Microstructural and histochemical analysis of aboveground organs of Centaurea cyanus used in herbal medicine.

Centaurea cyanus L. is a valuable source of many different bioactive substances. It is used in herbal medicine, but the structure of its organs used as raw material and secretory tissues has been insufficiently examined. The aim of this paper was to investigate the microstructure of C. cyanus flowers, bracts, stems and leaves with particular emphasis on secretory structures. Moreover, the main classes of secondary metabolites present in the secretion were identified and the taxonomic value of some micromorphological and anatomical features was analysed. Histochemical, micromorphological and ultrastructural analyses of aboveground organs of C. cyanus were carried out using light, fluorescence, scanning and transmission electron microscopy. The analyses revealed the presence of petal papillae and a characteristic cuticular pattern on the petals, stamens and stylar hairs. There were four types of non-glandular trichomes on the bracts, leaves and stem surfaces. The epidermal cells of the bracts contained prismatic calcium oxalate crystals. Two kinds of secretory structures, i.e. glandular trichomes and ducts, were observed in the C. cyanus organs. The glandular trichomes were located on the bract and stem surfaces, and the ducts were detected in the leaves and stems. Ultrastructural analyses of the epithelium of the ducts showed the presence of strongly osmiophilic insoluble phenolic material in vacuoles as well as moderately osmiophilic insoluble lipidic material in elaioplasts and vesicles. The results of histochemical assays showed a heterogeneous nature of the duct secretion, which contained essential oil, lipids, flavonoids, tannins and terpenes containing steroids.

7,8-Benzoflavone: a phytotoxin from root exudates of invasive Russian knapweed.

Root exudates from Acroptilon repens (Russian knapweed) were found to be phytotoxic and the phytotoxin in the exudate was identified as 7,8-benzoflavone (alpha-naphthoflavone), (1), not previously known as a natural product. In tests on growing seedlings both 1 and its isomer 5,6-benzoflavone (2) were phytotoxic. Flavone, a structural analog of 1 and a known granular leaf and stem exudate of other plant species, was also phytotoxic and more potent than 1 or 2.

Altered gene expression in three plant species in response to treatment with Nep1, a fungal protein that causes necrosis.

Nep1 is an extracellular fungal protein that causes necrosis when applied to many dicotyledonous plants, including invasive weed species. Using transmission electron microscopy, it was determined that application of Nep1 (1.0 micro g mL(-)(1), 0.1% [v/v] Silwet-L77) to Arabidopsis and two invasive weed species, spotted knapweed (Centaurea maculosa) and dandelion (Taraxacum officinale), caused a reduction in the thickness of the cuticle and a breakdown of chloroplasts 1 to 4 h after treatment. Membrane breakdown was most severe in cells closest to the surface of application. Differential display was used to isolate cDNA clones from the three species showing differential expression in response to Nep1 treatment. Differential gene expression was observed for a putative serpin (CmSER-1) and a calmodulin-like (CmCAL-1) protein from spotted knapweed, and a putative protein phosphatase 2C (ToPP2C-1) and cytochrome P-450 (ToCYP-1) protein from dandelion. In addition, differential expression was observed for genes coding for a putative protein kinase (AtPK-1), a homolog (AtWI-12) of wound-induced WI12, a homolog (AtLEA-1) of late embryogenesis abundant LEA-5, a WRKY-18 DNA-binding protein (AtWRKY-18), and a phospholipase D (AtPLD-1) from Arabidopsis. Genes showing elevated mRNA levels in Nep1-treated (5 micro g mL(-)(1), 0.1% [v/v] Silwet-L77) leaves 15 min after Nep1 treatment included CmSER-1 and CmCAL-1 for spotted knapweed, ToCYP-1 and CmCAL-1 for dandelion, and AtPK-1, AtWRKY-18, AtWI-12, and AtLEA-1 for Arabidopsis. Levels of mRNA for AtPLD-1 (Arabidopsis) and ToPP2C-1 (dandelion) decreased rapidly in Silwet-L77-treated plants between 15 min and 4 h of treatment, but were maintained or decreased more slowly over time in Nep1-treated (5 micro g mL(-)(1), 0.1% [v/v] Silwet-L77) leaves. In general, increases in mRNA band intensities were in the range of two to five times, with only ToCYP-1 in dandelion exceeding an increase of 10 times. The identified genes have been shown to be involved or are related to gene families that are involved in plant stress responses, including wounding, drought, senescence, and disease resistance.