Electrical penetration graphs indicate that tricin is a key secondary metabolite of rice,inhibiting phloem feeding of brown planthopper,Nilaparvata lugens

Tricin (5,7,4′‐trihydroxy‐3′,5′‐dimethoxyflavone) is a valuable secondary metabolite which is widely present in gramineous plants, including cultivated rice (Oryza sativa L.) (Poaceae). It can defend the rice plant against damage by the brown planthopper (BPH), Nilaparvata lugens (Stål) (Hemiptera: Delphacidae), one of the most important pests of rice. This study was conducted to elucidate the mechanisms of action of tricin on BPH feeding behavior. BPH feeding behavior in resistant (Rathu Heenati, RHT) and susceptible (Taichuang native 1, TN1) rice varieties and artificial diets was monitored using the electrical penetration graph (EPG) technique. Tricin concentrations in leaves of varieties RHT and TN1 were quantitatively analyzed by liquid chromatography, coupled to tandem mass spectrometric techniques. Six (NP and N1‐5) and four (NP, N1, N2, and N4) types of waveforms occurred during feeding on rice plants and artificial diets, respectively. The tricin concentration of rice varieties was correlated with total and average durations of N4. Moreover, EPG data indicated that tricin significantly increased the duration of non‐probing and pathway periods and strongly inhibited phloem ingestion (N4). The inhibition was strongly dose dependent, resulting in complete suppression of activity in the phloem region when the tricin concentration was increased to 1 g l^?1. This study revealed that tricin disturbed the feeding behavior of BPH mainly by increasing the non‐probe period and inhibiting phloem ingestion. We confirmed the hypothesis that tricin is a ‘stylet probing stimulant’ of rice planthoppers as proposed in previous studies. The information on the ecological effect of tricin from this study may be useful to clarify the resistance mechanism against BPH of RHT and other tricin‐containing rice varieties.     

Structure-function relationships of wheat flavone <Emphasis Type="Italic">O</Emphasis>-methyltransferase: Homology modeling and site-directed mutagenesis

Background  

Wheat (Triticum aestivum L.) O-methyltransferase (TaOMT2) catalyzes the sequential methylation of the flavone, tricetin, to its 3'-methyl- (selgin), 3',5'-dimethyl- (tricin) and 3',4',5'-trimethyl ether derivatives. Tricin, a potential multifunctional nutraceutical, is the major enzyme reaction product. These successive methylations raised the question as to whether they take place in one, or different active sites. We constructed a 3-D model of this protein using the crystal structure of the highly homologous Medicago sativa caffeic acid/5-hydroxyferulic acid O-methyltransferase (MsCOMT) as a template with the aim of proposing a mechanism for multiple methyl transfer reactions in wheat.     

Plastid genomes and deep relationships among the commelinid monocot angiosperms

The commelinid monocots comprise the orders Arecales (A), Commelinales (C), Poales sensu Angiosperm Phylogeny Group III (APGIII) (P), Zingiberales (Z), plus the unplaced family Dasypogonaceae (D), collectively containing numerous economically and ecologically important species and encompassing enormous morphological diversity. Commelinids are supported as monophyletic based on anatomy and molecular data; however, relationships among major commelinid groupings conflict among previous studies, representing a long‐standing problem in monocot systematics, with major implications for interpretations of character evolution. In more recent analyses, with whole‐plastome sampling largely focused on Poales, areas of conflict remain, suggesting the need for closer investigation of relationships and support. Here, we increased sampling of plastomes among non‐Poalean commelinid orders to investigate deep nodal support. Analysis of 83 plastid genes recovered relationships as ((A, D) (ZC, P)) with robust support, regardless of reconstruction method (parsimony/likelihood). However, conflict among genes was evident when grouped by genomic region. Cumulative analyses of genes ranked by decreasing numbers of informative characters indicated continued fluctuation in support, even as small genes were added to a nearly complete matrix, contrary to the expected pattern of stabilization in support. Topology tests among major commelinid groups suggested that the data were not powerful enough to reject all alternatives. This study provides clues to the limits of the plastid genome for resolving deep relationships among the commelinid monocots.     

A revised evolutionary history of Poales: origins and diversification

Poales represents more than one‐third of all monocotyledons (c. 20 000 species in 16 families) and constitutes a microcosm of the angiosperms. The extreme variation in species richness among the families of Poales is still not understood: Poaceae includes ～10 000 species, whereas six families have fewer than ten species. Here, using the largest phylogenetic analysis of Poales to date, molecular dating, ancestral reconstructions and diversification analyses, we develop a macro‐evolutionary and macro‐ecological approach to seek correlates for changing diversification patterns. We show that the poalean families diverged in the Late Cretaceous, a time of high levels of CO₂ and high rainfall. Our habitat reconstructions indicate that Poales inhabited open and dry habitats in this environment. We also demonstrate that lineages with CO₂‐concentrating mechanisms inhabiting dry and open environments exhibited higher diversification rates than C₃, shade and wet lineages. CO₂‐concentrating mechanisms counteract the effects of low atmospheric CO₂ and reduce phototranspiration. It is believed that the parallel evolution of C₄ and CAM (Crassulacean acid metabolism) photosynthesis in Poaceae, Cyperaceae and Bromeliaceae is an adaptation to changes in atmospheric CO₂ concentrations. Combinations of extrinsic and intrinsic factors might have played a role in shifts in diversification rates and may explain the variation in species richness in Poales. © 2014 The Linnean Society of London, Botanical Journal of the Linnean Society, 2014, 175 , 4–16.     

Antileishmanial Activity and Immunomodulatory Effects of Tricin Isolated from Leaves of Casearia arborea (Salicaceae)

Bioactivity‐guided fractionation of antileishmanial active extract from leaves of Casearia arborea led to isolation of three metabolites: tricin ( 1 ), 1′,6′‐di‐O‐β‐d ‐vanilloyl glucopyranoside ( 2 ) and vanillic acid ( 3 ). Compound 1 demonstrated the highest activity against the intracellular amastigotes of Leishmania infantum, with an IC₅₀ value of 56 μm . Tricin ( 1 ) demonstrated selectivity in mammalian cells (SI > 7) and elicited immunomodulatory effect on host cells. The present work suggests that tricin modulated the respiratory burst of macrophages to a leishmanicidal state, contributing to the parasite elimination. Therefore, the natural compound tricin could be further explored in drug design studies for leishmaniasis treatment.     

Implications of the Plastid Genome Sequence of Typha (Typhaceae, Poales) for Understanding Genome Evolution in Poaceae

Plastid genomes of the grasses (Poaceae) are unusual in their organization and rates of sequence evolution. There has been a recent surge in the availability of grass plastid genome sequences, but a comprehensive comparative analysis of genome evolution has not been performed that includes any related families in the Poales. We report on the plastid genome of Typha latifolia, the first non-grass Poales sequenced to date, and we present comparisons of genome organization and sequence evolution within Poales. Our results confirm that grass plastid genomes exhibit acceleration in both genomic rearrangements and nucleotide substitutions. Poaceae have multiple structural rearrangements, including three inversions, three genes losses (accD, ycf1, ycf2), intron losses in two genes (clpP, rpoC1), and expansion of the inverted repeat (IR) into both large and small single-copy regions. These rearrangements are restricted to the Poaceae, and IR expansion into the small single-copy region correlates with the phylogeny of the family. Comparisons of 73 protein-coding genes for 47 angiosperms including nine Poaceae genera confirm that the branch leading to Poaceae has significantly accelerated rates of change relative to other monocots and angiosperms. Furthermore, rates of sequence evolution within grasses are lower, indicating a deceleration during diversification of the family. Overall there is a strong correlation between accelerated rates of genomic rearrangements and nucleotide substitutions in Poaceae, a phenomenon that has been noted recently throughout angiosperms. The cause of the correlation is unknown, but faulty DNA repair has been suggested in other systems including bacterial and animal mitochondrial genomes.     

Tricin,a Flavonoid Monomer in Monocot Lignification

Tricin was recently discovered in lignin preparations from wheat (Triticum aestivum) straw and subsequently in all monocot samples examined. To provide proof that tricin is involved in lignification and establish the mechanism by which it incorporates into the lignin polymer, the 4′-O-β-coupling products of tricin with the monolignols (p-coumaryl, coniferyl, and sinapyl alcohols) were synthesized along with the trimer that would result from its 4′-O-β-coupling with sinapyl alcohol and then coniferyl alcohol. Tricin was also found to cross couple with monolignols to form tricin-(4′-O-β)-linked dimers in biomimetic oxidations using peroxidase/hydrogen peroxide or silver (I) oxide. Nuclear magnetic resonance characterization of gel permeation chromatography-fractionated acetylated maize (Zea mays) lignin revealed that the tricin moieties are found in even the highest molecular weight fractions, ether linked to lignin units, demonstrating that tricin is indeed incorporated into the lignin polymer. These findings suggest that tricin is fully compatible with lignification reactions, is an authentic lignin monomer, and, because it can only start a lignin chain, functions as a nucleation site for lignification in monocots. This initiation role helps resolve a long-standing dilemma that monocot lignin chains do not appear to be initiated by monolignol homodehydrodimerization as they are in dicots that have similar syringyl-guaiacyl compositions. The term flavonolignin is recommended for the racemic oligomers and polymers of monolignols that start from tricin (or incorporate other flavonoids) in the cell wall, in analogy with the existing term flavonolignan that is used for the low-molecular mass compounds composed of flavonoid and lignan moieties.Lignin, a complex phenylpropanoid polymer in the plant cell wall, is predominantly deposited in the cell walls of secondary-thickened cells (Vanholme et al., 2010). It is synthesized via oxidative radical coupling reactions from three prototypical monolignols, p-coumaryl, coniferyl, and sinapyl alcohols, differentiated by their degree of methoxylation ortho to the phenolic hydroxyl group. Considered within the context of the entire polymer, the main structural features of lignin can be defined in terms of its p-hydroxyphenyl (H), guaiacyl (G), and syringyl (S) units, derived respectively from these three monolignols (Ralph, 2010). Several novel monomers, all deriving from the monolignol biosynthetic pathway, have been found to incorporate into lignin in wild-type and transgenic plants. For example, monolignol acetate, p-hydroxybenzoate, and p-coumarate ester conjugates have all been shown to incorporate into lignin polymers and are the source of naturally acylated lignins (Ralph et al., 2004; Lu and Ralph, 2008); lignins derived solely from caffeyl alcohol were found in the seed coats of both monocot and dicot plants (Chen et al., 2012a, 2012b); lignins derived solely from 5-hydroxyconiferyl alcohol were found in a cactus (for example, in a member of the genera Astrophytum) seed coat (Chen et al., 2012a); a Medicago truncatula transgenic deficient in cinnamyl alcohol dehydrogenase exhibited a lignin that was overwhelmingly derived from hydroxycinnamaldehydes (instead of their usual hydroxycinnamyl alcohol analogs; Zhao et al., 2013); and iso-sinapyl alcohol was implicated as a monomer in caffeic acid O-methyltransferase down-regulated switchgrass (Panicum virgatum; Tschaplinski et al., 2012). These findings imply that plants are quite flexible in being able to use a variety of monomers during lignification to form the heterogenous lignin polymer. Most recently, and as addressed more fully here, the flavonoid tricin has been implicated as a monomer in monocot lignins (del Río et al., 2012). To our knowledge, tricin is the first monomer from outside the monolignol biosynthetic pathway to be implicated in lignification.Tricin [5,7-dihydroxy-2-(4-hydroxy-3,5-dimethoxyphenyl)-4H-chromen-4-one], a member of the flavonoid family, is recognized as a valuable human health compound due to its antioxidant, antiaging, anticancer, and cardioprotective potentials (Ogo et al., 2013). Tricin and its derivatives can be solvent extracted from monocot samples such as wheat (Triticum aestivum), oat bran (Avena sativa), bamboo (Leleba oldhami), sugarcane (Saccharum officinarum), and maize (Zea mays). Extracted compounds can take the form of tricin itself, 7-O-glycosylated tricin, or the flavonolignan in which tricin is 4′-O-etherified by putative coupling with coniferyl alcohol (Ju et al., 1998; Bouaziz et al., 2002; Wenzig et al., 2005; Duarte-Almeida et al., 2007; Van Hoyweghen et al., 2010; Nakano et al., 2011; Bottcher et al., 2013; Moheb et al., 2013).In 2012, we reported, to our knowledge, the first evidence that tricin was incorporated into lignin, as implicated by two previously unassigned correlation peaks at δ_C/δ_H 94.1/6.56 and 98.8/6.20 in a heteronuclear single-quantum coherence (HSQC) NMR spectrum from the whole cell wall and an isolated milled wood lignin of (unacetylated) wheat straw (del Río et al., 2012). The same evidence has now been found in the HSQC spectrum of wheat straw lignin isolated via different methods (Yelle et al., 2013; Zeng et al., 2013). Additional studies have verified the presence of tricin in lignin fractions from a variety of monocots, including bamboo (You et al., 2013), coconut coir (Cocos nucifera; Rencoret et al., 2013), maize, and others examined in our laboratories. The implication that tricin is the first phenolic from outside the monolignol biosynthetic pathway found to be integrated into the polymer has prompted further study with the aim of identifying and mechanistically delineating the role of tricin in lignin and its biosynthetic incorporation pathway.Tricin, unlike the monolignols that derive from the shikimate biosynthetic pathway (Sarkanen and Ludwig, 1971), is derived from a combination of the shikimate and acetate/malonate-derived polyketide pathways (Winkel-Shirley, 2001), as shown in Supplemental Figure S1. After p-coumaroyl-CoA is synthesized from p-coumaric acid by 4-coumarate:CoA ligase, it branches from the monolignol biosynthetic route to be transformed via chalcone synthase and chalcone isomerase into naringenin, the central precursor of most flavonoids. Naringenin is subsequently converted into apigenin by flavone synthase. Further hydroxylation at C-3′ and C-5′ followed by O-methylation furnishes tricin (Koes et al., 1994; Winkel-Shirley, 2001). The incorporation of tricin into lignin, therefore, suggests that an additional biosynthetic pathway, namely the polyketide pathway, may be associated with cell wall lignification in monocots.The revelation that tricin is incorporated into the lignin polymer was precipitated by closer study of signals found within the NMR spectra of various monocot samples. Before this discovery, tricin had not been noted in any lignin fractions, and although it is reasonable to anticipate compatibility based on its chemical structure, there is no direct and reliable evidence to date showing that tricin is able to react with monolignols through radical coupling; therefore, the efficiency and selectivity of the coupling reactions between tricin and various monolignols were also unknown. Synthetic model compounds that would facilitate the elucidation of the role of tricin within plant cell walls are desirable as aids to be used in a mechanistic study of flavonolignin generation. (We coin the term flavonolignin to describe the racemic oligomers and polymers of monolignols that start from tricin [or other flavonoids] in the cell wall, in analogy with the existing term flavonolignan that is used for the low-molecular mass compounds composed of flavonoid and lignan moieties that are presumably made in the cytoplasm [Begum et al., 2010; Niculaes et al., 2014; Dima et al., 2015]).The overall objective of this study is to demonstrate that tricin incorporates into the lignin polymer of monocots, with maize/corn stover as the representative experimental material. To this end, we have synthesized tricin and various model compounds in which tricin is conjugated to monolignols in the manner expected for the lignification process. Next, we verified whether these synthetic compounds could be made from their assumed precursors under the biomimetic radical conditions anticipated for lignification. Subsequently, NMR data generated from these synthetic and biomimetic coupling products were compared with NMR data from native maize stover lignin, including high-M_r fractions. We conclude that tricin is a monomer in monocot lignification and that, because little syringaresinol is found in maize lignin, tricin is functioning as a nucleation site that initiates lignin polymer chains.     

Microsporogenesis is simultaneous in the early‐divergent grass Streptochaeta,but successive in the closest grass relative,Ecdeiocolea

Simultaneous microsporogenesis is described for the first time in a grass, Streptochaeta spicata Schrad., a tropical Brazilian species that belongs in the early‐divergent subfamily Anomochlooideae. Microsporogenesis is successive in all other Poaceae examined so far, and most other members of the order Poales, to which grasses belong. The only other reports of simultaneous microsporogenesis in Poales are in Rapateaceae and some members of the cyperid clade (Juncaceae, Cyperaceae, Prionium and Thurnia). Among the graminids, Ecdeiocolea (the putative closest relative to Poaceae) is successive, as are Joinvillea, Flagellaria and all other Poaceae, indicating that the simultaneous condition is autapomorphic in Streptochaeta, though Anomochloa has yet to be examined. Anther wall development in Streptochaeta is of the reduced type, as also in another early‐divergent grass Pharus, though most other Poales, including most grasses, have the monocot type. In Streptochaeta, as in Pharus, the endothecium lacks thickenings, unlike other grasses that have a persistent endothecium with thickenings. The centrifixed anthers and nonplumose stigmas of Streptochaeta suggest entomophily.     

Tricin inhibits proliferation of human hepatic stellate cells in vitro by blocking tyrosine phosphorylation of PDGF receptor and its signaling pathways

4',5,7-Trihydroxy-3',5'-dimethoxyflavone (Tricin), a naturally occurring flavone, has anti-inflammatory potential and exhibits diverse biological activities including antigrowth activity in several human cancer cell lines and cancer chemopreventive effects in the gastrointestinal tract of mice. The present study aimed to investigate the biological actions of tricin on hepatic stellate cells (HSCs) in vitro, exploring its potential as a treatment of liver fibrosis, since HSC proliferation is closely related to the progression of hepatic fibrogenesis in chronic liver diseases leading to irreversible liver cirrhosis and hepatocellular carcinoma. Tricin inhibited platelet-derived growth factor (PDGF)-BB-induced cell proliferation by blocking cell cycle progression and cell migration in the human HSC line LI90 and culture-activated HSCs. It also reduced the phosphorylation of PDGF receptor β and the downstream signaling molecules ERK1/2 and Akt, which might be due to its tyrosine kinase inhibitor properties rather than inhibition of the direct binding between PDGF-BB and its receptor. Our findings suggest that tricin might be beneficial in HSC-targeting therapeutic or chemopreventive applications for hepatic fibrosis.     

Natural flavone tricin exerted anti-inflammatory activity in macrophage via NF-κB pathway and ameliorated acute colitis in mice

BackgroundUlcerative colitis is a subtype of inflammatory bowel disease, characterized by relapsing inflammation in the gastrointestinal tract with limited treatment options. Previous studies suggested that the natural compound tricin, a flavone isolated from rice bran, could suppress chemically-induced colitis in mice, while our recent study also demonstrated the anti-metastatic effect of tricin in colon tumor-bearing mice.Hypothesis/PurposeHere we further investigated the underlying mechanism of the inhibitory effects of tricin on lipopolysaccharides-activated macrophage RAW264.7 cells and explored the efficacy of tricin in acute colitis mouse model induced by 4.5% dextran sulfate sodium (DSS) for 7 days.MethodsTricin (75, 100, and 150 mg/kg) or the positive control drug sulfasalazine (200 mg/kg) were orally administered to mice for 7 days. Stool consistency scores, stool blood scores, and body weight were recorded daily. Disease activity index (DAI) was examined on day 7, and colon tissues were collected for biochemical analyses. The fecal microbiome of colitis mice after tricin treatment was characterized for the first time in this study using 16S rDNA amplicon sequencing.ResultsResults showed that tricin (50 µM) remarkably reduced nitric oxide production in lipopolysaccharides-activated RAW264.7 cells and the anti-inflammatory activity of tricin was shown to act through the NF-κB pathway. Besides, tricin treatment at 150 mg/kg significantly reversed colon length reduction, reduced myeloperoxidase activities and DAI scores, as well as restored the elevated myeloid-derived suppressive cells population in acute colitis mice. The influence from DSS on gut microbiota, such as the increased population of Proteobacteria phylum and Ruminococcaceae family, was shown to be relieved after tricin treatment.ConclusionOur present study firstly demonstrated that tricin ameliorated acute colitis by improving colonic inflammation and modulating gut microbiota profile, which supports the potential therapeutic use of tricin for colitis treatment.     

The First Complete Plastid Genome from Joinvilleaceae (J. ascendens; Poales) Shows Unique and Unpredicted Rearrangements

Joinvilleaceae is a family of tropical grass-like monocots that comprises only the genus Joinvillea. Previous studies have placed Joinvilleaceae in close phylogenetic proximity to the well-studied grass family. A full plastome sequence was determined and characterized for J. ascendens. The plastome was sequenced with next generation methods, fully assembled de novo and annotated. The assembly revealed two novel inversions specific to the Joinvilleaceae lineage and at least one novel plastid inversion in the Joinvilleaceae-Poaceae lineage. Two previously documented inversions in the Joinvilleaceae-Poaceae lineage and one previously documented inversion in the Poaceae lineage were also verified. Inversion events were identified visually and verified computationally by simulation mutations. Additionally, the loss and subsequent degradation of the accD gene in order Poales was explored extensively in Poaceae and J. ascendens. The two novel inversions along with changes in gene composition between families better delimited lineages in the Poales. The presence of large inversions and subsequent reversals in this small family suggested a high potential for large-scale rearrangements to occur in plastid genomes.     

Evaluation of tricin,a stylet probing stimulant of brown planthopper,in infested and non‐infested rice plants

The flavone, tricin (5,7,4′‐trihydroxy‐3′,5′‐dimethoxyflavone), is a valuable secondary metabolite that is common in gramineous plants, including cultivated rice (Oryza sativa). It can defend the rice plant against infestation by the brown planthopper (BPH), Nilaparvata lugens Stål, one of the most important pests of rice. This study evaluated the tricin concentration in infested and non‐infested rice plants. The results of the liquid chromatography coupled to tandem mass spectrometry (LC–MS/MS) quantitative analysis showed that the tricin concentration in rice leaves was significantly higher than in the stems and roots. The mass concentration of tricin in the leaves at the leaf stage was significantly higher than at the tiller and booting stages. The relationship between rice variety, BPH resistance and tricin concentration was investigated. There was a significant negative correlation between tricin concentration and the injury severity scores for rice varieties. Moreover, BPH infestation caused variations in tricin concentration among rice plants. High BPH infestation levels can significantly reduce the tricin concentration in rice plants. However, there is no significant effect of the length of infestation times on tricin concentrations in rice leaves. These results suggest that there may be an elicitor in BPH saliva, which is injected into rice plants during BPH infestation and triggers the tricin metabolic system. Future studies need to identify the elicitor and clarify the mechanism underlying tricin reduction in infested rice plants.     

Biodegradation of kraft lignin by a bacterial strain Comamonas sp. B-9 isolated from eroded bamboo slips

Aims: The aim was to obtain evidences for lignin degradation by unicellular bacterium Comamonas sp. B‐9. Methods and Results: Comamonas sp. B‐9 was inoculated into kraft lignin‐mineral salt medium (KL‐MSM) at pH 7·0 and 30°C for 7 days of incubation. The bacterial growth, chemical oxygen demand (COD) reduction, secretion of ligninolytic enzymes and productions of low‐molecular‐weight compounds revealed that Comamonas sp. B‐9 was able to degrade kraft lignin (KL). COD in KL‐MSM reduced by 32% after 7 days of incubation. The maximum activities of manganese peroxidase (MnP) of 2903·2 U l^?1 and laccase (Lac) of 1250 U l^?1 were observed at 4th and 6th day, respectively. The low‐molecular‐weight compounds such as ethanediol, 3, 5‐dimethyl‐benzaldehyde and phenethyl alcohol were formed in the degradation of KL by Comamonas sp. B‐9 based on GC‐MS analysis. Conclusions: This study confirmed that Comamonas sp. B‐9 could utilize KL as a sole carbon source and degrade KL to low‐molecular‐weight compounds. Significance and Impact of the Study: Comamonas sp. B‐9 may be useful in the utilization and bioconversion of lignin and lignin‐derived aromatic compounds in biotechnological applications. Meanwhile, using Comamonas sp. B‐9 in treatment of wastewater in pulp and paper industry is a meaningful work.     

Biogeography of the grasses (Poaceae): a phylogenetic approach to reveal evolutionary history in geographical space and geological time

The grasses (Poaceae) are the fifth most diverse family of angiosperms, including 800 genera and more than 10 000 species. Few phylogenetic studies have tried to investigate palaeo‐biogeographical and palaeo‐ecological scenarios that may have led to present‐day distribution and diversity of grasses at the family level. We produced a dated phylogenetic tree based on combined plastid DNA sequences and a comprehensive sample of Poaceae. Furthermore, we produced an additional tree using a supermatrix of morphological and molecular data that included all 800 grass genera so that ancestral biogeography and ecological habitats could be inferred. We used a likelihood‐based method, which allows the estimation of ancestral polymorphism in both biogeographical and ecological analyses for large data sets. The origin of Poaceae was retrieved as African and shade adapted. The crown node of the BEP + PACCMAD clade was dated at 57 Mya, in the early Eocene. Grasses dispersed to all continents by approximately 60 million years after their Gondwanan origin in the late Cretaceous. PACCMAD taxa adapted to open habitats as early as the late Eocene, a date consistent with recent phytolith fossil data for North America. C₄ photosynthesis first originated in Africa, at least for Chloridoideae in the Eocene at c. 30 Mya. The BEP clade members adapted to open habitats later than PACCMAD members; this was inferred to occur in Eurasia in the Oligocene. © 2010 The Linnean Society of London, Botanical Journal of the Linnean Society, 2010, 162 , 543–557.     

Flavonoids in leaves and inflorescences of australian Cyperus species

A survey of the flavonoids of some 92 species of Australian Cyperus, mainly of subtropical or tropical origin, has confirmed a correlation previously reported in this family between flavonoid pattern and plant geography. The pattern found was similar to that of African and South American Cyperaceae, particularly in the occurrence of the rare marker substance, luteolin 5-methyl ether. Tricin and luteolin are relatively common, in glycosidic form, in the leaves while the flavonol quercetin is infrequent. When present, quercetin occurs either in glycosidic form or free as a methyl ether. The 3-monomethyl and 3, 7-dimethyl ethers of kaempferol and quercetin and the 3, 7, ?-trimethyl ether of quercetin are reported for the first time from the Cyperaceae. The flavonoid pattern of inflorescences is distinct from that of the leaves in that tricin is not detectable and that luteolin 5-methyl ether appears to be replaced by 7, 3′, 4′-trihydroxyflavone. In addition, the aurone aureusidin is more commonly present than in the leaves and is occasionally accompanied by two further aurones. The glycoxanthones mangiferin and isomangiferin occur rarely in all three species examined in the section Haspani, i.e. in C. haspan, C. prolifer and C. tenuispica. In general, however, the flavonoid data do not offer any markers which separate off different sections within the genus; there are, however, some significant correlations between the frequency of the flavonoid classes and subgeneric groupings.     

Introduction of chemically labile substructures into Arabidopsis lignin through the use of LigD,the Cα‐dehydrogenase from Sphingobium sp. strain SYK‐6

Bacteria‐derived enzymes that can modify specific lignin substructures are potential targets to engineer plants for better biomass processability. The Gram‐negative bacterium Sphingobium sp. SYK‐6 possesses a Cα‐dehydrogenase (LigD) enzyme that has been shown to oxidize the α‐hydroxy functionalities in β–O–4‐linked dimers into α‐keto analogues that are more chemically labile. Here, we show that recombinant LigD can oxidize an even wider range of β–O–4‐linked dimers and oligomers, including the genuine dilignols, guaiacylglycerol‐β‐coniferyl alcohol ether and syringylglycerol‐β‐sinapyl alcohol ether. We explored the possibility of using LigD for biosynthetically engineering lignin by expressing the codon‐optimized ligD gene in Arabidopsis thaliana. The ligD cDNA, with or without a signal peptide for apoplast targeting, has been successfully expressed, and LigD activity could be detected in the extracts of the transgenic plants. UPLC‐MS/MS‐based metabolite profiling indicated that levels of oxidized guaiacyl (G) β–O–4‐coupled dilignols and analogues were significantly elevated in the LigD transgenic plants regardless of the signal peptide attachment to LigD. In parallel, 2D NMR analysis revealed a 2.1‐ to 2.8‐fold increased level of G‐type α‐keto‐β–O–4 linkages in cellulolytic enzyme lignins isolated from the stem cell walls of the LigD transgenic plants, indicating that the transformation was capable of altering lignin structure in the desired manner.     

Flavonoids naringenin chalcone,naringenin, dihydrotricin,and tricin are lignin monomers in papyrus

Recent studies demonstrate that several polyphenolic compounds produced from beyond the canonical monolignol biosynthetic pathways can behave as lignin monomers, participating in radical coupling reactions and being incorporated into lignin polymers. Here, we show various classes of flavonoids, the chalconoid naringenin chalcone, the flavanones naringenin and dihydrotricin, and the flavone tricin, incorporated into the lignin polymer of papyrus (Cyperus papyrus L.) rind. These flavonoids were released from the rind lignin by Derivatization Followed by Reductive Cleavage (DFRC), a chemical degradative method that cleaves the β-ether linkages, indicating that at least a fraction of each was integrated into the lignin as β-ether-linked structures. Due to the particular structure of tricin and dihydrotricin, whose C-3ʹ and C-5ʹ positions at their B-rings are occupied by methoxy groups, these compounds can only be incorporated into the lignin through 4ʹ–O–β bonds. However, naringenin chalcone and naringenin have no substituents at these positions and can therefore form additional carbon–carbon linkages, including 3ʹ– or 5ʹ–β linkages that form phenylcoumaran structures not susceptible to cleavage by DFRC. Furthermore, Nuclear Magnetic Resonance analysis indicated that naringenin chalcone can also form additional linkages through its conjugated double bond. The discovery expands the range of flavonoids incorporated into natural lignins, further broadens the traditional definition of lignin, and enhances the premise that any phenolic compound present at the cell wall during lignification could be oxidized and potentially integrated into the lignin structure, depending only on its chemical compatibility. This study indicates that papyrus lignin has a unique structure, as it is the only lignin known to date that integrates such a diversity of phenolic compounds from different classes of flavonoids. This discovery will open up new ways to engineer and design lignins with specific properties and for enhanced value.

A series of flavonoids incorporate into the rind lignin of papyrus, participating as monomers during lignification.     

Downregulation of p‐COUMAROYL ESTER 3‐HYDROXYLASE in rice leads to altered cell wall structures and improves biomass saccharification

p‐Coumaroyl ester 3‐hydroxylase (C3′H) is a key enzyme involved in the biosynthesis of lignin, a phenylpropanoid polymer that is the major constituent of secondary cell walls in vascular plants. Although the crucial role of C3′H in lignification and its manipulation to upgrade lignocellulose have been investigated in eudicots, limited information is available in monocotyledonous grass species, despite their potential as biomass feedstocks. Here we address the pronounced impacts of C3′H deficiency on the structure and properties of grass cell walls. C3′H‐knockdown lines generated via RNA interference (RNAi)‐mediated gene silencing, with about 0.5% of the residual expression levels, reached maturity and set seeds. In contrast, C3′H‐knockout rice mutants generated via CRISPR/Cas9‐mediated mutagenesis were severely dwarfed and sterile. Cell wall analysis of the mature C3′H‐knockdown RNAi lines revealed that their lignins were largely enriched in p‐hydroxyphenyl (H) units while being substantially reduced in the normally dominant guaiacyl (G) and syringyl (S) units. Interestingly, however, the enrichment of H units was limited to within the non‐acylated lignin units, with grass‐specific γ‐p‐coumaroylated lignin units remaining apparently unchanged. Suppression of C3′H also resulted in relative augmentation in tricin residues in lignin as well as a substantial reduction in wall cross‐linking ferulates. Collectively, our data demonstrate that C3′H expression is an important determinant not only of lignin content and composition but also of the degree of cell wall cross‐linking. We also demonstrated that C3′H‐suppressed rice displays enhanced biomass saccharification.     

Flavonoids and chemotaxonomy of the Commelinaceae

Diverse schemes have been proposed for the classification of the Commelinaceae on the basis of different characters. The leaf flavonoids of 152 species have been analysed. The flavone C-glycosides are the dominant compounds, found in 78% of the species examined. Flavonol O-glycosides are present in 28% of the species, quercetin being the most frequent aglycone. 6-Hydroxyluteolin was found mainly in Tradescantia but also in other species (10%). Tricin has been detected sporadically in less than 5% of the species. In spite of these variations the pattern of the family is very uniform, because the presence of C-glycoflavones and the distribution of the other compounds supports the scheme proposed by Rohweder who divided the family into Commelinieae and Tradescantieae on the basis of inflorescence characters.