Falcarindiol bound to the purified human PPARγ receptor with a Ki of 3.07 µM. In silico docking studies suggested a binding mode within the ligand binding site, where hydrogen bonds to Cys285 and Glu295 are predicted to be formed in addition to extensive hydrophobic interactions. Furthermore, falcarindiol further induced 3T3-L1 preadipocyte differentiation and enhanced the insulin-induced glucose uptake in differentiated 3T3-L1 adipocytes confirming effectiveness in cell models with endogenous PPARγ expression. In conclusion, we identified falcarindiol-type polyacetylenes as a novel class of natural partial PPARγ agonists, having potential to be further explored as pharmaceutical leads or dietary supplements.