Flavonoid diversification in different leaf compartments of Primula auricula (Primulaceae)

Populations of Primula auricula L. subsp. auricula from Austrian Alps were studied for flavonoid composition of both farinose exudates and tissue of leaves. The leaf exudate yielded Primula-type flavones, such as unsubstituted flavone and its derivatives, while tissue flavonoids largely consisted of flavonol 3-O-glycosides, based upon kaempferol (3, 4) and isorhamnetin (5–7). Kaempferol 3-O-(2″-O-β-xylopyranosyl-[6″-O-β-xylopyranosyl]-β-glucopyranoside) (3) and isorhamnetin 3-O-(2″-O-β-xylopyranosyl-[6″-O-β-xylopyranosyl]-β-glucopyranoside) (6) are newly reported as natural compounds. Remarkably, two Primula type flavones were also detected in tissues, namely 3′-hydroxyflavone 3′-O-β-glucoside (1) and 3′,4′-dihydroxyflavone 4′-O-β-glucoside (2), of which (1) is reported here for the first time as natural product. All structures were unambiguously identified by NMR and MS data. Earlier reports on the occurrence of 7,2′-dihydroxyflavone 7-O-glucoside (macrophylloside) in this species could not be confirmed. This structure was now shown to correspond to 3′,4′-dihydroxyflavone 4′-O-glucoside (2) by comparison of NMR data. Observed exudate variations might be specific for geographically separated populations. The structural diversification between tissue and exudate flavonoids is assumed to be indicative for different ecological roles in planta.     

Phenolic compounds from parasitic Sapria himalayana f. albovinosa and Sapria myanmarensis (Rafflesiaceae) in Myanmar

Three anthocyanins, four flavonols, three aromatic acids and five gallotannins were isolated from Sapria himalayana f. albovinosa in Myanmar. They were identified as cyanidin 3-O-glucoside (1), cyanidin 3-O-xyloside (2) and peonidin 3-O-glucoside (3) (anthocyanins), quercetin 3-O-glucoside (4), quercetin 7-O-glucoside (5), quercetin 3-O-glucuronide (6) and isorhamnetin 3-O-glucoside (7) (flavonols), ellagic acid (8), gallic acid (9) and ethyl gallate (10) (aromatic acids), and 1,2,4,6-tetragalloylglucose (11), 1,4,6-trigalloylglucose (12), 1,2,6-trigalloylglucose (13), 1,2,4-trigalloylglucose (14) and 1,6-digalloylglucose (15) (gallotannins) by UV, LC-MS, acid hydrolysis, NMR and/or HPLC comparisons with authentic samples. The chemical composition of S. myanmarensis was qualitatively the same with that of S. himalayana f. albovinosa. Phenolic compounds of the Rafflesiaceae species including Sapria, Rafflesia and Rhizanthes were isolated and identified in this survey for the first time.     

Anthocyanins and flavonols from the flowers of Amherstia nobilis endemic to Myanmar

Three anthocyanins (1–3) and eight flavonols (4–11) were isolated from the flowers of Amherstia nobilis endemic to Myanmar. Anthocyanins were identified as cyanidin 3-O-glucoside (1), 3-O-xyloside (2), and peonidin 3-O-glucoside (3). On the other hand, flavonols were identified as isorhamnetin 3-O-glucoside (4), 7-O-glucoside (5), 3,7-di-O-glucoside (6) and 3-O-rutinoside (7), quercetin 3-O-rutinoside (8) and 3-O-glucoside (9), and kaempferol 3-O-rutinoside (10) and 3-O-glucoside (11). Although an anthocyanin, pelargonidin 3-O-pentoside, has been reported from the flowers of A. nobilis, it was not found in this survey. The presence of flavonols in A. nobilis was reported in this survey for the first time. Flavonoid composition of Amherstia was chemotaxonomically compared with those of phylogenetically related genera Cynometra and Brownea.     

Secondary metabolites of Rubus caesius (Rosaceae)

The European dewberry (Rubus caesius) is a perennial shrub that is widely distributed in Europe but can also be found in North America. In folk medicine, the European dewberry is used to treat hyperglycaemia, diarrhoea and inflammation. LC-MS analysis of the European dewberry confirmed the presence of 35 compounds, mostly flavonoids, phenolic acids and derivatives of ellagic acid. Phytochemical analysis of R. caesius leaves led to the isolation of nine phenolics, namely: quercetin 3-O-β-D-rutinoside (1), kaempferol 3-O-β-D-glucuronide (2), quercetin 3-O-β-D-glucuronide (3), methyl brevifolincarboxylate (4), kaempferol 3-O-β-D-(6″-O-(E)-p-coumaroyl)-glucoside (5), kaempferol (6), quercetin (7), pedunculagin (8), and ellagic acid (9). Compounds 1–8 were isolated from this species for the first time. The chemophenetic significance was discussed.     

Flavonoids from the flowers of Adenium obesum (Forssk.) Roem. & Schult., Mandevilla sanderi (Hemsl.) Woodson,and Nerium oleander L. (Apocynaceae)

Twenty-two ornamental flowers from different Adenium obesum, Mandevilla sanderi, and Nerium oleander cultivars/seedlings were analyzed for the presence of anthocyanins, flavonols, and chlorogenic acid using nuclear magnetic resonance (NMR) and mass spectrometry (MS). Cyanidin 3-O-[6-O-(rhamnosyl)-galactoside] and cyanidin 3-O-(galactoside) were identified as the major and minor anthocyanins, respectively, in three A. obesum seedlings that had red and red-purple flowers.Cyanidin 3-O-[2-O-(xylosyl)-galactoside] was identified as the major anthocyanin, whereas cyanidin 3-O-[6-O-(rhamnosyl)-galactoside] and cyanidin 3-O-(galactoside) were identified as the minor anthocyanins in 8 M. sanderi cultivars that had red and red-purple flowers. Cyanidin 3-O-[6-O-(rhamnosyl)-galactoside] and cyanidin 3-O-(galactoside) were identified as the major anthocyanins, whereas cyanidin 3-O-[2-O-(xylosyl)-galactoside] was identified as the minor anthocyanin in 8 N. oleander cultivars with red and red-purple flowers. Low levels of anthocyanins were detected in the N. oleander and M. sanderi cultivars that had white flowers, and there were no anthocyanins detected in the N. oleander cultivars with yellow flowers. Chlorogenic acid and four flavonols, quercetin 3-O-[6-O-(rhamnosyl)-galactoside], quercetin 3-O-[6-O-(rhamnosyl)-glucoside], kaempferol 3-O-(galactoside), and kaempferol 3-O-[6-O-(rhamnosyl)-galactoside], were identified in the flowers from all 22 cultivars/seedlings investigated.     

Anthocyanin content of Tulipa species and cultivars and its impact on tepal colours

Flowers of tulips (17 species and 25 cultivars) were subjected to qualitative and relative quantitative examination for anthocyanins. Altogether five anthocyanins were identified as the 3-O-(6″-O-α-rhamnopyranosyl-β-glucopyranoside) of delphinidin (1), cyanidin (2) and pelargonidin (3), and the 3-O-[6″-O-(2‴-O-acetyl-α-rhamnopyranosyl)-β-glucopyranoside] of cyanidin (4) and pelargonidin (5). The pigments 1–5 represented 7%, 43%, 12%, 2% and 31%, respectively, of the total anthocyanin amount in the tepals of the Tulipa species, and 20%, 37%, 30%, 6% and 4%, respectively, in the cultivar tepals. Nearly 50% of the samples contained acetylated anthocyanins. The colours of the freeze-dried tepals described by the CIELab coordinates, hue angle (h_ab), saturation (C*), and lightness (L*) together with the anthocyanin content were subjected to multivariate analysis. All tepals classified with hue angles described as “blue nuances” were from cultivars. They contained 1 as the major anthocyanin, and no or just traces of pelargonidin derivatives. The species and cultivars having “magenta nuances” showed similar anthocyanin content with increased relative proportions of 2 at the expense of 1. Orange coloured tepals were to a large extent correlated with high relative proportions of the pelargonidin derivatives, 3 and 5. Acetylation of anthocyanins furnished a weak colour effect opposite to the bluing effect previously reported for anthocyanins with aromatic acyl groups. All six species belonging to the section Eichleres (subgenus Tulipa) were after principal component analysis grouped closely together. They were characterized by high concentrations of the pelargonidin derivatives 3 and 5, and orange petal nuances. However, within section Tulipa (subgenus Tulipa), considerable anthocyanin variation was observed. Species in the subgenus Eriostemones were generally characterized by the two anthocyanins 1 and 2, and no pelargonidin derivatives.     

Flavonoids from the leaves and flowers of the Himalayan Cathcartia villosa (Papaveraceae)

Five flavonols, four flavones and one C-glycosylflavone were isolated from the leaves of Cathcartia villosa which is growing in the Himalayan Mountains. They were characterized as quercetin 3-O-vicianoside (1), quercetin 7,4′-di-O-glucoside (3), quercetin 3-O-rutinoside (4), quercetin 3-O-glucoside (5), quercetin 3-O-arabinosylarabinosylglucoside (6) (flavonols), luteolin (7), luteolin 7-O-glucoside (8), apigenin (9), chrysoeriol (10) (flavones), and vicenin-2 (11) (C-glycosylflavone) by UV, LC-MS, acid hydrolysis, NMR and/or HPLC and TLC comparisons with authentic samples. On the other hand, two flavonols 1 and kaempferol 3-O-vicianoside (2) were isolated and identified from the flowers of the species. Flavonoids were reported from the genus Cathcartia in this survey for the first time. Their chemical characters were chemotaxonomically compared with those of related Papaveraceous genera, Meconopsis and Papaver.     

Anthocyanins in Caprifoliaceae

The qualitative and relative quantitative anthocyanin content of 19 species belonging to the genera Sambucus, Lonicera and Viburnum in the family Caprifoliaceae has been determined. Altogether 12 anthocyanins were identified; the 3-O-glucoside (2), 3-O-galactoside (5), 3-O-(6″-O-arabinosylglucoside) (7), 3-O-(6″-O-rhamnosylglucoside) (9), 3-O-(2″-O-xylosyl-6″-O-rhamnosylglucoside) (10), 3-O-(2″-O-xylosylgalactoside) (11), 3-O-(2″-O-xylosylglucoside) (12), 3-O-(2″-O-xylosylglucoside)-5-O-glucoside (14), 3-O-(2″-O-xylosyl-6″-O-Z-p-coumaroylglucoside)-5-O-glucoside (15) and 3-O-(2″-O-xylosyl-6″-O-E-p-coumaroylglucoside)-5-O-glucoside (16) of cyanidin, in addition to the 3-O-glucosides of pelargonidin and delphinidin (1 and 3). Pigment 7 is the first complete identification of the disaccharide vicianose, 6″-O-α-arabinopyranosyl-β-glucopyranose, linked to an anthocyanidin.     

Leaf flavonoid glycosides as chemosystematic characters in Ocimum

Thirty-one accessions of nine species belonging to three subgenera of Ocimum (basil, family Lamiaceae) were surveyed for flavonoid glycosides. Substantial infraspecific differences in flavonoid profiles of the leaves were found only in O. americanum, where var. pilosum accumulated the flavone C-glycoside, vicenin-2, which only occurred in trace amounts in var. americanum and was not detected in cv. Sacred. The major flavonoids in var. americanum and cv. Sacred, and also in all other species investigated for subgenus Ocimum, were flavonol 3-O-glucosides and 3-O-rutinosides. Many species in subgenus Ocimum also produced the more unusual compound, quercetin 3-O-(6″-O-malonyl)glucoside, and small amounts of flavone O-glycosides. The level of flavonol glycosides produced was reduced significantly in glasshouse-grown plants, but levels of flavone glycosides were unaffected. A single species investigated from subgenus Nautochilus, O. lamiifolium, had a different flavonoid glycoside profile, although the major compound was also a flavonol O-glycoside. This was identified as quercetin 3-O-xylosyl(1‴→2″)galactoside, using NMR spectroscopy. The species investigated from subgenus Gymnocimum, O. tenuiflorum (=O. sanctum), was characterised by the accumulation of flavone O-glycosides. These were isolated, and identified as the 7-O-glucuronides of luteolin and apigenin. Luteolin 5-O-glucoside was found in all nine species of Ocimum studied, and is considered to be a key character for the genus.     

Further studies of flavonols of Clibadium (compositae)

The flavonoids of an additional eight species of Clibadium have been determined. The compounds are derivatives of kaempferol, quercetin and quercetagetin. O-Methylated quercetagetin derivatives were found in several taxa with the possibility that 6-methoxykaempferol may also exist in one collection. Kaempferol and quercetin exist as 3-O-glucosides, galactosides, rhamnosides, rutinosides and diglucosides although not all glycosides occur in each taxon. Quercetagetin derivatives occur as 7-O-glucosides. Observations on these newly investigated species confirm previous work in the genus that three types of flavonoid profiles exist: (1) kaempferol and quercetin 3-glycosides; (2) kaempferol and quercetin 3-glycosides plus quercetagetin 7-glucoside; and (3) kaempferol and quercetin 3-glycosides plus quercetagetin 7-glucoside and O-methylated derivatives of quercetagetin.     

Acylated anthocyanins and flavonols from purple flowers of Dendrobium cv. ‘Pompadour’

Two rare anthocyanins, cyanidin 3-(6-malonylglucoside)-7,3′-di(6-sinapylglucoside) and the demalonyl derivative, were characterised as the purple floral pigments of Dendrobium cv. ‘Pompadour’. Nine known flavonol glycosides were also identified, including the 3-rutinoside-7-glucosides of kaempferol and quercetin. One new glycoside was detected: the ferulyl ester of quercetin 7-rutinoside-7-glucoside. These flavonoid patterns are typical for plants in the family Orchidaceae.     

The identification of flavonoids and the expression of genes of anthocyanin biosynthesis in the chrysanthemum flowers

In order to provide additional information on the coloration of chrysanthemum flowers, the flavonoid composition and the expression of six structural genes involved in anthocyanin pathway in the ray florets of a pink flowering (cv. H5) and two white flowering (cvs. Keikai and Jinba) Chrysanthemum grandiflorum cultivars were examined. HPLCDAD/ESI-MSⁿ analysis showed that cyanidin 3-O-(6″-O-malonylglucoside) and cyanidin 3-O-(3″,6″-O-dimalonylglucoside) were the two major flavonoids presented in H5, while white flowering cultivars contained flavones instead of anthocyanins. Nine flavone derivatives were detected in the three cultivars, the amount of each flavone varied upon cultivars, and seven of these were identified as luteolin 7-O-arabinosylglucuronide, apigenin 7-O-glucoside, luteolin 7-O-malonylglucoside, apigenin 7-O-malonylglucoside, chrysoeriol 7-O-malonylglucoside, acacetin 7-O-rutinoside and acacetin 7-O-malonylglucoside. The two white flowering cultivars showed similar total flavonoid content, which was about two fold higher than that in H5. A high expression of the genes encoding dihydroflavonol 4-reductase and 3-O-glucosyltransferase was detected only in H5 but not in Keikai or Jinba. Chalcone synthase, chalcone isomerase, flavanone 3-hydroxylase, and flavonoid 3′-hydroxylase were expressed in all flowers, suggesting that the lack of anthocyanin in white flowering cultivars cannot be due to any blockage of their expression.     

Anthocyanins from red flowers of Camellia cultivar 'Dalicha'

Five anthocyanins, cyanidin 3-O-(2-O-β-xylopyranosyl-6-O-(Z)-p-coumaroyl)-β-galactopyranoside (2), cyanidin 3-O-(2-O-β-xylopyranosyl-6-O-(E)-p-coumaroyl)-β-galactopyranoside (3), cyanidin 3-O-(2-O-β-xylopyranosyl-6-O-(E)-caffeoyl)-β-galactopyranoside (4), cyanidin 3-O-(2-O-β-xylopyranosyl-6-O-acetyl)-β-galactopyranoside (5), and cyanidin 3-O-(2-O-β-xylopyranosyl-6-O-acetyl)-β-glucopyranoside (6), together with the known cyanidin 3-O-(2-O-β-xylopyranosyl)-β-galactopyranoside (1), were isolated from red flowers of Camellia cultivar ‘Dalicha’ (Camellia reticulata) by chromatography using open columns. Their structures were subsequently determined on the basis of spectroscopic analyses, i.e., ¹H NMR, ¹³C NMR, HMQC, HMBC, HR ESI-MS and UV-vis.     

Flavonoids and phenolic acids from the aerial parts of Alyssum alyssoides L. (Brassicaceae)

Two phenolic acids (1 and 2) and seven flavonoids (3–9) were isolated from the aerial parts of Alyssum alyssoides (Brassicaceae). All these compounds (1–9) were isolated from this particular species for the first time. Their structures were identified, on the basis of MS and NMR spectra as: p-hydroxy-benzoic acid (1), 3-methoxy-4-hydroxybenzoic acid (vanillic acid) (2), kaempferol 3-O-β-D-glucopyranoside (astragalin) (3), kaempferol 3-O-(6″-α-L-rhamnopyranosyl)-β-D-glucopyranoside (nicotiflorin) (4), quercetin 3-O-β-D-glucopyranoside (isoquercetin) (5), quercetin 3-O-β-D-galactopyranoside (hyperoside) (6), isorhamnetin 3-O-β-D-glucopyranoside (7), isorhamnetin 3-O-β-D-galactopyranoside (8) and isorhamnetin 3-O-(6″-α-L-rhamnopyranosyl)-β-D-glucopyranoside (narcissin) (9). The chemotaxonomic significance of these compounds was summarized.     

Flavonoids of Mayaca fluviatilis (Mayacaceae)

Mayaca is an aquatic monocot of the monogeneric family Mayacaceae. The flavonol glycosides quercetin 3-O-glucoside, quercetin 3-O-rutinoside, and kaempferol 3-O-glucoside, and the flavone luteolin 5-O-glucoside were found in methanolic leaf extracts. The presence of flavonol and flavone O-glycosides sets the Mayacaceae apart from the Commelinaceae, which accumulates predominantly flavone C-glycosides.     

Anthocyanins of grasses

The anthocyanin content of 23 grass species (Poaceae) belonging to five subfamilies has been determined. Altogether 11 anthocyanins were identified; the 3-(6″-malonylglucosides) and 3-glucosides of cyanidin, peonidin and delphinidin, the 3-(3″,6″-dimalonylglucoside), 3-(6″-rhamnosylglucoside) and 3-(6″-glucosylglucoside) of cyanidin, in addition to peonidin 3-(dimalonylglucoside) and delphinidin 3-(6″-rhamnosylglucoside). Anthocyanins acylated with one and/or two malonic acid moieties dominated the anthocyanin profiles of all the species in the subfamilies Pooideae and Panicoideae. On the other hand, the 3-glucoside and 3-rutinoside of cyanidin were the major anthocyanins in Sinarundinaria murielae (subfamily Bambusoideae) and Molinia caerulea (subfamily Arundinoideae), while the 3-glucosides of cyanidin and peonidin were the principal anthocyanins in rice, Oryza sativum (subfamily Oryzoideae). Pelargonidin derivatives and free anthocyanidins have previously been reported to occur in several Poaceae species, however, not identified in any of the species included in this survey.     

Dihydrochalcone glucosides and antioxidant activity from the roots of Anneslea fragrans var. lanceolata

Bioassay-guided fractionation of the roots of Anneslea fragrans var. lanceolata led to the isolation of four dihydrochalcone glucosides, davidigenin-2′-O-(6″-O-4″′-hydroxybenzoyl)-β-glucoside (1), davidigenin-2′-O-(2″-O-4″′-hydroxybenzoyl)-β-glucoside (2), davidigenin-2′-O-(3″-O-4″′-hydroxybenzoyl)-β-glucoside (3), and davidigenin-2′-O-(6″-O-syringoyl)-β-glucoside (4), and 13 known compounds. The structures were identified by means of spectroscopic analysis. Davidigenin-2′-O-(6″-O-syringoyl)-β-glucoside (4), 1-O-3,4-dimethoxy-5-hydroxyphenyl-6-O-(3,5-di-O-methylgalloyl)-β-glucopyranoside (5), lyoniresinol (10), and syringic acid (13) showed ABTS [2,2′-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid)] cation radical scavenging activity, with SC₅₀ values of 52.6 ± 5.5, 26.0 ± 0.7, 6.0 ± 0.2, and 27.5 ± 0.6 μg/mL in 20 min, respectively. Lyoniresinol (10), isofraxidin (12), and syringic acid (13) also showed DPPH [1,1-diphenyl-2-picrylhydrazyl] radical scavenging activity, with SC₅₀ values of 8.4 ± 1.8, 51.6 ± 2.2, and 4.3 ± 0.7 μg/mL in 30 min, respectively.     

Acylated flavonoid tetraglycoside from Planchonia careya leaves

Phytochemical investigations of the aqueous extract of Planchonia careya leaves revealed two known flavonol glycosides, kaempferol 3-O-gentiobioside (1) and quercetin 3-O-glucoside (isoquercitrin) (2), and a novel acylated kaempferol tetraglycoside, kaempferol 3-O-[α-rhamnopyranosyl(1 → 3)-(2-O-p-coumaroyl)]-β-glucopyranoside, 7-O-[α-rhamnopyranosyl-(1 → 3)-(4-O-p-coumaroyl)]-α-rhamnopyranoside (3). Structural elucidation was achieved using UV, NMR, and mass spectrometry.     

(2S)-Prenylflavanones and taraxerane triterpenoids from Mallotus mollissimus

Three prenylflavanones, (2S)-5,7-dihydroxy-4′-methoxy-8-(3″,3″-dimethylallyl)flavanone (3), (2S)-5,4′-dihydroxy-7-methoxy-6-(3″,3″-dimethylallyl)flavanone (6), 8-prenylnaringenin (11), and a new epimeric pair (2″S/2″R)-(2S)-5,7-dihydroxy-4′-methoxy-6-(2″-hydroxy-3″-methylbut-3″-enyl)flavanones (4a/4b) were isolated together with taraxerone, taraxerol, epitaraxerol, β-sitosterol, oleanolic acid, 1-O-docosanoyl glycerol, apigenin, and apigenin 7-O-β-D-glucopyranoside from the MeOH extract of the leaves of Mallotus mollissimus. The structures of the isolated compounds were determined on the basis of 1D/2D NMR and HR-MS spectroscopic data; the 2S configuration of the prenylflavanones 3, 4, and 6 was deduced from CD spectroscopic data. The presence of three taraxerane triterpenoids reinforces the inclusion of M. mollissimus (syn. Croton mollissimus) in Mallotus genus. Among species of Mallotus the occurrence of the (2S)-prenylflavanones 3, 4, and 6 is confined to M. mollissimus.     

Acylated anthocyanidin 3-sophoroside-5-glucosides from Ajuga reptans flowers and the corresponding cell cultures

Four anthocyanins from Ajuga reptans flowers and its cell cultures were isolated, and a fifth was also characterized by HPLC-mass spectrometry. By means of chemical and spectroscopic analyses, their structures were identified as delphinidin 3-(p-coumaroyl-feruloyl)sophoroside-5-malonylglucoside, delphinidin 3-(diferuloyl)sophoroside-5-malonylglucoside, and cyanidin 3-(di-p-coumaroyl)sophoroside-5-glucoside, respectively. The other two were tentatively identified as delphinidin 3-(diferuloyl)sophoroside-5-glucoside and cyanidin 3-(feruloyl-p-coumaroyl)sophoroside-5-malonylglucoside. In neutral aqueous solution, the crude extract from A. reptans flower cell cultures and the major anthocyanin cyanidin 3-(di-p-coumaroyl)sophoroside-5-malonylglucoside were more stable than cyanidin 3-glucoside, and also prevented more efficiently peroxidation than did the latter. A. reptans flower cell culture anthocyanins may have a potential as natural colorants for food utilities or other purposes.