Gabriella Pasqua - Academia.edu (original) (raw)
Papers by Gabriella Pasqua
European journal of histochemistry : EJH, 2003
The accumulation of essential oils in Angelica archangelica subsp. archangelica roots at differen... more The accumulation of essential oils in Angelica archangelica subsp. archangelica roots at different developmental stages was investigated through histochemical and chemical analyses. Roots less than 1 mm in diameter showed a primary diarch structure and two primary secretory ducts in the pericycle. These ducts were ephemeral and probably became dysfunctional early on. Oil accumulation was observed only in the secondary secretory ducts formed by cambium activity and located in the secondary phloem. Gas chromatographic analyses revealed that only taproots exceeding 5 mm in diameter contained a high concentration of alpha- and beta-phellandrene, which appreciably influence the oil's aroma.
Plant Physiology and …, 2002
To better understand the biosynthesis of Camptotheca acuminata alkaloids, the effect on camptothe... more To better understand the biosynthesis of Camptotheca acuminata alkaloids, the effect on camptothecin production of feeding with potential precursors of biosynthesis was studied (i.e., tryptamine and loganin combined, secologanin, and strictosidine). Two key enzymes in alkaloid biosynthesis [i.e., tryptophan decarboxylase (TDC; EC 4.1.1.28) and strictosidine synthase (STR; EC 4.3.3.2)] were also studied. The analyses were conducted using a C. acuminata CG1 cell line that does not produce alkaloids, which could be useful in better understanding the biosynthetic pathway and in identifying possible limiting factors. The activity of TDC was 5 pkat mg-1; the activity of STR was 1.1 pkat mg -1 . Feeding with strictosidine revealed that this precursor is easily biotransformed by two enzymes (i.e., a hydroxylase and a dehydrogenase) in hydroxystrictosidine and didehydrostrictosidine, but camptothecin was never detected. The indole pathway and the low level of STR activity could be limiting factors in the production of camptothecin in the cell line used.
Plant Physiology and Biochemistry, 2006
The metabolic profile of secondary products in calli and cell suspension cultures of Camptotheca ... more The metabolic profile of secondary products in calli and cell suspension cultures of Camptotheca acuminata Decaisne was investigated and compared to that of the leaves and roots taken from the plant. Neither in vitro system produced the anticancer quinoline alkaloid camptothecin (CPT); they accumulated discrete quantities of polyhydroxylated triterpenoids, different from those found in the plant organs, and ellagic acid derivatives. Nine ellagic acid derivatives (1a-1d and 2a-2e) and eight triterpenoid acids (3a-3e and 4a-4c) were isolated and characterised. All the identified triterpenes were related to ursane- or oleanane-type skeletons and their concentrations rose to 4.5% in suspended cells.
Plant Biosystems - An International Journal Dealing with all Aspects of Plant Biology, 2015
Centaurea cineraria subsp. circae is an endemic plant with a distribution area limited to Circeo ... more Centaurea cineraria subsp. circae is an endemic plant with a distribution area limited to Circeo mountain (Lazio, Italy), whose population was estimated in a very low number of individuals. The aim of this work is to investigate ex situ conservation strategies such as achene collection and in vitro plant propagation, that will permit to carry out restoration programs. The test carried out on the achenes demonstrated that only 5.5% of them were morphologically healthy. Seed germination tests showed that seeds do not display dormancy and that germination does not require pretreatments. The higher germination rate (67.5%) was observed under a photoperiod of 12 h/12 h (light/dark) and temperature regime +20 °C/+10 °C. The in vitro studies demonstrated that micropropagation, acclimatization and the transfer outdoors of C. cineraria subsp. circae are not particularly difficult: 74% of shoot explants in MS medium added with 0.5 mg/L benzylaminopurine (BAP) and 2 mg/L kinetin (Kin) formed multiple shoots; 100% of shoots rooted in MS medium added with 0.5 mg/L IBA and over 90% survived the acclimatization phase. After been transferred outdoors the totality of in vitro propagated plants bloomed and appeared morphologically indistinguishable from wild plants. Preliminary chemical analyses showed a similar profile for in vitro propagated and wild plants.
British Journal of Nutrition, 2010
Consumption of grape seed extract (GSE) is widely marketed as a dietary supplement and is conside... more Consumption of grape seed extract (GSE) is widely marketed as a dietary supplement and is considered safe for human health. Nevertheless, the analytical composition of GSE from different grape cultivars, growing in special agronomic constraints, differs greatly in flavan-3-ols content. The major concern with GSE studies is a lack of availability of uniformly standardised preparations, which raises an important question whether different GSE samples have comparable activity and trigger the same mechanisms of action on a given biological system. Therefore, it is tempting to speculate that GSE, obtained from different cultivars, could exert differentiated anticancer effects. The focus of the present study is to determine the selective biological efficacy of GSE obtained from three different sources on the human colon cancer cell line Caco-2. Irrespective of its source, high doses of GSE induced a significant inhibition on Caco-2 cell growth. Moreover, apoptosis was enhanced through both caspase-dependent and caspase-independent mechanisms, leading to an early apoptosis-inducing factor release and, further, to a dramatic increase in caspase 7 and 3 activity. However, a significant difference in apoptotic rates induced by the three grape sources clearly emerged when treating cancer cells with low and intermediate GSE concentrations (25 and 50 mg/ml).
Plant Biosystems, 2008
This is the first report on the in vitro asymbiotic germination of Orchis mascula L. seeds, colle... more This is the first report on the in vitro asymbiotic germination of Orchis mascula L. seeds, collected in the Simbruini Natural Park (Italy). Different pre-inoculum treatments and various culture media were tested to induce germination and to improve embryo development. The Orchimax medium supplemented with benzyladenine and active charcoal proved to be the best in promoting seed germination. The time course of protocorm development is described.
Hypericum perforatum L. is a medicinal plant commonly used worldwide for the treatment of mild an... more Hypericum perforatum L. is a medicinal plant commonly used worldwide for the
treatment of mild and moderate depression due to its wide range of bioactive
compounds. H. perforatum regenerated roots have been proposed as an efficacious in
vitro system to biosynthesize pharmaceutically useful secondary metabolites. Up to
now the effects of the increase of biomass density and/or chitosan elicitation in
addressing the metabolism of H. perforatum regenerated roots towards specific
metabolic pathways have not yet been studied in depth. A metabolomic platform, which
integrates a NMR-based metabolic profiling and ANOVA-simultaneous component
analysis, has been applied in order to characterize the changes of the primary and
secondary metabolism of H. perforatum regenerated roots during cultivation in a
confined growth environment and in response to chitosan elicitation.
NMR -based metabolomics allowed the primary and secondary metabolism of H.
perforatum regenerated roots to be characterized and the effects due to the increasing
biomass density or to the elicitation process to be determined. The most significant
metabolic variations involved levels of amino acids, carbohydrates, phenylpropanoids,
lipids, and isoprenoids. With the biomass growth, the levels of tryptophan, fructose,
shikimic acid, and epicatechin increased, whereas γ-aminobutyric acid and histidine
decreased. In response to chitosan elicitation, valine, isoleucine, glutamine, γ-aminobutyric acid, fructose, sucrose, poly-unsaturated fatty acids, epicatechin,
xanthones, dimethylallyl-pyrophosphate, and stigmasterol increased, while histidine
decreased.
The metabolic profiling of regenerated roots shows how the cultures respond to
different stress conditions, providing useful information for designing optimal
bioprocesses to produce mixture of secondary metabolites having potentially different
bioactivities.
Poly(lactic-co-glycolic) acid (PLGA)- based NPs are currently considered among the most promising... more Poly(lactic-co-glycolic) acid (PLGA)-
based NPs are currently considered among the most
promising drug carriers, nevertheless their use in
plants has never been investigated. In this work, for the
first time, we demonstrated the ability of PLGA NPs to
cross the plant cell wall and membrane of Vitis
vinifera cell cultures and grapevine-pathogenic fungi.
By means of fluorescence microscopy, we established
that PLGA NPs can enter in grapevine leaf tissues
through stomata openings and that they can be
absorbed by the roots and transported to the shoot
through vascular tissues. TEM analysis on cultured
cells showed that NPs B 50 nm could enter cells,
while bigger ones remained attached to the cell wall.
Viability tests demonstrated that PLGA NPs were not
cytotoxic for V. vinifera-cultured cells. The cellular
uptake of PLGA NPs by some important grapevinepathogenic
fungi has also been observed, thus suggesting
that PLGA NPs could be used to deliver
antifungal compounds within fungal cells. Overall the
results reported suggest that such NPs may play a key
role in future developments of agrobiotechnologies, as
it is currently happening in biomedicine.
For the first time, grape seed extracts (GSEs), obtained from wine and table cultivars of Vitis v... more For the first time, grape seed extracts (GSEs), obtained from wine and table cultivars of Vitis vinifera L., cultured in experimental
fields of Lazio and Puglia regions of Italy and grown in different agronomic conditions, have been tested on 43 Candida species
strains.We demonstrated a significant correlation between the content of the flavan-3-ols in GSEs extracts, with a polymerization
degree ≥4, and anti-Candida activity.Moreover, we demonstrated thatGSEs, obtained from plants cultured with reduced irrigation,
showed a content of polymeric flavan-3-ols >250mg/gwith geometric mean MICvalues between 5.7 and 20.2 mg/L against Candida
albicans reference strains. GSE, showing 573mg/g of polymeric flavan-3-ols, has been tested in an experimental murine model of
vaginal candidiasis by using noninvasive in vivo imaging technique. The results pointed out a significant inhibition of Candida
albicans load 5 days after challenge.These findings indicate that GSEs with high content of polymeric flavan-3-ols can be used in
mucosal infection as vaginal candidiasis.
The aim of this work was to evaluate the effect of stilbenes from different cultivars of Vitis v... more The aim of this work was to evaluate the effect of stilbenes from different cultivars of
Vitis vinifera on tumour proliferation. Extracts were obtained from elicited V. vinifera
cell cultures and characterised by HPLC/DAD/MS. Cell growth was evaluated in four
human cancer cell lines and in normal human fibroblasts. The cells were exposed to the
extracts or to trans-resveratrol, used as reference molecule, for 48 h, at 1–10 mM
concentrations of total stilbenoids. All the extracts exhibited antiproliferative activity,
mediated by modulation of the cell cycle and induction of cytotoxicity in cancer but not
in normal cell lines, and positively correlated with the content in dimeric stilbenoids.
The Alphonse Lavalle´e extract was the most active, and the obtained stilbenoid fraction
resulted 8–10 times more active than trans-resveratrol. Extracts from V. vinifera cell
cultures could represent new sources of active stilbenoid compounds to be further
assayed in in vivo studies for their antitumoural properties.
High-performance liquid chromatography coupled with a UV photodiode-array detector and an electro... more High-performance liquid chromatography coupled with a UV photodiode-array detector and an electrospray mass spectrometer (HPLC-ES-MS) was used to analyze the phytochemical constituents of an extract of sour orange (Citrus aurantium L., family Rutaceae). Eight flavonoids were identified, as isonaringin (1), naringin (3), hesperidin (5), neohesperidin (6), naringenin (2), hesperitin (4), nobiletin (8) and tangeritin . The permethoxylated flavones tangeritin (7) and nobiletin (8) showed the most intense protonated molecules. This method can be used to detect trace amounts of these compounds in an orange extract. © 1997 Elsevier Science B. V.
Plant Science, 2003
Methanolic extracts from cell suspension cultures, calli, and in vitro regenerated shoots and roo... more Methanolic extracts from cell suspension cultures, calli, and in vitro regenerated shoots and roots of Hypericum perforatum cv. Topas have been evaluated for their ability to produce active metabolites (hypericins, hyperforins and flavonoids). Biosynthesis of hypericins is connected with the formation of secretory structures (black globules) in regenerated vegetative buds. A further degree of leaf development is necessary to stimulate the production of hyperforins and flavonoids. Xanthones are the main metabolic products in suspension cells, undifferentiated calli and roots regenerated from plantlets or formed by callus. No xanthones are detected in the aerial parts of regenerated plantlets accumulating hypericins, hyperforins and flavonoids. #
British Journal of Nutrition, 2010
Consumption of grape seed extract (GSE) is widely marketed as a dietary supplement and is conside... more Consumption of grape seed extract (GSE) is widely marketed as a dietary supplement and is considered safe for human health. Nevertheless, the analytical composition of GSE from different grape cultivars, growing in special agronomic constraints, differs greatly in flavan-3-ols content. The major concern with GSE studies is a lack of availability of uniformly standardised preparations, which raises an important question whether different GSE samples have comparable activity and trigger the same mechanisms of action on a given biological system. Therefore, it is tempting to speculate that GSE, obtained from different cultivars, could exert differentiated anticancer effects. The focus of the present study is to determine the selective biological efficacy of GSE obtained from three different sources on the human colon cancer cell line Caco-2. Irrespective of its source, high doses of GSE induced a significant inhibition on Caco-2 cell growth. Moreover, apoptosis was enhanced through both caspase-dependent and caspase-independent mechanisms, leading to an early apoptosis-inducing factor release and, further, to a dramatic increase in caspase 7 and 3 activity. However, a significant difference in apoptotic rates induced by the three grape sources clearly emerged when treating cancer cells with low and intermediate GSE concentrations (25 and 50 mg/ml).
Rapid Communications in Mass Spectrometry, 2010
Cell suspension cultures of Vitis vinifera L., particularly after elicitation, have shown the abi... more Cell suspension cultures of Vitis vinifera L., particularly after elicitation, have shown the ability to biosynthesize several stilbenoids. In this work the application of specific tandem mass spectrometry (MS/MS) experiments and the integration with data from UV-vis spectra, available only for the main compounds of the extract, allowed the detection of over 20 different stilbenoids. Up to eight monoglicosides, both monomers and dimers, a high number of dimer forms (up to 17), belonging to the group of viniferins and to their oxidized derivatives, were identified in the extract of Malvasia cell cultures. Moreover, the selectivity and sensitivity of the method allowed detection also of a few cis derivatives (up to 3) present in very low concentrations and presumably produced by the light exposure during treatment of the sample. To the best of the authors' knowledge, there are no literature data on MS/MS experiments targeted to simultaneously study monomeric and dimeric stilbenoids, both as free and glycosilated forms, in a complex mixture from V. vinifera cell suspension cultures.
Planta Medica, 2002
A new compound was isolated from calli of Taxus baccata L. and assigned the structure 3beta,11-di... more A new compound was isolated from calli of Taxus baccata L. and assigned the structure 3beta,11-dihydroxy-12-methoxyabieta-8,11,13-triene-7-one. Two other metabolites were identified as 3-oxocryptojaponol and taxamairein C, both previously isolated from Taxus mairei.
Plant Science, 2002
Thin cell layers (TCL) of Nicotiana tabacum L. cv. Samsun were cultured in vitro to evaluate the ... more Thin cell layers (TCL) of Nicotiana tabacum L. cv. Samsun were cultured in vitro to evaluate the effect of the pH of the culture medium and ionic uptake on vegetative organogenesis. The chromatographic analysis showed important variations during culture in the concentrations of the ions in the tissues of the explants grown at different pH levels. After 25 days of culture, shoots with regular development were obtained especially from the explants grown on the media with an initial pH (pH i ) of 4.0 and 5.0. At pH i 7.0, the shoots were often grouped into tufts, lacking apical dominance and vitreous in appearance, probably due to a high uptake of ammonium in the first 48 h of culture and a low level of calcium in the first 24 h of culture. Regardless of the origin, (i.e. subepidermal chlorenchyma cells or wound callus generated by the outermost parenchyma layers) vegetative buds differed according to the pH i level. Direct superficial origin was observed only at pH i 4.0 and 5.0, whereas indirect cicatritial origin was observed at all pH levels. #
Plant Science, 2003
The activity and substrate specificity of latex lipases from Euphorbia species (E. characias , E.... more The activity and substrate specificity of latex lipases from Euphorbia species (E. characias , E. wulfenii , E. pinea , E. myrsinites and E. dendroides ) were investigated. High lipolytic activity was found only in E. characias and for the first time in E. wulfenii latex. For both species the lipolytic activity on various triglycerides, and under different temperature and pH conditions, in both crude latex and in partially purified enzymes was quantified. Optimised extraction and purification methods permitted the recovery of the enzymatic fraction with high lipolytic activity. This fraction is probably constituted by a pool of different lipolytic enzymes. Finally, lipolytic activity was also measured for E. characias and E. wulfenii during vegetative and reproductive stages. #
Plant Science, 2007
The objective of the present work was to contribute to the understanding of the physiological rol... more The objective of the present work was to contribute to the understanding of the physiological role of latex lipolytic activity in Euphorbia characias. To this end, the acid and basic lipolytic activity of E. characias latex, as well as the substrate specificity on various triacylglycerols, were measured during the plant's vegetative and reproductive stages. Both activities appeared to increase during the reproductive stage and to peak at the beginning of the vegetative stage, when new leaves and branches are formed. For the first time, the phospholipolytic and esterase activity of E. characias latex is also reported. An extraction method in aqueous medium with the zwitterionic detergent CHAPS was successfully used to extract lipolytic activity from latex. Extraction permitted the selective recovery of a single protein spot, with a molecular weight of 37 kDa, and presumably made of several acid isoforms which retained both lipolytic and phospholipolytic activity. The biochemical results suggest that lipolytic and phospholipolytic activity could depend on a single hydrolytic enzyme with several isoforms, equally expressed throughout the biological cycle of the plant. On the basis of the obtained results, we hypothesise that the E. characias latex lipase should be considered as an aspecific acylhydrolase with a combined lipase/phospholipase A activity. #
Plant Science, 2002
This study was carried out to investigate how the emission of isoprenoids that follows wounding o... more This study was carried out to investigate how the emission of isoprenoids that follows wounding of pine needles is restrained and how this can be associated to wound sealing processes and to defense mechanisms against pathogenic attacks. Needles of two Mediterranean pines (Pinus pinea and Pinus halepensis ) emitted a high amount of monoterpenes immediately after wounding but the emission became undetectable within 24 h. Histochemical analysis revealed that the purple stain coloring the isoprenoids in fresh needle sections turned black 24 h after wounding the needles. This suggests that isoprenoids accumulated at the wounding sites. Sections below the wounds showed that the ducts were still full of isoprenoids stained in purple. This excluded the possibility that the emission from wounded needles ceases because the ducts were empty. Chromatographic analysis of the resin covering the wounds indicated a high amount of sesquiterpenes 1 h after wounding and a considerable increase in oxygenated isoprenoids 24 h after wounding. Our results indicate that the emission from wounded needles may be rapidly limited and eventually stopped by the accumulation of oxidation products of the same emitted isoprenoids. This mechanism is similar to that used to seal trunk wounding and may constitute a physical and chemical barrier against pest attacks. #
Protoplasma, 2005
In this paper, a system of laticifers in Camptotheca acuminata Decne (Nyssaceae) is described. La... more In this paper, a system of laticifers in Camptotheca acuminata Decne (Nyssaceae) is described. Laticifers were already present in the leaf primordia of the shoot apex. In the mature leaves, laticifers were found in the midrib and in the larger veins, both in the parenchymatic region delimited by vascular bundles and in the cortex just external to the phloem. In the stem, laticifers were present in both the primary and secondary body, running parallel to the longitudinal axis. They were located in the pith and in the cortex proximal to the phloem. No laticifers were found in the roots. The histochemical analyses indicated that the main compounds accumulated in laticifers were phenols. Neutral lipids and fatty acids were also present. Ultrastructural observations showed osmiophilic globules both in the vacuoles and in the peripheral regions of the cytoplasm of the laticifer cells. Plastids were present, although altered, with some parallel membranes and lacking starch grains. The discovery in C. acuminata of a laticifer system, which had never been described for the order Cornales, could be of taxonomic value, also considering that this order has traditionally represented one of the most problematic groups of flowering plants.
European journal of histochemistry : EJH, 2003
The accumulation of essential oils in Angelica archangelica subsp. archangelica roots at differen... more The accumulation of essential oils in Angelica archangelica subsp. archangelica roots at different developmental stages was investigated through histochemical and chemical analyses. Roots less than 1 mm in diameter showed a primary diarch structure and two primary secretory ducts in the pericycle. These ducts were ephemeral and probably became dysfunctional early on. Oil accumulation was observed only in the secondary secretory ducts formed by cambium activity and located in the secondary phloem. Gas chromatographic analyses revealed that only taproots exceeding 5 mm in diameter contained a high concentration of alpha- and beta-phellandrene, which appreciably influence the oil's aroma.
Plant Physiology and …, 2002
To better understand the biosynthesis of Camptotheca acuminata alkaloids, the effect on camptothe... more To better understand the biosynthesis of Camptotheca acuminata alkaloids, the effect on camptothecin production of feeding with potential precursors of biosynthesis was studied (i.e., tryptamine and loganin combined, secologanin, and strictosidine). Two key enzymes in alkaloid biosynthesis [i.e., tryptophan decarboxylase (TDC; EC 4.1.1.28) and strictosidine synthase (STR; EC 4.3.3.2)] were also studied. The analyses were conducted using a C. acuminata CG1 cell line that does not produce alkaloids, which could be useful in better understanding the biosynthetic pathway and in identifying possible limiting factors. The activity of TDC was 5 pkat mg-1; the activity of STR was 1.1 pkat mg -1 . Feeding with strictosidine revealed that this precursor is easily biotransformed by two enzymes (i.e., a hydroxylase and a dehydrogenase) in hydroxystrictosidine and didehydrostrictosidine, but camptothecin was never detected. The indole pathway and the low level of STR activity could be limiting factors in the production of camptothecin in the cell line used.
Plant Physiology and Biochemistry, 2006
The metabolic profile of secondary products in calli and cell suspension cultures of Camptotheca ... more The metabolic profile of secondary products in calli and cell suspension cultures of Camptotheca acuminata Decaisne was investigated and compared to that of the leaves and roots taken from the plant. Neither in vitro system produced the anticancer quinoline alkaloid camptothecin (CPT); they accumulated discrete quantities of polyhydroxylated triterpenoids, different from those found in the plant organs, and ellagic acid derivatives. Nine ellagic acid derivatives (1a-1d and 2a-2e) and eight triterpenoid acids (3a-3e and 4a-4c) were isolated and characterised. All the identified triterpenes were related to ursane- or oleanane-type skeletons and their concentrations rose to 4.5% in suspended cells.
Plant Biosystems - An International Journal Dealing with all Aspects of Plant Biology, 2015
Centaurea cineraria subsp. circae is an endemic plant with a distribution area limited to Circeo ... more Centaurea cineraria subsp. circae is an endemic plant with a distribution area limited to Circeo mountain (Lazio, Italy), whose population was estimated in a very low number of individuals. The aim of this work is to investigate ex situ conservation strategies such as achene collection and in vitro plant propagation, that will permit to carry out restoration programs. The test carried out on the achenes demonstrated that only 5.5% of them were morphologically healthy. Seed germination tests showed that seeds do not display dormancy and that germination does not require pretreatments. The higher germination rate (67.5%) was observed under a photoperiod of 12 h/12 h (light/dark) and temperature regime +20 °C/+10 °C. The in vitro studies demonstrated that micropropagation, acclimatization and the transfer outdoors of C. cineraria subsp. circae are not particularly difficult: 74% of shoot explants in MS medium added with 0.5 mg/L benzylaminopurine (BAP) and 2 mg/L kinetin (Kin) formed multiple shoots; 100% of shoots rooted in MS medium added with 0.5 mg/L IBA and over 90% survived the acclimatization phase. After been transferred outdoors the totality of in vitro propagated plants bloomed and appeared morphologically indistinguishable from wild plants. Preliminary chemical analyses showed a similar profile for in vitro propagated and wild plants.
British Journal of Nutrition, 2010
Consumption of grape seed extract (GSE) is widely marketed as a dietary supplement and is conside... more Consumption of grape seed extract (GSE) is widely marketed as a dietary supplement and is considered safe for human health. Nevertheless, the analytical composition of GSE from different grape cultivars, growing in special agronomic constraints, differs greatly in flavan-3-ols content. The major concern with GSE studies is a lack of availability of uniformly standardised preparations, which raises an important question whether different GSE samples have comparable activity and trigger the same mechanisms of action on a given biological system. Therefore, it is tempting to speculate that GSE, obtained from different cultivars, could exert differentiated anticancer effects. The focus of the present study is to determine the selective biological efficacy of GSE obtained from three different sources on the human colon cancer cell line Caco-2. Irrespective of its source, high doses of GSE induced a significant inhibition on Caco-2 cell growth. Moreover, apoptosis was enhanced through both caspase-dependent and caspase-independent mechanisms, leading to an early apoptosis-inducing factor release and, further, to a dramatic increase in caspase 7 and 3 activity. However, a significant difference in apoptotic rates induced by the three grape sources clearly emerged when treating cancer cells with low and intermediate GSE concentrations (25 and 50 mg/ml).
Plant Biosystems, 2008
This is the first report on the in vitro asymbiotic germination of Orchis mascula L. seeds, colle... more This is the first report on the in vitro asymbiotic germination of Orchis mascula L. seeds, collected in the Simbruini Natural Park (Italy). Different pre-inoculum treatments and various culture media were tested to induce germination and to improve embryo development. The Orchimax medium supplemented with benzyladenine and active charcoal proved to be the best in promoting seed germination. The time course of protocorm development is described.
Hypericum perforatum L. is a medicinal plant commonly used worldwide for the treatment of mild an... more Hypericum perforatum L. is a medicinal plant commonly used worldwide for the
treatment of mild and moderate depression due to its wide range of bioactive
compounds. H. perforatum regenerated roots have been proposed as an efficacious in
vitro system to biosynthesize pharmaceutically useful secondary metabolites. Up to
now the effects of the increase of biomass density and/or chitosan elicitation in
addressing the metabolism of H. perforatum regenerated roots towards specific
metabolic pathways have not yet been studied in depth. A metabolomic platform, which
integrates a NMR-based metabolic profiling and ANOVA-simultaneous component
analysis, has been applied in order to characterize the changes of the primary and
secondary metabolism of H. perforatum regenerated roots during cultivation in a
confined growth environment and in response to chitosan elicitation.
NMR -based metabolomics allowed the primary and secondary metabolism of H.
perforatum regenerated roots to be characterized and the effects due to the increasing
biomass density or to the elicitation process to be determined. The most significant
metabolic variations involved levels of amino acids, carbohydrates, phenylpropanoids,
lipids, and isoprenoids. With the biomass growth, the levels of tryptophan, fructose,
shikimic acid, and epicatechin increased, whereas γ-aminobutyric acid and histidine
decreased. In response to chitosan elicitation, valine, isoleucine, glutamine, γ-aminobutyric acid, fructose, sucrose, poly-unsaturated fatty acids, epicatechin,
xanthones, dimethylallyl-pyrophosphate, and stigmasterol increased, while histidine
decreased.
The metabolic profiling of regenerated roots shows how the cultures respond to
different stress conditions, providing useful information for designing optimal
bioprocesses to produce mixture of secondary metabolites having potentially different
bioactivities.
Poly(lactic-co-glycolic) acid (PLGA)- based NPs are currently considered among the most promising... more Poly(lactic-co-glycolic) acid (PLGA)-
based NPs are currently considered among the most
promising drug carriers, nevertheless their use in
plants has never been investigated. In this work, for the
first time, we demonstrated the ability of PLGA NPs to
cross the plant cell wall and membrane of Vitis
vinifera cell cultures and grapevine-pathogenic fungi.
By means of fluorescence microscopy, we established
that PLGA NPs can enter in grapevine leaf tissues
through stomata openings and that they can be
absorbed by the roots and transported to the shoot
through vascular tissues. TEM analysis on cultured
cells showed that NPs B 50 nm could enter cells,
while bigger ones remained attached to the cell wall.
Viability tests demonstrated that PLGA NPs were not
cytotoxic for V. vinifera-cultured cells. The cellular
uptake of PLGA NPs by some important grapevinepathogenic
fungi has also been observed, thus suggesting
that PLGA NPs could be used to deliver
antifungal compounds within fungal cells. Overall the
results reported suggest that such NPs may play a key
role in future developments of agrobiotechnologies, as
it is currently happening in biomedicine.
For the first time, grape seed extracts (GSEs), obtained from wine and table cultivars of Vitis v... more For the first time, grape seed extracts (GSEs), obtained from wine and table cultivars of Vitis vinifera L., cultured in experimental
fields of Lazio and Puglia regions of Italy and grown in different agronomic conditions, have been tested on 43 Candida species
strains.We demonstrated a significant correlation between the content of the flavan-3-ols in GSEs extracts, with a polymerization
degree ≥4, and anti-Candida activity.Moreover, we demonstrated thatGSEs, obtained from plants cultured with reduced irrigation,
showed a content of polymeric flavan-3-ols >250mg/gwith geometric mean MICvalues between 5.7 and 20.2 mg/L against Candida
albicans reference strains. GSE, showing 573mg/g of polymeric flavan-3-ols, has been tested in an experimental murine model of
vaginal candidiasis by using noninvasive in vivo imaging technique. The results pointed out a significant inhibition of Candida
albicans load 5 days after challenge.These findings indicate that GSEs with high content of polymeric flavan-3-ols can be used in
mucosal infection as vaginal candidiasis.
The aim of this work was to evaluate the effect of stilbenes from different cultivars of Vitis v... more The aim of this work was to evaluate the effect of stilbenes from different cultivars of
Vitis vinifera on tumour proliferation. Extracts were obtained from elicited V. vinifera
cell cultures and characterised by HPLC/DAD/MS. Cell growth was evaluated in four
human cancer cell lines and in normal human fibroblasts. The cells were exposed to the
extracts or to trans-resveratrol, used as reference molecule, for 48 h, at 1–10 mM
concentrations of total stilbenoids. All the extracts exhibited antiproliferative activity,
mediated by modulation of the cell cycle and induction of cytotoxicity in cancer but not
in normal cell lines, and positively correlated with the content in dimeric stilbenoids.
The Alphonse Lavalle´e extract was the most active, and the obtained stilbenoid fraction
resulted 8–10 times more active than trans-resveratrol. Extracts from V. vinifera cell
cultures could represent new sources of active stilbenoid compounds to be further
assayed in in vivo studies for their antitumoural properties.
High-performance liquid chromatography coupled with a UV photodiode-array detector and an electro... more High-performance liquid chromatography coupled with a UV photodiode-array detector and an electrospray mass spectrometer (HPLC-ES-MS) was used to analyze the phytochemical constituents of an extract of sour orange (Citrus aurantium L., family Rutaceae). Eight flavonoids were identified, as isonaringin (1), naringin (3), hesperidin (5), neohesperidin (6), naringenin (2), hesperitin (4), nobiletin (8) and tangeritin . The permethoxylated flavones tangeritin (7) and nobiletin (8) showed the most intense protonated molecules. This method can be used to detect trace amounts of these compounds in an orange extract. © 1997 Elsevier Science B. V.
Plant Science, 2003
Methanolic extracts from cell suspension cultures, calli, and in vitro regenerated shoots and roo... more Methanolic extracts from cell suspension cultures, calli, and in vitro regenerated shoots and roots of Hypericum perforatum cv. Topas have been evaluated for their ability to produce active metabolites (hypericins, hyperforins and flavonoids). Biosynthesis of hypericins is connected with the formation of secretory structures (black globules) in regenerated vegetative buds. A further degree of leaf development is necessary to stimulate the production of hyperforins and flavonoids. Xanthones are the main metabolic products in suspension cells, undifferentiated calli and roots regenerated from plantlets or formed by callus. No xanthones are detected in the aerial parts of regenerated plantlets accumulating hypericins, hyperforins and flavonoids. #
British Journal of Nutrition, 2010
Consumption of grape seed extract (GSE) is widely marketed as a dietary supplement and is conside... more Consumption of grape seed extract (GSE) is widely marketed as a dietary supplement and is considered safe for human health. Nevertheless, the analytical composition of GSE from different grape cultivars, growing in special agronomic constraints, differs greatly in flavan-3-ols content. The major concern with GSE studies is a lack of availability of uniformly standardised preparations, which raises an important question whether different GSE samples have comparable activity and trigger the same mechanisms of action on a given biological system. Therefore, it is tempting to speculate that GSE, obtained from different cultivars, could exert differentiated anticancer effects. The focus of the present study is to determine the selective biological efficacy of GSE obtained from three different sources on the human colon cancer cell line Caco-2. Irrespective of its source, high doses of GSE induced a significant inhibition on Caco-2 cell growth. Moreover, apoptosis was enhanced through both caspase-dependent and caspase-independent mechanisms, leading to an early apoptosis-inducing factor release and, further, to a dramatic increase in caspase 7 and 3 activity. However, a significant difference in apoptotic rates induced by the three grape sources clearly emerged when treating cancer cells with low and intermediate GSE concentrations (25 and 50 mg/ml).
Rapid Communications in Mass Spectrometry, 2010
Cell suspension cultures of Vitis vinifera L., particularly after elicitation, have shown the abi... more Cell suspension cultures of Vitis vinifera L., particularly after elicitation, have shown the ability to biosynthesize several stilbenoids. In this work the application of specific tandem mass spectrometry (MS/MS) experiments and the integration with data from UV-vis spectra, available only for the main compounds of the extract, allowed the detection of over 20 different stilbenoids. Up to eight monoglicosides, both monomers and dimers, a high number of dimer forms (up to 17), belonging to the group of viniferins and to their oxidized derivatives, were identified in the extract of Malvasia cell cultures. Moreover, the selectivity and sensitivity of the method allowed detection also of a few cis derivatives (up to 3) present in very low concentrations and presumably produced by the light exposure during treatment of the sample. To the best of the authors' knowledge, there are no literature data on MS/MS experiments targeted to simultaneously study monomeric and dimeric stilbenoids, both as free and glycosilated forms, in a complex mixture from V. vinifera cell suspension cultures.
Planta Medica, 2002
A new compound was isolated from calli of Taxus baccata L. and assigned the structure 3beta,11-di... more A new compound was isolated from calli of Taxus baccata L. and assigned the structure 3beta,11-dihydroxy-12-methoxyabieta-8,11,13-triene-7-one. Two other metabolites were identified as 3-oxocryptojaponol and taxamairein C, both previously isolated from Taxus mairei.
Plant Science, 2002
Thin cell layers (TCL) of Nicotiana tabacum L. cv. Samsun were cultured in vitro to evaluate the ... more Thin cell layers (TCL) of Nicotiana tabacum L. cv. Samsun were cultured in vitro to evaluate the effect of the pH of the culture medium and ionic uptake on vegetative organogenesis. The chromatographic analysis showed important variations during culture in the concentrations of the ions in the tissues of the explants grown at different pH levels. After 25 days of culture, shoots with regular development were obtained especially from the explants grown on the media with an initial pH (pH i ) of 4.0 and 5.0. At pH i 7.0, the shoots were often grouped into tufts, lacking apical dominance and vitreous in appearance, probably due to a high uptake of ammonium in the first 48 h of culture and a low level of calcium in the first 24 h of culture. Regardless of the origin, (i.e. subepidermal chlorenchyma cells or wound callus generated by the outermost parenchyma layers) vegetative buds differed according to the pH i level. Direct superficial origin was observed only at pH i 4.0 and 5.0, whereas indirect cicatritial origin was observed at all pH levels. #
Plant Science, 2003
The activity and substrate specificity of latex lipases from Euphorbia species (E. characias , E.... more The activity and substrate specificity of latex lipases from Euphorbia species (E. characias , E. wulfenii , E. pinea , E. myrsinites and E. dendroides ) were investigated. High lipolytic activity was found only in E. characias and for the first time in E. wulfenii latex. For both species the lipolytic activity on various triglycerides, and under different temperature and pH conditions, in both crude latex and in partially purified enzymes was quantified. Optimised extraction and purification methods permitted the recovery of the enzymatic fraction with high lipolytic activity. This fraction is probably constituted by a pool of different lipolytic enzymes. Finally, lipolytic activity was also measured for E. characias and E. wulfenii during vegetative and reproductive stages. #
Plant Science, 2007
The objective of the present work was to contribute to the understanding of the physiological rol... more The objective of the present work was to contribute to the understanding of the physiological role of latex lipolytic activity in Euphorbia characias. To this end, the acid and basic lipolytic activity of E. characias latex, as well as the substrate specificity on various triacylglycerols, were measured during the plant's vegetative and reproductive stages. Both activities appeared to increase during the reproductive stage and to peak at the beginning of the vegetative stage, when new leaves and branches are formed. For the first time, the phospholipolytic and esterase activity of E. characias latex is also reported. An extraction method in aqueous medium with the zwitterionic detergent CHAPS was successfully used to extract lipolytic activity from latex. Extraction permitted the selective recovery of a single protein spot, with a molecular weight of 37 kDa, and presumably made of several acid isoforms which retained both lipolytic and phospholipolytic activity. The biochemical results suggest that lipolytic and phospholipolytic activity could depend on a single hydrolytic enzyme with several isoforms, equally expressed throughout the biological cycle of the plant. On the basis of the obtained results, we hypothesise that the E. characias latex lipase should be considered as an aspecific acylhydrolase with a combined lipase/phospholipase A activity. #
Plant Science, 2002
This study was carried out to investigate how the emission of isoprenoids that follows wounding o... more This study was carried out to investigate how the emission of isoprenoids that follows wounding of pine needles is restrained and how this can be associated to wound sealing processes and to defense mechanisms against pathogenic attacks. Needles of two Mediterranean pines (Pinus pinea and Pinus halepensis ) emitted a high amount of monoterpenes immediately after wounding but the emission became undetectable within 24 h. Histochemical analysis revealed that the purple stain coloring the isoprenoids in fresh needle sections turned black 24 h after wounding the needles. This suggests that isoprenoids accumulated at the wounding sites. Sections below the wounds showed that the ducts were still full of isoprenoids stained in purple. This excluded the possibility that the emission from wounded needles ceases because the ducts were empty. Chromatographic analysis of the resin covering the wounds indicated a high amount of sesquiterpenes 1 h after wounding and a considerable increase in oxygenated isoprenoids 24 h after wounding. Our results indicate that the emission from wounded needles may be rapidly limited and eventually stopped by the accumulation of oxidation products of the same emitted isoprenoids. This mechanism is similar to that used to seal trunk wounding and may constitute a physical and chemical barrier against pest attacks. #
Protoplasma, 2005
In this paper, a system of laticifers in Camptotheca acuminata Decne (Nyssaceae) is described. La... more In this paper, a system of laticifers in Camptotheca acuminata Decne (Nyssaceae) is described. Laticifers were already present in the leaf primordia of the shoot apex. In the mature leaves, laticifers were found in the midrib and in the larger veins, both in the parenchymatic region delimited by vascular bundles and in the cortex just external to the phloem. In the stem, laticifers were present in both the primary and secondary body, running parallel to the longitudinal axis. They were located in the pith and in the cortex proximal to the phloem. No laticifers were found in the roots. The histochemical analyses indicated that the main compounds accumulated in laticifers were phenols. Neutral lipids and fatty acids were also present. Ultrastructural observations showed osmiophilic globules both in the vacuoles and in the peripheral regions of the cytoplasm of the laticifer cells. Plastids were present, although altered, with some parallel membranes and lacking starch grains. The discovery in C. acuminata of a laticifer system, which had never been described for the order Cornales, could be of taxonomic value, also considering that this order has traditionally represented one of the most problematic groups of flowering plants.