Phytochemistry (v.94, #C)

Recent advances in the molecular genetics of pyridine alkaloid production and regulation in Nicotiana tabacum are reviewed.Alkaloids represent an extensive group of nitrogen-containing secondary metabolites that are widely distributed throughout the plant kingdom. The pyridine alkaloids of tobacco (Nicotiana tabacum L.) have been the subject of particularly intensive investigation, driven largely due to the widespread use of tobacco products by society and the role that nicotine (16) (see Fig. 1) plays as the primary compound responsible for making the consumption of these products both pleasurable and addictive. In a typical commercial tobacco plant, nicotine (16) comprises about 90% of the total alkaloid pool, with the alkaloids nornicotine (17) (a demethylated derivative of nicotine), anatabine (15) and anabasine (5) making up most of the remainder. Advances in molecular biology have led to the characterization of the majority of the genes encoding the enzymes directly responsible the biosynthesis of nicotine (16) and nornicotine (17), while notable gaps remain within the anatabine (15) and anabasine (5) biosynthetic pathways. Several of the genes involved in the transcriptional regulation and transport of nicotine (16) have also been elucidated. Investigations of the molecular genetics of tobacco alkaloids have not only provided plant biologists with insights into the mechanisms underlying the synthesis and accumulation of this important class of plant alkaloids, they have also yielded tools and strategies for modifying the tobacco alkaloid composition in a manner that can result in changing the levels of nicotine (16) within the leaf, or reducing the levels of a potent carcinogenic tobacco-specific nitrosamine (TSNA). This review summarizes recent advances in our understanding of the molecular genetics of alkaloid biosynthesis in tobacco, and discusses the potential for applying information accrued from these studies toward efforts designed to help mitigate some of the negative health consequences associated with the use of tobacco products.
Keywords: Nicotiana tabacum; Solanaceae; Nicotine; Nornicotine; Tobacco-specific nitrosamines; Alkaloid biosynthetic genes;

Domain-swapping and deletion analysis established that the overall folding of RZs governed by the N-terminal RRM domain and the C-terminal glycine-rich region, as well as the size of the disordered C-terminal glycine-rich region, are crucial for RNA chaperone activity of RZs.Despite the increase in understanding of RNA chaperone activity of zinc finger-containing glycine-rich RNA-binding proteins (RZs) during the cold adaptation process, the structural features relevant to the RNA chaperone activity of RZs still largely remain to be established. To investigate the structural determinants important for the RNA chaperone activity of RZs, domain-swapping and deletion analyses was carried out to assess the contribution of the N-terminal zinc finger RNA-recognition motif (RRM) domain and the C-terminal glycine-rich region of wheat (Triticum avestivum) and rice (Oryza sativa) RZs to RNA chaperone activity. Although the amino acid sequence similarity among wheat TaRZ2, wheat TaRZ3, and rice OsRZ1 was high, only TaRZ2 had RNA chaperone activity as evidenced by complementation ability in cold-sensitive Escherichia coli mutant cell under cold stress and in vivo and in vitro nucleic acid-melting activity. Domain-swapping and deletion analysis demonstrated that the overall folding of RZs governed by the N-terminal RRM domain and the C-terminal glycine-rich region, as well as the size of the disordered C-terminal glycine-rich region, are crucial for the RNA chaperone activity of RZs. Collectively, these results indicate that a specific modular arrangement of RRM domain and the disordered C-terminal region determines the RNA chaperone activity of RZs in cells.
Keywords: Oryza sativa; Triticum avestivum; Poaceae; RNA-binding protein; RNA chaperone; RZ protein; Zinc finger glycine-rich RNA-binding protein;

Homology model of Avicennia marina enoyl-CoA hydratase mainly comprises α-helical regions. Glu106 is the conserved catalytic residue while Glu86 is replaced by Gly. Asp114 is proposed as the second catalytic residue. There were structural variations in mobile loop and formation and loss of interactions between active site and substrate.Enoyl-CoA hydratase catalyzes the hydration of 2-trans-enoyl-CoA into 3-hydroxyacyl-CoA. The present study focuses on the correlation between the functional and structural aspects of enoyl-CoA hydratase from Avicennia marina. We have used bioinformatics tools to construct and analyze 3D homology models of A. marina enoyl-CoA hydratase (AMECH) bound to different substrates and inhibitors and studied the residues involved in the ligand–enzyme interaction. Structural information obtained from the models was compared with those of the reported crystal structures. We observed that the overall folds were similar; however, AMECH showed few distinct structural changes which include structural variation in the mobile loop, formation and loss of certain interactions between the active site residues and substrates. Some changes were also observed within specific regions of the enzyme. Glu106 is almost completely conserved in sequences of the isomerases/hydratases including AMECH while Glu86 which is the other catalytic residue in most of the isomerases/hydratases is replaced by Gly and shows no interaction with the substrate. Asp114 is located within 4 Å distance of the catalytic water which makes it a probable candidate for the second catalytic residue in AMECH. Another prominent feature of AMECH is the presence of structurally distinct mobile loop having a completely different coordination with the hydrophobic binding pocket of acyl portion of the substrate.
Keywords: Homology modeling; Structure prediction; Substrate interactions; Salt stress;

Abolishing activity against ascorbate in a cytosolic ascorbate peroxidase from switchgrass by Frank A. Kovacs; Gautam Sarath; Kyle Woodworth; Paul Twigg; Christian M. Tobias (45-52).
Cytosolic ascorbate peroxidase from switchgrass was cloned and expressed to obtain purified active monomeric protein. R172S mutation removed ascorbate activity but maintained aromatic substrate reactivity.Switchgrass (Panicum virgatum L.) is being developed as a bioenergy species. Recently an early version of its genome has been released permitting a route to the cloning and analysis of key proteins. Ascorbate peroxidases (APx) are an important part of the antioxidant defense system of plant cells and present a well studied model to understand structure–function relationships. Analysis of the genome indicates that switchgrass encodes several cytosolic ascorbate peroxidases with apparent varying levels of tissue expression. A major cytosolic ascorbate peroxidase was thus selected for further studies. This gene was cloned and expressed in Escherichia coli cells to obtain purified active protein. Full heme incorporation of the enzyme was achieved utilizing slow growth and supplementing the media with 5-aminolevulinic acid. The enzyme was observed to be monomeric in solution via size exclusion chromatography. Activity toward ascorbate was observed that was non-Michaelis–Menten in nature. A site-directed mutant, R172S, was made in an attempt to differentiate activity against ascorbate versus other substrates. The R172S protein exhibited negligible ascorbate peroxidase activity, but showed near wild type activity toward other aromatic substrates.
Keywords: Ascorbate; Peroxidase; Panicum virgatum; Poaceae; Mutant; ABTS;

Biochemical and immunological characterization of a recombinantly-produced antifungal cysteine proteinase inhibitor from green kiwifruit (Actinidia deliciosa) by Milica Popovic; Uros Andjelkovic; Lidija Burazer; Buko Lindner; Arnd Petersen; Marija Gavrovic-Jankulovic (53-59).
Heterologously produced cysteine proteinase inhibitor (rCPI) from green kiwifruit possesses antifungal properties against Alternaria radicina and Botrytis cinerea confirmed by measuring fungal growth and microscopical observation.Plant proteinase inhibitors are considered important defense molecules against insect and pathogen attack. The cysteine proteinase inhibitor (CPI) from green kiwifruit (Actinidia deliciosa) belongs to the cystatin family and shows potent antifungal activity (in vitro and in vivo). However, the low abundance of this molecule in fruit (6 μg/g of fresh fruit) seems to limit further investigations on the interaction between phytocystatin and photopathogenic fungi. In this paper the cDNA of the kiwi CPI was expressed in Escherichia coli. Fifteen N-terminal amino acids were identified by Edman degradation, and 77% of the rCPI primary structure was confirmed by mass fingerprint. The structural homology of recombinant CPI (rCPI) to its natural counterpart has been clearly demonstrated in immunological assays (immunoblot and ELISA inhibition). Biological activity of rCPI was demonstrated in inhibition assay with cysteine proteinase papain (EC50 2.78 nM). In addition, rCPI reveals antifungal properties toward pathogenic fungi (Alternaria radicina and Botrytis cinerea), which designates it as an interesting model protein for the exploration of plant phytocystatins – pathogen interactions. Understanding the molecular mechanisms of natural plant resistance could lead to the development of ecologically safe fungicides for controlling post-harvest diseases and maintaining food quality.
Keywords: Cysteine proteinase inhibitor; Phytocystatin; Antifungal activity; Kiwifruit;

Vacuolar iron transporter (CcVIT) was critical for blue coloration of cornflower and amino acid replacement (A236E) reduced the activity to give purple color petals.The blue petal color of the cornflower (Centaurea cyanus) is caused by protocyanin, a kind of metalloanthocyanin, which is a self-assembled supramolecular metal complex pigment. Protocyanin is composed of six molecules of anthocyanin, six molecules of flavone, one ferric ion, and one magnesium ion. The ferric ion is essential for blue color development. Here, we identify the vacuolar iron transporter gene (CcVIT) from the blue petals of C. cyanus and its function is identified and characterized. The CcVIT transcript was observed only in the petals. Its amino acid sequence is highly homologous to the Arabidopsis thaliana (AtVIT1) and Tulipa gesneriana (TgVit1) vacuolar iron transporters. Heterologous expression of the CcVIT gene in yeast indicated that the corresponding gene product transports ferrous ion into vacuoles. Analysis of purple mutant-line petals clarified that the anthocyanin and flavone components were the same as those found in plants with blue petals, but the amount of iron ions in the colored cells decreased, and consequently the amount of blue protocyanin was reduced. The CcVIT gene was expressed even in purple mutant petals, however, an amino acid substitution (A236E) occurred in that case. This change in the CcVIT gene sequence also resulted in loss of iron transport activity. The CcVIT protein thus plays a critical role in the blue coloration of cornflower petals.
Keywords: Centaurea cyanus; Asteraceae; Amino acid substitution; Blue flower coloration; Metalloanthocyanin; Protocyanin; Iron distribution; Vacuolar iron transporter;

Identification of the thiamin salvage enzyme thiazole kinase in Arabidopsis and maize by Mohammad Yazdani; Rémi Zallot; Meral Tunc-Ozdemir; Valérie de Crécy-Lagard; David K. Shintani; Andrew D. Hanson (68-73).
Plants, like microorganisms, can salvage the thiazole moiety of thiamin for reuse in thiamin synthesis. Plant genes encoding the key salvage enzyme thiazole kinase were identified by genomics, genetics, and biochemistry.The breakdown of thiamin (vitamin B1) and its phosphates releases a thiazole moiety, 4-methyl-5-(2-hydroxyethyl)thiazole (THZ), that microorganisms and plants are able to salvage for re-use in thiamin synthesis. The salvage process starts with the ATP-dependent phosphorylation of THZ, which in bacteria is mediated by ThiM. The Arabidopsis and maize genomes encode homologs of ThiM (At3g24030 and GRMZM2G094558, respectively). Plasmid-driven expression of either plant homolog restored the ability of THZ to rescue Escherichia coli thiM deletant strains, showing that the plant proteins have ThiM activity in vivo. Enzymatic assays with purified recombinant proteins confirmed the presence of THZ kinase activity. Furthermore, ablating the Arabidopsis At3g24030 gene in a thiazole synthesis mutant severely impaired rescue by THZ. Collectively, these results show that ThiM homologs are the main source of THZ kinase activity in plants and are consequently crucial for thiamin salvage.
Keywords: Arabidopsis thaliana; Brassicaceae; Zea mays; Poaceae; Thiamin; Thiazole; Salvage; Comparative genomics;

Molecular and biochemical characterization of the jasmonic acid methyltransferase gene from black cottonwood (Populus trichocarpa) by Nan Zhao; Jianzhuang Yao; Minta Chaiprasongsuk; Guanglin Li; Ju Guan; Timothy J. Tschaplinski; Hong Guo; Feng Chen (74-81).
A black cottonwood (Populus trichocarpa) gene (PtJMT1) encoding jasmonic acid methyltransferase was isolated and biochemically characterized. Escherichia coli-expressed PtJMT1 was active with both jasmonic acid (JA) and benzoic acid, with the former being a more preferred substrate, to produce their corresponding methyl esters methyl jasmonate (MeJA) and methyl benzoate. Two residues in the active site of PtJMT1 were demonstrated to be critical for determining the relative specific activities of PtJMT1 with these two substrates. A number of biotic stress factors were found to induce the expression of PtJMT1, suggesting that this gene has a role in poplar defense against biotic stresses. Phylogenetic analysis provides further insight into the evolution of JMT genes.Methyl jasmonate is a metabolite known to be produced by many plants and has roles in diverse biological processes. It is biosynthesized by the action of S-adenosyl-l-methionine:jasmonic acid carboxyl methyltransferase (JMT), which belongs to the SABATH family of methyltransferases. Herein is reported the isolation and biochemical characterization of a JMT gene from black cottonwood (Populus trichocarpa). The genome of P. trichocarpa contains 28 SABATH genes (PtSABATH1 to PtSABATH28). Recombinant PtSABATH3 expressed in Escherichia coli showed the highest level of activity with jasmonic acid (JA) among carboxylic acids tested. It was therefore renamed PtJMT1. PtJMT1 also displayed activity with benzoic acid (BA), with which the activity was about 22% of that with JA. PtSABATH2 and PtSABATH4 were most similar to PtJMT1 among all PtSABATHs. However, neither of them had activity with JA. The apparent Km values of PtJMT1 using JA and BA as substrate were 175 μM and 341 μM, respectively. Mutation of Ser-153 and Asn-361, two residues in the active site of PtJMT1, to Tyr and Ser respectively, led to higher specific activity with BA than with JA. Homology-based structural modeling indicated that substrate alignment, in which Asn-361 is involved, plays a role in determining the substrate specificity of PtJMT1. In the leaves of young seedlings of black cottonwood, the expression of PtJMT1 was induced by plant defense signal molecules methyl jasmonate and salicylic acid and a fungal elicitor alamethicin, suggesting that PtJMT1 may have a role in plant defense against biotic stresses. Phylogenetic analysis suggests that PtJMT1 shares a common ancestor with the Arabidopsis JMT, and functional divergence of these two apparent JMT orthologs has occurred since the split of poplar and Arabidopsis lineages.
Keywords: Poplar; Populus trichocarpa; Salicaceae; Benzoic acid; SABATH family; Substrate-specificity evolution;

Metabolomic analysis of isonitrosoacetophenone-induced perturbations in phenolic metabolism of Nicotiana tabacum cells by Ntakadzeni E. Madala; Paul A. Steenkamp; Lizelle A. Piater; Ian A. Dubery (82-90).
Isonitrosoacetophenone, a xenobiotic oxime, affects the shikimate-, phenylpropanoid- and flavonoid pathways in tobacco cells.Plants have developed biochemical and molecular responses to adapt to different stress environments. One of the characteristics of the multi-component defence response is the production of defence-related metabolites. Plant defences can be triggered by various stimuli, including synthetic or naturally occurring molecules, especially those derived from pathogens. In the current study, Nicotiana tabacum cell suspensions were treated with isonitrosoacetophenone (INAP), a subcomponent of a plant-derived stress metabolite with anti-fungal and anti-oxidant properties, in order to investigate the effect thereof on cellular metabolism. Subsequent metabolomic-based analyses were employed to evaluate changes in the metabolome. UPLC–MS in conjunction with multivariate data analyses was found to be an appropriate approach to study the effect of chemical inducers like INAP on plant metabolism in this model system. Principal component analysis (PCA) indicated that INAP is capable of inducing time-dependent metabolic perturbations in the cultured cells. Orthogonal projection to latent structures discriminant analysis (OPLS-DA) revealed metabolites of which the levels are affected by INAP, and eight of these were tentatively annotated from the mass spectral data and online databases. These metabolites are known in the context of plant stress- and defence responses and include benzoic- or cinnamic acid derivatives that are either glycosylated or quinilated as well as flavonoid derivatives. The results indicate that INAP affects the shikimate-, phenylpropanoid- and flavonoid pathways, the products of which may subsequently lead to an anti-oxidant environment in vivo.
Keywords: Biotransformation; Induced-defences; Isonitrosoacetophenone; 2-Keto-2-phenyl-acetaldoxime; Metabolism; Metabolomics; Multivariate statistics; Nicotiana tabacum; Xenobiotics;

Medullary CBSCs in the canopy lichens Lobaria pulmonaria and Lobaria scrobiculata peak in seasons with most invertebrate herbivores; the cortical CBSC follows sun exposure at spatial scales.Acetone-extractable carbon based secondary compounds (CBSCs) were quantified in two epiphytic lichens to study possible effects of external factors (season and aspect) on secondary chemistry and to relate defense investments to biomass growth and changes in specific thallus mass (STM). At the end of four separate annual cycles starting in each of the four seasons, the cyanolichen Lobaria scrobiculata and the cephalolichen Lobaria pulmonaria (green algae as the primary photobiont and with localized Nostoc in internal cephalodia) were monitored in their natural forest habitats and after being transplanted at three contrasting aspects in open sites. Season strongly influenced most CBSCs. Medullary CBSCs in both species were twice as high in summer as in winter. Aspect hardly affected major CBSCs, whereas transplantation from forest to clear-cut slightly reduced these compounds. No major CBSCs in any species showed a trade-off with growth rate. Dry matter- as well as thallus area-based medullary CBSC contents increased with STM. The cortical usnic acid strongly increased with growth rate and followed spatial, but not seasonal variations in light exposure. Maximal CBSC levels during seasons with most herbivores is consistent with the hypothesis inferring that herbivory is a major selective force for CBSCs. Lack of trade-off between growth and defence investments suggests that these two processes do not compete for photosynthates.
Keywords: Lobaria pulmonaria; Lobaria scrobiculata; Canopy lichens; Aspect; Seasonal variation; Growth rate; Lichen compounds; Stictic acid; Usnic acid;

The typical defense compounds of Plantaginaceae are the iridoid glycosides, which retard growth and/or enhance mortality of non-adapted herbivores. In plants, glycosidic defense compounds and hydrolytic enzymes often form a dual defense system, in which the glycosides are activated by the enzymes to exert biological effects. Yet, little is known about the activating enzymes in iridoid glycoside-containing plants. To examine the role of plant-derived β-glucosidases in the dual defense system of two common plantain species, Plantago lanceolata and Plantago major, we determined the concentration of iridoid glycosides as well as the β-glucosidase activity in leaves of different age. To investigate the presence of other leaf metabolites potentially involved in plant defense, we used a metabolic fingerprinting approach with ultra-high performance liquid chromatography coupled with time-of-flight-mass spectrometry. According to the optimal defense hypothesis, more valuable parts such as young leaves should be better protected than less valuable parts. Therefore, we expected that both, the concentrations of defense compounds as well as the β-glucosidase activity, should be highest in younger leaves and decrease with increasing leaf age. Both species possessed β-glucosidase activity, which hydrolyzed aucubin, one of the two most abundant iridoid glycosides in both plant species, with high activity. In line with the optimal defense hypothesis, the β-glucosidase activity in both Plantago species as well as the concentration of defense-related metabolites such as iridoid glycosides correlated negatively to leaf age. When leaf extracts were incubated with bovine serum albumin and aucubin, SDS–PAGE revealed a protein-denaturing effect of the leaf extracts of both plantain species, suggesting that iridoid glycosides and plant β-glucosidase interact in a dual defense system.
Keywords: Plantago major; Plantago lanceolata; Plantaginaceae; Iridoid glycosides; β-Glucosidase; Dual defense system; Metabolic fingerprinting; Optimal defense theory; Protein denaturation;

Flavonoids from the cocoon of Rondotia menciana by Chikara Hirayama; Hiroshi Ono; Yan Meng; Toru Shimada; Takaaki Daimon (108-112).
Larvae of Rondotia menciana uptake flavonol glycosides from Morus alba leaf. Main components of the R. menciana cocoon were flavonol galactosides, which were not present in its host plant. The result suggests that conjugation reaction of flavonoids with galactose occurs in the insect.Two flavonol glycosides along with four known flavonoids were isolated from the cocoon of the mulberry white caterpillar, Rondotia menciana (Lepidoptera: Bombycidae: Bombycinae), a closely related species of the domesticated silkworm Bombyx mori, both of which feed on leaves of mulberry (Morus alba). The two glycosides were characterized as quercetin 3-O-β-d-galactopyranosyl-(1 → 3)-β-d-galactopyranoside and kaempferol 3-O-β-d-galactopyranosyl-(1 → 3)-β-d-galactopyranoside, based on spectroscopic data and chemical evidence. The flavonol galactosides found in the cocoon were not present in the host plant, nor in the cocoon of the silkworm, B. mori. Notably, flavonol glucosides, which are the main constituents of cocoon flavonoids in B. mori mori, were not found in the R. menciana cocoon. The present result strongly suggests that R. menciana is quite unique in that they predominantly use an UDP-galactosyltransferase for conjugation of dietary flavonoids, whereas UDP-glucosyltransferases are generally used for conjugation of plant phenolics and xenobiotics in other insects.
Keywords: Morus alba; Moraceae; Cocoon; Flavonol galactosides; Flavonoids; Conjugation; Rondotia menciana; Bombyx mori;

Inducibility of chemical defences by two chewing insect herbivores in pine trees is specific to targeted plant tissue, particular herbivore and defensive trait by Xoaquín Moreira; Lina Lundborg; Rafael Zas; Amparo Carrillo-Gavilán; Anna-Karin Borg-Karlson; Luis Sampedro (113-122).
Bark feeding by the weevil, but not folivory by the caterpillar, increased total terpenes in those tissues targeted by the herbivores. Both pine species showed similar responses. Some particular monoterpenes appeared more responsive.There is increasing evidence that plants can react to biotic aggressions with highly specific responses. However, few studies have attempted to jointly investigate whether the induction of plant defences is specific to a targeted plant tissue, plant species, herbivore identity, and defensive trait. Here we studied those factors contributing to the specificity of induced defensive responses in two economically important pine species against two chewing insect pest herbivores. Juvenile trees of Pinus pinaster and P. radiata were exposed to herbivory by two major pest threats, the large pine weevil Hylobius abietis (a bark-feeder) and the pine processionary caterpillar Thaumetopoea pityocampa (a folivore). We quantified in two tissues (stem and needles) the constitutive (control plants) and herbivore-induced concentrations of total polyphenolics, volatile and non-volatile resin, as well as the profile of mono- and sesquiterpenes. Stem chewing by the pine weevil increased concentrations of non-volatile resin, volatile monoterpenes, and (marginally) polyphenolics in stem tissues. Weevil feeding also increased the concentration of non-volatile resin and decreased polyphenolics in the needle tissues. Folivory by the caterpillar had no major effects on needle defensive chemistry, but a strong increase in the concentration of polyphenolics in the stem. Interestingly, we found similar patterns for all these above-reported effects in both pine species. These results offer convincing evidence that induced defences are highly specific and may vary depending on the targeted plant tissue, the insect herbivore causing the damage and the considered defensive compound.
Keywords: Hylobius abietis; Pinus pinaster; Pinus radiata; Thaumetopoea pityocampa; Needles; Phloem; Resin; Monoterpenes; Sesquiterpenes; Phenolics;

Flower color polymorphism in Iris lutescens (Iridaceae): Biochemical analyses in light of plant–insect interactions by Hui Wang; Lucie Conchou; Jean-Marie Bessière; Guillaume Cazals; Bertrand Schatz; Eric Imbert (123-134).
The different accumulation of delphinidin derivatives is mainly responsible for flower color difference between purple- and yellow-flowered I. lutescens, while the color polymorphism is not associated with significant difference in the scent emitted between the two morphs.We describe a flower color polymorphism in Iris lutescens, a species widespread in the Northern part of the Mediterranean basin. We studied the biochemical basis of the difference between purple and yellow flowers, and explored the ecological and evolutionary consequences of such difference, in particular visual discrimination by insects, a potential link with scent emitted and the association between color and scent. Anthocyanins were found to be present in much greater concentrations in purple flowers than in yellow ones, but the anthocyanin composition did not differ between color morphs. Likewise, no quantitative difference in anthocyanin content was found between vegetative tissues of the two morphs. Floral anthocyanins were dominated by delphinidin 3-O-(p-coumaroylrutinoside)-5-O-glucoside (also called delphanin) and its aliphatic derivatives. Small amounts of delphinidin 3-O-(p-caffeoylrutinoside)-5-O-glucoside and its aliphatic derivatives were also characterized. Based on a description of bumblebees’ (one of the main pollinators of I. lutescens) color perception, purple and yellow flowers of I. lutescens could be visually discriminated as blue and blue-green, respectively, and likely by a wide variety of other insects. The overall chemical composition of the scent produced was not significantly different between morphs, being dominated by terpenoids, mainly myrcene, (E)-β-ocimene and limonene. A slight color-scent correlation was nevertheless detected, consistent with the shared biosynthetic origin of both pigments and volatile compounds. Therefore in this species, the difference in the amounts of pigments responsible for flower color difference seems to be the major difference between the two morphs. Pollinators are probably the main selective agent driving the evolution of flower color polymorphism in I. lutescens, which represents a suitable species for investigating how such polymorphism is maintained.
Keywords: Flower color polymorphism; Iris lutescens; Anthocyanins; Floral scent; Terpenoids; Plant–insect interactions; Color-scent association;

Thiamine treatments alleviate aphid infestations in barley and pea by Afaf M. Hamada; Lisbeth M.V. Jonsson (135-141).
Thiamine is a vitamin synthesized in plant leaves. We show here that aphid life span and reproduction is reduced on plants treated with additional thiamine.Treatment of plants with thiamine (Vitamin B1) has before been shown to activate plant defence against microorganisms. Here, we have studied the effects of thiamine treatments of plants on aphid reproduction and behaviour. The work was mainly carried out with bird cherry-oat aphid (Rhopalosiphum padi L.) on barley (Hordeum vulgare L.). Aphid population growth and aphid acceptance on plants grown from seeds soaked in a 150 μM thiamine solution were reduced to ca. 60% of that on control plants. R. padi life span and the total number of offspring were reduced on barley plants treated with thiamine. Healthy aphids and aphids infected with the R. padi virus were similarly affected. Spraying or addition of thiamine at 150 μM to nutrient solutions likewise resulted in reduced aphid population growth to ca. 60%, as did plant exposure to thiamine odour at 4 mM. Thiamine treatments resulted in reduced aphid population growth also when tested with grain aphid (Sitobion avenae F.) on barley and pea aphid (Acyrthosiphon pisum H.) on pea (Pisum sativum L.). There was no direct effect of thiamine on aphid reproduction or thiamine odour on aphid behaviour, as evaluated using artificial diets and by olfactometer tests, respectively.Two gene sequences regulated by salicylic acid showed higher transcript abundance and one gene sequence regulated by methyl jasmonate showed lower transcript abundance in thiamine-treated plants but not in control plants after aphid infestation. These results suggest that the aphid antibiosis and antixenosis effects may be related to priming of defence, but more studies are needed to explain the effects against aphids.
Keywords: Hordeum vulgare; Pisum sativum; Acyrthosiphon pisum; Rhopalosiphum padi; Sitobion avenae; Aphid resistance; Thiamine; Vitamin B1;

Effect of aluminum on metabolism of organic acids and chemical forms of aluminum in root tips of Eucalyptus camaldulensis Dehnh. by Takashi Ikka; Tsuyoshi Ogawa; Donghua Li; Syuntaro Hiradate; Akio Morita (142-147).
Eucalyptus camaldulensis can detoxify Al by forming Al–citrate complexes, and this is achieved through citrate accumulation via suppression of the citrate decomposition pathway.Eucalyptus (Eucalyptus camaldulensis) has relatively high resistance to aluminum (Al) toxicity than the various herbaceous plants and model plant species. To investigate Al-tolerance mechanism, the metabolism of organic acids and the chemical forms of Al in the target site (root tips) in Eucalyptus was investigated. To do this, 2-year old rooted cuttings of E. camaldulensis were cultivated in half-strength Hoagland solution (pH 4.0) containing Al (0, 0.25, 0.5, 1.0, 2.5 and 5.0 mM) salts for 5 weeks; growth was not affected at concentrations up to 2.5 mM even with Al concentration reaching 6000 μg g−1 DW. In roots, the citrate content also increased with increasing Al application. Concurrently, the activities of aconitase and NADP+-isocitrate dehydrogenase, which catalyze the decomposition of citrate, decreased. On the other hand, the activity of citrate synthase was not affected at concentrations up to 2.5 mM Al. 27Al-NMR spectroscopic analyses were carried out where it was found that Al–citrate complexes were a major chemical form present in cell sap of root tips. These findings suggested that E. camaldulensis detoxifies Al by forming Al–citrate complexes, and that this is achieved through Al-induced citrate accumulation in root tips via suppression of the citrate decomposition pathway.
Keywords: Eucalyptus; Eucalyptus camaldulensis; Myctaceae; Aluminum tolerance; Organic acid metabolism; 27Al-NMR analysis; Al–citrate complex;

Covariation within and among geographically structured chemotypes of Eucalyptus tricarpa shed light on biosynthesis.Display OmittedAustralian members of the Myrtaceae produce large quantities of ecologically and economically important terpenes and display abundant diversity in both yield and composition of their oils. In a survey of the concentrations of leaf terpenes in Eucalyptus tricarpa (L.A.S. Johnson) L.A.S. Johnson & K.D. Hill, which were previously known from few samples, exceptional variability was found in composition. The aim was to characterize the patterns of variation and covariation among terpene components in this species and to use this information to enhance our understanding of their biosynthesis. There were marked discontinuities in the distributions of numerous compounds, including the overall proportions of mono- and sesquiterpenes, leading us to delineate three distinct chemotypes. Overall, positive covariation predominated, but negative covariation suggested competitive interactions involved in monoterpene synthesis. Two groups of covarying monoterpenes were found, each of which was positively correlated with a group of sesquiterpenes and negatively correlated with the alternate sesquiterpene group. These results imply substantial cross-talk between mono- and sesquiterpene biosynthesis pathways. However, only those compounds hypothesized to share final carbocation intermediates or post-processing steps were strongly positively correlated within chemotypes. This suggests that the broader patterns of covariation among groups of compounds may result from co-regulation of multiple biosynthetic genes, controlling the complex terpene profiles of the chemotypes of Eucalyptus.
Keywords: Eucalyptus; Myrtaceae; Terpene; Chemotype; Population differentiation; Biosynthetic constraint;

Secondary metabolites from the aerial parts of Centaurea pannonica (Heuff.) Simonk. from Serbia and their chemotaxonomic importance by Tanja Milošević Ifantis; Slavica Solujić; Dragana Pavlović-Muratspahić; Helen Skaltsa (159-170).
Structure of sesquiterpene lactones (13) isolated from the C. pannonica non polar extract.Display OmittedThe non polar extract of Centaurea pannonica (Heuff.) Simonk., growing wild in Serbia, was studied and twenty-five compounds including 14 sesquiterpene lactones, 7 flavonoids, 3 lignans and 1 phenylpropanoid glycoside were isolated. All compounds were isolated for the first time from this species. Among them, one germacranolide 2α-hydroxy, 8-dehydroxy 15-O-methacrylate salonitenolide (1) and two guaianolides 2α,8α-dihydroxy-dehydrocostus lactone (2) and pannonin (3) are new natural compounds. The structures of the compounds were established on the basis of spectroscopic analyses (UV, IR, HREIMS and 1D & 2D NMR). The chemical profile of C. pannonica, which belongs to the “Centaurea jacea” group, was compared to previously studied taxa of the same group and used to assess the phylogenetic relationships in the group.
Keywords: C. pannonica (Heuff.) Simonk.; Centaurea L.; Centaurea jacea group; NMR; Guaianolides; Flavonoids; Lignans; Chemotaxonomy;

Molecular classification of the natural exudates of the rosids by Joseph B. Lambert; Eric W. Donnelly; Eric A. Heckenbach; Connor L. Johnson; Michael A. Kozminski; Yuyang Wu; Jorge A. Santiago-Blay (171-183).
Phylogenetic tree at the ordinal level of the rosid clade.Exudates of the rosid clade of the eudicots have been surveyed and characterized by carbon-13 and proton nuclear magnetic resonance spectroscopy. Of 554 samples divided roughly equally between the subclades fabids and malvids, about two-fifths are resins, a third gums, one-ninth gum resins, one-twelfth kinos, and the remaining not affiliated with these four main molecular classes. Two small new molecular classes, respectively from the Clusiaceae (xanthics) and the Zygophyllaceae (guaiacs), are identified and described.
Keywords: Exudates; Fabids; Gums; Malvids; Nuclear magnetic resonance spectroscopy; Resins; Rosids;

Dichapetalins from Dichapetalum species and their cytotoxic properties by Christophe Long; Yannick Aussagues; Nicolas Molinier; Laurence Marcourt; Laure Vendier; Arnaud Samson; Valérie Poughon; Patrick B. Chalo Mutiso; Frédéric Ausseil; François Sautel; Paola B. Arimondo; Georges Massiot (184-191).
Six dichapetalins named dichapetalins N–S were isolated from Dichapetalum mombuttense, D. zenkeri and D. leucosia.Six dichapetalins named dichapetalins N–S were isolated from Dichapetalum mombuttense, Dichapetalum zenkeri and Dichapetalum leucosia. They were accompanied in the same plants by the known dichapetalins A, B, C, I, L and M. The structures of the compounds were elucidated by 1D and 2D NMR experiments and mass spectrometry. They all possessed the dammarane skeleton substituted at position C-3 by a C6–C2 unit forming a 2-phenylpyran moiety. All contained a lactone ring in the side chain except dichapetalins O, Q and R, in which this ring was replaced by a lactol. Dichapetalin Q and R were also the first dichapetalins bearing a tertiary methyl and a double bond instead of the cyclopropane of the dammaranes. All these compounds were assayed against cancer cell lines HCT116 and WM 266-4 and displayed cytotoxic and anti-proliferative activities in the 10–6 to 10–8  M range.
Keywords: Dichapetalum mombuttense; Dichapetalum leucosia; Dichapetalum ruhlandii; Dichapetalum zenkeri; Dichapetalum eickii; Dichapetalaceae; Dammarane; Cytotoxicity;

Terpenes and polyacetylenes from cultivated Artemisia granatensis boiss (Royal chamomile) and their defensive properties by Alejandro F. Barrero; M. Mar Herrador del Pino; Adriana González Portero; Pilar Arteaga Burón; Jesús F. Arteaga; Jesús Burillo Alquézar; Carmen Elisa Díaz; Azucena González Coloma. (192-197).
Artemisia granatensis was cultivated in artificial systems and some of the compounds isolated from ethanolic extracts showed interesting insect antifeedant effects. Artemisia granatensis, an endemic endangered plant species from Sierra Nevada (Spain) has been successfully cultivated in artificial systems (plants in artificial soil and transformed in vitro roots) to generate enough plant biomass (aerial and root) to allow for its chemical and biological study and at the same time to provide with methods for the sustainable production of the plant and its metabolites. A eudesmanolide (17) along with six sesquiterpenes (1116), nine monoterpenes (210), one nor-monoterpene (1), three acetylenic spiroacetal enoleters (1820) and one coumarin (21) have been identified from the aerial plant ethanolic extract. Acetylenic spiroacetal enoleters 1819 and coumarins 2123 have been isolated from the transformed root ethanolic extract. These extracts and some isolated compounds or mixtures of them have been tested for their insect antifeedant effects against Spodoptera littoralis, Myzus persicae and Rhopalosiphum padi. Significant antifeedant properties were determined for the aerial plant extract, spiroacetals 1920 and secoguaianolides 13  +  14 and 16.
Keywords: Artemisia granatensis; Cultivation; Polyacetylenes; Terpenes; Antifeedant effects;

Thielavins A, J and K: α-Glucosidase inhibitors from MEXU 27095, an endophytic fungus from Hintonia latiflora by José Rivera-Chávez; Martín González-Andrade; María del Carmen González; Anthony E. Glenn; Rachel Mata (198-205).
From the endophytic fungus MEXU 27097, isolated from Hintonia latiflora, three tridepsides were obtained and characterized as thielavins A, J and K. In vitro, in silico and in vivo studies confirmed their α-glucosidase inhibitory properties.Display OmittedBioassay-guided fractionation of the bio-active organic extract obtained from solid-media culture of MEXU 27095, an endophytic fungus isolated from the Mexican medicinal plant Hintonia latiflora (Rubiaceae), led to separation of three tridepsides which were identified as thielavins A, J and K. All three compounds inhibited Saccharomyces cerevisieae α-glucosidase (αGHY) in a concentration-dependent manner with IC50 values of 23.8, 15.8, and 22.1 μM, respectively. Their inhibitory action was higher than that of acarbose (IC50  = 545 μM), used as a positive control. Kinetic analysis established that the three compounds acted as non-competitive inhibitors with ki values of 27.8, 66.2 and 55.4 μM, respectively (α = 1.0, 1.2, 0.7, respectively); acarbose behaved as competitive inhibitor with a ki value of 156.1 μM. Thielavin J inhibited the activity of α-glucosidase from Bacillus stearothermophilus (αGHBs) with an IC50 of 30.5 μM, being less active than acarbose (IC50  = 0. 015 μM); in this case, compound (2) (ki  = 20.0 μM and α = 2.9) and acarbose (ki  = 0.008 μM and α = 1.9) behaved as non-competitive inhibitors. Docking analysis predicted that all three thielavins and acarbose bind to homologated αGHBs and to αGHY (PDB: 3A4A) in a pocket close to the catalytic site for maltose and isomaltose, respectively. The α-glucosidase inhibitory properties of thielavin K (3) were corroborated in vivo since it induced a noted antihyperglycemic action during an oral sucrose tolerance test (3.1, 10.0 and 31.6 mg/kg) in normal and nicotinamide–streptozotocin diabetic mice. In addition, at a dose of 10 mg/kg, it provoked a moderate hypoglycemic activity in diabetic mice.
Keywords: Type II diabetes mellitus; Hintonia latiflora; Endophytic fungus; Thielavins; α-Glucosidase; Enzymatic inhibition; Molecular docking; Oral sucrose tolerance test;

Galactofuranose-rich polysaccharides from Trebouxia sp. induce inflammation and exacerbate lethality by sepsis in mice by Yanna D. Rattmann; Simone M. Malquevicz-Paiva; Marcello Iacomini; Lucimara M.C. Cordeiro (206-210).
This study demonstrated the in vivo proinflammatory effects of polymers of galactofuranose. Trebouxia sp. is a genus of green algae that is a symbiotic partner of lichenized fungi. Previous studies conduced demonstrated that Trebouxia sp. is able to produce galactofuranose-rich polysaccharides (β-d-galactofuranan, mannogalactofuranan), which were able to activate macrophages in vitro. The present study was proposed to investigate the effects of SK10 polysaccharides fraction from Trebouxia sp. on the model of polymicrobial sepsis induced by cecal ligation and puncture in mice in vivo. The subcutaneous administration of SK10 increased the late mortality rate by 20%, stimulated neutrophil accumulation in lungs (indirectly measured through myeloperoxidase activity) and also Interleukin-1β, creatinine and glucose serum levels. Moreover this study demonstrates the in vivo proinflammatory effects of polymers of galactofuranose and that they can act as pathogen-associated molecular patterns being highly recognized by the immune system of mammals, even if they come from a non-pathogenic microorganism.
Keywords: Trebouxia sp.; Trebouxiophyceae; Galactofuranose-rich polysaccharides; Pro-inflammatory activity; Sepsis;

Three new β-dihydroagarofuranoid sesquiterpenes were identified from Celastrus paniculatus, and the mode of mechanism of the major constituent (1α,2α,8β,9β)-1,8-bis(acetyloxy)-2,9-bis(benzoyloxy)-14-hydroxy-β-dihydroagarofuran in inducing autophagy and apoptosis was characterized. Celastrus paniculatus is a traditional medicinal plant with diverse pharmacological activities. To identify its bioactive constituents, three new β-dihydroagarofuranoid sesquiterpenes were isolated from the whole plant, of which the major constituent is (1α,2α,8β,9β)-1,8-bis(acetyloxy)-2,9-bis(benzoyloxy)-14-hydroxy-β-dihydroagarofuran. It was assessed for its antiproliferative activity, and it suppressed the viability of MCF-7 breast cancer cells with an IC50 of 17 ± 1 μM. This growth inhibition was, in part, attributable to apoptosis. Moreover, this drug treatment led to LC3B-II accumulation, indicative of autophagy. Western blot analysis established its ability to target a broad range of signaling effectors related to survival and cell cycle progression, including Akt, NF-κB, p53, and MAP kinases. In addition, flow cytometry analysis indicates increased reactive oxygen species production in response to this compound. Taken together, these findings suggest a pleiotropic mode of mechanism that underlies the antiproliferative activity of this compound in MCF-7 breast cancer cells.
Keywords: Celastrus paniculatus; Celastraceae; Sesquiterpenes; Autophagy; Apoptosis;

Bioactivity against Bursaphelenchus xylophilus: Nematotoxics from essential oils, essential oils fractions and decoction waters by Jorge M.S. Faria; Pedro Barbosa; Richard N. Bennett; Manuel Mota; A. Cristina Figueiredo (220-228).
Ruta graveolens, Satureja montana and Thymbra capitata essential oils showed high nematotoxic activities against Bursaphelenchus xylophilus. Activities from fractions containing hydrocarbon- or oxygen-containing molecules suggested additive and/or synergic relations.The Portuguese pine forest has become dangerously threatened by pine wilt disease (PWD), caused by the pinewood nematode (PWN), Bursaphelenchus xylophilus. Synthetic chemicals are the most common pesticides used against phytoparasitic nematodes but its use has negative ecological impacts. Phytochemicals may prove to be environmentally friendly alternatives. Essential oils (EOs) and decoction waters, isolated from 84 plant samples, were tested against B. xylophilus, in direct contact assays. Some successful EOs were fractionated and the fractions containing hydrocarbons or oxygen-containing molecules tested separately. Twenty EOs showed corrected mortalities ⩾96% at 2 μL/mL. These were further tested at lower concentrations. Ruta graveolens, Satureja montana and Thymbra capitata EOs showed lethal concentrations (LC100) < 0.4 μL/mL. Oxygen-containing molecules fractions showing corrected mortality ⩾96% did not always show LC100 values similar to the corresponding EOs, suggesting additive and/or synergistic relationships among fractions. Nine decoction waters (remaining hydrodistillation waters) revealed 100% mortality at a minimum concentration of 12.5 μL/mL. R. graveolens, S. montana and T. capitata EOs are potential environmentally friendly alternatives for B. xylophilus control given their high nematotoxic properties. Nematotoxic activity of an EO should be taken in its entirety, as its different components may contribute, in distinct ways, to the overall EO activity.
Keywords: Bursaphelenchus xylophilus; Pinus pinaster; Essential oils; Hydrocarbon molecules; Oxygen-containing molecules; Decoction waters; Nematotoxic; 2-Undecanone; Carvacrol; γ-Terpinene;

Biotransformation of oleanolic and maslinic acids by Rhizomucor miehei by Antonio Martinez; Francisco Rivas; Alberto Perojil; Andres Parra; Andres Garcia-Granados; Antonia Fernandez-Vivas (229-237).
The biotransformation of oleanolic (1) and maslinic (2) acids with Rhizomucor miehei, are described. The main action of the microorganism on both substrates was hydroxylation at C-30 (36). With oleanolic acid (1), R. miehei also hydroxylated at C-7 (4) or at C-1 (5), whereas with maslinic acid (2) the microorganism also produced 11-ene or 11α,12α-epoxy derivatives with an 28,13β-olide group, and an 11-oxoolean-12-ene derivative.Microbial transformation of oleanolic acid by Rhizomucor miehei produced three metabolites. A known compound, a 30-hydroxyl derivative (queretaroic acid), and two 7β,30- and 1β,30-dihydroxylated metabolites, respectively. The action of the same fungus (R. miehei) on maslinic acid produced an olean-11-en-28,13β-olide derivative, a metabolite hydroxylated at C-30, an 11-oxo derivative, and two metabolites with an 11α,12α-epoxy group, hydroxylated or not at C-30. Their structures were elucidated by extensive analyses of their spectroscopic data, and also by chemical correlations.
Keywords: Biotransformation; Biohydroxylation; Triterpenoids; Oleanolic acid; Maslinic acid; Rhizomucor miehei;

Phytochemical constituents of Sarracenia purpurea L. (pitcher plant) by Asim Muhammad; Pierre S. Haddad; Tony Durst; John Thor Arnason (238-242).
Isolated new compounds from the pitcher plant. The structures of these new compounds were identified by using spectroscopic technique such as, 1 and 2 D-NMR, Mass spectrometryFrom the leaves of Sarracenia purpurea, collected in Mistissini, Quebec, Canada, four goodyerosides and three phenolics and nine known compounds, were isolated. The structures of the compounds were determined by mass spectrometry, including HRMS, and by 1D and 2D NMR spectroscopy.
Keywords: Pitcher plant; Sarracenia purpurea; Sarraceniaceae; Chemical components; 1 and 2D NMR spectroscopy;

NMR reveals tannin type and content in intact heartwood and bark, and is applicable in extraction and cultivar optimization, and identification of new source species. 13C NMR is an effective method of characterizing proanthocyanidin (PAC) tannins in quebracho (Schinopsis lorentzii) heartwood and black wattle (Acacia mearnsii) bark, before and after commercial extraction. The B-rings of the constituent flavan-3-ols, catechols (quebracho) or pyrogallols (wattle), are recognized in unprocessed source materials by “marker” signals at ca. 118 or 105 ppm, respectively. NMR allows the minimum extraction efficiency to be calculated; ca. 30%, and ca. 80%, for quebracho heartwood and black wattle bark, respectively. NMR can also identify PAC tannin (predominantly robinetinidin), and compare tannin content, in bark from other acacia species; tannin content decreases in the order A. mearnsii, Acacia pycnantha (87% of A. mearnsii), Acacia dealbata and Acacia decurrens (each 74%) and Acacia karroo (30%). Heartwood from an underexploited PAC tannin source, Searsia lancea, taxonomically close to quebracho, shows abundant profisetinidin and catechin PACs. NMR offers the advantage of being applicable to source materials in their native state, and has potential applications in optimizing extraction processes, identification of tannin sources, and characterization of tannin content in cultivar yield improvement programmes.
Keywords: Schinopsis lorentzii; Acacia mearnsii; Acacia pycnantha; Acacia dealbata; Acacia decurrens; Acacia karoo; Searsia lancea; Condensed tannin; Extraction; In situ identification; Solid state NMR;

Casbane diterpenoids from the roots of Euphorbia pekinensis by Wei-Wei Tao; Jin-Ao Duan; Yu-Ping Tang; Nian-Yun Yang; Jian-Ping Li; Ye-Fei Qian (249-253).
Four casbane diterpenoids, together with three known related compounds were isolated from the roots of Euphorbia pekinensis. Isolated compounds were evaluated for their cytotoxic activity against seven human cancer cell lines by a WST-8 assay.Four casbane diterpenoids, together with three known related compounds were isolated from the roots of Euphorbia pekinensis. Their structures were elucidated on the basis of spectroscopic studies and comparison with the known related compounds. In addition, the absolute configuration of three compounds was determined by the modified Mosher’s method. The isolated compounds were evaluated for their cytotoxic activity against seven human cancer cell lines by a WST-8 assay.
Keywords: Euphorbia pekinensis; Euphorbiaceae; Casbane diterpenoid; Cytotoxic activity; Cancer cell lines;

Rhamnogalactofuranan from the microalga Myrmecia biatorellae, symbiotic partner of Lobaria linita by Lucimara M.C. Cordeiro; Flávio Beilke; Vanessa de Fátima Reinhardt; Guilherme L. Sassaki; Marcello Iacomini (254-259).
This rhamnogalactofuranan had an (1 → 3)-linked β-d-galactofuranosyl main-chain, substituted at O-6 by single units of β-d-Galf, or by side-chains of 2-O- and 2,4-di-O-linked α-l-Rhap units.A structural study of the cell wall polysaccharides of Myrmecia biatorellae, the symbiotic algal partner of the lichenized fungus Lobaria linita was carried out. It produced a cold-water insoluble rhamnogalactofuranan, with a (1 → 3)-linked β-d-galactofuranosyl main-chain, substituted at O-6 by single units of β-d-Galf, or by side-chains of 2-O- and 2,4-di-O-linked α-l-Rhap units. The structure of the polysaccharide was established by chemical and NMR spectroscopic analysis.
Keywords: Myrmecia biatorellae; Symbiotic microalgae; Lichen; Lobaria linita; Polysaccharide; Rhamnogalactofuranan;

Agarofuran sesquiterpenes from Schaefferia argentinensis by Manuela E. García; Rubén D. Motrich; Beatriz L. Caputto; Marianela Sánchez; Jorge A. Palermo; Ana Estévez-Braun; Angel G. Ravelo; Viviana E. Nicotra (260-267).
Sixteen dihydro-β-agarofuran sesquiterpenes were isolated from the aerial parts of Schaefferia argentinensis. The in vitro antiproliferative activity of the major sesquiterpenes was examined against T47D, MCF7, and MDA human cancer cell lines. The activity observed for the tested compounds is marginal.Sixteen dihydro-β-agarofuran sesquiterpenes were isolated from the aerial parts of Schaefferia argentinensis Speg. Their structures were determined by a combination of 1D and 2D NMR and MS techniques. The in vitro antiproliferative activity of the major sesquiterpenes was examined in T47D, MCF7, and MDA-MB231 human cancer cell lines, but was found to be marginal.
Keywords: Schaefferia argentinensis; Celastraceae; Phytochemistry; Antiproliferative activity; Dihydro-β-agarofuran sesquiterpene;

Phenanthrenes, 9,10-dihydrophenanthrenes, bibenzyls with their derivatives, and malate or tartrate benzyl ester glucosides from tubers of Cremastra appendiculata by Yang Wang; Shu-Hong Guan; Yu-Hui Meng; Yi-Bei Zhang; Chun-Ru Cheng; Yang-Yang Shi; Rui-Hong Feng; Feng Zeng; Zhi-Yuan Wu; Jing-Xian Zhang; Min Yang; Xuan Liu; Qing Li; Xiao-Hui Chen; Kai-Shun Bi; De-An Guo (268-276).
Seven 9,10-dihydrophenanthrene derivatives and four malate benzyl ester glucosides, along with 23 known compounds were isolated from tubers of Cremastra appendiculata. Among the compounds obtained, one compound showed moderate while five showed weak cytotoxic activity against the A549 cell line.Eleven previously unknown compounds and 23 known compounds, including 20 phenanthrene or 9,10-dihydrophenanthrene derivatives, five bibenzyls, seven malate or tartrate benzyl ester glucosides, adenosine and gastrodin were isolated from tubers of Cremastra appendiculata. Among the obtained compounds, two are the first isolated dimers with one phenanthrene or bibenzyl unit connected to C-3 of 2,3,4,5-tetrahydro-phenanthro[2,1-b]furan moiety. In addition, 33 of these compounds were evaluated in vitro for their cytotoxic activity against two cancer cell lines. Among the compounds examined, one compound showed moderate cytotoxic activity, while five showed weak cytotoxic activity against the A549 cell line.
Keywords: Cremastra appendiculata; Orchidaceae; Bibenzyl; Phenanthrene; 9,10-Dihydrophenanthrene; 2,3,4,5-Tetrahydro-phenanthro[2,1-b]furan; Malate benzyl ester glucoside; Tartrate benzyl ester glucoside;

Plant-derived juvenile hormone III analogues and other sesquiterpenes from the stem bark of Cananga latifolia by Heejung Yang; Hye Seong Kim; Eun Ju Jeong; Piseth Khiev; Young-Won Chin; Sang Hyun Sung (277-283).
From the stem bark of C. latifolia, six sesquiterpenes (two linear and four cyclic) as well as a juvenile hormone III (JH III), was isolated, as well as two linear and four cyclic sesquiterpenes.Juvenile hormone III (JH III) is a larval metamorphosis-regulating hormone present in most insect species. JH III was first isolated from the plant, Cyperus iria L., but the presence of JH III has not been reported in other plant species. In the present study, proof of the existence of JH III and its analogues from Cananga latifolia was established. From an aqueous MeOH extract of C. latifolia stem bark, six compounds were isolated along with nine known compounds. These were identified by using spectroscopic analyses as: (2E,6E,10R)-11-butoxy-10-hydroxy-3,7,11-trimethyldodeca-2,6-dienoic acid methyl ester, (2E,6E)-3,7,11-trimethyl-10-oxododeca-2,6-dienoic acid methyl ester, (2E)-3-methyl-5-[(1S,2R,6R)-1,2,6-trimethyl-3-oxocyclohexyl]-pent-2-enoic acid methyl ester, 1β-hydroxy-3-oxo-4β, 5α,7α-H-eudesmane 11-O-α-l-rhamnopyranoside, 4-epi-aubergenone 11-O-2′,3′-di-O-acetyl-α-l-rhamnopyranoside and 4-epi-aubergenone 11-O-2′,4′-di-O-acetyl-α-l-rhamnopyranoside. Three of the previously known compounds, (2E,6E,10R)-10-hydroxy-3,7,11-trimethyldodeca-2,6,11-trienoaic acid methyl ester, (2E,6E,10R)-10,11-dihydroxy-3,7,11-trimethyldodeca-2,6-dienoic acid and (2E,6S)-3-methyl-6-hydroxy-6-[(2R,5R)-5-(2-hydroxypropan-2-yl)-2-methyltetrahydrofuran-2-yl]-hex-2-enoaic acid methyl ester have now been found in a plant species. Ultra performance liquid chromatography–quadruple time-of-flight mass spectroscopy (UPLC-QTOF/MS) analysis of the chemical constituents of C. latifolia showed that several were predominant in the sub-fractions of a C. latifolia stem bark extract.
Keywords: Cananga latifolia; Annonaceae; Juvenile hormone III; Sesquiterpene;

Staghorn sumac fruits were found to contain a rich assortment of anthocyanins, including pyranoanthocyanins displaying distinctive UV–Vis absorption spectra. Structures were characterized by UPLC–ESI-MS and NMR spectroscopy.The six major anthocyanins found in the burgundy coloured fruits of Staghorn sumac (Rhus typhina L.) were isolated and the structures of four compounds were determined by NMR spectroscopic methods as being: 7-O-methyl-delphinidin-3-O-(2″galloyl)-β-d-galactopyranoside; 7-O-methyl-cyanidin-3-O-(2″galloyl)-β-d-galactopyranoside; 7-O-methyl-delphinidin-3-O-(2″′galloyl)-β-d-galactopyranoside-4-vinyl-catechol-3″-O-β-d-glucopyranoside; and 7-O-methyl-cyanidin-3-O-(2″′galloyl)-β-d-galactopyranoside-4-vinyl-catechol-3″-O-β-d-glucopyranoside, respectively. Additionally, two related anthocyanin compounds, cyanidin-3-O-(2″galloyl)-β-d-galactopyranoside and 7-O-methyl-cyanidin-3-O-β-d-galactopyranoside were also recovered, with NMR spectroscopic values closely matching previous reports from other plant species. The prevalence of 7-O-methyl anthocyanins and their galloylated derivatives in sumac is highly unusual, and warrants special attention. Additionally, the in planta occurrence of two 7-O-methyl-pyranoanothocyanin-vinyl-catechol aglycones, Sumadin A and Sumadin B, and their derivatives is noted. To our knowledge, E-ring glycosylated vinyl-catechol pyranoanthocyanins were previously unknown.
Keywords: Staghorn sumac; Rhus typhina; Anacardiaceae; Vinyl-catechol; Pyranoanthocyanin; Anthocyanin; Sumadin;