Effect of the pre-treatment of the samples on the natural substances extraction from Helianthus annuus L. using supercritical carbon dioxide.

The extraction of bioactive compounds from sunflowers (Helianthus annuus L.) with supercritical carbon dioxide has been studied. The samples were treated in four different ways and the effects of two factors (pressure and temperature) were investigated at 100, 500 bar and 35, 50 degrees C. The best yields were obtained using a high temperature and a high pressure (50 degrees C and 500 bar). The dry samples produced better extraction yields than the moist samples. The bioactivities of the extracts were compared for the samples treated in different ways. The best activity profiles were obtained for the moist samples extracted at 35 degrees C and 500 bar.

First European interlaboratory study of the analysis of benzoxazinone derivatives in plants by liquid chromatography.

Six laboratories from four different countries participated in the first European interlaboratory comparison exercise within the framework of the "Fate and toxicity of allelochemicals (natural plant toxins) in relation to environment and consumer" (FATEALLCHEM) European Union Project. The study, organized between November 2002 and March 2003, involved the analyses of seven benzoxazinone derivatives in two standard solutions and one purified extract of root material. Results are reported from the first phase of the study that examined the variability associated with different detection methods and different laboratories. The analytical strategies were based on liquid chromatography (LC) with diode array detection, LC coupled to mass spectrometry (MS) and LC coupled to tandem MS. When data from all laboratories were pooled, the relative standard deviation values ranged from 2 to 14% for the determination of target compounds in standard solutions, and between 19 and 47% for the analysis in root material. Comparison of the three detection techniques leads to the conclusion that MS approaches are the most accurate and precise techniques for the determination of benzoxazinone derivatives at ng/microL level in plant material.

Alfalfa (Medicago sativa L.) flavonoids. 2. Tricin and chrysoeriol glycosides from aerial parts.

Ten flavone glycosides have been isolated and identified in aerial parts of alfalfa. These included six tricin, one 3'-O-methyltricetin, and three chrysoeriol glycosides. Most of these compounds were acylated with ferulic, coumaric, or sinapic acids, and acylation occurred on the terminal glucuronic acid. Eight of these compounds, including 7-O-beta-D-glucuronopyranosyl-3'-O-methyltricetin, 7-O-beta-D-glucuronopyranosyl-4'-O-beta-D-glucuronopyranosidechrysoeriol, 7-O-[2'-O-feruloyl-beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranoside]chrysoeriol, 7-O-[2'-O-feruloyl-[beta-D-glucuronopyranosyl(1-->3)]-O-beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranoside]chrysoeriol, 7-O-[2'-O-sinapoyl-beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranoside]tricin, 7-O-[2'-O- feruloyl-beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranoside]tricin, 7-O-[2'-O-p-coumaroyl-beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranoside]tricin, and 7-O-[2'-O-feruloyl-[beta-D-glucuronopyranosyl(1-->3)]-O-beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranoside]tricin, have not been reported previously in the plant kingdom. Two previously identified alfalfa flavones, 7-O-beta-D-glucuronopyranosidetricin and 7-O-[beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranoside]tricin, were also isolated.

Acylated apigenin glycosides from alfalfa (Medicago sativa L.) var. Artal.

Three flavones, including 4'-O-[2'-O-E-feruloyl-O-beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranoside]apigenin, 7-O-beta-D-glucuronopyranosyl-4'-O-[2'-O-E-feruloyl-O-beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranoside]apigenin and 7-O-beta-D-glucuronopyranosyl-4'-O-[2'-O-p-E-coumaroyl-O-beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranoside]apigenin have been identified in alfalfa var. Artal. The known flavone 7-O-[2-O-E-feruloyl-[beta-D-glucuronopyranosyl(1-->3)]-O-beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucurono-pyranoside] apigenin was also isolated. The structures of these compounds were deduced on the basis of their spectral data.

Sesquiterpene lactones with potential use as natural herbicide models. 2. guaianolides.

A structure-activity study to evaluate the effect of 17 guaianolide sesquiterpene lactones (in a range of 100-0.001 microM) on the growth and germination of several mono- and dicotyledon target species is accomplished. Results are compared with those obtained in the same bioassay with an internal standard, the commercial herbicide Logran, to validate the results with a known active formulation and to compare the results with a commercial product to test their potential use as natural herbicide models. Specific conditions for the selective mono- or polyhydroxylation of guaianolides using the SeO(2)/tert-butyl hydroperoxide system are presented and discussed. The high regio- and stereoselectivities of the reaction are explained through the specific structural requirements of the bulky first adduct formed during the ene reaction. These compounds appear to have deeper effects on the growth of either monocots or dicots than the previously tested germacranolides. Otherwise, the lactone group seems to be necessary for the activity, though it does not necessarily need to be unsaturated. However, the presence of a second and easily accessible unsaturated carbonyl system greatly enhances the inhibitory activity. Lipophilicity and the stereochemistry of the possible anchoring sites are also crucial factors for the activity. Finally, the levels of growth inhibition obtained with some compounds on dicots or monocots are totally comparable to those of Logran and allow proposing them as lead compounds.

Bioactive carotanes from Trichoderma virens.

Four new metabolites with carotane skeletons, trichocaranes A (1), B (2), C (3), and D (4), were isolated from Trichoderma virens and their structures established by the interpretation of NMR and mass spectroscopic data. The trichocaranes significantly inhibited the growth of etiolated wheat coleoptiles: 40% at 10(-4) M with 1 and 2 and 86% at 10(-3) M with 3.

Search for a standard phytotoxic bioassay for allelochemicals. Selection of standard target species.

In the search for a standard bioassay of phytotoxicity for allelochemicals, 22 commercial varieties of eight plant species [four dicotyledons: lettuce (Compositae), carrot (Umbelliferae), cress (Cruciferae), tomato (Solanaceae); and four monocotyledons: onion (Liliaceae), barley, wheat, and corn (Gramineae)] proposed as models for the most common weed families have been tested at different pH and solution volumes per set conditions. Nine commercial varieties selected as standard target species (STS) were tested with standard commercial herbicides to ensure their sensitivity to phytotoxic compounds. Results are discussed to establish the proper growth requirements, and sensitivity of commercial seeds of STS and to find the most suitable commercial herbicides that allow comparison with an internal standard to validate the response of potential allelochemicals.

Natural and synthetic podolactones with potential use as natural herbicide models.

A collection of 11 natural and synthetic podolactones have been tested as allelochemicals in a range between 10(-4) and 10(-9) M, and their potential use as natural herbicide models is discussed. Their effects on the germination and growth of the dicots Lactuca sativa (cv. Nigra and cv. Roman), Lepidium sativum, and Lycopersicon esculentum and the monocots Allium cepa, Hordeum vulgare, and Triticum aestivum as standard target species have been studied. An important inhibitory effect on the germination and growth of all tested species (average = 90%) was produced by compounds 9-11 at 10(-4) M. The specific structural requirements related to their activities are studied. On the basis of these results, their use as potential natural herbicide models is proposed.

Dehydrozaluzanin C: a potent plant growth regulator with potential use as a natural herbicide template.

The natural product dehydrozaluzanin C (former DHZ) is a sesquiterpene lactone obtained from different weeds of the Compositae family. Its potential as a plant growth regulator has been evaluated by using a phytotoxic allelopathic bioassay, where the commercial herbicide Logran is used as internal reference. The evaluation is made based on their effects on germination and growth over several dicotyledon and monocotyledon species. The activity was tested in the range of 1000-0.001 microM. In almost all cases, DHZ was more active than the internal reference at 1000 microM, and its activity fell below the level of the internal reference at 100 microM. These results confirm DHZ as a potent plant growth regulator and a good candidate for the development of new herbicide models.

Fungicidal activity of natural and synthetic sesquiterpene lactone analogs.

Fungicidal activity of 36 natural and synthetic sesquiterpene lactones with guaianolide, trans, trans-germacranolide, cis, cis-germacranolide, melampolide, and eudesmanolide carbon skeletons was evaluated against the phytopathogenic fungi Colletotrichum acutatum, C. fragariae, C. gloeosporioides, Fusarium oxysporum, Botrytis cinerea, and Phomopsis sp. Dose-response data for the active compounds dehydrozaluzanin C, dehydrocostuslactone, 5alpha-hydroxydehydrocostuslacone, costunolide, and zaluzanin C are presented. A new 96-well microbioassay procedure for fast and easy evaluation of antifungal activity was used to compare these compounds with commercial fungicide standards. Some structure-activity conclusions are also presented.

Sunflower sesquiterpene lactone models induce Orobanche cumana seed germination.

Six sunflower sesquiterpene lactone models which share structural features of the lactone rings of strigol and its synthetic analogues, the GR family, with different conformational flexibilities were tested as Orobanche cumana germination stimulants. Among them, parthenolide and 3,5-dihydroxydehydrocostus-lactone significantly increased O. cumana germination, presenting higher activity than GR-24, used as a standard in the germination bioassay. The effect of these two compounds is species-specific, showing no germination stimulant activity on other Orobanche spp. tested (O. crenata, O. ramosa and O. aegyptiaca). Data presented are discussed in terms of a structure-activity relationship.

Novel bioactive breviane spiroditerpenoids from Penicillium brevicompactum Dierckx.

The structures of four new, naturally occurring bioactive spiroditerpenoids, (+)-breviones B, C, D, and E, potential allelopathic agents, have been determined from extracts of semisolid fermented Penicillium brevicompactum Dierckx. The structures display the novel breviane spiroditerpenoid skeleton. Structure elucidation was performed by chemical transformations and by homo- and heteronuclear 2D-NMR spectral data. On the basis of combined studies of the theoretical conformations and NOEDIFF data, their relative stereochemistry is proposed. A mixed biogenesis for this novel family of spiroditerpenoids is tendered. The levels of activity shown by breviones B, C, and E in the etiolated wheat coleoptiles bioassay, especially breviones E (100% inhibition) and C (80% inhibition) both at 10(-4) M, suggest them as lead compounds for new agrochemicals.

Sesquiterpene lactones with potential use as natural herbicide models (I): trans,trans-germacranolides.

A structure-activity study to evaluate the effect of the trans, trans-germacranolide sesquiterpene lactones costunolide, parthenolide, and their 1,10-epoxy and 11,13-dihydro derivatives (in a range of 100-0.001 microM) on the growth and germination of several mono and dicotyledon target species is accomplished. Results are compared with those obtained in the same bioassay with an internal standard, the commercial herbicide Logran, to validate the results with a known active formulation and to compare the results with a commercial product to test their potential use as natural herbicide models. These compounds appear to have a more selective effects on the radicle growth of monocotyledons. Certain factors such as the presence of nucleophile-acceptor groups and their accessibility enhance the inhibitory activity. The levels of radicle inhibition obtained with some compounds on wheat are totally comparable to those of Logran and allow to propose them as lead compounds.

Allelopathic potential of menthofuran monoterpenes fromCalamintha ashei.

A reversed-phase HPLC analysis was used to separate and quantify five menthofuran monoterpenes inCalamintha ashei leaf soaks and washes. (+)-Evodone and desacetylcalaminthone were the major constituents of both soaks and washes. Concentrations of (+)-evodone and desacetylcalaminthone were as high as 0.66 and 0.74 mM, respectively, in leaf soaks. The highest concentration of monoterpenes in leaf washes obtained by misting was 0.021 mM. Aqueous solubilities of the menthofurans were determined to exceed concentrations required for growth inhibition. Bioassays of individualCalamintha monoterpenes demonstrated effects on germination as low as 0.05 mM for (+)-evodone. An equimolar mixture of desacetylcalaminthone and (+)-evodone reducedRudbeckia hirta germination by 17% at a combined concentration of 0.025 mM forLeptochloa dubia. Confirmation of allelopathic effects byCalamintha ashei will require long-term bioassays ofCalamintha menthofurans on the growth of native sandhill species under conditions comparable to the harsh environment of the Florida scrub.

Allelochemicals fromPilocarpus goudotianus leaves.

The effect on germination, shoot, and root growth by bergapten, xanthotoxin, imperatorin, xanthyletin, xanthoxyletin, luvangetin, donatin and alloxanthoxyletol fromPilocarpus goudotianus leaves, onLactuca sativa var. nigra seedlings has been evaluated. A structure-activity correlation is discussed based on the bioassay results. Furanocoumarins appear to be the most active compounds in comparison with pyrano- and simple coumarins. The presence of an oxygenated function at C-8 decreases the germination effect in furano- and pyranocoumarins, while C-5 substituents do not cause significant changes on the activity.

Just how insoluble are monoterpenes?

Prior generalizations about the ecological roles of monoterpenes may be misleading if based on the presumed insolubility of monoterpenes in water. We determined the aqueous solubility of 31 biologically active monoterpenes by gas chromatography. While hydrocarbons were of low solubility (< 35 ppm), oxygenated monoterpenes exhibited solubilities one or two orders of magnitude higher, with ranges of 155-6990 ppm for ketones and of 183-1360 ppm for alcohols. Many monoterpenes are phytotoxic in concentrations under 100 ppm, well below the saturated aqueous concentrations of oxygenated monoterpenes. Therefore, even dilute, unsaturated solutions of monoterpenes, occurring naturally in plant tissues and soil solutions, may act as potent biological inhibitors.

13 C NMR of coumarins. III-Simple coumarins.

13 C NMR data for several simple coumarins are reported. The γsyn effect of a 5-methoxy substituent on the C-4 chemical shift was observed and measured. The effects of an 8-iodo substituent on the chemical shifts of the carbons of the coumarinic skeleton are discussed.