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International audience; The need for new anthelmintic with no chemical residues is becoming urgent. In a program aiming at the evaluation of plant as sources of new active molecules, the anthelmintic activities of the essential oils (EOs) obtained from either Zanthoxylum zanthoxyloides seeds or Newbouldia laevis leaves were evaluated against Strongyloides ratti by analyzing the results of two in vitro bioassays. These two plants and their tested parts were retained after an ethnopharmacology survey that confirmed their use by small-scale farmers for treatment of small ruminants affected by digestive helminths. The plants were harvested in Benin, and their EO were obtained by hydrodistillation. The EO yield of extraction was 0.65% (w/w) of for Z. zanthoxyloides seeds and 0.05% (w/w) for N. laevis. The chemical compositions of the two EOs were analyzed by gas chromatography coupled with mass spectrometry. The major constituents of the EO from Z. zanthoxyloides consisted of the following compounds: gamma-terpinene (18 %), undecane (15 %), valencene (8.3 %), decanal (8.3 %), and 3-carene (6.7 %). In contrast, the major constituents of the EO from N. laevis leaves consisted of the following compounds: beta-caryophyllene (36 %) and eugenol (5.8 %). An egg-hatching inhibition (EHI) assay was developed and a larval migration inhibition assay was used on S. ratti to examine the effects of the EOs and to evidence their inhibitory concentrations (IC50 and IC90) values on this nematode. Furthermore, the toxicity of the two EOs on Vero cell line was evaluated. When tested on S. ratti egg hatching, the two EOs resulted in similar IC50 values (19.5 and 18.2 mu g/ml for Z. zanthoxyloides and N. laevis, respectively), which were about sevenfold higher than that of the control (thiabendazole, IC50 = 2.5 mu g/ml). Larval migration was inhibited at similar concentrations for: Z. zanthoxyloides (IC50 = 46 mu g/ml), N. laevis (IC50 = 51 mu g/ml), and the control [levamisole (IC50 = 36 mu g/ml)]. No cytotoxicity was found on Vero cells because both EOs had IC50 values higher than 50 mu g/ml. Therefore, we have concluded that the EOs from two plants, used in folk medicine, may contain compounds with anthelmintic activity and could be used as improved traditional medicines or, at least, as food additives in a combined treatment for the control of helminth infections.

The aim of this work is the isolation of anti-leishmanial compounds from the ethyl acetate extracts of the bark of HEDYOSMUM ANGUSTIFOLIUM. We have successfully isolated and characterized five sesquiterpenes: one new compound (oxyonoseriolide, 1), one compound isolated for the first time from a natural source (hedyosmone, 2), and three known sesquiterpenes (onoseriolide, 3; chloranthalactone A, 4; and spathulenol, 5) that had not been previously isolated from H. ANGUSTIFOLIUM. The biological activities of 1- 5 showed that onoseriolide ( 3) was the most active compound against axenic amastigotes from LEISHMANIA AMAZONENSIS and L. INFANTUM. Moreover, it was still active on the intramacrophagic amastigotes of L. INFANTUM. The isolated compounds have also been tested on PLASMODIUM FALCIPARUM and against various mammalian cell lines.

The structure of the secretory tissues and the essential oil of the leaves of Croton campestris St Hil. were studied. The oil isolated by hydrodistillation was analyzed by GC and GC/MS and was found to contain 75 components accounting for 92.9% of the total oil, with the major components being caryophyllene oxide (29.9%) and humulene oxide II (8.0%). Growth inhibitory activity of the active compounds in solution was evaluated by measuring Minimal Inhibitory Concentration using a broth micro-method. The Minimal Inhibitory Concentration of the leaf oil was 1.56 µg/mL for Staphylococcus aureus, 6.25 µg/mL for Enterrococcus hirae and 6.25 µg/mL for Candida albicans.

Bolivianine, a novel sesterpene with an unprecedented skeleton, has been isolated from the trunk bark of Hedyosmum angustifolium (Chloranthaceae), with isobolivianine, an isomer formed under acidic conditions. The structure and relative stereochemistry were elucidated on the basis of spectroscopic data. A hypothesis for biogenesis was made.

Liquid chromatography (LC) coupled to negative electrospray ionisation (ESI) tandem mass spectrometry (MS/MS) was used for the rapid and sensitive identification of flavonoid compounds in Agauria salicifolia. The leaf flavonoid content in individual of A. salicifolia originating from population with contrasted ecogeographical situation and morphological characteristics was found to be variable qualitatively and highly variable quantitatively. Identification of the compounds was carried out by interpretation of UV, MS and MS/MS spectra. Fourteen flavonoids were identified, all of which had not previously been reported in Agauria spp. Two flavonol-O-glucuronides were found to differentiate the two populations.

Twelve naturally occurring glucosinolates displaying alkenyl, hydroxylated, methylsulphinyl, aromatic and indole side chains were investigated by both negative and positive ion electrospray ionisation-tandem mass spectrometry (ESI-MS/MS). In order to resolve the MS/MS spectra obtained from the anion and cation molecular ions of glucosinolates, the different fragments were investigated by MSn experiments using an ion trap spectrometer. The MS3 spectra obtained permitted possible fragmentation schemes to be proposed. These were supported by accurate mass measurements of some characteristic diagnostic ions with the help of a quadrupole time-of-flight instrument. The negative ion ESI-MS/MS behaviour of the different glucosinolates investigated in this study confirmed previously described patterns and revealed new interesting structural informative fragments. Some are common to all the glucosinolates and others are highly specific for a type of variable side chain. The positive ion ESI-MS/MS fragments obtained from the [MNa+Na]+ or [MK+K]+ molecular ions did not provide complementary specific diagnostic ions. Nevertheless, when compared with the negative ion mode, the daughter ions appeared more homogenous and with a better relative abundance for all of the 12 compounds studied. Moreover, the positive ion mode appeared to be more efficient than the negative mode for the study of methoxylated glucosinolates and should be useful to detect the glucosinolates present as organic salts in crude plant extracts.

A novel and very unusual azaanthracene alkaloid, 1-aza-7,8,9,10-tetramethoxy-4-methyl-2-oxo-1,2-dihydroanthracene ( 1) and a new diastereoisomer of the bis-benzylisoquinoline alkaloid rodiasine, 1 S,1' R-rodiasine ( 2), as well as the alkaloids O-methylpunjabine ( 3) and O-methylmoschatoline ( 4) have been isolated from Pseudoxandra cuspidata bark, used in French Guiana as an antimalarial. Their structures were elucidated by spectroscopic analyses, especially 2D-NMR techniques (ADEQUATE and NOESY). We found that the antimalarial activity of this bark was mostly due to bis-benzylisoquinoline 1 S,1' R-rodiasine ( 2) (IC (50)= 1 microM) also displaying a low cytotoxicity.

The bark of Croton campestris provided three furano-clerodane named, respectively, velamone, velamolone acetate and velamolone. Fractions containing these diterpenoids were investigated for their molluscicidal activity against Bulinus truncatus , one of the aquatic snail vectors of schistosomiasis. A dichloromethanic extract of root barks, rich in furano-clerodane and molluscicidal at 20 ppm was promising as natural molluscicide. Enriched fraction with velamone gave 80% mortality from 25 ppm, while its LD 100 was at 50 ppm. The LD 80 was ca. at 45 ppm, while LD 100 was at 60 ppm for the enriched fraction containing velamolone acetate. The velamolone-enriched fraction was less efficient and gave a mortality of 90% at 90 ppm. The pure compounds, velamone and velamolone acetate were active at 100% at 3 and 6 ppm, respectively. Velamolone showed a molluscicidal activity at 20 ppm.

The dried roots of Ferula hermonis yielded three new daucanes, (1R,4R)-4-hydroxydauca-7-ene-6-one (1), (1R,4R)-4-hydroxydauca-7-ene-6,9-dione (2), and (1R,3S,8S)-3-ethoxy-8-angeloyloxydauca-4-en-9-one (3), together with the three known sesquiterpenes, ferutinin, teferidin, and (+)-alpha-bisabolol. The structures of compounds 1-3 were elucidated on the basis of spectroscopic evidence. The effect of these compounds on the proliferation of estrogen-dependent MCF-7 cells was evaluated, and it was found that compounds 1 and 3 exhibited proliferative activity, whereas 2 showed an antiproliferative effect.

Four flavonoids, two terpenoids and a mixture of three polyprenols have been isolated from the leaves of Pilocarpus trachylophus.

Galipea officinalis Hancock, a Venezuelan shrubby tree which is acclaimed in folk medicine for its many healing properties, is the only species of the genus to contain tetrahydroquinoline alkaloids. A GC-MS method has been developed in order to analyse the essential oil, hexane and chloroform extracts of the trunk bark of this plant, without prior derivatisation of the alkaloidal components. A study of the MS fragmentation patterns of the components permitted the identification of five new minor quinoline alkaloids together with the known alkaloids. In addition, the method could also be used for the characterisation of alkaloids within the genus Galipea. Copyright © 2001 John Wiley & Sons, Ltd.

The isolation of 6-O-sinapoyl sucrose (1) from Iberis amara seeds and an evaluation of its antioxidative properties in comparison with sinapic acid and ascorbic acid are reported.

The iridoid 2′-coumaroyl mussaenosidic acid, isolated from Avicennia germinans (Avicenniaceae), was analysed by NMR and molecular modeling: two isomeric forms were confirmed. In addition, the known compound 1 (6S, 9R) roseoside was isolated for the first time from this species of Avicennia.

Three furano-diterpenes have been isolated from the dichloromethane extract of Croton campestris St Hil. roots. Their structures have been established by spectroscopic methods. The compounds were named velamone (ent-15,16-epoxy-2-oxo-3,13(16),14-clerodatrien), velamolone (ent-15,16-epoxy-20-hydroxy-2-oxo-3,13(16),14-clerodatrien) and velamolone acetate (ent-15,16-epoxy-20-acetoxy-2-oxo-3,13(16),14-clerodatrien).

This paper describes an efficient synthetic route to pentalongin (1H-naphtho[2,3-c]pyran-5,10-dione), a naphthoquinoid isolated from the aerial parts of Mitracarpus scaber, and to the structurally-related compound psychorubrin(3-hydroxy-1H-3, 4-dihydronaphtho[2,3-c]pyran-5,10-dione).

A new iridoid glucoside, namely 2′(4′′- hydroxycinnamoyl) mussaenosidic acid, has been isolated from the leaves of Avicennia germinans (Gabon) along with 2′-caffeoyl mussaenosidic acid. The structure of the new compound was established by spectroscopic methods.

International audience; Three new spermidine alkaloids and two known compounds were isolated from the leaves of Androya decaryi. Their structures were elucidated on the basis of their spectroscopic data (NMR and mass spectrometry), by X-Ray diffraction and by comparison with literature values. Evaluation of the in vitro antiplamosdial properties of the isolated compounds revealed they did not possess any significant activity.

The familyVerbenaceae sensu stricto is chemically characterized by the accumulation of 10-hydroxy or 8α-methyl-8β-hydroxy derivatives of 4-carboxylated iridoids compounds. The former were supposed to be characteristic of some genera of tribeLantaneae, once were never found in another tribe. This paper reports the occurrence of theviridoside (10-hydroxy-iridoid) in leaves ofVerbenoxylum reitzii (tribeCitharexyleae) and its taxonomic implication.

Dans le but d’aborder le mecanisme d’action des extraits acetoniques et ethanoliques de Newbouldia laevis (Bignoniaceae) et de Zanthoxylum zanthoxyloides (Rutaceae), leur effet inhibiteur a ete evalue in vitro sur la migration larvaire de Haemonchus contortus . Le test d’inhibition de la migration larvaire (LMI) a ete applique sur les larves infestantes (L 3 ), âgees de 2 a 3 mois incubees avec des extraits vegetaux a differentes concentrations : 150, 300, 600 et 1200 μg/mL mis ou non en contact avec la polyvinylpolypyrrolidone (PVPP). Un temoin negatif (tampon PBS) a ete inclus dans chaque test. L’observation sous microscope et le denombrement des L 3 ayant migre par rapport au nombre total de larves deposees dans l’insert ont permis de calculer le taux de la migration larvaire. Les extraits de Newbouldia laevis et de Zanthoxylum zanthoxyloides inhibent in vitro la migration larvaire de Haemonchus contortus . Cet effet est dose-dependant (p

Seventeen natural C-glycosylflavones have been analysed using positive desorption chemical ionization mass spectrometry with ammonia as reagent gas. This technique provides the molecular weight, information for the identification of the aglycone, the sugar sequence, and the characterization of the C-sugar (hexose, pentose). The distinction between mono-C-glycosylflavones, di-C-glycosylflavones, C-glycosylflavone 2″-O-glycosides, and di-C-glycosylflavone 2‴-O-glycosides is also obtained.

In the search for new molluscicidal natural products, the activity of the chloroform extract of the root barks of Caryopteris x clandonensis was tested. The LC(100) was

Five new neoclerodane diterpenoids, rel-(3R,4S,5R,7R,8S,9R,10S,12R,20S)-7-acetoxy-15,16,12,20-diepoxy-3,4-dihydroxy-20-methoxyneocleroda-13(16),14-diene (1), rel-(3R,4S,5R,7R,8S,9R,10S,12R,20S)-7-acetoxy-15,16,12,20-diepoxy-3,4,20-trihydroxyneocleroda-13(16),14-diene (2), rel-(3R,4S,5S,6R,7S,8S, 9R,10S,12R,20S)-6,7-diacetoxy-3,4,15,16,12,20-triepoxy-20-hydroxyneocleroda-13(16),14-diene (3), rel-(3R,4S,5R,7R,8S,9R,10S,12R,20S)-7-acetoxy-3,4,15,16,12,20-triepoxy-20-hydroxyneocleroda-13(16),14-diene (4), and rel-(3R,4S,5R,7R,8S,9R,10S,12R,20R)-7,20-diacetoxy-3,4,15,16,12,20-triepoxyneocleroda-13(16),14-diene (5), have been isolated from the bark of Croton eluteria. The structures of the compounds 1-5 (cascarillins E-I) were determined by spectroscopic data interpretation.

International audience; Abstract: GC-FID and GC-MS analysis of essential oil from oregano leaves (Origanum compactum) resulted in the identification of 46 compounds, representing more than 98% of the total composition. Carvacrol was the predominant compound (36.46%), followed by thymol (29.74%) and p-cymene (24.31%). Serial extractions with petroleum ether, ethyl acetate, ethanol, and water were performed on aerials parts of Origanum compactum. In these extracts, different chemical families were characterized: polyphenols (gallic acid equivalent 21.2 to 858.3 g/kg), tannins (catechin equivalent 12.4 to 510.3 g/kg), anthocyanins (cyanidin equivalent 0.38 to 5.63 mg/kg), and flavonoids (quercetin equivalent 14.5 to 54.7 g/kg). The samples (essential oil and extracts) were subjected to a screening for antioxidant (DPPH and ABTS assays) and antimalarial activities and against human breast cancer cells. The essential oil showed a higher antioxidant activity with an IC50= 2 \textpm 0.1 mg/L. Among the extracts, the aqueous extract had the highest antioxidant activity with an IC50= 4.8 \textpm 0.2 mg/L (DPPH assay). Concerning antimalarial activity, Origanum compactum essential oil and ethyl acetate extract showed the best results with an IC50 of 34 and 33 mg/mL, respectively. In addition, ethyl acetate extract (30 mg/L) and ethanol extract (56 mg/L) showed activity against human breast cancer cells (MCF7). The oregano essential oil was considered to be nontoxic.

Three native glucosinolates have been isolated from seeds of Brassica juncea using mild methods for their separation. The compounds were identified by various NMR techniques.

Three furanoid clerodanes have been isolated from the stem bark of Croton eluteria Bennett. Their structures have been established by spectroscopic methods. The compounds were named cascarillin B (7α-acetoxy-3,4,15,16-diepoxy-12-oxo-cleroda-13(16),14-dien-20-al), cascarillin C (7α-acetoxy-15,16,12,20-diepoxy-20-hydroxy-cleroda-3,4,13(16),14-triene) and cascarillin D (7α-acetoxy-3,4,15,16-diepoxy-cleroda-13(16),14-dien-20-al).

In continuation of our phytochemical studies on Caryopteris x clandonensis (Lamiaceae), three further iridoids were isolated from the methanolic extract of the stems. Their structures were established by 1D and 2D NMR and MS analysis as a C-6 epimer of 8-O-acetylharpagide (6-epi-8-O-acetylharpagide), a derivative of harpagide which contained the unusual feature of a 3',4' seco-glycopyranosyl moiety (clandonoside II) and a methyl cetal of 8-O-acetylharpagide aglucone hydrate named clandonensine.

Introduction In our work, we evaluate the potential antioxidant, antimalarial activity and also activity against human breast cancer cells (MCF7) of Argan fruit extracts using in vitro models to validate the traditional use of this plant. Its chemical composition was also studied to begin the understanding of its activities, waiting to find the structure-activity relationship. Results Polyphenols (89.4–218.5 eqGallic acid (mg/g dry)), tannins (39.3–214.0 eqCatechin (mg/g dry)), flavonoids (3.4–11.1 eqQuercetin (mg/g dry)) and anthocyanins (0.74–10.92 eqCyanindin (μg/g dry)) were quantified. A good (ethyl acetate and decoction) and moderate (petroleum ether) antioxidant activity were obtained for DPPH (IC50 32.3–600.8 μg/ml) and ABTS (IC50 11.9–988.8 μg/ml) assays. In addition, we found a good antimalarial activity (IC50 35 to >100 μg/ml) and human breast cancer cells activity (IC50 42 to >100 μg/ml). Conclusions The ethyl acetate extract and the decoction show interesting antimalarial and antioxidant activities. The results indicate a good correlations between anthocyanins quantitiy and the potential antioxidant (R2=0.9867) and also to antimalarial activity (R2=0.8175).

Two tetrahydroquinoline alkaloids have been isolated from the bark of Galipea officinalis Hancock. Their structures have been established by spectroscopic methods. The compounds were named angustureine (1,2,3,4-tetrahydro-2(n-pentyl)-1-methylquinoline) and galipeine (2-[(3′-hydroxy-4′-methoxy-phenylethyl)]-2,3,4-tetrahydro-1-methylquinoline).

Two new iridoid keto-glycosides, clandonoside and 8- O -acetylclandonoside, together with their hydrates, were isolated from the stems of Caryopteris × Clandonensis besides the known harpagide and 8- O -acetylharpagide. The structure of clandonoside and 8- O -acetylclandonoside were established as harpagide-aglucone-1- O -β- d -ribohexo-3-ulopyranoside and 8- O -acetylharpagide-aglucone-1- O -β- d -ribohexo-3-ulopyranoside, respectively, by spectroscopic methods. This is the first report of a C-1 linked β- d -ribohexo-3-ulopyranose (β- d -ketohexose) iridoid.

Schizozygia coffaeoides Baill. (Apocynaceae) is a monotypic shrub used in Kenyan folk medicine for treatment of various infections as skin diseases caused by fungi [1]. Schizogane alkaloids represent a small group of hexacyclic N-acyl compounds that were first isolated from the roots of S. coffaeoides by Renner and co-workers [2, 3] but with doubts on the structure of two of the isolated compounds (isoschizogamine and isoschizogaline). Both structures were later revised [4, 5] and named isoschizoganes in which a five-membered (as in 2 and 3) rather than a six-membered (as in 1) lactam ring is present, thus revealing a hexacyclic skeleton of N-acyl tetrahydroquinoline type with an aminal moiety. We present here the key 13C-NMR signals to distinguish schizoand isoschizoganes alkaloids and also in vitro antimalarial and cytotoxic evaluation of crude CH2Cl2 extracts of roots and stems of S. coffaeoides and 3 isoand shizogane alkaloids isolated from the CH2Cl2 roots extract Antimalarial and cytotoxic activities of schizogane alkaloids isolated from Schizozygia coffaeoides

The fragmentations of hydroxylated flavanones, chalcones and dihydrochalcones were investigated by direct loop injection using an ion trap mass spectrometry equipped with atmospheric pressure chemical ionization (APCI) probe. Some of them have been isolated from the leaves of Piper hostmannianum var. berbicense and standards were used to confirm their fragmentation behaviour. In negative ion mode, fragmentations of these three types of flavonoids revealed specific diagnostic ions which allowed us to identify aglycones in a crude plant extract. The major fragment ion obtained in MS/MS experiment for methoxylated chalcones is the neutral loss of a methyl radical whereas a H(2)O molecule is lost in the case of methoxylated dihydrochalcones. Methoxylated chalcones and flavanones isomers could be differentiated by the relative intensity ratio of [M-H-CH(3)]*(-) and [M-H-C(2)H(2)O](-) ions. Based on UV and MS data, a decision tree that includes UV lambda(max) absorptions and MS/MS diagnostic ions was built in order to obtain structural information of unknown compounds present in the extract. This tree was used to identify flavonoids in the ethyl acetate extract of P. hostmannianum var. berbicense leaves after analysis by high-performance liquid chromatography-diode array detection-atmospheric pressure chemical ionization ion trap multistage mass spectrometry. A total of 11 flavonoids were tentatively characterized based on the MS fragmentations pattern observed in MS(n) experiments.

Bolivianine, a novel sesterpene with an unprecedented skeleton, has been isolated from the trunk bark of Hedyosmum angustifolium (Chloranthaceae), with isobolivianine, an isomer formed under acidic conditions. The structure and relative stereochemistry were elucidated on the basis of spectroscopic data. A hypothesis for biogenesis was made.

Vincent Roumy, Gaston Garcia-Pizango, Andrea-Luz Gutierrez-Choquevilca, L. Ruiz, Valérie Jullian, et al.. Amazonian plants from Peru used by Quechua and Mestizo to treat malaria with evaluation of their activity. Journal of Ethnopharmacology, 2007, 112, pp.482-489. ⟨10.1016/j.jep.2007.04.009⟩. ⟨hal-00384336⟩; International audience; Indigenous Quechua and Mestizo populations from distinct areas in Loreto, Peru, were interviewed about traditional medication for the treatment of malaria. An ethnographic survey concerning the native theory of illness aetiology in the specific case of malaria permitted the elaboration of an efficient ethnopharmacological enquiry. The survey took place on three main zones corresponding to villages on the Napo and the Pastaza rivers (for the Quechua), and in the surroundings of Iquitos (for the Mestizos) and led to the collection of 14 plants. Serial extractions in hexane, dichloromethane, and methanol were performed on the different parts of the plants collected. The extracts were then tested for antiplasmodial activity in vitro. Seven plants displayed antiplasmodial activity (IC50 from 2 to 25 µg/mL) and usually low cytotoxicity, indicating their antiplasmodial specificity. The results give scientific validation to the traditional medical knowledge of Quechua and Mestizo populations from Loreto and confirm a source of potentially active plants.

Tapirira guianensis is a common tree used in traditional medicine in French Guiana against several infectious diseases (malaria, leishmaniasis, bacteria, etc.). The bioassay-guided purification of CH2Cl2 bark extract led to the isolation of four cyclic alkyl polyol derivatives: 4,6,2′-trihydroxy-6-[10′(Z)-heptadecenyl]-1-cyclohexen-2-one (1a), 1,4,6-trihydroxy-1,2′-epoxy-6-[10′(Z)-heptadecenyl]-2-cyclohexene (1b), 1,4,5,2′-tetrahydroxy-1-[10′(Z)-heptadecenyl]-2-cyclohexene (2), and 1,3,4,6-tetrahydroxy-1,2′-epoxy-6-[10′(Z)-heptadecenyl]-cyclohexane (3). The structures were established on the basis of 1D and 2D NMR analyses. The anti-leishmanial, anti-plasmodial, anti-bacterial (on Staphylococcus aureus, Staphylococcus epidermidis and Escherichia coli), and anti-fungal (on Candida albicans) activities of the extracts and of these original compounds were evaluated. Two showed medicinal interest supporting the traditional uses of the plant. The structures were established through spectral analyses of the isolates and their derivatives.

The bioassay-guided purification of an n-hexane extract from the leaves of Piper hostmannianum var. berbicense led to the isolation of four monoterpene or prenyl-substituted dihydrochalcones (1a, 1b, 2, 3) as well as the known compounds 2 ',6 '-dihydroxy-4 '-methoxydihydrochalcone (4), linderatone (5), strobopinin (6), adunctin E (7) and (-)-methyllinderatin (8). Their structures were established on the basis of NMR and X-ray analysis. (-)-Methyllinderatin, linderatone and 2 ',6 '-dihydroxy-4 '-methoxydihydrochalcone exhibited the most potent antiplasmodial activity with IC50 values of 5.64, 10.33 and 12.69 mu M, respectively against both chloroquine-sensitive and resistant strains of Plasmodium falciparum (F32, FcBl). The activity of (-)-methyllinderatin was confirmed in vivo against Plasmodium vinckei petteri in mice (80% of reduction of parasitemia) at a dose of 20 mg/kg/day.

Three native glucosinolates have been isolated from seeds of Brassica juncea using mild methods for their separation. The compounds were identified by various NMR techniques.

A new coumarin glucoside, 3′(S)-hydroxy-2′,2′-dimethyl-dihydropyranocoumarin-8-β- d -glucopyranosyl, one coumarin, five furanocoumarins, three bicoumarins, three quinoline alkaloids and one sinapoyl sucrose derivative have been isolated from the roots of Ruta corsica.

Three furanoid clerodanes have been isolated from the stem bark of Croton eluteria Bennett. Their structures have been established by spectroscopic methods. The compounds were named cascarillin B (7alpha-acetoxy-3,4,15,16-diepoxy-12-oxo-cleroda-13(16),14-dien-20-al), cascarillin C (7alpha-acetoxy-15,16,12,20-diepoxy-20-hydroxy-cleroda-3,4,13(16),14-triene) and cascarillin D (7alpha-acetoxy-3,4,15,16-diepoxy-cleroda-13(16),14-dien-20-al).

An ion-pair HPLC-ESI-MS-MS method has been developed for the direct and rapid characterization of isoquinoline alkaloids in a crudely purified extract of the aerial parts of Eschscholtzia californica (Papaveraceae). This plant was chosen because of its increasing use in pharmaceutical industries and because its well known alkaloid composition allows the optimization of the experimental procedure through an on-line analytical sequence. Thus, 14 isoquinoline alkaloids of different types were detected and characterized. The identities of these compounds were confirmed unambigously by their fragmentation and UV spectra obtained by LC-diode-array detection. Various experiments including tandem mass spectrometry and in-orifice collision induced dissociation were performed and prove that MS-MS is a very efficient technique to identify these compounds. An explanation for each isoquinoline alkaloid type MS-MS fragmentation pattern is proposed and indicates similar neutral and/or radical losses. The order of the fragmentation depended on the type of compound but the lost fragments were similar.

Western countries observed an increased interest for the use of herbal medicines because of their supposed safety in contrast to chemical drugs. Wild germander (Teuchrium chamaedrys L) was traditionally used as a folk medicine for its for and antiseptic properties. In 1991, germander consumed to treat obesity, caused an epidemic of cytolytic hepatitis. Thirty cases of hepatotoxicity were first reported including cases with positive rechallengge. For these patients, an early recurrence was observed despite lack of other features of hypersensitivity (Castot and Larrey, 1992). In mice, germander toxicity required CYP3A-dependent metabolism (Loeper et al 1994). More specifically, the metabolic activation of the furan ring of the diterpenoid teucrin A (TA) (Kouzi et al 1994). TA- toxicity was via CYP3A-generated electrophilic metabolites that were detoxified by glutathione conjugation, depleted cellular thiols and caused apoptosis in isolated rat hepatocytes atocytes (Lekehal et al 1996; Fau et al 1997). However, these death processes did not explain the immunologial cal features observed in several cases of germander-induced hepatitis. To explain this immune process, patient’s sera consuming germander tea in great quantity were Tested by Western blot analysis. They contained autoantibodies directed against human microsomal epoxide hydrolase (hmEH), that was located both, in the endoplasmic reticulum and the plasma membrane (PM) of human hepatocyte and hmEH- expressing yeast. Germander-induced autoantibodies (GIAA) were shown to recognize hmEH on the cell surface. To implicate hmEH in the metabolic activation of TA we used a “humanized” yeast strain expressing human P450-reductase and cytochrome b5 transformed with CYP3A4 and/or hmEH cDNAs. TA was metabolized by CYP3A4 into a and hmEH with TA inactivated hmEH in a time-dependent-manner, in agreement with formation of reactive teuchrin A-metabolite that could covalently alter and inhibit hmEH. This modified enzyme may bypass the immunologic tolerance that normally exists for hmEH.

A new neo -clerodane diterpenoid has been isolated from the stem bark of Croton levatii . The structure of crovatin was established from its NMR and X-ray data as methyl ent -(8 R ,10β)-3,19 S : 15,16:12 S ,20 R : 19,20-tetra epoxy-cleroda-13(16),14-dien-18β-oate.

Germander, a plant used in folk medicine, caused an epidemic of cytolytic hepatitis in France. In about half of these patients, a rechallenge caused early recurrence, suggesting an immunoallergic type of hepatitis. Teucrin A (TA) was found responsible for the hepatotoxicity via metabolic activation by CYP3A. In this study, we describe the presence of anti-microsomal epoxide hydrolase (EH) autoantibodies in the sera of patients who drank germander teas for a long period of time. By Western blotting and immunocytochemistry, human microsomal EH was shown to be present in purified plasma membranes of both human hepatocytes and transformed spheroplasts and to be exposed on the cell surface where affinity-purified germander autoantibodies recognized it as their autoantigen. Immunoprecipitation of EH activity by germander-induced autoantibodies confirmed this finding. These autoantibodies were not immunoinhibitory. The plasma membrane-located EH was catalytically competent and may act as target for reactive metabolites from TA. To test this hypothesis CYP3A4 and EH were expressed with human cytochrome P450 reductase and cytochrome b(5) in a "humanized" yeast strain. In the absence of EH only one metabolite was formed. In the presence of EH, two additional metabolites were formed, and a time-dependent inactivation of EH was detected, suggesting that a reactive oxide derived from TA could alkylate the enzyme and trigger an immune response. Antibodies were found to recognize TA-alkylated EH. Recognition of EH present at the surface of human hepatocytes could suggest an (auto)antibody participation in an immune cell destruction.

Although cytochrome P-450 (CYP)-generated reactive metabolites can cause hepatocyte apoptosis, the mechanism of this effect is incompletely understood. In the present study, we assessed the hepatotoxicity of skullcap, a diterpenoid-containing herbal remedy. Male rat hepatocytes were incubated for 2 hours with skullcap diterpenoids (100 microg/mL). This treatment decreased cell glutathione and protein thiols and increased cell [Ca(2+)]. This activated Ca(2+)-dependent tissue transglutaminase, forming a cross-linked protein scaffold, and also opened the mitochondrial permeability transition pore, causing outer mitochondrial membrane rupture, increased cytosolic cytochrome c, activation of procaspase 3, internucleosomal DNA fragmentation, and ultrastructural features of apoptosis. Cell death was increased by a CYP3A inducer (dexamethasone) or a sulfur amino acid-deficient diet increasing glutathione depletion. In contrast, cell death was prevented by decreasing CYP3A activity (with troleandomycin), preventing glutathione depletion (with cysteine or cystine), blocking Ca(2+)-modulated events (with calmidazolium), preventing mitochondrial permeability transition (with cyclosporin A), or inhibiting caspase 3 (with acetyl-Asp-G u-Va-Asp-a dehyde). Both calmidazolium and cyclosporin A also prevented the increase in cytosolic cytochrome c and procaspase 3 activation. In conclusion, CYP3A activates skullcap diterpenoids into reactive metabolites that deplete cellular thiols and increase cell [Ca(2+)]. This activates Ca(2+)-dependent transglutaminase and also opens the mitochondrial permeability transition pore, causing outer mitochondrial membrane rupture, cytochrome c release, and caspase activation. Preventing mitochondrial permeability transition pore opening and/or caspase activity blocks apoptosis, showing the fundamental role of these final events in metabolite-mediated hepatotoxicity.

The structure and absolute configuration of fraxinellonone isolated from Fagaropsis glabra was established as 3α-(3′-furanyl)-3α,7-dimethyl-4,5-dihydro-1,6-isobenzofurandione, by NMR and CD comparison with fraxinellone. Complete carbon assignments for fraxinellone and isofraxinellone were achieved by 13 C- 1 H COSY experiments.

International audience; A new iridoid glucoside, namely 2'-caffeoyl mussaenosidic acid has been isolated from leaves of Avicennia germinans along with the known compounds 2'-cinnamoyl mussaenosidic acid and verbascoside. The structure of the new compound was established by spectroscopic methods.