Chemotaxonomic markers in Digitalideae (Plantaginaceae)

In a chemosystematic investigation of Digitalideae (Plantaginaceae), the water-soluble part of extracts of two species of Digitalis, two species of Isoplexis, as well as Erinus alpinus and Lafuentea rotundifolia were studied with regard to their content of main carbohydrates, iridoids and caffeoyl phenylethanoid glycosides (CPGs). Digitalis and Isoplexis contained sorbitol, cornoside and a number of other phenylethanoid glycosides including the new tyrosol beta-D-mannopyranoside, sceptroside but were found to lack iridoid glucosides. Erinus contained mainly glucose, the new 8,9-double bond iridoid, erinoside, and a number of known iridoid glucosides including two esters of 6-rhamnopyranosylcatalpol, as well as the CPG poliumoside. Finally, Lafuentea was characterized by the presence of glucose, aucubin and cryptamygin B but apparently lacked CPGs. The chemosystematic significance of the isolated compounds is discussed.     

Phylogeny and classification of Oleaceae based on rps16 and trnL-F sequence data

Phylogenetic relationships among 76 species of Oleaceae, representing all 25 recognized genera of the family, were assessed by a cladistic analysis of DNA sequences from two noncoding chloroplast loci, the rps16 intron and the trnL-F region. Consensus trees from separate and combined analyses are congruent and agree well with nonmolecular data (chromosome numbers, fruit and wood anatomy, leaf glycosides, and iridoids). The two debated genera Dimetra and Nyctanthes, previously suggested to belong to Verbenaceae (sensu lato) or Nyctanthaceae, are shown to belong to Oleaceae, sister to the hitherto genus incertae sedis Myxopyrum. This clade is also supported by anatomical and chemical data. The subfamily Jasminoideae is paraphyletic, and a new classification is presented. The subfamily level is abandoned, and the former Jasminoideae is split into four tribes: Myxopyreae (Myxopyrum, Nyctanthes, and Dimetra), Fontanesieae (Fontanesia), Forsythieae (Abeliophyllum and Forsythia), and Jasmineae (Jasminum and Menodora). The tribe Oleeae (previous subfamily Oleoideae) is clearly monophyletic, comprising the subtribes Ligustrinae (Syringa and Ligustrum), Schreberinae status novus (Schrebera and Comoranthus), Fraxininae status novus (Fraxinus), and Oleinae (12 drupaceous genera). An rps16 sequence obtained from Hesperelaea, known only from the type specimen collected in 1875, confirmed the placement of this extinct taxon in the subtribe Oleinae.     

Chemical Traits of Hemiparasitism in Odontites luteus

The study of the monoterpene glycosides content of Odontites luteus has shown the presence of a total of fifteen iridoid glucosides. The presence of compounds 1  –  5 and 7  –  10 is perfectly on‐line with both the biogenetic pathway for iridoids biosynthesis in Lamiales and the current botanical classification of the species. On the other side, the presence of compounds like agnuside ( 6 ), adoxosidic acid ( 11 ), monotropein ( 12 ), 6,7‐dihydromonotropein ( 13 ), methyl oleoside ( 14 ) and methyl glucooleoside ( 15 ) is of high interest because, first of all, they have never been reported before in Lamiales. In second instance, the majority of the last compounds are formally derived from a different biogenetic pathway which involves deoxyloganic acid/loganin and led to the formation of decarboxylated iridoid showing the 8β‐configuration. Furthermore, a second abnormality was found during our study and this regards compounds 14 and 15 which are seco‐iriodids and thus not typical for this family. The presence of these unusual compounds, biogenetically not related to species belonging to Lamiales, is a clear evidence of the metabolites transfer from the hosts. In fact, the collection area is also populated by species belonging to Oleaceae and Ericaceae which could be the possible hosts since the biosynthesis of seco‐iridoids and or iridoids related to deoxyloganic acid/loganin pathway, with the 8β‐configuration, is well documented in these species.     

Veronica: Iridoids and cornoside as chemosystematic markers

The iridoid glucoside, ajugol, and the phenylethanoid glucoside, cornoside, have been isolated from species of Veronica (Plantaginaceae) for the first time. The presence of these compounds has been screened in 18 plant accessions belonging to 15 species of Veronica (Plantaginaceae), by isolation or NMR spectroscopy of crude extracts. In addition, the distribution of iridoids in the genus has been reviewed, using mainly the published data of isolated compounds. Using the recent expansion and reclassification of the genus based on DNA-sequence results as the model, we find that the genus is rather homogeneous with regard to the distribution of iridoid glucosides, aucubin and/or catalpol as well as 6-O-esters of catalpol being universally present in 10 of the 12 subgenera for which data exist. Only the two subgenera Pocilla and Chamaedrys deviate from this pattern. Pocilla is heterogeneous; in this subgenus, species in subsect. Agrestes contain the standard iridoid garniture, while species in subsect. Biloba do not contain the 6-O-esters of catalpol, but ajugol instead. Veronica intercedens (subsect. Subracemosae) differs from the remainder of the subgenus in only containing 5-hydroxylated iridoids (melittoside and globularifolin) and is so far the only species within the genus in which such compounds have been detected. These chemical differences are clearly reflected in the DNA-based phylogram of the subgenus. Subg. Chamaedrys appears homogeneous in lacking iridoids or only containing these in small amounts, but instead half of the investigated species contained the phenylethanoid glucoside cornoside. The distribution of this compound in angiosperms is reviewed; cornoside often substitutes iridoid glucosides in plants where these are expected to be present. The chemical results of Veronica fit in very well with the phylogenetic implications of the DNA-sequence results.     

Iridoids in Hydrangeaceae

The content of glycosides in Kirengeshoma palmata and Jamesia americana (Hydrangeaceae) have been investigated. The former contains loganin and secoiridoids, including the alkaloid demethylalangiside. The latter contains no iridoids, but the known glucosides arbutin, picein and prunasin. In order to futher investigate the chemotaxonomy of the family Hydrangeaceae, the distribution of the iridoid and secoiridoid glucosides as well as the known biosynthetic pathways to these compounds have been reviewed. However, only a few genera of the family has been investigated. Loganin, secologanin, and derivatives of these are common. The genus Deutzia is characteristic in containing more structurally simple iridoids in which C-10 has been lost during biosynthesis. Such compounds have so far only been reported from the genus Mentzelia (Loasaceae). The taxonomic relationships between Hydrangeaceae and the closely related Cornaceae and Loasaceae is discussed and found to agree well with recent DNA sequence results.     

Iridoids from the roots of Triosteum pinnatifidum

A chemical investigation of the roots of Triosteum pinnatifidum led to the isolation of 10 iridoids, elucidated as triohimas A–C, naucledal, secologanin dimethyl acetal, grandifloroside, sweroside, loganin, vogeloside and (E)-aldosecologanin. Most of the compounds were derived from loganin or secologanin with a glucose moiety at C-1 position. The results indicate a close relationship between the two genera Triosteum and Lonicera, and support the viewpoint that the iridoids derived from loganin or secologanin could be the chemotaxonomic markers of the Caprifoliaceae family.     

Chemotaxonomy of Plantago. Iridoid glucosides and caffeoyl phenylethanoid glycosides

Data for 34 species of Plantago (Plantaginaceae), including subgen. Littorella (= Littorella uniflora), have been collected with regard to their content of iridoid glucosides and caffeoyl phenylethanoid glycosides (CPGs). In the present work, 21 species were investigated for the first time and many known compounds were found together with three new iridoid glucosides. Of these, arborescoside and arborescosidic acid, both of the uncommon type with an 8,9-double bond, were present in several species, while 6-deoxymelittoside was found only in P. subulata. The known compounds deoxyloganic acid, caryoptoside and rehmannioside D were isolated from the genus for the first time. The earlier reported occurrence of sorbitol in the family was confirmed, and this compound was shown by NMR spectroscopy to be the main sugar in the three species investigated for this. The combined data show that CPGs are present in all species investigated. With regard to the iridoids, the distribution patterns showed a good correlation with the classification of Rahn. Thus, aucubin is typical for the whole genus, while bartsioside and catalpol as well as 5-substituted iridoids are each characteristic for a subgenus in the family. Finally, the close relationship between Plantago and Veronica suggested by chloroplast DNA sequence analysis. could be corroborated by the common occurrence of the rare 8,9-unsaturated iridoids in these two genera.     

Diversification of an ancient theme: hydroxynitrile glucosides

Many plants produce cyanogenic glucosides as part of their chemical defense. They are alpha-hydroxynitrile glucosides, which release toxic hydrogen cyanide (HCN) upon cleavage by endogenous plant beta-glucosidases. In addition to cyanogenic glucosides, several plant species produce beta- and gamma-hydroxynitrile glucosides. These do not release HCN upon hydrolysis by beta-glucosidases and little is known about their biosynthesis and biological significance. We have isolated three beta-hydroxynitrile glucosides, namely (2Z)-2-(beta-D-glucopyranosyloxy)but-2-enenitrile and (2R,3R)- and (2R,3S)-2-methyl-3-(beta-D-glucopyranosyloxy)butanenitrile, from leaves of Ribesuva-crispa. These compounds have not been identified previously. We show that in several species of the genera Ribes, Rhodiola and Lotus, these beta-hydroxynitrile glucosides co-occur with the L-isoleucine-derived hydroxynitrile glucosides, lotaustralin (alpha-hydroxynitrile glucoside), rhodiocyanosides A (gamma-hydroxynitrile glucoside) and D (beta-hydroxynitrile glucoside) and in some cases with sarmentosin (a hydroxylated rhodiocyanoside A). Radiolabelling experiments demonstrated that the hydroxynitrile glucosides in R. uva-crispa and Hordeum vulgare are derived from L-isoleucine and L-leucine, respectively. Metabolite profiling of the natural variation in the content of cyanogenic glucosides and beta- and gamma-hydroxynitrile glucosides in wild accessions of Lotus japonicus in combination with genetic crosses and analyses of the metabolite profile of the F2 population provided evidence that a single recessive genetic trait is most likely responsible for the presence or absence of beta- and gamma-hydroxynitrile glucosides in L. japonicus. Our findings strongly support the notion that the beta- and gamma-hydroxynitrile glucosides are produced by diversification of the cyanogenic glucoside biosynthetic pathway at the level of the nitrile intermediate.     

Production of recombinant proteins by filamentous fungi

The initial focus of recombinant protein production by filamentous fungi related to exploiting the extraordinary extracellular enzyme synthesis and secretion machinery of industrial strains, including Aspergillus, Trichoderma, Penicillium and Rhizopus species, was to produce single recombinant protein products. An early recognized disadvantage of filamentous fungi as hosts of recombinant proteins was their common ability to produce homologous proteases which could degrade the heterologous protein product and strategies to prevent proteolysis have met with some limited success. It was also recognized that the protein glycosylation patterns in filamentous fungi and in mammals were quite different, such that filamentous fungi are likely not to be the most suitable microbial hosts for production of recombinant human glycoproteins for therapeutic use. By combining the experience gained from production of single recombinant proteins with new scientific information being generated through genomics and proteomics research, biotechnologists are now poised to extend the biomanufacturing capabilities of recombinant filamentous fungi by enabling them to express genes encoding multiple proteins, including, for example, new biosynthetic pathways for production of new primary or secondary metabolites. It is recognized that filamentous fungi, most species of which have not yet been isolated, represent an enormously diverse source of novel biosynthetic pathways, and that the natural fungal host harboring a valuable biosynthesis pathway may often not be the most suitable organism for biomanufacture purposes. Hence it is expected that substantial effort will be directed to transforming other fungal hosts, non-fungal microbial hosts and indeed non microbial hosts to express some of these novel biosynthetic pathways. But future applications of recombinant expression of proteins will not be confined to biomanufacturing. Opportunities to exploit recombinant technology to unravel the causes of the deleterious impacts of fungi, for example as human, mammalian and plant pathogens, and then to bring forward solutions, is expected to represent a very important future focus of fungal recombinant protein technology.     

Iridoids from the aerial parts of Verbena littoralis (Verbenaceae)

The iridoids, 6S-hydroxy-8S-methyl-4-methylene-hexahydro-cyclopenta[c]pyran-3-one and 6S,9S-dihydroxy-8S-methyl-4-methylene-hexahydro-cyclopenta[c]pyran-3-one, were isolated from the aerial parts of Verbena littoralis. Their structures and stereochemistry were elucidated by means of NMR spectral data analysis. Both compounds showed moderate in vitro activity against gram positive and negative bacteria as well as moderate in vivo intestinal peristaltic action in mouse. The iridoids also showed moderate free radical scavenging activity against l,l-diphenyl-2-picrylhydrazyl (DPPH) as well as antioxidant activity, the latter being evidenced by redox properties measured using E1CD-HPLC.     

Tissue distribution, core biosynthesis and diversification of pyrrolizidine alkaloids of the lycopsamine type in three Boraginaceae species

Three species of the Boraginaceae were studied: greenhouse-grown plants of Heliotropium indicum and Agrobacterium rhizogenes transformed roots cultures (hairy roots) of Cynoglossum officinale and Symphytum officinale. The species-specific pyrrolizidine alkaloid (PA) profiles of the three systems were established by GC-MS. All PAs are genuinely present as N-oxides. In H. indicum the tissue-specific PA distribution revealed the presence of PAs in all tissues with the highest levels in the inflorescences which in a flowering plant may account for more than 70% of total plant alkaloid. The sites of PA biosynthesis vary among species. In H. indicum PAs are synthesized in the shoot but not roots whereas they are only made in shoots for C. officinale and in roots of S. officinale. Classical tracer studies with radioactively labelled precursor amines (e.g., putrescine, spermidine and homospermidine) and various necine bases (trachelanthamidine, supinidine, retronecine, heliotridine) and potential ester alkaloid intermediates (e.g., trachelanthamine, supinine) were performed to evaluate the biosynthetic sequences. It was relevant to perform these comparative studies since the key enzyme of the core pathway, homospermidine synthase, evolved independently in the Boraginaceae and, for instance, in the Asteraceae [Reimann, A., Nurhayati, N., Backenkohler, A., Ober, D., 2004. Repeated evolution of the pyrrolizidine alkaloid-mediated defense system in separate angiosperm lineages. Plant Cell 16, 2772-2784.]. These studies showed that the core pathway for the formation of trachelanthamidine from putrescine and spermidine via homospermidine is common to the pathway in Senecio ssp. (Asteraceae). In both pathways homospermidine is further processed by a beta-hydroxyethylhydrazine sensitive diamine oxidase. Further steps of PA biosynthesis starting with trachelanthamidine as common precursor occur in two successive stages. Firstly, the necine bases are structurally modified and either before or after this modification are converted into their O(9)-esters by esterification with one of the stereoisomers of 2,3-dihydroxy-2-isopropylbutyric acid, the unique necic acid of PAs of the lycopsamine type. Secondly, the necine O(9)-esters may be further diversified by O(7)- and/or O(3')-acylation.     

Flower color-flower scent associations in polymorphic Hesperis matronalis (Brassicaceae)

Floral scent emission rate and composition of purple and white flower color morphs of Hesperis matronalis (Brassicaceae) were determined for two populations and, for each, at two times of day using dynamic headspace collection and GC-MS. The floral volatile compounds identified for this species fell into two main categories, terpenoids and aromatics. Principal component analysis of 30 compounds demonstrated that both color morphs emitted more scent at dusk than at dawn. Color morphs varied in chemical composition of scent, but this differed between populations. The white morphs exhibited significant differences between populations, while the purple morphs did not. In the white morphs, one population contains color-scent associations that match expectations from classical pollination syndrome theory, where the flowers have aromatic scents, which are expected to maximize night-flying moth pollinator attraction; in the second population, white morphs were strongly associated with terpenoid compounds. The potential impact that pollinators, conserved biosynthetic pathways, and the genetics of small colonizing populations may have in determining population-specific associations between floral color and floral scent are discussed.     

Chemotaxonomy of Veroniceae and its allies in the Plantaginaceae

In a chemosystematic investigation of tribe Veroniceae (Plantaginaceae), representatives of Camptoloma, Sibthorpia, Veronica subg. Pentasepalae and subg. Hebe, Veronicastrum, Wulfenia, and the related Ellisiophyllum and Globularia were examined for non-flavonoid glycosides. From the 14 species studied, 28 different iridoid glucosides and 10 caffeoyl phenylethanoid glucosides (CPGs), as well as salidroside and arbutin were isolated and characterized by NMR; of these, five compounds were previously unknown. It was found that the representatives of Veroniceae, as well as Globularia, were characterized by mannitol, aucubin, catalpol and catalpol esters. Each of the three studied species of Veronica subg. Hebe contained at least one of the 6-O-catalpol esters typical for Veronica s. str. (verminoside), supporting the inclusion of Hebe in Veronica. However, their main constituents were esters of 6-O-rhamnopyranosylcatalpol; a CPG, hebeoside (2'-beta-xylopyranosyl-verbascoside) was isolated from V. (Hebe) salicifolia. The two species of Veronicastrum also contained 6-O-rhamnopyranosylcatalpol esters, including the previously unknown 2',3'- and 3',4'-dicinnamoyl derivatives and, in contrast to the earlier reports, they lacked 6-O-catalpol esters. The main iridoid constituents in the three investigated species of Wulfenia were 10-O-aucubin and 10-O-catalpol esters (isoscrophularioside or globularin) while baldaccioside (10-O-cinnamoyl asystasioside E) was isolated from W. baldaccii. Globularia vulgaris contained 10-O-catalpol esters (e.g., globularin) and, in addition, asperuloside together with its benzoyl analogue named besperuloside. The representatives of Sibthorpia and Ellisiophyllum were almost completely devoid of iridoids; this, however, together with the CPGs present implied a close relationship between the two genera. Camptoloma lyperiiflorum lacked hexitols but contained esters of 6-O-rhamnopyranosylcatalpol different from those found in Veroniceae but known from Buddleja, Scrophularia and Verbascum (Scrophulariaceae s. str.).     

Chemical constituents from Ligustrum robustum Bl

Ligustrum robustum Bl. (syn. Ligustrum purpurascens Y. C. Yang), a species growing in the southwest of China is used as traditional Chinese tea named Ku-Ding-Cha. 16 compounds (1–16) were isolated from the species L. robustum in present investigation and seven of them (1, 2, 11–14, 16) were firstly reported from this species, compound 14 was firstly reported from the genus Ligustrum and the family Oleaceae. The chemotaxonomic significance of these compounds were summarized.     

Phytoalexins and phytoanticipins from the wild crucifers Thellungiella halophila and Arabidopsis thaliana: rapalexin A, wasalexins and camalexin

Investigation of phytoalexin production using abiotic elicitation showed that the phytoalexin rapalexin A was produced by both Thellungiella halophila and Arabidopsis thaliana, but while A. thaliana produced camalexin, T. halophila produced wasalexins A and B and methoxybrassenin B. Considering that the genome of T. halophila is being sequenced currently and that the wasalexin pathway present in T. halophila is expected to involve a number of genes also present in Brassica species, our discovery should facilitate the isolation of genes involved in biosynthetic pathways of phytoalexins of the most economically important crucifer species.     

Geraniol synthases from perilla and their taxonomical significance

Geraniol synthases were isolated from five pure strains of Perilla citriodora and Perilla frutescens which vary in essential oil type, the main compounds of which were citral, elsholtziaketone, perillaketone, and perillene, respectively. This result supports the putative biosynthetic pathways of these three furylalkenes which are all produced by way of citral. Nucleotide sequences of geraniol synthases from three oil types of P. citriodora were identical, and almost the same as the sequence from P. frutescens, a species with twice the chromosome number of P. citriodora. This identity in sequence between P. citriodora and P. frutescens, together with other previous results, indicates that P. frutescens was formed as an amphidiploid of P. citriodora and an unknown wild species.     

Benzoxazinoids and iridoid glucosides from four Lamium species

A new class of compounds for the plant family Lamiaceae, benzoxazinoids, was found in Lamium galeobdolon. From the aerial parts of the species were isolated the new 2-O-beta-D-glucopyranosyl-6-hydroxy-2H-1,4-benzoxazin-3(4H)-one (6-hydroxy blepharin) together with four known benzoxazinoids, DHBOA-Glc, blepharin, DIBOA, DIBOA-Glc, as well as harpagide, 8-O-acetyl-harpagide and salidroside. Eight known iridoid glucosides, 24-epi-pterosterone and verbascoside were isolated from Lamium amplexicaule, L. purpureum and L. garganicum. The iridoids, 5-deoxylamiol and sesamoside, as well as the phytoecdysone, 24-epi-pterosterone, were found for the first time for the genus Lamium. The phytochemical data are discussed from a systematic and evolutionary point of view.     

Molecular phylogeny of Phoma and allied anamorph genera: Towards a reclassification of the Phoma complex

The present generic concept of Phoma is broadly defined, with nine sections being recognised based on morphological characters. Teleomorph states of Phoma have been described in the genera Didymella, Leptosphaeria, Pleospora and Mycosphaerella, indicating that Phoma anamorphs represent a polyphyletic group. In an attempt to delineate generic boundaries, representative strains of the various Phoma sections and allied coelomycetous genera were included for study. Sequence data of the 18S nrDNA (SSU) and the 28S nrDNA (LSU) regions of 18 Phoma strains included were compared with those of representative strains of 39 allied anamorph genera, including Ascochyta, Coniothyrium, Deuterophoma, Microsphaeropsis, Pleurophoma, Pyrenochaeta, and 11 teleomorph genera. The type species of the Phoma sections Phoma, Phyllostictoides, Sclerophomella, Macrospora and Peyronellaea grouped in a subclade in the Pleosporales with the type species of Ascochyta and Microsphaeropsis. The new family Didymellaceae is proposed to accommodate these Phoma sections and related anamorph genera. The present study demonstrated that Phoma radicina, the type species of Phoma sect. Paraphoma and Phoma heteromorphospora, the type species of Phoma sect. Heterospora can be assigned to the Phaeosphaeriaceae and Leptosphaeriaceae respectively.     

Unusual iridoid glycosides in Veronica sects. Hebe and Labiatoides

In a chemosystematic investigation of three Southern hemisphere species of Veronica, namely the Australian Veronica derwentiana Andrews and Veronica perfoliata R.Br. (formerly Derwentia species), and the New Zealand Veronica catarractae G. Forster (formerly a species of Parahebe), the water-soluble constituents were isolated and identified by spectroscopic methods. Apart from other iridoid glucosides common to the genus, three unusual substituted benzoyl esters of aucubin (derwentiosides A–C) were obtained from V. derwentiana and a chlorinated iridoid glycoside (catarractoside) from V. catarractae in addition to other iridoids common to the genus. The chemical profile of V. perfoliata is similar to that of Northern hemisphere species of Veronica because of the presence of characteristic 6-O-catalpol esters. The profile of V. derwentiana is unique, since 6-O-esters of aucubin rather than of catalpol dominate, however, the acyl groups are the same as those present in catalpol esters found in some other Veronica sections. V. catarractae also contains one of the catalpol esters characteristic of Veronica, but in addition three 6-O-rhamnopyranosyl substituted iridoid glycosides, one of which is 6-O-rhamnopyranosylcatalpol. Esters of the latter compound are previously only known from the more derived species in recent phylogenetic trees of sect. Hebe to which V. catarractae now also belongs, but as a more basal member.     

Evidence for common machinery utilized by the early and late RNA localization pathways in Xenopus oocytes

In Xenopus, an early and a late pathway exist for the selective localization of RNAs to the vegetal cortex during oogenesis. Previous work has suggested that distinct cellular mechanisms mediate localization during these pathways. Here, we provide several independent lines of evidence supporting the existence of common machinery for RNA localization during the early and late pathways. Data from RNA microinjection assays show that early and late pathway RNAs compete for common localization factors in vivo, and that the same short RNA sequence motifs are required for localization during both pathways. In addition, quantitative filter binding assays demonstrate that the late localization factor Vg RBP/Vera binds specifically to several early pathway RNA localization elements. Finally, confocal imaging shows that early pathway RNAs associate with a perinuclear microtubule network that connects to the mitochondrial cloud of stage I oocytes suggesting that motor driven transport plays a role during the early pathway as it does during the late pathway. Taken together, our data indicate that common machinery functions during the early and late pathways. Thus, RNA localization to the vegetal cortex may be a regulated process such that differential interactions with basal factors determine when distinct RNAs are localized during oogenesis.