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Seven essential oil samples of two endemic species of Malagasy sage, Salvia sessilifolia Baker and Salvia leucodermis Baker, were investigated via GC(RI), GC-MS and 13 C NMR spectrometry. In total, 81compounds were identified accounting for 93.5% to 98.7% of the total composition. The main constituents for the both species were ( E )-β-caryophyllene (29.2% to 60.1%), myrcene (1.2% to 21.7%), α-humulene (5.2% to 19.7%), ( E )-nerolidol (0.8% to 15.5%) and caryophyllene oxide (1.4% to 10.8%). Ethnobotanical survey of 46 informants revealed that decoctions of leafy twigs and chewed leaves were usually used. Due to the repeated fires, over-harvesting and grazing, the populations of S. sessilifolia and S. leucodermis are drastically fragmented. These risk factors led to threats to the habitats of the target species. Salvia sessilifolia Baker and Salvia leucodermis Baker are proposed to be classified as endangered species.

An experimental procedure using 1 H NMR was developed and validated to quantify 5-hydroxymethyl-2(5H)-furanone, a valuable chemical synthon ((S)-enantiomer), in a dichloromethane extract of Helleborus lividus subsp. corsicus leaves. This method, using vanillin as the internal standard, exhibited a perfect linearity of measurements ( R  = 1) associated with very good accuracy (relative errors comprised between -1.62% and 4.25%) and precision (reproducibility 30.51 mg ± 0.4%). The limit of detection and the limit of quantitation have been measured at 0.14 mg and 0.59 mg, respectively. The experiment time is very short since a single analysis is at the minute level. 5-Hydroxymethyl-2(5H)-furanone accounted for nearly 85% in the dichloromethane extract of 2  = 1) associated with very good accuracy (relative errors comprised between -1.62% and 4.25%) and precision (reproducibility 30.51 mg ± 0.4%). The limit of detection and the limit of quantitation have been measured at 0.14 mg and 0.59 mg, respectively. The experiment time is very short since a single analysis is at the minute level. 5-Hydroxymethyl-2(5H)-furanone accounted for nearly 85% in the dichloromethane extract of H. lividus leaves (1.7% of the mass of fresh leaves). This plant represents an important and natural source of (S)-5-hydroxymethyl-2(5H)-furanone (main enantiomer; determined using a GC chiral analysis).corsicus leaves (1.7% of the mass of fresh leaves). This plant represents an important and natural source of (S)-5-hydroxymethyl-2(5H)-furanone (main enantiomer; determined using a GC chiral analysis)., Competing Interests: The authors declare that they have no conflicts of interest., (Copyright © 2022 Thomas Maroselli et al.)

The root and stem bark essential oils from Isolona dewevrei (Annonaceae), growing wild in Côte d'Ivoire, were investigated for the first time, using a combination of chromatographic [CC, GC(RI)] and spectroscopic [MS, 13 C-NMR] techniques. A new natural germacrone was isolated by repetitive column chromatography carried out on a stem bark oil sample. Its structure was elucidated as germacra-1(10),4(15),5-trien-8-one by 1 D, 2 D-NMR and QTOF-MS. Ninety-six components accounting for 95.5 ± 0.8% and 95.8 ± 1.0%, respectively, for the root and stem bark essential oil samples were identified. The major compounds of root oil were cyperene (19.7 ± 1.6%) and camphene (10.1 ± 2.1%), followed by 5-isopentenylindole (6.4 ± 2.4%), β-elemene (3.9 ± 0.3%), ( Z )-α-bisabolene (3.2 ± 1.2%) and γ-gurjunene (3.2 ± 0.3%). The stem bark oil was also dominated by cyperene (29.2 ± 4.1%), followed by β-elemene (6.2 ± 1.1%), ( Z )-α-bisabolene (3.8 ± 1.0%), ( E )-β-caryophyllene (3.3 ± 0.7%) and α-copaene (2.8 ± 1.1%).

This study made it possible to characterise by GC (in combination with retention indices), GC-MS and 13 C NMR, the chemical composition of the essential oils from Solanum rugosum and Solanum erianthum , two Solanaceae of the Ivorian flora. The two essential oils were characterised by a very high proportion of sesquiterpenes. Specifically, the essential oil of S. rugosum was dominated by (E)- β -caryophyllene (33.7%), β -elemol (19.8%) and germacrene D (14.4%), while that of S. erianthum was mainly composed of α -humulene (38.6%), β -elemol (17.8%) and (E)- β -caryophyllene (16.7%). The chemical composition of Solanum rugosum is described here for the first time.

This study aimed to investigate the chemical composition of the leaf essential oil from Ivoirian Isolona dewevrei . A combination of chromatographic and spectroscopic techniques (GC(RI), GC-MS and 13 C-NMR) was used to analyze two oil samples (S1 and S2). Detailed analysis by repetitive column chromatography (CC) of essential oil sample S2 was performed, leading to the isolation of four compounds. Their structures were elucidated by QTOF-MS, 1D and 2D-NMR as (10βH)-1β,8β-oxido-cadin-4-ene ( 38 ), 4-methylene-(7αH)-germacra-1(10),5-dien-8β-ol ( cis -germacrene D-8-ol) ( 52 ), 4-methylene-(7αH)-germacra-1(10),5-dien-8α-ol ( trans -germacrene D-8-ol) ( 53 ) and cadina-1(10),4-dien-8β-ol ( 56 ). Compounds 38 , 52 and 53 are new, whereas NMR data of 56 are reported for the first time. Lastly, 57 constituents accounting for 95.5% (S1) and 97.1% (S2) of the whole compositions were identified. Samples S1 and S2 were dominated by germacrene D (23.6 and 20.5%, respectively), followed by germacrene D-8-one (8.9 and 8.7%), (10βH)-1β,8β-oxido-cadin-4-ene (7.3 and 8.7), 4-methylene-(7αH)-germacra-1(10),5-dien-8β-ol (7.8 and 7.4%) and cadina-1(10),4-dien-8β-ol (7.6 and 7.2%). Leaves from I. dewevrei produced sesquiterpene-rich essential oil with an original chemical composition, involving various compounds reported for the first time among the main components. Integrated analysis by GC(RI), GC-MS and 13 C-NMR appeared fruitful for the knowledge of such a complex essential oil.

The investigation of the stem essential oil of Laggera pterodonta (DC.) Sch. Bip. ex Oliv. (Asteraceae) from Côte d'Ivoire was carried out, using a combination of chromatographic (GC-RI, CC, pc-GC) and spectroscopic (GC-MS, 13 C NMR) techniques. This study led to the identification of fifty constituents of which two new natural compounds 7β,11β-epoxy-eudesman-4α-ol and 7α,11α-epoxy-eudesman-4α-ol. Their structures were elucidated by 1 D and 2 D NMR spectroscopy after pc-GC purifying. Finally, 98.9% of the whole composition of the oil was identified with a high amount of 2,5-dimethoxy- p -cymene (78.9%). The other significant components were α-humulene (6.2%), ( E )-β-caryophyllene (1.7%), thymyl methyl oxide (1.7%), α-phellandrene (1.5%), p -cymene (1.2%), (3αH,4βH,6αH,1αMe)-1,6-epoxy-3-hydroxycarvotanacetone angelic acid ester (1.1%) and 10- epi -γ-eudesmol (1.0%).

Three new compounds, a dihydrobenzofuran (coumaran) derivative (compound 1 ) and two pterocarpans (compounds 2 and 3 ) were isolated from a root extract of Calicotome villosa growing wild in Corsica. Their structures were elucidated using 1D and 2D NMR spectroscopy and MS/MS as 2-(1-methylethenyl)-5-hydroxy-6-carbomethoxy-2,3-dihydro-benzofuran, 4,9-dihydroxy-3-methoxy-2-dimethylallylpterocarpan, and 4,9-dihydroxy-3',3'-dimethyl-2,3-pyranopterocarpan.

The essential oils of six endemic Malagasy Helichrysum species were investigated by GC (RI), GC-MS and 13 C NMR spectrometry. In total, 153 compounds were identified accounting for 90.8% to 99.9% of the total composition. The main constituents were α-pinene for H . benthamii , 1,8-cineole for H. dubardii , (E)-β-caryophyllene for H . indutum , and H . bojerian um . H. diotoides essential oil was characterized by the presence of two lilac alcohols and four lilac acetates whereas H. hirtum essential oil exhibited an atypical composition with 7β- H -silphiperfol-5-ene, 7-epi-subergorgiol, and 7-epi-silphiperfol-5-en-13-oic acid as major components., Competing Interests: The authors declare no conflict of interest.

The chemical composition of 44 leaf oil samples of Laggera pterodonta (DC.) Sch.Bip. ex Oliv. (Asteraceae) from Côte d'Ivoire was investigated, using combination of chromatographic (GC-FID) and spectroscopic (GC/MS, 13 C-NMR) techniques. Two oil samples chosen according to their chromatographic profiles were submitted to column chromatography and all fractions of CC were analyzed by GC-FID, GC/MS and 13 C-NMR. In total, 83 components accounting for 96.5 to 99.4 % of the whole chemical composition were identified. Significant variations were observed within terpene classes: monoterpene hydrocarbons (0.4-22.7 %), oxygenated monoterpenes (32.9-54.9 %), sesquiterpene hydrocarbons (18.6-38.3 %) and oxygenated sesquiterpenes (3.5-38.4 %). Thus, the 44 compositions were subjected to hierarchical cluster analysis (HCA) and principal component analysis (PCA). Two groups were differentiated according to their composition. All the samples contained 2,5-dimethoxy-p-cymene, α-humulene and (E)-β-caryophyllene among the main components. Other components were present at appreciable contents and allowed differentiation of two groups: sabinene and germacrene D for Group I; 10-epi-γ-eudesmol and eudesm-7(11)-en-4α-ol for Group II. All the samples collected in Eastern Côte d'Ivoire constituted Group I, while samples collected in the Central area of the country constituted Group II., (© 2019 Wiley-VHCA AG, Zurich, Switzerland.)

Leaves of Ilex aquifolium L. have been used for their therapeutic properties. In previous studies, components contained in the leaves were first isolated by various chromatographic techniques. Then, quantitation of oleanolic and ursolic acids, which are responsible for the biological and therapeutic activities of the plant, was performed by HPLC, HPTLC, and somewhat by GC-MS. Our objective was to develop a simple method that allows the identification of compounds contained in the leaves of Corsican I. aquifolium and to quantify ursolic and oleanolic acids. Leaves were successively extracted with hexane and dichloromethane. The extracts were chromatographed on silica gel and the fractions of column chromatography submitted to 13 C-NMR analysis, following a computerized method developed in the laboratory. 13 C-NMR allowed the identification of various triterpenes including ursolic acid and oleanolic acid. Quantitation of both acids was achieved, for the first time, by 1 H-NMR after validation of the method (accuracy, precision, linearity, limit of detection and limit of quantitation). Ursolic and oleanolic acids accounted for 55.3% and 20.8% of the dichloromethane extract, respectively. This represents 1.3% and 0.5% of the mass of dried leaves. 1 H-NMR spectroscopy appeared as a powerful tool for a rapid quantitation of biologically active compounds from I. aquifolium .

Root and flower essential oils of Cyanthillium cinereum (L.) H. Rob. (Synonym Vernonia cinerea (L.) Less.) (Asteraceae) collected in Southern Côte d'Ivoire was investigated using a combination of chromatographic and spectroscopic techniques. The root oil composition was dominated by trans -β-bergamotene (20.7%), β-elemene (19.0%), cyperene (10.6%), germacrene A (7.1%) and β-pinene (3.8%), whereas γ-humulene (31.0%), ( E )-β-caryophyllene (17.0%), trans -β-bergamotene (7.7%), β-pinene (7.5%) and ( E )-β-farnesene (6.0%) were the major components of flower oil. Two new compounds bearing the trans -β-bergamotene framework were identified: trans -β-bergamotenone and ( E )- trans -β-bergamotenol.

The chemical composition of twenty-five essential oil samples from the aerial parts of two Malagasy endemic species Billburttia capensoides Sales & Hedge and B. vaginoides Sales & Hedge, were investigated for the first time. Based on chromatographic profiles, three selected samples were investigated using GC(RI), GC-MS and 13 C NMR. The content of the main components varied drastically from sample to sample: p-mentha-1,3,8-triene (0.2-52.7%), terpinolene (2.8-40.7%) and dill apiole (0.0-22.2%). Statistical analysis of the 25 oil compositions allowed the distinction of two well-differentiated groups. Samples of group I contained mainly p-mentha-1,3,8-triene while the Group II was dominated by terpinolene and dill apiole.

Twenty-three resin samples have been obtained by tapping from individual Pinus pinaster adult trees grown in Corsica and submitted to acido-basic partition. Identification and quantitative determination of resin acids has been carried out using 13 C-NMR spectroscopy following a method developed by our group. The main components were dehydroabietic acid (up to 37.6 %), levopimaric acid (up to 35.5 %) and abietic acid (up to 24.7 %). A lignan, pinoresinol, has been identified in some samples. Within the 23 compositions, submitted to k-means analysis and Principal Component Analysis, two clusters have been perfectly differentiated, whose compositions were dominated by dehydroabietic acid (Group I, M=23.5 %, SD=6.3) and levopimaric acid (Group II, M=21.2 %; SD=6.2), respectively. Both compositions have been observed in the three locations of harvest., (© 2019 Wiley-VHCA AG, Zurich, Switzerland.)

The composition of Enantia polycarpa Engl. & Diels leaf essential oil has been investigated for the first time using a combination of chromatographic and spectroscopic techniques. The compositions of 52 leaf essential oil samples have been subjected to statistical analysis, hierarchical cluster analysis (HCA) and principal component analysis (PCA). Four groups were differentiated, of which the compositions were dominated by β-elemene and germacrene B (Group III, 22/52 samples); germacrene D (Group I, 16/52 samples); β-cubebene (Group IV, 8/52 samples) and by germacrene B and germacrene D (Group II, 6/52 samples). A special attention was brought to the quantification of the thermolabile components, germacrene A, germacrene B and germacrene C, as well as that of their rearranged compounds, β-elemene, γ-elemene and δ-elemene. 13 C-NMR data of β-cubebene have been provided., (© 2018 Wiley-VHCA AG, Zurich, Switzerland.)

The chemical investigation of an ethyl acetate extract (EtOAc) obtained from Laurencia obtusa , collected in Corsica, allowed for the identification of three new compounds ( 1 , 2 , and 4 ) and six known compounds. Compounds 1 to 4 were isolated and fully characterized by a detailed spectroscopic analysis. Compounds 1 and 2 are two C 15 -acetogenins sharing the same ring system: a tetrahydropyran linked by a methylene to a tetrahydrofuran ring. Compound 1 exhibits a bromoallene unit whereas compound 2 possesses an uncommon α-bromo-α,β-unsaturated aldehyde terminal unit. Compound 4 is the first diterpene exhibiting a 19(4 → 3) abeo -labdane skeleton isolated from a Laurencia species. Isolation of concinndiol (compound 3 ) together with compound 4 suggests a common biosynthetic origin. Additionally, five known compounds, namely sagonenyne, laurene, α-bromocuparene, microcladallene A, and β-snyderol were identified in chromatographic fractions by NMR analysis using a computerized method that was developed in our laboratory., Competing Interests: The authors declare no conflict of interest.

Generic and specific determination among the Laurencia complex is a challenging task. DNA barcoding combined with phenotypic investigations are mandatory for species differentiation. In this study, two morphologically different members of the Laurencia complex were investigated using untargeted 1 H-NMR-based metabolomics. Twenty-one population samples were collected in order to evaluate both temporal and geographical homogeneity. Data obtained from 1 H-NMR analysis followed by statistical analysis allowed a clear separation of all the samples into two groups. DNA mitochondrial tests confirmed this pattern and identified the two species as Laurenciella sp. and Laurencia obtusa. In addition, metabolites responsible of this discrimination were investigated directly in crude extracts by 13 C-NMR using an in-house computer-assisted method. The combination of both untargeted ( 1 H) and targeted ( 13 C) NMR-based metabolomic approaches proves to be a powerful and complementary approach to discriminate species from the Laurencia complex., (© 2017 Wiley-VHCA AG, Zurich, Switzerland.)

The aim of this survey was to determine the chemical composition of essential oils (EO) of five conifers acclimated in Corsica by GC(RI), GC-MS and 13 C NMR. L. decidua needle and wood EOs contained as majors components: α- and β-pinenes, germacrene D (needles) and bornyl acetate (wood). The EOs of needles, wood and cones of P. menziesii were characterised by β- and α-pinenes, terpinen-4-ol, sabinene, terpinolene (needles and wood), Δ-3-carene (wood) and limonene (cones). Needles and wood EOs of P. ponderosa contained as major components: β- and α-pinenes, Δ-3-carene (wood) and estragole (needles). S. giganteum EOs of foliage and wood were rather similar and dominated by α-pinene, and safrole. The EOs of leaf, wood and cones from C. japonica were very similar, and exhibited α-pinene, sabinene, β-elemol and kaurene as major constituents. It appeared that EO compositions of some species were different from reported literature data.

A new C 15 -acetogenin, sagonenyne ( 20 ), exhibiting an unusual single tetrahydropyran ring was isolated from an ethyl acetate extract of Laurencia obtusa collected on the Corsican coastline. Its structure was established by detailed NMR spectroscopic analysis, mass spectrometry, and comparison with literature data. Twenty-three known compounds were identified in the same extract by means of column chromatography steps, using a 13 C-NMR computer aided method developed in our laboratory. In addition to sesquiterpenes, which represent the main chemical class of this extract, diterpenes, sterols, and C 15 -acetogenins were identified. The crude extract was submitted to a cytotoxicity assay and was particularly active against THP-1 cells, a human leukemia monocytic cell line.

The composition of four samples of essential oil isolated from the leaves of individual Monodora crispata trees growing wild in an Ivorian forest (Adiopodoum6) were investigated by a combination of chromatographic (GC(RI)) and spectrometric (GC-MS, "C NMR) techniques. In total, fifty-seven compounds accounting for 92.3-98.7% of the whole composition were identified. These oils were characterized by the preeminence of sesquiterpene hydrocarbons and the content of the main components varied substantially from sample to sample. Two samples were largely dominated by germacrene D (67.3% and 76.3%, respectively), a third sample was represented by germacrene D (28.3%), germacrene C (14.3%), δ-elemene (12.1%) and β-elemene (9.3%) and the fourth sample was characterized by 3-dimethylallylindole (15.1%), germacrene D (12.0%) and cyperene (7.8%). C NMR spectroscopy was particularly efficient for the identification of heat-sensitive compounds.

The chemical composition of trunk bark oil from Cleistopholis patens (Benth.) Engl. & Diels, growing wild in Côte d'Ivoire, has been investigated by GC (FID) in combination with retention indices, GC/MS and 13 C-NMR. Moreover, one oil sample has been subjected to CC and all the fractions analyzed by GC (RI) and 13 C-NMR. In total, 61 components have been identified, including various sesquiterpene esters scarcely found in essential oils. 13 C-NMR was particularly efficient for the identification of a component not eluted on GC and for the quantification of heat-sensitive compounds. Then, 36 oil samples, isolated from trunk bark harvested in six Ivoirian forests have been analyzed. The content of the main components varied drastically from sample to sample: (E)-β-caryophyllene (0.4 - 69.1%), β-pinene (0 - 57%), α-phellandrene (0 - 33.2%), α-pinene (0.1 - 30.6%), β-elemol (0.1 - 29.9%), germacrene D (0 - 25.4%), juvenile hormone III (0 - 22.9%), germacrene B (0 - 20.6%) and sabinene (tr-20.3%). Statistical analysis, hierarchical clustering and principal components analysis, carried out on the 36 compositions evidenced a fair chemical variability of the stem bark oil of this species. Indeed, three clusters have been distinguished: the composition of group I (ten samples) was dominated by β-pinene and α-pinene, group II (nine samples) was represented by α-phellandrene and p-cymene and group III (16 samples) by β-elemol. A sample displayed an atypical composition dominated by (E)-β-caryophyllene., (© 2017 Wiley-VHCA AG, Zurich, Switzerland.)

Forty-two essential oil samples were isolated from leaves of Xylopia rubescens harvested in three forests of Southern Ivory Coast. All the samples have been submitted to GC-FID and the retention indices (RIs) of individual components have been measured on two capillary columns of different polarity. In addition, 20 oil samples, selected on the basis of their chromatographic profile, were also analyzed by 13 C-NMR and 24 components (78.0 - 92.4% of the whole compositions) have been identified. The content of the main components varied drastically from sample to sample: furanoguaia-1,4-diene (5.7 - 54.1%), furanoguaia-1,3-diene (1.1 - 10.5%), (8Z,11Z,14Z)-heptadeca-8,11,14-trien-2-one (4.3 - 16.0%), and (E)-β-caryophyllene (1.7 - 17.3%). Hierarchical cluster and principal components analysis of the 42 oil compositions allowed the distinction of two well-differentiated groups of unequal importance within the oil samples. Oil samples of the main group (Group II) contained mainly furanoguaia-1,4-diene (mean [M] = 43.1%; standard deviation [SD] = 3.2%) while furanoguaia-1,3-diene (M = 8.4%; SD = 0.9%) and (8Z,11Z,14Z)-heptadeca-8,11,14-trien-2-one (M = 7.1%; SD = 1.5%) were present at appreciable contents. The composition of Group I was dominated by furanoguaia-1,4-diene (M = 17.0%; SD = 8.5%), (8Z,11Z,14Z)-heptadeca-8,11,14-trien-2-one (M = 10.2%; SD = 2.4%) and (E)-β-caryophyllene (M = 9.5%; SD = 5.3%)., (© 2017 Wiley-VHCA AG, Zurich, Switzerland.)

In order to get better knowledge about the volatiles produced by Xanthocyparis vietnamensis, a species recently discovered in Vietnam, its wood oil has been analyzed by a combination of chromatographic (GC, CC) and spectroscopic (GC-MS, (13)C-NMR) techniques. Forty components that accounted for 87.9% of the oil composition have been identified. The composition is dominated by nootkatene (20.7%), 11,12,13-tri-nor-eremophil-1(10)-en-7-one (17.2%), γ-eudesmol (5.1%), nootkatone (4.7%), valencene (3.5%) and 13-nor-eremophil-1(10)-en-11-one (2.6%). The structure of two new compounds-10-epi-nor-γ-eudesmen-11-one and 12-hydroxy-isodihydroagarofuran-has been elucidated, while 11,12,13-tri-nor-eremophil-1(10)-en-7-ol is reported as a natural product for the first time. The composition of X. vietnamensis wood oil varied drastically from those of leaf oils, dominated by hedycaryol (34.4%), phyllocladene (37.8%) or by pimara-6(14)-15-diene (19.4%).

The genus Myrtus L. (Myrtaceae family) comprises two species, Myrtus communis L. (known as common myrtle) growing wild all around the Mediterranean basin and Myrtus nivellei Batt. and Trab. (known as Saharan myrtle), found in central Sahara. Only one country, Algeria, hosts both species, M. communis in the North and M. nivellei in the South. The aim of this review was to collect, summarize, and compare the main results reported relative to the essential oils isolated from aerial parts of both species: botanical aspects, habitat, traditional use, chemical composition, new compounds, antimicrobial activity, antioxidant activity, anti-inflammatory effect, and insecticidal activity. Both essential oils have potential applications in human health., (© 2016 Verlag Helvetica Chimica Acta AG, Zürich.)

The composition of oil samples isolated from needles of Pinus halepensis growing in three locations in Corsica (Saleccia, Capo di Feno, and Tre Padule) has been investigated by combination of chromatographic (GC with retention indices) and spectroscopic (MS and (13)C-NMR) techniques. In total, 35 compounds that accounted for 77 - 100% of the whole composition have been identified. α-Pinene, myrcene, and (E)-β-caryophyllene were the major component followed by α-humulene and 2-phenylethyl isovalerate. Various diterpenes have been identified as minor components. 47 Oil samples isolated from pine needles have been analyzed and were differentiated in two groups. Oil samples of the first group (15 samples) contained myrcene (M = 28.1 g/100 g; SD = 10.6) and (E)-β-caryophyllene (M = 19.0 g/100 g; SD = 2.2) as major components and diterpenes were absent. All these oil samples were isolated from pine needles harvested in Saleccia. Oil samples of the second group (32 samples) contained mostly (E)-β-caryophyllene (M = 28.7 g/100 g; SD = 7.9), α-pinene (M = 12.3 g/100 g; SD = 3.6), and myrcene (M = 11.7 g/100 g; SD = 7.3). All these oil samples were isolated from pine needles harvested in Capo di Feno and Tre Padule., (© 2016 Verlag Helvetica Chimica Acta AG, Zürich.)

The chemical composition of 45 essential oil samples isolated from the leaves of Polyalthia oliveri harvested in three Ivoirian forests was investigated by GC-FID (retention indices measured on two columns of different polarities), and by (13) C-NMR, following a method developed in our laboratory. In total, 41 components were identified. The content of the main components varied drastically from sample to sample: (E)-β-caryophyllene (1.2 - 50.8%), α-humulene (0.6 - 47.7%), isoguaiene (0 - 27.9%), alloaromadendrene (0 - 24.7%), germacrene B (0 - 18.3%), δ-cadinene (0.4 - 19.3%), and β-selinene (0.2 - 18.5%). The analysis of six oil samples selected in function of their chromatographic profiles is reported in detail. The 45 oil compositions were submitted to hierarchical cluster and principal components analysis, which allowed the distinction of three groups within the oil samples. The compositions of the oils from group I (15 samples) and II (12 samples) were dominated by (E)-β-caryophyllene and α-humulene, respectively. Oil samples of group III (18 samples) needed to be partitioned into four subgroups III.1-III.4 whose compositions were dominated by alloaromadenrene, isoguaiene, germacrene B, and δ-cadinene, respectively., (© 2016 Verlag Helvetica Chimica Acta AG, Zürich.)

One oil sample isolated from aerial parts of Pulicaria mauritanica Coss. from Western Algeria has been analyzed by GC(RI), GC-MS and ¹³C NMR. In total, 21 components, accounting for 97.0% of the oil, were identified. Then, 36 oil samples coming from plants harvested at two flowering periods in three locations were analyzed by GC(RI) and ¹³C NMR. Although all the oil samples exhibited similar composition, dominated by carvotanacetone (89.2-96.1%), the yield of essential oil varied drastically from sample to sample (0.35-1.44%), depending on the location of harvest. The essential oil displayed moderate antimicrobial effect against bacteria, yeast and filamentous fungi (MIC = 2-4 µL/mL).

Deodarone [2,2,6-trimethyl-6-(4-methylcyclohex-3-enyl)-tetrahydro-4-pyrone] is a sesquiterpene tetrahydro-γ-pyrone related to bisabolene and atlantone, first isolated from Cedrus deodora essential oil. With respect to the stereochemistry of the asymmetric carbons C4 and C8, two diastereoisomers may be distinguished. Identification and quantification of both diastereoisomers in wood and tar oils from C. atlantica has been achieved using 13C NMR spectroscopy, in combination with GC (polar column). The contents of (4R,8R)- and (4R,8S)-deodarone varied between 1.1-2.8% and 1.0-3.0%, respectively.

The objective was to investigate the yield and chemical composition of 50 essential oil samples isolated from leaves and flowers of Tetraclinis articulata harvested in eight locations (coastal township and highlands) of Tlemcen Province (North-Western Algeria). Essential oil yields varied drastically from sample to sample (0.03 to 0.86%, w/w). No direct correlation was observed between the yield and the altitude of the harvest areas. The oils consisted mainly of monoterpenes: α-pinene (9.2-56.5%), bornyl acetate (1.2-45.1%), camphor (0.5-40.3%), borneol (0.2-12.9%), limonene (3.6-12.5%), and myrcene (1.6-9.7%). Sesquiterpenes were represented by germacrene D (up to 14.2%) and (E)-β-caryophyllene (up to 13.3%). PCA analysis of the data allowed the distinction of two groups within the samples. The composition of group I (9 samples) was dominated by camphor, (Mean = 30.9%) followed by α-pinene (M = 19.1%) and bornyl acetate (M = 11.4%). Group II was divided into two sub-groups. Samples of sub-group IIA (8 samples) contained mainly α-pinene (M = 45.4%). Samples of the largest group IIB (33 samples) were characterized by similar contents of α-pinene (M = 28.2%) and bornyl acetate (M = 24.5%) and the occurrence of camphor to a lesser extent (M = 10.0%).

The chemical composition of a leaf oil sample from Ivoirian Xylopia staudtii Engler & Diels (Annonaceae) has been investigated by a combination of chromatographic [GC(RI)] and spectroscopic (GC-MS, 13C NMR) techniques. Thirty-five components that accounted for 91.8% of the whole composition have been identified. The oil composition was dominated by the furanoguaiadienes furanoguaia-1,4-diene (39.0%) and furanoguaia-1,3-diene(7.5%), and by germacrene D (17.5%). The composition of twelve other leaf oil samples demonstrated qualitative homogeneity, but quantitative variability. Indeed, the contents of the major components varied substantially: furanoguaia-1,4-diene (24.7-51.7%) and germacrene D (5.9-24.8%). The composition of X. staudtii leaf oil is close to that of X. rubescens leaf oil but varied drastically from those of the essential oils isolated from other Xylopia species. 13C NMR spectroscopy appeared as a powerful and complementary tool for analysis of sesquiterpene-rich essential oils.

In Algeria, Myrtus communis L. is distributed throughout the Tell Atlas and the coastal regions of Algiers and Constantine. The leaves are used in respiratory disorders, diarrhea and hemorrhoids. The aims of this work were to evaluate the antifungal and anti-inflammatory potential of well characterized essential oils (EO). Since EO can be applied by inhalation, dermal application and oral administration, we used several mammalian cell lines to assess safe bioactive doses. The chemical composition of two samples was investigated by GC-FID, GC-MS and (13)C NMR spectroscopy. Monoterpene derivatives are the main compounds: α-pinene (50.8 and 33.6%), 1,8-cineole (21.9 and 13.3%), linalool (2.7 and 14.8%), and linalyl acetate (0.5 and 9.5%). The antifungal evaluation revealed that the oils were more active against Cryptococcus neoformans (yeast) and Epidermophyton floccosum, Microsporum canis, Trichophyton rubrum (dermatophytes). The anti-inflammatory potential was evaluated using an in vitro model of lipopolysaccharide (LPS)-stimulated macrophages. Assessment of cell viability was made through the MTT assay. Both oils were able to significantly inhibit NO production, without affecting cell viability, in concentrations up to 0.64 mg/mL. These promising results, disclose bioactive concentrations of Myrtle essential oils with a safety profile suggesting a potential oral and topical application or use by inhalation., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

The chemical composition of Vietnamese oil samples of the aerial parts of Clausena indica (Dalz.) Oliver and C. anisum-olens (Blanco) Merryll have been investigated using a combination of chromatographic and spectroscopic techniques. C. indica essential oil contained mainly terpinolene (53.9 and 56.1%), and myristicin (17.9 and 7.3%), whereas the major components of C. anisun-olens essential were citronellal (22.8%), geranial (21.4%) and neral (16.8%). The compositions of the investigated samples have been compared with those of essential oils from various origins.

The composition of the essential oil isolated from cones of Pinus halepensis grown in Corsica has been investigated by a combination of chromatographic (CC, GC) and spectroscopic (MS, 13C NMR) techniques. In total, 48 compounds that accounted for 95.5% of the whole composition have been identified. α-Pinene (47.5%) was the major component followed by myrcene (11.0%), (E)-β-caryophyllene (8.3%) and caryophyllene oxide (5.9%). Various diterpenes have been identified by 13C NMR in the fractions of CC. Fifteen oil samples isolated from cones harvested in three forests have been analyzed and two groups of similar importance have been differentiated within the 15 compositions. Oil samples of the first group contained α-pinene (mean 45.0 g/100 g, SD = 5.5) as the major component. The composition of samples of the second group was dominated by myrcene (mean 30.3 g/100g, SD = 9.0) and α-pinene (mean 24.6 g/100 g, SD = 3.1).

The composition of eight oil samples isolated from leaves of Eucalyptus bosistoana F. Muell., acclimatized in Tamanrasset (southern Algeria, Saharan climate), has been investigated by GC (retention indices), GC-MS and 13C NMR spectroscopy. T wo groups may be distinguished. The five samples of group I have their composition dominated by p-cymene (32.0-39.5%), cryptone (11.5-15.6%), 1,8-cineole (7.8-10.5%) and spathulenol (6.8-16.5%). The three oil samples of group II contained mainly 1,8-cineole (55.3-63.9%) and alpha-pinene (11.6-12.1%).

Three fruit oil samples of Ammodaucus leucotrichus Cosson & Durieu from Algerian Sahara were obtained by hydrodistillation and analyzed by GC(RI), GC-MS and 13C NMR spectroscopy. The main compounds were perillaldehyde (87.0-87.9%) and limonene (7.4-8.2%). The antimicrobial effect of the essential oil was evaluated against bacteria, yeasts and filamentous fungi. High antibacterial activity was observed against Escherichia coli, Staphylococcus aureus. Enterobacter cloaceae, Bacillus cereus and Salmonella typhimurium, with MIC values between 0.5-1.0 microL/mL. Fungal strains were also sensitive to the essential oil (MIC values: 0.25-0.75 microL/mL).The most potent activity was observed against the filamentous fungi, Fusarium oxysporum and Aspergillusflavus (0.25-0.50 microL/mL).

The chemical composition of 42 essential-oil samples isolated from the leaves of Xylopia quintasii harvested in three Ivoirian forests was investigated by GC-FID, including the determination of retention indices (RIs), and by (13) C-NMR analyses. In total, 36 components accounting for 91.9-92.6% of the oil composition were identified. The content of the main components varied drastically from sample to sample: (E)-β-caryophyllene (0.9-56.9%), (Z)-β-ocimene (0.3-54.6%), β-pinene (0.8-27.9%), α-pinene (0.1-22.8%), and furanoguaia-1,4-diene (0.0-17.6%). The 42 oil compositions were submitted to hierarchical cluster and principal components analysis, which allowed the distinction of three groups within the oil samples. The composition of the oils of the major group (22 samples) was dominated by (E)-β-caryophyllene. The oils of the second group (12 samples) contained β-pinene and α-pinene as the principal compounds, while the oils of the third group (8 samples) were dominated by (Z)-β-ocimene, germacrene D, (E)-β-ocimene, and furanoguaia-1,4-diene. The oil samples of Group I and II came from clay-soil forests, while the oil samples belonging to Group III were isolated from leaves harvested in a sandy-soil forest., (Copyright © 2014 Verlag Helvetica Chimica Acta AG, Zürich.)

Croton kimosorum Leandri is an endemic species to Madagascar. The chemical composition of aerial parts, leaf and stem oils is reported for the first time. Analysis was carried out by combination of chromatographic (CC, GC), spectroscopic and spectrometric (MS, 13C NMR) techniques. In total, 76 compounds have been identified. Essential oil isolated from aerial parts contained mainly linalool (21.6%), sabinene (10.4%), 1,8-cineole (6.3%), beta-pinene (6.2%), (E)-beta-caryophyllene (5.9%), terpinen-4-ol (4.8%), geraniol (4,5%) and germacrene D (2.3%). In comparison with the first sample, the composition of leaf and stem oils varied slightly, while essential oil isolated by vapor distillation from a semi-industrial still exhibited similar composition.

The composition of root bark oil from Cleistopholis glauca Pierre ex Engler & Diels growing wild in Ivory Coast was investigated by GC (in combination with retention indices) and "3C NMR spectroscopy after partition o f hydrocarbons a nd oxygenated compounds on silica g el. Thirty-one compounds havebeen identified. C. glauca produces a sesquiterpene-rich oil, patchoulenone (33.5%), cyperene (9.5%) and germacrene D (6.6%) being the main components. Special attention was paid to the identification and quantification ofgermacrene C (a heat-sensitive compound) and &-elemene, which were achieved by a combination of GC(FID) and 13C NMR spectroscopy. The composition of C. glauca root bark and leaf oils differed drastically.

In the course of our ongoing work on the chemical characterization of Taxus baccata L. growing wild in Corsica, we have developed and validated a method for direct quantification of taxane derivatives by (13)C NMR using 10-deacetylbaccatin III as reference compound and 1,6-hexanediol as internal standard. We have observed good accuracy (relative errors between 0.3% and 3.5%), linearity (R(2) = 0.999) and precision (reproducibility 8.5 mg ± 1.1%) of the measurements. The experimental procedure was applied to the quantification of six identified taxanes in a fraction of chromatography of a methanol extract of T. baccata leaves. This method can be applied to other compounds bearing the taxane skeleton., (Copyright © 2013 John Wiley & Sons, Ltd.)

The chemical composition of 48 leaf oil samples isolated from individual plants of Cleistopholis patens (Benth.) Engl. et Diels harvested in four Ivoirian forests was investigated by GC-FID (determination of retention indices), GC/MS, and (13) C-NMR analyses. The main components identified were β-pinene (traces-59.1%), sabinene (traces-54.2%), (E)-β-caryophyllene (0.3-39.3%), linalool (0.1-38.5%), (E)-β-ocimene (0.1-33.2%), germacrene D (0.0-33.1%), α-pinene (0.1-32.3%), and germacrene B (0-21.2%). The 48 oil compositions were submitted to hierarchical clustering and principal components analyses, which allowed the distinction of three groups within the oil samples. The oil composition of the major group (GroupI, 33 samples) was dominated by (E)-β-caryophyllene and linalool. The oils of Group II (eight samples) contained mainly β-pinene and α-pinene, while those of Group III (seven samples) were dominated by sabinene, limonene, and β-phellandrene. Moreover, the compositions of the Ivoirian C. patens leaf oils differed from those of Nigerian and Cameroonian origins., (Copyright © 2013 Verlag Helvetica Chimica Acta AG, Zürich.)

Introduction: Germacrenes A-C are secondary metabolites produced by various plants. They are sesquiterpene hydrocarbons bearing the (E,E)-1,5-cyclodecadiene structure known to undergo thermal rearrangement through a [3.3]-sigmatropic reaction. Such a rearrangement was evidenced by comparing the contents of a given germacrene and the corresponding elemene calculated by GC-flame ionisation detection (FID) with the relative intensities of the signals of both molecules in the (13) C-NMR spectrum of the mixture, recorded at room temperature., Objective: To develop a protocol to identify and quantify germacrenes A, B and C and in parallel the corresponding elemenes, using a combination of GC-FID and (13) C-NMR and then provide a correct analysis of Cleistopholis patens essential oils., Methods: The essential oil was submitted to GC-FID, GC-retention index, GC-MS and (13) C-NMR analyses. The relative percentages of every couple of germacrene and elemene measured by GC-FID were summed. Then, the relative ratio of the mean intensities of the signals of the protonated carbons of a given germacrene and the corresponding elemene was calculated. The contents of both compounds were obtained by combining GC-FID and (13) C-NMR data., Results: The true content of germacrene A/β-elemene, germacrene B/γ-elemene and germacrene C/δ-elemene in leaf and root oils from C. patens was evaluated by combination of GC-FID and (13) C-NMR data. Correct analysis of the essential oils was provided., Conclusion: Combined analysis of essential oil including (13) C-NMR without isolation of the components, appeared really efficient to identify and quantify germacrene isomers and in parallel elemene isomers in essential oils., (Copyright © 2013 John Wiley & Sons, Ltd.)

Ethnopharmacologic Relevance: Myrtus nivellei Batt. & Trab. (Myrtaceae) known as Sahara myrtle is appreciated by the Touaregs as medicinal plant. Infusion of leaves is employed against diarrhea and blennorrhea. Crushed leaves added to oil or to butter ointment have been traditionally used for the treatment of dermatosis. Aim of the study is to consider the traditional medicinal uses and the lack of scientific studies on their biological activities, the present study was designed to elucidate the chemical composition, the antifungal activity of its essential oils against fungi responsible for human infections, as well as, its cytotoxicity in the mammalian keratinocytes., Materials and Methods: Chemical analysis of Myrtus nivellei essential oil isolated by hydrodistillation of aerial parts (leaves and flowers), was carried out using a combination of chromatographic (CC, GC with retention indices) and spectroscopic techniques (MS, (13)C NMR, 2D NMR). The antifungal activity was evaluated by using broth macrodilution methods for yeasts and filamentous fungi. Cytotoxicity was tested in HaCaT keratinocytes through the MTT assay., Results: Ten samples coming from two localities of harvest were investigated. The chemical composition was largely dominated by 1,8-cineole (33.6-50.4%) and limonene (17.5-25.0%). The structure of two new compounds bearing the isoamylcyclopentane skeleton has been elucidated. The oil was more active against Cryptococcus neoformans with MIC of 0.16μL/mL followed by dermatophytes, with MICs of 0.64 and 1.25μL/mL. Furthermore, evaluation of cell viability showed no cytotoxicity in HaCaT keratinocytes at concentrations up to 1.25µL/mL., Conclusions: The composition of Myrtus nivellei oil differed from that of Myrtus communis. The structure of two di-nor-sesquiterpenoids has been elucidated. It was possible to find appropriate doses of Myrtus nivellei oil with both antifungal activity and very low detrimental effect on keratinocytes. These findings add significant information to the pharmacological activity of Myrtus nivellei essential oils, specifically to its antifungal properties, thus justifying and reinforcing the use of this plant in traditional medicine., (Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.)

The volatile components isolated from leaves of Vepris madagascarica (Baillon) H. Perier (Rutaceae), an endemic species of north-eastern, sub-humid forests of Madagascar, were investigated by GC (Retention Indices), GC-MS and 13C NMR spectroscopy. Oil samples obtained on laboratory and industrial scales exhibited similar composition, dominated by phenylpropanoids. (E)-anethole (78.2% and 78.6%) was the major component followed by estragole (15.6% and 15.4%). In addition, trunk bark oil also contained (E)-anethole as its major component (84.6%), as well as various sesquiterpenes in low contents.

Essential oils (EO, 15 collective samples and 47 individual samples) of Thymbra capitata collected from Moroccan Western Rif were analyzed using gas chromatography (GC) in combination with retention indices (RI), gas chromatography-mass spectrometry (GC-SM) and 13C NMR spectroscopy. Twenty components were identified. Carvacrol (68.2%-85.9%) was by far the major component of all the samples, while the content of thymol (0.1-0.3%) was very low. Other components present in appreciable amounts were gamma-terpinene (up to 8.9%), p-cymene (up to 7.1%), linalool (up to 4.4%) and (E)-beta-caryophyllene (up to 4.1%). In contrast, the yield of EO varied drastically from sample to sample (0.5-3.7%). No correlation could be established between yield of EO and altitude, pH, chemical composition and granularity of the soil. Cultivation under controlled conditions is suggested to improve the quantitative characteristics of carvacrol-rich Moroccan T. capitata.

The essential oil isolated from Asarum cordifolium C. E. C. Fischer recently discovered in Vietnam, and A. glabrum Merr., an endangered species listed as vulnerable in the Red Data Book of Vietnam, have been analyzed by a combination of chromatographic and spectroscopic techniques including 13C NMR spectroscopy. The composition of A. cordifolium essential oil, investigated for the first time, was dominated by elemicin (82.5%). The essential oil isolated from A. glabrum contained safrole (41.9%) as its major component and was characterized by the diversity of phenylpropanoids contained in this oil (10 compounds).

The composition of 55 samples of essential oil isolated from the aerial parts of wild growing Myrtus communis L. harvested in 16 locations from East to West Algeria were investigated by GC (determination of retention indices) and (13) C-NMR analyses. The essential oils consisted mainly of monoterpenes, α-pinene (27.4-59.2%) and 1,8-cineole (6.1-34.3%) being the major components. They were also characterized by the absence of myrtenyl acetate. The compositions of the 55 oils were submitted to k-means partitioning and principal component analysis, which allowed the distinction of two groups within the oil samples, which could be subdivided into two subgroups each. Groups I (78% of the samples) and II were differentiated on the basis of the contents of α-pinene, linalool, and linalyl acetate. Subgroups IA and IB could be distinguished by their contents of α-pinene and 1,8-cineole. Subgroups IIA and IIB differed substantially in their contents of 1,8-cineole and limonene. All the samples contained 3,3,5,5,8,8-hexamethyl-7-oxabicyclo[4.3.0]non-1(6)-ene-2,4-dione (up to 4.9%)., (Copyright © 2013 Verlag Helvetica Chimica Acta AG, Zürich.)

The chemical composition of 48 essential-oil samples isolated from the leaves of Xylopia aethiopica harvested in six Ivoirian forests was investigated by GC-FID and (13) C-NMR analyses. In total, 23 components accounting for 82.5-96.1% of the oil composition were identified. The composition was dominated by the monoterpene hydrocarbons β-pinene (up to 61.1%) and α-pinene (up to 18.6%) and the sesquiterpene hydrocarbon germacrene D (up to 28.7%). Hierarchical cluster and principal component analyses allowed the distinction of two groups on the basis of the β-pinene and germacrene D contents. The chemical composition of the oils of Group I (38 oil samples) was clearly dominated by β-pinene, while those of Group II (10 samples) were characterized by the association of β-pinene and germacrene D. The leaves collected in the four inland forests produced β-pinene-rich oils (Group I), while the oil samples belonging to Group II were isolated from leaves harvested in forests located near the littoral., (Copyright © 2012 Verlag Helvetica Chimica Acta AG, Zürich.)

The chemical composition of 50 samples of leaf oil isolated from Algerian Juniperus phoenicea var. turbinata L. harvested in eight locations (littoral zone and highlands) was investigated by GC-FID (in combination with retention indices), GC/MS, and (13) C-NMR analyses. The composition of the J. phoenicea var. turbinata leaf oils was dominated by monoterpenes. Hierarchical cluster and principal component analyses confirmed the chemical variability of the leaf oil of this species. Indeed, three clusters were distinguished on the basis of the α-pinene, α-terpinyl acetate, β-phellandrene, and germacrene D contents. In most oil samples, α-pinene (30.2-76.7%) was the major compound, associated with β-phellandrene (up to 22.5%) and α-terpinyl acetate (up to 13.4%). However, five out of the 50 samples exhibited an atypical composition characterized by the predominance of germacrene D (16.7-22.7%), α-pinene (15.8-20.4%), and α-terpinyl acetate (6.1-22.6%)., (Copyright © 2012 Verlag Helvetica Chimica Acta AG, Zürich.)

Introduction: In the course of on-going work on the characterisation of aromatic plants from the Ivory Coast we investigated the composition of the root oil from Xylopia aethiopica., Objectives: The aim of this work was to investigate the chemical composition of X. aethiopica root oil and elucidate the structure of two new compounds., Methodology: Analysis of the essential oil was carried out using a combination of chromatographic (CC, GC with retention indices) and spectroscopic techniques (MS, (13)C-NMR, 2D-NMR)., Results: Twenty seven components, accounting for 95.6% of the whole composition, were identified including various compounds for which spectroscopic data were absent on commercial computerised MS libraries. Three compounds are reported for the first time as natural compounds and the structure of two new compounds, 4,4-dimethyl-2-vinylcyclohexene and endo-5-methoxy-3-patchoulene, has been elucidated using extensive two-dimensional NMR spectroscopy., Conclusion: The composition of X. aethiopica root oil is dominated by two dimethylvinylcyclohexene isomers. It differs drastically from the composition of leaf and fruit oils of the same plant. The combination of analytical techniques appeared crucial for a fruitful analysis., (Copyright © 2012 John Wiley & Sons, Ltd.)

Identification and quantitative determination of individual components of resin collected on the trunk of 28 Cedrus atlantica trees, grown in Corsica, has been carried out using 13C NMR spectroscopy. Eight resin acids bearing either the pimarane or abietane skeleton, two monoterpene hydrocarbons and four oxygenated neutral diterpenes have been identified, as well as three lignans, scarcely found in resins. Three groups could be distinguished within the 28 resin samples. The nine samples of Group I had their composition dominated by diterpene acids (33.7-45.8%), with abietic acid (6.2-18.7%) and isopimaric acid (5.1-12.6%) being the major components. The four samples of Group II contained resin acids (main components) and lignans in moderate amounts (up to 10.3%). Conversely, lignans (38.8-63.8%) were by far the major components of the 15 samples of Group III. Depending on the sample, the major component was pinoresinol (18.1-38.9%), lariciresinol (17.2-33.7%) or lariciresinol 9'-acetate (16.9-29.1%). Finally, due to the high biological interest in lignans, a rapid procedure, based on 1H NMR spectroscopy, was developed for quantification of lignans in resins of C. atlantica.

The composition of a volatile extract of Eryngium duriaei subsp. juresianum, signalised by the antifungal activity (MIC values=0.16-0.32 μL mL(-1)) against several dermatophyte species (Trichophyton mentagrophytes, T. rubrum, Epidermophyton floccosum; T. verrucosum, T. mentagrophytes var interdigitale, Microsporum canis and M. gypseum) was established following a combined methodology of GC, GC-MS and an exclusive (13)C NMR technique that does not require prior isolation of compounds. Twenty-five components were identified accounting 84.6% of the whole composition. Major compound was found to be α-neocallitropsene (26.0%) although the dominance of caryophyllane derived compounds, the most probable responsible for the antifungal activity, namely isocaryophyllen-14-al (16.2%), 14-hidroxy-β-caryophyllene (13.4%), caryophyllene oxide (7.6%) and E-β-caryophyllene (6.3%)., (Copyright © 2010 Elsevier B.V. All rights reserved.)