Acylated hydroxycinnamic acid glucosides from flowers of Telekia Speciosa

One new derivative of ferulic acid (1), two new caffeic acid derivatives (2 and 3) and three known derivatives of caffeic acid: 6-O-(E)-caffeoyl-glucopyranose (4), (E)-caffeic acid 4-O-β-glucopyranoside (5) and 5-caffeoylquinic acid (chlorogenic acid, 6) were isolated from a butanolic fraction of extract from Telekia speciosa flowers. Moreover, the flavonol glucoside–patulitrin (7) was identified in the analyzed extract. Structures of (E)-ferulic acid 4-O-β-(6-O-2-hydroxyisovaleryl)-glucopyranoside (1), (E)-caffeic acid 4-O-β-(6-O-2-hydroxyisovaleryl)-glucopyranoside (2) and (E)-caffeic acid 4-O-β-(6-O-3-hydroxy-2-methylpropanoyl)-glucopyranoside (3) were elucidated by 1D and 2D NMR, HRESIMS and other spectral analyses.     

Hydroxycinnamic acid spermidine amides from pollen of corylus avellana L.

Two hydroxyannamic acid amides from the pollen of Corylus avellana L. have been identified as (E)-caffeoyl-(E)-feruloylspermidine and di-(E)-feruloylspermidine on the basis of ¹H NMR, ¹³C NMR and mass spectral data.     

Accumulation and enzymatic synthesis of 2-O-acetyl-3-O-(p-coumaroyl)-meso-tartaric acid in spinach cotyledons

2-O-Acetyl-3-O-[(E)-p-coumaroyl]-meso-tartaric acid was isolated from cotyledons of Spinacia oleracea and its structure elucidated and characterized with the aid of TLC, HPLC, FAB MS and ¹H NMR. Accumulation and enzymatic synthesis of the diester are described, proceeding first via 1-O-(p-coumaroyl)-β-glucose in the formation of p-coumaroyltartaric acid and second via acetyl-CoA in the formation of 2-O-acetyl-3-O-[(E)-p-coumaroyl]-meso-tartaric acid. Some properties of the CoA-thioester-dependent acyltransferase activity were studied.     

12-disinapolylglucose accumulated in cotyledons of dark-grown raphanus sativus seedlings

A new sinapic acid ester has been isolated and characterized as 1(E),2(E)-di-O-sinapoyl-β-d-glucopyranoside from cotyledons of dark-grown red radish (Raphanus sativus) seedlings. Its structure was elucidated by negative ion fast atom bombardment mass spectrometry, ¹H and ¹³C NMR spectra and enzymatic determination of the glucose moiety. A possible biosynthetic mechanism for the formation of this new ester is discussed in which the energy-rich acyl glucoside 1-O-sinapoyl-β-d-glucose acts as the acyl donor in a sinapoyl transfer to the hydroxyl group at C-2 of the glucose moiety of another molecule of 1-O-sinapoyl-β-d-glucose (‘disproportionation’).     

Linoleate 9R-dioxygenase and allene oxide synthase activities of Aspergillus terreus

Oxygenation of linoleic acid by Aspergillus terreus was studied with LC-MS/MS. 9(R)-Hydroperoxy-10(E),12(Z)-octadecadienoic acid (9R-HpODE) was identified along with 10(R)-hydroxy-8(E),12(Z)-octadecadienoic acid and variable amounts of 8(R)-hydroxy-9(Z),12(Z)-octadecadienoic acid. 9R-HpODE was formed from [11S-²H]18:2n − 6 with loss of the deuterium label, suggesting antarafacial hydrogen abstraction and oxygenation. Two polar metabolites were identified as 9-hydroxy-10-oxo-12(Z)-octadecenoic acid (α-ketol) and 13-hydroxy-10-oxo-11(E)-octadecenoic acid (γ-ketol), likely formed by spontaneous hydrolysis of an unstable allene oxide, 9(R),10-epoxy-10,12(Z)-octadecadienoic acid. α-Linolenic acid and 20:2n − 6 were oxidized to hydroperoxy fatty acids at C-9 and C-11, respectively, but α- and γ-ketols of these fatty acids could not be detected. The genome of A. terreus lacks lipoxygenases, but contains genes homologous to 5,8-linoleate diol synthases and linoleate 10R-dioxygenases of aspergilli. Our results demonstrate that linoleate 9R-dioxygenase linked to allene oxide synthase activities can be expressed in fungi.     

Metabolomics-oriented isolation and structure elucidation of 37 compounds including two anthocyanins from Arabidopsis thaliana

In order to conduct metabolomic studies in a model plant for genome research, such as Arabidopsis thaliana (Arabidopsis), it is a prerequisite to obtain structural information for the isolated metabolites from the plant of interest. In this study, we isolated metabolites of Arabidopsis in a relatively non-targeted way, aiming at the construction of metabolite standards and chemotaxonomic comparison. Anthocyanins (5 and 7) called A8 and A10 were isolated and their structures were elucidated as cyanidin 3-O-[2-O-(β-d-xylopyranosyl)-6-O-(4-O-(β-d-glucopyranosyl)-E-p-coumaroyl)-β-d-glucopyranoside]-5-O-[6-O-(malonyl)-β-d-glucopyranoside] and cyanidin 3-O-[2-O-(2-O-(E-sinapoyl)-β-d-xylopyranosyl)-6-O-(4-O-(β-d-glucopyranosyl)-E-p-coumaroyl)-β-d-glucopyranoside]-5-O-[β-d-glucopyranoside] from analyses of 1D NMR, 2D NMR (¹H NMR, NOE, ¹³C NMR, HMBC and HMQC), HRFABMS, FT-ESI-MS and GC-TOF-MS data. In addition, 35 known compounds, including six anthocyanins, eight flavonols, one nucleoside, one indole glucosinolate, four phenylpropanoids and a derivative, together with three indoles, one carotenoid, one apocarotenoid, three galactolipids, two chlorophyll derivatives, one steroid, one hydrocarbon, and two dicarboxylic acids, were also isolated and identified from their spectroscopic data.     

Polyphenols isolated from antiradical extracts of Mallotus metcalfianus

Six flavonoids including two new flavones, luteolin 7-O-(4″-O-(E)-coumaroyl)-β-glucopyranoside), chrysoeriol-7-O-(4″-O-(E)-coumaroyl)-β-glucopyranoside) and a mixture of two pairs of diastereoisomeric flavonolignans, (±)-hydnocarpin 7-O-(4″-O-(E)-coumaroyl)-β-glucopyranoside)/(±)-hydnocarpin-D 7-O-(4″-O-(E)-coumaroyl)-β-glucopyranoside) with a 2:1 ratio were isolated from the whole plant of Mallotus metcalfianus Croizat, in addition to 10 known compounds. Their structures were evaluated on the basis of different spectroscopic methods, including extensive 1D and 2D NMR spectroscopy. Some extracts have moderate antimicrobial properties and interesting antiradical (DPPH) activity, as well as some compounds isolated from this species. Tannins were also identified in some active extracts.     

Phenolic glycosides from Cucumis melo var. inodorus seeds

A new phenolic glycoside (E)-4-hydroxycinnamyl alcohol 4-O-(2′-O-β-d-apiofuranosyl)(1″ → 2′)-β-d-glucopyranoside (1) was isolated and identified from Cucumis melo seeds together with benzyl O-β-d-glucopyranoside (2), 3,29-O-dibenzoylmultiflor-8-en-3α,7β,29-triol (3) and 3-O-p-amino-benzoyl-29-O-benzoylmultiflor-8-en-3α,7β,29-triol (4). Their structures were elucidated by extensive NMR experiments including ¹H–¹H (COSY, TOCSY, ROESY) and ¹H–¹³C (HSQC and HMBC) spectroscopy and chemical evidence. The multiflorane triterpene esters were identified as new melon constituents.     

3-O-feruloyl-4-O-caffeoylquinic acid from coffee beans

A new phenolic ester was isolated from unroasted robusta coffee beans (Coffea canephora) by HPLC. The isolated compound was identified as an ester of caffeic acid and ferulic acid with quinic acid (3-O-feruloyl-4-O-caffeoylquinic acid) using ¹H NMR and mass spectroscopy.     

Occurrence of bergenin phenylpropanoates in Vatica bantamensis

We isolated five bergenin phenylpropanoates, i.e., 11-O-(E)-sinapate (1), 11-O-(E)-ferulate (2), 11-O-(Z)-ferulate (3), 11-O-(E)-coumalate (4), and 11-O-(Z)-coumalate (5), and three bergenin hydroxybenzoates, i.e., 11-O-syringate (6), 11-O-vanillate (7), and 11-O-p-hydroxybenzoate (8), along with bergenin (9), from the leaves of Vatica bantamensis. Moreover, we identified the geometrical isomerization between 2 and 3. These structures were characterized by nuclear magnetic resonance (NMR). This is the first report that shows the occurrence of bergenin phenolic acid esters in dipterocarpaceaeous plants.     

Caffeic acid derivatives and further compounds from Espeletia barclayana Cuatrec. (Asteraceae,Espeletiinae)

This work describes the isolation of seven (1–7) secondary metabolites from Espeletia barclayana (Asteraceae, Espeletiinae) and their identification by spectroscopic (NMR) and spectrometric (MS) techniques. Ten (8–17) additional compounds were identified based on their retention times, high-resolution mass spectrometry data, and comparison with reference substances or data from literature. The systematic significance of some of the identified substances – the sesquiterpene lactone longipilin acetate, four caffeoylquinic acids and two tri-caffeoylaltraric acids – is discussed with the aim of providing insights into the complex relationships among Espeletiinae taxa and its closest relatives. Five of the isolated metabolites [5-O-(E)-caffeoylquinic acid (1), 1,3-di-O-(E)-caffeoylquinic acid (2), 1,5-di-O-(E)-caffeoylquinic acid (3), 3,4-di-O-(E)-caffeoylquinic acid (4) and 3-O-methylquercetin 7-O-β-glucopyranoside (7)] constitute new reports for the genus Espeletia and for the subtribe Espeletiinae. Chemical data suggest that Espeletiinae might have a closer relationship with Smallanthus than with Ichthyothere, i.e., the two genera suggested to be the sister groups of Espeletiinae based on molecular markers.     

Proline-glutamate chimera’s side chain conformation directs the type of β-hairpin structure

Our aim was to study the impact of two proline chimeras, containing a glutamic acid side chain in cis- or trans-configuration, on secondary structure formation. We further investigated to what extent the configuration of the side chain contributes to the overall peptide conformation. We used a 10 residue peptide (IYSNPDGTWT) that forms a β-hairpin in water. The turn-forming proline was substituted with either a cis- or trans-proline-glutamic acid chimera, resulting in the peptides IYSNP ^{cis -E} DGTWT (P1_P ^cis-E ) and IYSNP ^{trans -E} DGTWT (P1_P ^trans-E ). We studied the conformation of the modified peptides by circular dichroism (CD) and NMR-spectroscopy, and SEC/static light scattering (SLS) analysis. NMR analysis reveals that the modified peptides maintain the β-hairpin conformation in aqueous solution. At 5 °C and pH 4.3, the peptide (P1_P ^cis-E ) was found to adopt two coexisting β-hairpin conformations (2:2 β-hairpin, and 3:5 β-hairpin). In contrast to that, the peptide (P1_P ^trans-E ) adopts a 2:2 β-hairpin that exists in equilibrium with a 4:4 β-hairpin conformation. The adoption of ordered β-hairpin structures for both modified peptides could be confirmed by CD spectroscopy, while SEC/SLS analysis showed a monomeric oligomerization state for all three investigated peptides. With the combination of several NMR methods, we were able to elucidate that even small alterations in the side chain conformation of the proline-glutamate chimera (cis or trans) can significantly influence the conformation of the adopted β-hairpin.     

Study of the 3-hydroxy eicosanoyl-coenzyme A dehydratase and (E)-2,3 enoyl-coenzyme A reductase involved in acyl-coenzyme A elongation in etiolated leek seedlings

(R,S)-[1-¹⁴C]3-Hydroxy eicosanoyl-coenzyme A (CoA) has been chemically synthesized to study the 3-hydroxy acyl-CoA dehydratase involved in the acyl-CoA elongase of etiolated leek (Allium porrum L.) seedling microsomes. 3-Hydroxy eicosanoyl-CoA (3-OH C20:0-CoA) dehydration led to the formation of (E)-2,3 eicosanoyl-CoA, which has been characterized. Our kinetic studies have determined the optimal conditions of the dehydration and also resolved the stereospecificity requirement of the dehydratase for (R)-3-OH C20:0-CoA. Isotopic dilution experiments showed that 3-hydroxy acyl-CoA dehydratase had a marked preference for (R)-3-OH C20:0-CoA. Moreover, the very-long-chain synthesis using (R)-3-OH C20:0-CoA isomer and [2-¹⁴C]malonyl-CoA was higher than that using the (S) isomer, whatever the malonyl-CoA and the 3-OH C20:0-CoA concentrations. We have also used [1-¹⁴C]3-OH C20:0-CoA to investigate the reductant requirement of the enoyl-CoA reductase of the acyl-CoA elongase complex. In the presence of NADPH, [1-¹⁴C]3-OH C20:0-CoA conversion was stimulated. Aside from the product of dehydration, i.e. (E)-2,3 eicosanoyl-CoA, we detected eicosanoyl-CoA resulting from the reduction of (E)-2,3 eicosanoyl-CoA. When we replaced NADPH with NADH, the eicosanoyl-CoA was 8- to 10-fold less abundant. Finally, in the presence of malonyl-CoA and NADPH or NADH, [1-¹⁴C]3-OH C20:0-CoA led to the synthesis of very-long-chain fatty acids. This synthesis was measured using [1-¹⁴C]3-OH C20:0-CoA and malonyl-CoA or (E)-2,3 eicosanoyl-CoA and [2-¹⁴C]malonyl-CoA. In both conditions and in the presence of NADPH, the acyl-CoA elongation activity was about 60 nmol mg⁻¹ h⁻¹, which is the highest ever reported for a plant system.     

Isolation and characterization of the chemical constituents from Plumeria rubra

Rubranonoside (=7-O-α-l-rhamnopyranosyl-4′-O-β-d-glucopyranosylnaringenin; (1), a new flavanone glycoside, rubranin (=(2S,3S,4R)-2-{[(2R,16E)-2-hydroxyhexaeico-16-en]amino}octadecane-1,3,4-triol-1-O-β-d-glucopyranoside; (2), a new sphingolipid, rubradoid (plumieridine-1-O-β-d-galactopyranoside; (3), a new iridoid galactoside, rubrajaleelol (4) and rubrajaleelic acid (5), two new nor-terpenoids together with known iridoids: 1-α-plumieride (6), plumieride p-Z-coumarate (7) and plumieride-p-E-coumarate (8) have been isolated from the EtOAc-soluble fraction of the MeOH extract of Plumeria rubra. Their structures were assigned from ¹H, ¹³C NMR spectra and 2D NMR analyses (COSY, NOESY, HMQC and HMBC experiments) in combination with HRMS experiments and comparison with literature data of related compounds. All the isolates (1–8) were tested for their antioxidant, antiurease, cytotoxic and phytotoxic activities and were found almost inactive.     

Coumaroyl-, caffeoyl- and feruloyltartronates and their accumulation in mung bean

Three new plant constituents were isolated from the primary leaves of Vigna radiata (= Phaseolus aureus) and their structures elucidated and characterized with the aid of negative-ion fast atom bombardment mass spectrometry (FAB MS), ¹H NMR and UV spectroscopy, thin-layer, gas-liquid and high performance liquid chromatography. The new conjugates are (E)-p-coumaroyl-, (E)-caffeoyl- and (E)-feruloyltartronic acids. Their structures were unequivocally confirmed by comparison with synthetic material. The metabolism of the new hydroxycinnamic acid conjugates in young plants of Vigna radiata is described.     

Exploration of antimicrobial and antioxidant potential of newly synthesized 2,3-disubstituted quinazoline-4(3H)-ones

A series of 2-(chloromethyl)-3-(4-methyl-6-oxo-5-[(E)-phenyldiazenyl]-2-thioxo-5,6-dihydropyrimidine-1(2H)-yl)quinazoline-4(3H)-ones 9a-j was synthesized by treating 2-(chloroacetyl)amino benzoic acid with 3-amino-6-methyl-5-[(E)-phenyldiazenyl]-2-thioxo-2,5-dihydropyrimidine-4(3H)-one 8a-j and was screened for in vitro antibacterial activities against a representative panel of Gram-positive and Gram-negative bacteria. The compounds were synthesized in excellent yields and the structures were corroborated on the basis of IR, ¹H NMR, Mass and elemental analysis data. All the synthesized compounds elicited the potent inhibitory action against all the tested bacterial stains. Furthermore, in order to explore the antioxidant potential of newly synthesized compounds, the free radical scavenging activity measurement were performed by the 1,1-diphenyl-2-picryl-hydrazyl (DPPH) assay method. It is revealed from the antioxidant screening results that the compounds 9c and f manifested profound antioxidant potential.     

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.     

Phytochemical and chemotaxonomic study of Pulsatilla cernua (Thunb.) Bercht. ex J. Presl

Twenty-two compounds were isolated from the 70% EtOH–H₂O extract of Pulsatilla cernua (Thunb.) Bercht. ex J. Presl roots, and their structures were determined based on ¹H NMR, ¹³C NMR and MS spectroscopic data, including (+)-pinoresinol (1), matairesinol (2), 4-ethoxycinnamic acid (3), p-hydroxy ethyl cinnamate (4), 3-(4′-methoxyphenyl)-2(E)-propenoic acid (5), methyl 4-hydroxycinnamate (6), radicol (7), cryptomeridiol (8), fraxinellone (9), diolmycin B2 (10), hederagonic acid (11), hederagenin (12), oleanolic acid (13), 3-O-α-L-arabinopyranosyl-oleanolic acid (14), hederagenin 3-O-α-L-arabinopyranoside (15), 3-O-[α-L-rhamnopyranosyl-(1→2)-α-L-arabinopyranosyl] oleanolic acid (16), hederasaponin B (17), kizutasaponin K₁₂ (18), patrinia saponin H3 (19), hederacholichiside F (20), cernuoside A (21) and cernuoside B (22). Eight compounds (3–10) were isolated and identified from the genus Pulsatilla for the first time.     

Formation of (-⊃-1′,2′-epi-2-cis-xanthoxin acid from a precursor of abscisic acid

Tomato shoots and avocado mesocarp supplied with (±)-[2-¹⁴C]-5-(1,2-epoxy-2,6,6-trimethylcyclohexyl)-3-methylpenta-cis-2-trans-4-dienoic acid metabolize it into (+)-abscisic acid and a more polar material that was isolated and identified as (?)-epi-1′(R),2′(R)-4′(S)-2-cis-xanthoxin acid. The (+)-1′(S),2′(S)-4′(S)-2-cis-xanthoxin acid recently synthesized from natural violaxanthin, has the 1′,2′-epoxy group on the opposite side of the ring to that of the 4′(S)-hydroxyl group and the compound is rapidly converted into (+)-abscisic acid. The 1′,2′-epoxy group of (?)-1′,2′-epi-2-cis-xanthoxin acid is on the same side of the ring as the 4′(S) hydroxyl group: the compound is not metabolized into abscisic acid. The configuration of the 1′,2′-epoxy group probably controls whether or not the 4′(S) hydroxyl group can be oxidized. (+)-2-cis-Xanthoxin acid is probably not a naturally occurring intermediate because a ‘cold trap’, added to avocado fruit forming [¹⁴C]-labelled abscisic acid from [2-¹⁴C]mevalonate, failed to retain [¹⁴C] label.     

Biochemical Characterization of the Oxygenation of Unsaturated Fatty Acids by the Dioxygenase and Hydroperoxide Isomerase of Pseudomonas aeruginosa 42A2

We have studied oxygenation of fatty acids by cell extract of Pseudomonas aeruginosa 42A2. Oleic acid ((9Z)-18:1) was transformed to (10S)-hydroperoxy-(8E)-octadecenoic acid ((10S)-HPOME) and to (7S,10S)-dihydroxy-(8E)-octadecenoic acid (7,10-DiHOME). Experiments under oxygen-18 showed that 7,10-DiHOME contained oxygen from air and was formed sequentially from (10S)-HPOME by isomerization. (10R)-HPOME was not isomerized. The (10S)-dioxygenase and hydroperoxide isomerase activities co-eluted on ion exchange chromatography and on gel filtration with an apparent molecular size of ∼50 kDa. 16:1n-7, 18:2n-6, and 20:1n-11 were also oxygenated to 7,10-dihydroxy fatty acids, and (8Z)-18:1 was oxygenated to 6,9-dihydroxy-(7E)-octadecenoic acid. A series of fatty acids with the double bond positioned closer to ((6Z)-18:1, (5Z,9Z)-18:2) or more distant from the carboxyl group ((11Z)-, (13Z)-, and (15Z)-18:1) were poor substrates. The oxygenation mechanism was studied with [7S-²H]18:1n-9, [7R-²H]18:2n-6, and [8R-²H]18:2n-6 as substrates. The pro-R hydrogen at C-8 was lost in the biosynthesis of (10S)-HPODE, whereas the pro-S hydrogen was lost and the pro-R hydrogen was retained at C-7 during biosynthesis of the 7,10-dihydroxy metabolites. Analysis of the fatty acid composition of P. aeruginosa revealed relatively large amounts of (9E/Z)-16:1 and (11E/Z)-18:1 and only traces of 18:1n-9. We found that (11Z)-18:1 (vaccenic acid) was transformed to (11S,14S)-dihydroxy-(12E)-octadecenoic acid and to a mixture of 11- and 12-HPOME, possibly due to reverse orientation of (11Z)-18:1 at the active site compared with oleic acid. The reaction mechanism of the hydroperoxide isomerase suggests catalytic similarities to cytochrome P450.