RESUMO
Different plant parts of Ligusticopsis wallichiana (family: Apiaceae) are widely used as traditional medicines. Although many volatile constituents are already identified from the leaves of L. wallichiana, there is no detailed report on the nonvolatile constituents. In the present study, we aimed to isolate and identify the major chemical constituents from the leaves. Bhutkesoside A (1), falcarindiol (2), ferulic acid (3), cnidioside A (4), quercetin 3-O-ß-D-glucopyranoside (5), rutin (6), 4'-O-methylquercetin 3-O-ß-D-glucopyranoside (7), scopoletin (8), umbelliferone (9), eugenol 4-O-ß-D-glucopyranoside (10) and pumilaside A (11) were isolated from the 70% MeOH extract. The structures of isolated compounds were elucidated on the basis of 1H- and 13C-NMR spectroscopic data. Compounds 4-11 are reported for the first time from L. wallichiana. Compounds 5 and 6 showed potent free radical-scavenging activity.
RESUMO
The rhizomes of Astilbe rivularis, commonly known as 'Thulo Okhati' are widely used in Nepal as tonic for uterine and menstrual disorders. In our preliminary study, the 70% MeOH extract of the rhizomes showed potent antioxidant activity. Hence, present study was aimed for the isolation of potent antioxidant constituents. Bergenin (1), 11-O-galloylbergenin (2), (+)-catechin (3), (-)-catechin (4), (-)-afzelechin (5), (-)-epiafzelechin (6) and 2-(ß-D-glucopyranosyloxy)-4-hydroxylbenzenacetonitrile (7) were isolated from the rhizomes. Structures of these compounds were elucidated on the basis of spectroscopic methods. All these isolated compounds were evaluated for their in vitro antioxidant activity by 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging assay. 11-O-Galloylbergenin (2), (+)-catechin (3), (-)-catechin (4), (-)-afzelechin (5) and (-)-epiafzelechin (6) showed potent antioxidant activity.
Assuntos
Antioxidantes/química , Antioxidantes/farmacologia , Saxifragaceae/química , Antioxidantes/isolamento & purificação , Benzopiranos/química , Benzopiranos/isolamento & purificação , Benzopiranos/farmacologia , Compostos de Bifenilo/química , Catequina/isolamento & purificação , Catequina/farmacologia , Avaliação Pré-Clínica de Medicamentos/métodos , Flavonoides/química , Flavonoides/isolamento & purificação , Flavonoides/farmacologia , Ácido Gálico/análogos & derivados , Ácido Gálico/química , Ácido Gálico/isolamento & purificação , Ácido Gálico/farmacologia , Espectroscopia de Ressonância Magnética , Estrutura Molecular , Nepal , Fenóis/química , Fenóis/isolamento & purificação , Fenóis/farmacologia , Picratos/química , Extratos Vegetais/química , Rizoma/químicaRESUMO
Four diarylpentanoids (1-4), two phenylpropanoids (5-6), three biflavonoids (7-9), two lignans (10-11) and a coumarin (12) were isolated from the roots of Diploniorpha sikokiana (Franchet & Savatier) Honda (Family: Thymelaeaceae). Similarly, two phenylpropanoids (5-6), five biflavonoids (7-9, 13, 14), three lignans (11, 15, 16), a flavonoid (17) and two coumarins (12, 18) were isolated from the stems, and five flavonoids (17, 19-22) from the leaves. Among them, 1,5-diphenyl-l-pentanone (1), 1,5-diphenyl-2-penten-l-one (2), 3(S)-hydroxy-1,5-diphenylpentane (3), stelleranol (8), (-)-syringaresinol (10), (-)-pinoresinol 4,4'-di-O-ß-D-glucopyranoside (11), apiosylskimmin (12), syringaresinol 4-O-ß-D-glucopyranoside (16), quercetin 3-O-α-L-rhamnopyranoside (19), kaempferol 3-0-α-L-rhamnopyranoside (20) and tiliroside (21) were isolated for the first time from D. sikokiana.
Assuntos
Thymelaeaceae/química , Componentes Aéreos da Planta/química , Raízes de Plantas/químicaRESUMO
Shrub Chaste Tree Fruit (SCTF) is defined as the fruits of Vitex rotundifolia L. f. and V. trifolia L. and has been used as a component of some traditional Japanese medicines (Kampo formulations). Agnus Castus Fruit (ACF) is defined as the dried ripe fruits of V. agnus-castus L.; it is used in traditional European medicines, but is becoming popular in Japan as both an over-the-counter drug and as an ingredient in health foods for treating premenstrual syndrome (PMS). To ensure the efficacy and safety of both SCTF and ACF products, it is important to precisely authenticate their botanical origins and to clearly distinguish between SCTF and ACF. Therefore, we tried to identify SCTF-specific marker compounds based on LC/MS metabolic analysis. The multivariate analysis of LC/MS data from SCTF and ACF samples furnished candidate marker compounds of SCTF. An SCTF-specific marker was isolated from SCTF crude drugs and identified as 3-O-trans-feruloyl tormentic acid on the basis of spectroscopic data from NMR and MS. Since avoiding contamination from closely related species is a significant requirement for pharmaceuticals of natural origin, this information will be valuable for the quality control of both SCTF and ACF products from the viewpoint of regulatory science.
Assuntos
Biomarcadores/análise , Cromatografia Líquida/métodos , Espectrometria de Massas por Ionização por Electrospray/métodos , Vitex/classificação , Espectroscopia de Ressonância Magnética , Especificidade da Espécie , Vitex/químicaRESUMO
Agnus Castus Fruit is defined in the European Pharmacopoeia as the dried ripe fruit of Vitex agnus-castus. In Europe it is used as a medicine targeting premenstrual syndrome and climacteric disorder. In Japan, Agnus Castus Fruit is becoming popular as a raw material for over-the-counter drugs and health food products, though its congenic species, Vitex rotundifolia and Vitex trifolia, have been used as Shrub Chaste Tree Fruit in traditional medicines. Therefore, it is important to discriminate these Vitex plants from the viewpoint of regulatory science. Here we tried to identify putative marker compounds that distinguish between Agnus Castus Fruit and Shrub Chaste Tree Fruit. We analyzed extracts of each crude drug by liquid chromatography-mass spectrometry, and performed differential analysis by comparison of each chromatogram to find one or more peaks characteristic of Agnus Castus Fruit. A peak was isolated and identified as an equilibrium mixture of new compounds named chastol (1) and epichastol (1a). The planar structures of 1 and 1a were determined spectroscopically. Their relative configurations were revealed by nuclear Overhauser effect spectroscopy and differential nuclear Overhauser effect-NMR data. Since avoiding contamination from closely related species is needed for the quality control of natural pharmaceuticals, this information will be valuable to establish a method for the quality control of both, Agnus Castus Fruit and Shrub Chaste Tree Fruit products.
Assuntos
Diterpenos/isolamento & purificação , Vitex/química , Vitex/classificação , Cromatografia Líquida , DNA de Plantas , Europa (Continente) , Frutas/química , Japão , Plantas Medicinais/química , Plantas Medicinais/classificação , Controle de Qualidade , Análise de Sequência de DNA , Especificidade da Espécie , Espectrometria de Massas em Tandem , Vitex/genéticaRESUMO
Two new diacetylene glycosides: bhutkesoside A (1) and B (2), along with 10 known compounds, i.e. falcarindiol (3), chlorogenic acid (4), 5-O-p-coumaroyl-quinic acid (5), 3,5-di-O-caffeoyl-quinic acid (6), 4-hydroxy-7-methoxy-phenylethanol (7), ferulic acid (8), dehydrodiconiferyl alcohol-4-O-ß-d-glucopyranoside (9), 5,7-dihydroxy-2-methylchromone-7-O-rutinoside (10), schumanniofioside B (11) and marmesinin (12) were isolated from the roots of Ligusticopsis wallichiana (DC) Pimenov & Kljuykov (Apiaceae), commonly known as 'Bhutkesh' in Nepal. The structures were determined on the basis of spectroscopic data. Compounds 4 and 6 showed potent antioxidant activity on DPPH free radical scavenging assay.
Assuntos
Apiaceae/química , Sequestradores de Radicais Livres/química , Sequestradores de Radicais Livres/farmacologia , Glicosídeos/isolamento & purificação , Compostos de Bifenilo , Glicosídeos/química , Glicosídeos/farmacologia , Hidrólise , Espectroscopia de Ressonância Magnética , Nepal , Picratos , Extratos Vegetais/química , Extratos Vegetais/farmacologia , Raízes de Plantas/química , Relação Estrutura-AtividadeRESUMO
A new flavonoid, dhasingreoside (1) and seven known compounds, quercetin 3-O-ß-D-galacturonopyranoside (2), quercetin 3-O-ß-D-galactopyranoside (3), quercetin 3-O-ß-D-glucuronopyranoside (4), quercetin 3-O-α-L-rhamnopyranoside (5), (-)-epicatechin (6), salicylic acid (7) and gaultherin (8), have been isolated from the shade-dried stems and leaves of Gaultheria fragrantissima, commonly known as 'Dhasingre' in Nepal. The structures were elucidated on the basis of physical, chemical and spectroscopic methods. Among known compounds, five compounds (3-6 and 8) were isolated for the first time from G. fragrantissima. In vitro antioxidant activity of all the isolated compounds was evaluated by 1,1-diphenyl-2-picrylhydrazyl free radical-scavenging assay. Dhasingreoside (1) and other compounds (2-6) showed significant free radical-scavenging activity.
Assuntos
Flavonoides/química , Sequestradores de Radicais Livres/química , Gaultheria/química , Flavonoides/isolamento & purificação , Sequestradores de Radicais Livres/isolamento & purificação , Nepal , Folhas de Planta/química , Caules de Planta/químicaRESUMO
A new C-glucosyl-alpha-hydroxydihydrochalcone derivative, (aS)-3'-C-ff-glucopyranosyl-a,3,4,2'4'-pentahydroxydihydrochalcone, named as bijayasaline (1) was isolated from the heartwood of Bijayasal (Pterocarpus marsupium) and characterized on the basis of NMR and circular dichroism (CD) spectral data. Bijayasaline (1) showed potent DPPH free radical scavenging activity.
Assuntos
Chalconas/química , Pterocarpus/química , Madeira/química , Compostos de Bifenilo/química , Sequestradores de Radicais Livres/química , Sequestradores de Radicais Livres/farmacologia , Estrutura Molecular , Picratos/químicaRESUMO
Eleven phenolic compounds, quercetin (1), quercetin 3-O-ß-d-galactopyranoside (2), quercetin 3-O-(6â³-O-galloyl)-ß-d-galactopyranoside (3), quercetin 3-O-(6â³-O-caffeoyl)-ß-d-galactopyranoside (4), quercetin 3-O-ß-d-glucopyranoside (5), rutin (6) quercetin 3-O-α-l-arabinopyranoside (7), quercetin 3-O-α-l-arabinofuranoside (8), protocatechulic acid (9), gallic acid (10) and chlorogenic acid (11), were isolated from the flowers of Aconogonon molle, a Nepalese medicinal plant. Structures of these compounds were elucidated on the basis of spectroscopic methods. All these compounds were isolated for the first time from flowers, and five compounds (4, 5, 8, 9 and 11) were isolated for the first time from A. molle. All of these isolated compounds were evaluated for their in vitro antioxidant activity by using the 1,1-diphenyl-2-picrylhydrazyl radical-scavenging method. Quercetin (1), quercetin glycosides (2-8) and gallic acid (10) exhibited potent antioxidant activity.
Assuntos
Antioxidantes/isolamento & purificação , Antioxidantes/farmacologia , Compostos de Bifenilo/farmacologia , Sequestradores de Radicais Livres/isolamento & purificação , Sequestradores de Radicais Livres/farmacologia , Fenóis/isolamento & purificação , Fenóis/farmacologia , Picratos/farmacologia , Plantas Medicinais/química , Polygonaceae/química , Antioxidantes/química , Ácido Clorogênico/isolamento & purificação , Flavonoides/química , Flavonoides/isolamento & purificação , Flavonoides/farmacologia , Flores/química , Sequestradores de Radicais Livres/química , Glucosídeos/química , Glucosídeos/isolamento & purificação , Glucosídeos/farmacologia , Glicosídeos/química , Glicosídeos/isolamento & purificação , Glicosídeos/farmacologia , Estrutura Molecular , Nepal , Fenóis/análise , Fenóis/química , Extratos Vegetais/química , Folhas de Planta/química , Quercetina/análogos & derivados , Quercetina/química , Quercetina/isolamento & purificação , Quercetina/farmacologiaRESUMO
Three new glycosides: thotneoside A (quercetin 3-O-(6â³-O-phenylacetyl)-ß-D-galactopyranoside) (1), thotneoside B (quercetin 3-O-(6â³-O-phenylacetyl)-ß-D-glucopyranoside) (2) and thotneoside C (3-methyl-2-butenoic acid 1-O-ß-D-glucopyranoside) (3), together with nine known compounds; quercetin (4), quercetin 3-O-ß-D-galactopyranoside (5), quercetin 3-O-(6â³-O-galloyl)-ß-D-galactopyranoside (6), quercetin 3-O-ß-D-galacturonopyranoside (7), quercetin 3-O-ß-D-glucuronopyranoside (8), quercetin 3-O-α-L-rhamnopyranoside (9), rutin (10), quercetin 3-O-α-L-arabinopyranoside (11) and 2,4,6-trihydroxyacetophenone 2-O-ß-D-glucopyranoside (12) have been isolated from the shade dried leaves of Aconogonon molle, commonly known as "Thotneâ³ in Nepal. The structures were elucidated on the basis of chemical and spectroscopic methods. All of these compounds were isolated for the first time from A. molle and their in vitro antioxidant activity was evaluated by 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical scavenging assay. Quercetin (4) and its glycosides (1-2, 5-11) showed potent free radical scavenging activity.
Assuntos
Antioxidantes/química , Antioxidantes/farmacologia , Glicosídeos/química , Glicosídeos/farmacologia , Folhas de Planta/química , Polygonaceae/química , Antioxidantes/isolamento & purificação , Compostos de Bifenilo/química , Radicais Livres/química , Glicosídeos/isolamento & purificação , Picratos/química , Quercetina/química , Quercetina/isolamento & purificação , Quercetina/farmacologiaRESUMO
A new flavone C-glycoside, 5-O-acetyl-2"-alpha-rhamnopyranosylisovitexin (1), was isolated from the aerial parts of Lychnis senno Siebold et Zucc. (Family Caryophyllaceae), along with 2"-alpha-rhamnopyranosylisovitexin (2), 2"-a-rhamnopyranosylisoorientin (3) and isoorientin 3'-mehyl ether (4). All these compounds were isolated for the first time from L. senno and their antioxidative activity was analyzed by the 2,2'-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS) radical-scavenging method. Compound 3 showed moderate antioxidative activity.
Assuntos
Flavonas/isolamento & purificação , Sequestradores de Radicais Livres/isolamento & purificação , Glicosídeos/isolamento & purificação , Lychnis/química , Flavonas/química , Glicosídeos/química , Estrutura Molecular , Plantas Medicinais/químicaRESUMO
A new dammarane-type triterpenoid saponin, chikusetsusaponin VII (1), and nineteen known triterpenoid saponins, ginsenoside Rb1 (2), ginsenoside Rb3 (3), ginsenoside Rc (4), ginsenoside Rd (5), ginsenoside Re (6), ginsenoside Rg1 (7), ginsenoside Rg2 (8), ginsenoside Rh1 (9), notoginsenoside R1 (10), notoginsenoside R2 (11), notoginsenoside Fe (12), chikusetsusaponin IVa (13), chikusetsusaponin IV (14), chikusetsusaponin V (15), chikusetsusaponin VI (16), chikusetsusaponin FK6 (17), gypenoside XVII (18), 28-desglucosylchikusetsusaponin IV (19), and zingibroside R1 (20), were isolated from rhizomes, taproots, and lateral roots of Panax japonicus C. A. Meyer, so-called "Satsuma-ninjin," grown in southern Miyazaki Prefecture, Japan. The structure of new chikusetsusaponin VII was elucidated on the basis of spectral and physicochemical evidence. Although the chemical composition of the rhizome was found to be similar to that of the "Chikusetsu-ninjin," the saponin composition of lateral root of "Satsuma-ninjin" was found to be close to that of lateral root of P. ginseng. The total yield of oleanolic acid saponins of the taproot was less than that of rhizome, but the total yield of dammarane-type saponins of the taproot was found to be similar to that of rhizome.
Assuntos
Ginsenosídeos/química , Panax/química , Saponinas/química , Terpenos/química , Japão , Raízes de Plantas/química , Rizoma/químicaRESUMO
A new flavonol triglycoside, myricetin-3-O-[ß-xylopyranosyl-(1-2)-α-rhamnopyranoside]-4'-O-α -rhamnopyranoside, named as bayarin (1), was isolated from the aerial parts of Zizyphus incurva Roxb. (Rhamnaceae) along with eight known compounds: six flavonoids and two saponins. All of these compounds were isolated for the first time from this plant. Structures of these compounds were elucidated on the basis of spectroscopic data.
Assuntos
Flavonoides/isolamento & purificação , Saponinas/isolamento & purificação , Ziziphus/química , Flavonoides/química , Estrutura Molecular , Plantas Medicinais/química , Saponinas/químicaRESUMO
Four new dammarane-type triterpenoid saponins such as chikusetsusaponin LM3 (1), chikusetsusaponin LM4 (2), chikusetsusaponin LM5 (3), chikusetsusaponin LM6 (4), and twenty known triterpenoid saponins such as ginsenoside Rb3 (5), ginsenoside Rc (6), ginsenoside Rd (7), ginsenoside Re (8), ginsenoside Rg1 (9), ginsenoside F3 (10), ginsenoside F5 (11), ginsenoside F6 (12), chikusetsusaponin IVa (13), chikusetsusaponin V (14), chikusetsusaponin L5 (15), chikusetsusaponin L9a (16), chikusetsusaponin L9bc (17), chikusetsusaponin L10 (18), chikusetsusaponin FK2 (19), chikusetsusaponin FK6 (20), chikusetsusaponin FK7 (21), chikusetsusaponin FT1 (22), chikusetsusaponin LM1 (23), and chikusetsusaponin LM2 (24), were isolated from the leaves of Panax japonicus C. A. MEYER collected in Miyazaki prefecture, Japan. The structures of new chikusetsusaponins were elucidated on the basis of spectral and physicochemical evidences.
Assuntos
Ginsenosídeos/química , Panax/química , Saponinas/química , Triterpenos/química , Sequência de Carboidratos , Ginsenosídeos/isolamento & purificação , Dados de Sequência Molecular , Folhas de Planta/química , Saponinas/isolamento & purificação , Triterpenos/isolamento & purificação , DamaranosRESUMO
Two new C-methyl flavonoids, diplomorphanins A (1) and B (2) were isolated from the aerial parts of Diplomorpha canescens (MEISN.) C. A. MEYER along with a known compound, farrerol 7-O-ß-D-glucopyranoside (3). Structures of these compounds were determined on the basis of spectroscopic data.
Assuntos
Flavonoides/química , Thymelaeaceae/química , Flavonoides/isolamento & purificação , Espectroscopia de Ressonância Magnética , Conformação Molecular , Componentes Aéreos da Planta/químicaRESUMO
The new flavonoid, pilloin 5-O-ß-D-glucopyranoside (1), was isolated from the stems of Diplomorpha ganpi (Family: Thymelaeaceae) together with 19 known compounds. The structures of these compounds were determined on the basis of spectroscopic data.
Assuntos
Flavonoides/isolamento & purificação , Glucosídeos/isolamento & purificação , Caules de Planta/química , Thymelaeaceae/química , Flavonoides/química , Glucosídeos/química , Espectroscopia de Ressonância Magnética , Estrutura MolecularRESUMO
Four new dammarane-type triterpenoid saponins, chikusetsusaponin FT(1) (1), chikusetsusaponin FT(2) (2), chikusetsusaponin FT(3) (3), chikusetsusaponin FT(4) (4), and six known triterpenoid saponins, chikusetsusaponin FK(4) (8), chikusetsusaponin FK(5) (9), chikusetsusaponin FK(2) (10), chikusetsusaponin FK(3) (11), chikusetsusaponin LN(4) (12), and chikusetsusaponin IVa (14), were isolated from the fruits of Panax japonicus C. A. MEYER, collected in Toyama prefecture, Japan, and five new dammarane-type triterpenoid saponins, chikusetsusaponin FT(1) (1), chikusetsusaponin FT(3) (3), chikusetsusaponin FT(4) (4), chikusetsusaponin FH(1) (5), chikusetsusaponin FH(2) (6), and eight known triterpenoid saponins, ginsenoside Re (7), chikusetsusaponin FK(5) (9), chikusetsusaponin FK(2) (10), chikusetsusaponin FK(3) (11), chikusetsusaponin LN(4) (12), 28-desglucosylchikusetsusaponin IVa (13), chikusetsusaponin IVa (14), and chikusetsusaponin V (15), were isolated from the fruits of P. japonicus C. A. MEYER, collected in Hokkaido, Japan. The structures of new chikusetsusaponins were elucidated on the basis of chemical and physicochemical evidences.
Assuntos
Frutas/química , Panax/química , Saponinas/química , Triterpenos/química , Japão , Espectroscopia de Ressonância Magnética , Modelos Moleculares , Estrutura MolecularRESUMO
Two new dammarane-type triterpenoid saponins, chikusetsusaponin LM1 (1), chikusetsusaponin LM2 (2), and three known triterpenoid saponins, ginsenoside Re (3), ginsenoside Rg1 (4), ginsenoside F3 (5), were isolated from the leaves of P. japonicus C. A. Meyer collected in Miyazaki prefecture, Japan. The structures of new chikusetsusaponins were elucidated on the basis of spectroscopic and physicochemical evidences.
Assuntos
Panax/química , Saponinas/isolamento & purificação , Triterpenos/isolamento & purificação , Japão , Estrutura Molecular , Folhas de Planta/química , Saponinas/química , Triterpenos/químicaRESUMO
One new C-methyl flavanone glucoside, farrerol 4'-O-ß-D-glucopyranoside (1) was isolated from the aerial parts of Diplomorpha canescens (MEISN.) C. A. MEYER. Fourteen known phenolic compounds such as farrerol (2), luteolin 7-methyl ether 5-O-ß-D-glucopyranoside (3), (-)-pinoresinol (4), (-)-lariciresinol (5), (-)-dihydrosesamin (6), (±)-dehydrodiconiferyl alcohol (7), rutarensin (8), umbelliferone (9), coniferyl aldehyde (10), sinapyl aldehyde (11), p-coumaric acid methyl ester (12), p-hydroxybenzaldehyde (13), p-hydroxyacetophenone (14) and syringaldehyde (15) were also isolated for the first time from this plant. Structure of 1 was determined on the basis of spectroscopic data including two dimensional (2D)-NMR, circular dichroism (CD) and by the application of Klyne's rule.
Assuntos
Flavonoides/química , Glucosídeos/química , Fenóis/química , Thymelaeaceae/química , Dicroísmo Circular , Flavonoides/isolamento & purificação , Glucosídeos/isolamento & purificação , Espectroscopia de Ressonância Magnética , Conformação Molecular , Fenóis/isolamento & purificação , Componentes Aéreos da Planta/químicaRESUMO
Seven new dammarane-type triterpenoid saponins, chikusetsusaponin FK1 (1), chikusetsusaponin FK2 (2), chikusetsusaponin FK3 (3), chikusetsusaponin FK4 (4), chikusetsusaponin FK5 (5), chikusetsusaponin FK6 (6), and chikusetsusaponin FK7 (7), and eleven known triterpenoid saponins, ginsenoside Rb3 (9), ginsenoside Rc (10), chikusetsusaponin VI (11), ginsenoside Re (12), ginsenoside Rg1 (13), pseudo-ginsenoside RS1 (14), notoginsenoside R1 (15), chikusetsusaponin L5) (17), chikusetsusaponin L10 (18), chikusetsusaponin IVa (19), and chikusetsusaponin V (20), were isolated from the fruits of Panax japonicus C. A. MEYER, collected in Kumamoto prefecture, Japan, and two new dammarane-type triterpenoid saponin, chikusetsusaponin FK5 (5) and chikusetsusaponin FM1 (8), and five known triterpenoid saponins, ginsenoside Rb3 (9), ginsenoside Rc (10), ginsenoside Re (12), ginsenoside Rg1 (13), and floralquinquenoside E (16), were isolated from the fruits of P. japonicus C. A. MEYER, collected in Miyazaki prefecture, Japan. The structures of new chikusetsusaponins were elucidated on the basis of chemical and physicochemical evidences.