Micropropagation of Salvia wagneriana Polak and hairy root cultures with rosmarinic acid production

Salvia wagneriana Polak is a tropical species native to Central America, well adapted to grow in the Mediterranean basin for garden decoration. Micropropagation has been assessed from axillary shoots of adult plants using a Murashige and Skoog basal medium, with the addition of 1.33-μM 6-benzylaminopurine for shoot proliferation; the subsequent rooting phase occurred in plant growth regulator-free medium. The plants were successfully acclimatised with high survival frequency. Hairy roots were induced after co-cultivation of leaf lamina and petiole fragments with Agrobacterium rhizogenes and confirmed by PCR. The establishment and proliferation of the selected HRD3 line were obtained in hormone-free liquid medium and the production of rosmarinic acid (RA) was evaluated after elicitation. The analysis of RA was performed by LC-ESI-DAD-MS in the hydroalcoholic extracts. The addition of casein hydrolysate increased the RA production, whereas no enrichment was observed after the elicitation with jasmonic acid

Establishment of in vitro tissue cultures from Echinacea angustifolia D.C. adult plants for the production of phytochemical compounds

SUMMARY The establishment of in vitro cultures of Echinacea angustifolia D.C. was obtained directly from sections of flower stalks of adult plants. The shoot formation was obtained from this plantmaterial placed on a modified MS basal medium named CH supplemented with 0.5 mg L1 6 benzylaminopurine (BA). The in vitro propagation procedure of E. angustifolia consisted of three distinct phases: an initial regeneration phase fromstalk sections (IP shoots on basal mediumwith 0.25 mg L1 BA), an elongation phase on active charcoal and an axillary proliferation of the shoots (AP shoots on basal medium with 0.5 mg L1 BA).Regenerating calli were established from leaves of in vitro shoots cultured on CH medium supplemented with 3 mg L1 BA and 0.5 mg L1 indole-3-butyric acid (IBA). Developed shoots from the callus cultures were subcultured on the CH medium with 0.5 mg L1 BA (leaf regenerated shoots: LR shoots). The secondary metabolite content of the in vitro plant material was compared with that of the greenhouse growing plants. The quali-quantitative LC-DAD-ESI-MS analysis on the extracts from axillary proliferation shoots (AP shoots) showed significant production of caffeic acid derivatives while leaf callus and LR shoots, accumulated mainly alkamides. These results showed that the proper choice of the procedures for in vitro multiplication allowed us to obtain plant biomass able to produce the active compounds typical of E. angustifolia plants

Daidzein Production and HeLa Cytotoxicity of Bituminaria bituminosa hairy root Cultures

Bituminaria bituminosa (L.) C.H. Stirt is a perennial species widely distributed in the Mediterranean basin and the Canary Islands. This species is used in folk medicine and currently has considerable pharmaceutical interest for its content in phenylpropanoids, furanocoumarins and pterocarpans. In vitro cultures (shoots and hairy roots) have been performed to obtain plant material useful for the production of these metabolites. Hairy root cultures were successfully established after inoculation of hypocotyls with the LBA 9402 A. rhizogenes strain. The HRPB3 line was selected for further analysis and elicited with chitosan and salicylic acid. All the HRPB3 cultures showed higher polyphenol content and greater DPPH-antioxidant activity than shoots cultured in vitro. The presence of isoflavone daidzein was detected in the hairy root extracts. The cytotoxic effect of HR extracts has been further tested on HeLa cells: the salicylic acid elicited HR exhibited good antiproliferative effects

Physiological and Biochemical Adaptive Traits in Leaves of Four Citrus Species Grown in an Italian Charterhouse

Citrus trees are a very important crops that are cultivated worldwide, but not much knowledge is known about the ecophysiological responses to climatic changes in trees under natural conditions. The aim of this study was to investigate their adaptive capacity in response to seasonal phenological and environmental changes. The trial included Citrus trees (sweet orange, bitter orange, lemon, mandarin) growing under non-regular cropping conditions in a Monumental Charterhouse in Tuscany, in a subtropical Mediterranean climate with hot summer conditions. During a 1-year field trial, we determined the variations in chlorophyll fluorescence parameters and leaf biochemical traits (content of chlorophylls and carotenoids, total phenolic content (TPC), total antioxidant capacity (TAC), and total non-structural carbohydrates). In all Citrus spp., interspecific mean values of photochemical efficiency peaked during the summer, while a marked photoinhibition occurred in the winter in concomitance with higher interspecific mean values of leaf TPC, TAC, and non-structural carbohydrates. The trees showed the pivotal role played by photosynthetic acclimation as a survival strategy to tolerate abiotic stress in the climate change hotspot of Mediterranean environment. This study is included in a wider project aimed at a new valorization of Citrus trees as genetic resource and its by-products with added-value applications for innovative functional foods

Production of Curcuminoids in different in vitro organs of Curcuma longa.

Curcuma longa L. (turmeric) is one of the most important spice and safe food additives. Its main constituents, curcuminoids, showed anti-inflammatory, antitumor and antioxidant effects. In the present work, an in vitro propagation method was developed to achieve selected plant organs with quantified curcuminoid content. I n vitro plants were obtained from sprouting buds as primary explants. The major curcuminoid constituents, such as curcumin (CUR), demethoxycurcumin (DEM), and bis-demethoxycurcumin (bis-DEM) were examined in different organs by LC-DAD-ESI-MS. A significant production of curcumin (more than 260 μg g−1fresh weight) was obtained from in vitro microrhizomes, especially grown in a Murashige and Skoog medium (MS) supplemented with kinetin (0.1 mg L−1), α-naphthaleneacetic acid (NAA, 1 mg L−1), sucrose (6%), agar (5%) and activated charcoal (0.1%). The analyzed microrhizomes showed reduced amounts of DEM and bis-DEM in comparison with CUR levels. In addition a shoot culture line was suitable to biosynthesize curcuminoids, in a ratio very similar to that identified in the fresh rhizomes of parent plants. This study represents the first direct quantification of curcuminoids in turmeric in vitro shoots and microrhizomes to be used in dietary supplements

Morphogenetic changes in essential oil composition of Hypericum perforatum during the course of ontogenesis.

Bertoli, Alessandra/0000-0002-0701-5163 WOS: 000291275300010 PubMed: 21639687 Context: In the past few years, an increasing interest in the volatile secondary metabolites of Hypericum perforatum L. (Guttiferae) has been arising. Objective: The present study is a contribution to better understand the relationship between the morphological variations and volatile composition during the phenological cycle. Materials and methods: Leaves at the stages of vegetative, floral budding, flowering and green capsule, buds, full opened flowers and green capsules were assayed for essential oil (EO) components by gas chromatography-flame ionization detector (GC-FID) and GC-mass spectrometry (MS). Results: Significant amounts of sesquiterpenes (oxygenated 26--50% and hydrocarbons 20--40%) and oxygenated hydrocarbons (13--38%) characterized the all analyzed samples showing peculiar fluctuations during the seven phenological stages. Although monoterpenes were present in much lower amounts (monoterpene hydrocarbons 0.4--6%; oxygenated monoterpenes 0.8--6%) they were considered also important discrimination for several stages. The green capsules and the full opened flowers collected at flowering stage were clearly distinguished in terms of EO compositions from the other samples. Discussion: For the first time, the EO composition of Turkish wild Hypericum perforatum was monitored by the hydrodistillation of different plant organs collected at different seven stages in order to point out the modification of target volatiles related to each phenological step. Conclusions: Based on the EO composition monitored during these seven morphological stages by GC-MS, principal component analysis and cluster analysis, significant metabolite modifications were observed during the phenological cycle which involved the levels of specific volatile target compounds belonging to the chemical classes of hydrocarbons, monoterpenes and sesquiterpenes

Evaluation of volatile constituents of Cochlospermum angolense.

The chemical composition of the essential oils obtained from the leaves and roots of Cochlospermum angolense (Welw) growing wild in Angola was analyzed for the first time by capillary gas chromatography (GC) and gas chromatography/mass spectrometry (GC-MS). The investigation led to the identification of 67 and 130 compounds from the leaves and roots, respectively. Both oils were strongly characterized by the presence of sesquiterpenoids (68.8% in the leaves and 53.2% in the roots), while monoterpenoids were present in minor percentages (9.8% in the leaves and 26.2% in the root). The main constituents of the leaves were germacrene D (9.4%), α-cadinol (7.4%) and 10- epi-cubenol (6.2%), while the most abundant compounds in the root essential oil were the sesquiterpenes β-caryophyllene (19.7%) and isoborneol (6.6%). The analysis by HS-SPME of the roots, leaves, fruits and seeds were also reported for the first time. Different volatile profiles were detected

Essential oils and volatile emission of eight South African species of Helichrysum grown in uniform environmental conditions

Abstract This paper reported for the first time the aroma profile and essential oil composition of eight species of Helichrysum endemic in South Africa but grown in Italy (CREA-Sanremo collection). The volatiles of all the studied Helichrysum were dominated by monoterpene hydrocarbons, except for H. basalticum, where sesquiterpene hydrocarbons was the main class of compounds with β-caryophyllene (13.7%) and α-guaiene (11.9%) as major components. Despite the great diversity of compounds obtained in the volatile emission (more than 130), only four compounds were responsible for at least 34% of the identified fraction. These compounds were: sabinene (16.0%), β-caryophyllene and α-guaiene in H. basalticum; sabinene (57.7%) in H. foetidum; (Z)-β-ocimene (34.6%), tricyclene (16.2%) and myrcene (10.0%) in H. incanatum; myrcene (29.3%), tricyclene (21.5%) and limonene in H. indicum; tricyclene (32.1%) and (E)-3-hexanol acetate (20.1%) in H. montanum; limonene (10.3%), sabinene (8.9%), 1-decene (7.6%) and 1-hexanol (7.0%) in H. mutabile; α- and β-pinene in H. patulum (27.6% and 44.9%, respectively) and α-pinene (36.3%) and α-fanchene(15.6%) in H. setosum. The EOs composition of these species was also different from each other with sesquiterpene compounds as the prevalent class. Valerianol (16.3%, os, in H. basalticum), ledol (16.3%, os, in H. foetidum), β-caryophyllene (11.0% and 13.4%, sh, in H. indicum and H. patulum, respectively), viridiflorol (18.3%, os, in H. montanum) and valerianol (30.1%, os in H. setosum) were found to be the main constituents. It is worthy to highlighting that the H. incarnatum EO showed a similar behaviour to that of the spontaneous emission with a predominance of monoterpene hydrocarbons (60.7% in EO vs 81.4% in VOC), both represented by (Z)-β-ocimene as main constituent

Sensitivity of entomopathogenic fungi and bacteria to plants secondary metabolites, for an alternative control of Rhipicephalus (Boophilus) microplus in cattle

Boophilus (Rhipicephalus) microplus is a one host hard tick widespread in warm climates worldwide, responsible for great economic losses. To avoid resistance in ticks population, induced by the repeated administration of conventional acaricides and/or the presence of residues in the environment in meat and in milk, an alternative approach can be achieved using entomopathogenic microorganisms such as fungi and bacteria, or essential oils (EOs). The aim of the present study was to evaluate the in vitro sensitivity of Beauveria bassiana, Metarhizium anisopliae, Scopulariopsis sp, Bacillus thuringiensis and Proteus mirabilis to Eucalyptus globulus, Lavandula hybrida, Pelargonium graveolens EOs and to their main constituents such as lynalool, linalyl-acetate, geraniol, citronellol and 1,8 cineole. EOs has been chemically characterized by GC-MS. Fungal isolates were tested by a microdilution assay to achieve minimal inhibitory concentration (MIC) of both EOs and main components. The sensitivity of bacteria was evaluated by an agar disk diffusion. The results obtained show the feasibility of an integrate approach for an eco-friendly control of R. microplus by use of both entomopathogenic fungi and P. graveolens EO. L. hybrida could be an interesting alternative when B. bassiana is not employed. Conversely, a combined use of B. thuringiensis and EOs would not be advisable in the integrate control of ticks