Effects of the macroalga Asparagopsis taxiformis and oregano leaves on methane emission, rumen fermentation, and lactational performance of dairy cows.

Asparagopsis taxiformis (AT) is a source of multiple halogenated compounds and, in a limited number of studies, has been shown to decrease enteric CH4 emission in vitro and in vivo. Similarly, oregano has been suggested as a potential CH4 mitigating agent. This study consisted of 2 in vitro and 2 in vivo experiments. Experiment (Exp.) 1 was aimed at establishing the effect of AT on CH4 emission in vitro. Two experiments (Exp. 2 and 3) with lactating dairy cows were conducted to determine the antimethanogenic effect of AT and oregano (Exp. 3) in vivo. Another experiment (Exp. 4) was designed to investigate stability of bromoform (CHBr3) in AT over time. In Exp. 3, 20 Holstein cows were used in a replicated 4 × 4 Latin square design with four 28-d periods. Treatments were basal diet (control) or basal diet supplemented with (dry matter basis) 0.25% AT (LowAT), 0.50% AT (HighAT), or 1.77% oregano (Origanum vulgare L.) leaves. Enteric gas emissions were measured using the GreenFeed system (C-Lock Inc., Rapid City, SD), and rumen samples were collected for fermentation analysis using the ororuminal technique. In Exp.1 (in vitro), relative to the control, AT (at 1% dry matter basis, inclusion rate) decreased CH4 yield by 98%. In Exp. 3, HighAT decreased average daily CH4 emission and CH4 yield by 65% and 55%, respectively, in experimental periods 1 and 2, but had no effect in periods 3 and 4. The differential response to AT among experimental periods was likely a result of a decrease in CHBr3 concentration in AT over time, as observed in Exp. 4 (up to 84% decrease in 4 mo of storage). In Exp. 3, H2 emission was increased by AT and, as expected, the proportion of acetate in the total volatile fatty acids in the rumen was decreased and those of propionate and butyrate were increased by HighAT compared with the control. Compared with the control, HighAT decreased dry matter intake, milk yield, and energy-corrected milk yield in Exp. 3. Milk composition was not affected by treatment, except lactose percentage and yield were decreased by HighAT. Concentrations of iodine and bromide in milk were increased by HighAT compared with the control. Milk CHBr3 concentration and its organoleptic characteristics were not different between control and HighAT. Oregano had no effect on CH4 emission or lactational performance of the cows in Exp. 3. Overall, AT included at 0.50% in the ration of dairy cows can have a large mitigation effect on enteric CH4 emission, but dry matter intake and milk production may also decrease. There was a marked decrease in the CH4 mitigation potential of AT in the second half of Exp. 3, likely resulting from CHBr3 decay over time.

Antifungal activities of cytochalasins produced by Diaporthe miriciae, an endophytic fungus associated with tropical medicinal plants.

In the present study, we evaluated the antifungal potential of cytochalasins produced by Diaporthe taxa against phytopathogenic fungi. Using molecular methods, seven endophytic fungal strains from the medicinal plants Copaifera pubiflora and Melocactus ernestii were identified as Diaporthe miriciae, while two isolates were identified to the genus level (Diaporthe sp.). All crude extracts of Diaporthe species produced via solid-state fermentation were evaluated by 1H NMR analyses. Crude extracts of the isolates D. miriciae UFMGCB 6350, 7719, 7646, 7653, 7701, 7772, and 7770 and Diaporthe sp. UFMGCB 7696 and 7720 were demonstrated to produce highly functionalized compounds. The extracts of D. miriciae UFMGCB 7719 and 6350 were selected as representative Diaporthe samples and subjected to bioassay-directed fractionation to isolate cytochalasins H and J. Cytochalasins H and J were evaluated for activities against the fungal plant pathogens Colletotrichum fragariae, Colletotrichum gloeosporioides, Colletotrichum acutatum, Botrytis cinerea, Fusarium oxysporum, Phomopsis obscurans, and Phomopsis viticola using microdilution broth assays. Cytochalasins H and J exhibited the most potent activities against the Phomopsis species tested. Our results showed that Diaporthe species were potential producers of different cytochalasins, which exhibit potential for controlling fungal diseases in planta and (or) maintaining antagonism.

Multiple activities of insect repellents on odorant receptors in mosquitoes.

Several lines of evidence suggest that insect repellent molecules reduce mosquito-host contacts by interacting with odorants and odorant receptors (ORs), thereby ultimately affecting olfactory-driven behaviours. We describe the molecular effects of 10 insect repellents and a pyrethroid insecticide with known repellent activity on two highly specific Aedes aegypti (Diptera: Culicidae) ORs, AaOR2 + AaOR7 and AaOR8 + AaOR7, exquisitely sensitive to key mosquito attractants indole and (R)-(-)-1-octen-3-ol, expressed in oocytes of Xenopus (Anura: Pipidae). Our study demonstrates that insect repellents can both inhibit odorant-evoked currents mediated by ORs and independently elicit currents in the absence of odorants. All of the repellents had effects on one or both ORs; most of these compounds were selective inhibitors and showed a high degree of specificity in their capacity to activate the two ORs. These results show that a range of insect repellents belonging to structurally diverse chemical classes modulate the function of mosquito ORs through multiple molecular mechanisms.

Repellency of callicarpenal and intermedeol against workers of imported fire ants (Hymenoptera: Formicidae).

Callicarpenal and intermedeol are two insect-repellent terpenoids isolated from leaves of American beautyberry (Callicarpa americana L.; Verbenaceae) and Japanese beautyberry (Callicarpa japonica Thunb.). The repellency of these two terpenoids against workers of red imported fire ants, Solenopsis invicta Buren, black imported fire ants, Solenopsis richteri Forel, and a hybrid of these two species was evaluated using digging bioassays. In a multiple choice digging bioassay using two colonies from each species and their hybrid, callicarpenal showed significant repellency at concentration as low as 50 ppm against both red imported fire ant colonies and 6.25 ppm against all black imported fire ant and hybrid colonies. Intermedeol showed significant repellency at concentration as low as 1.50 ppm against both red imported fire ant colonies and 6.25 ppm against all black imported fire ant and hybrid colonies. In total, 15 colonies, five colonies from each species and the hybrid, were tested on callicarpenal and intermedeol at 50 ppm in a two-choice digging bioassay. Both callicarpenal and intermedeol showed repellency against all colonies, and intermedeol showed significantly greater repellency than callicarpenal against both species and their hybrid.

Isolation and identification of antifungal and antialgal alkaloids from Haplophyllum sieversii.

Bioassay-guided fractionation of the hexane/ethyl acetate/water (H/EtOAc/H2O) crude extract of the aerial parts of Haplophyllum sieversii was performed because of preliminary screening data that indicated the presence of growth inhibitory components against Colletotrichum fragariae, Colletotrichum gloeosporioides, and Colletotrichum acutatum. Fractionation was directed using bioautographical methods resulting in the isolation of the bioactive alkaloids flindersine, anhydroevoxine, haplamine, and a lignan eudesmin. These four compounds were evaluated for activity against C. fragariae, C. gloeosporioides, C. acutatum, Botrytis cinerea, Fusarium oxysporum, and Phomopsis obscurans in a dose-response growth-inhibitory bioassay at 50.0, 100.0, and 150.0 microM. Of the four compounds tested, flindersine demonstrated the highest level of antifungal activity. Additionally, flindersine, eudesmin, and haplamine were screened against the freshwater phytoplanktons Oscillatoria perornata, Oscillatoria agardhii, Selenastrum capricornutum, and Pseudanabaena sp. (strain LW397). Haplamine demonstrated selective inhibition against the odor-producing cyanobacterium O. perornata compared to the activity against the green alga S. capricornutum, with lowest observed effect concentration values of 1.0 and 10.0 microM, respectively.

Isolation and identification of mosquito bite deterrent terpenoids from leaves of American (Callicarpa americana) and Japanese (Callicarpa japonica) beautyberry.

Essential oil extracts from Callicarpa americana and Callicarpa japonica were investigated. Bioassay-guided fractionation of C. americana extracts using the yellow fever mosquito, Aedes aegypti, led to the isolation of alpha-humulene, humulene epoxide II, and intermedeol and a newly isolated terpenoid (callicarpenal). Similar work involving C. japonica resulted in the isolation of an additional compound, spathulenol, as well as the four compounds isolated from C. americana. Structure elucidation was performed on all isolated compounds using a combination of gas chromatography-mass spectrometry-electron ionization, high-resolution liquid chromatography-MS-electrospray ionization, and one- and two-dimensional NMR experiments. Heretofore, 13,14,15,16-tetranorclerodane, callicarpenal, has never been identified from natural sources. Complete (1)H and (13)C NMR assignment data are provided for this compound. In bite deterrent studies, spathulenol, intermedeol, and callicarpenal showed significant repellent activity against A. aegypti and Anopheles stephensi.

Bioactive crude plant seed extracts from the NCAUR oilseed repository.

Over four-hundred crude extracts from 202 plant species distributed among 131 plant families were evaluated for their bioactivity against brine shrimp (Artemia salina). Activity was determined for both the organic (CH2Cl2:MeOH) and aqueous extracts against A. salina in a 96 well-plate assay. Of the greater than four-hundred extracts tested, 21 organic and 6 aqueous extracts demonstrated potent cytotoxic activity (LC50 = < 100 microg/ml). Three of these organic extracts (Crateva religiosa, Diospyros dichrophylla, and Olax subscorpioidea) were chosen for chemical investigations due to their high activity and a lack of prior investigations. Chemical analysis of these extracts resulted in the isolation of oleanolic acid (1) and 4-epi-hederagenin (2) from C. religiosa, isodiospyrin (3) from D. dichrophylla, and santalbic acid (4) from O. subscorpioidea.

Antimycobacterial plant terpenoids.

Abstract. Tuberculosis (TB), mainly caused by Mycobacterium tuberculosis, is the leading killer among all infectious diseases worldwide and is responsible for more than two million deaths annually. For over thirty years no antitubercular agents with new mechanisms of action have been developed. The recent increase in the number of multi-drug resistant clinical isolates of M. tuberculosis has created an urgent need for the discovery and development of new antituberculosis leads. This review covers recent reports on plant-derived terpenoids that have demonstrated moderate to high activity in in vitro bioassays against M. tuberculosis. In this review, mono-, sesqui-, di- and triterpenes, and sterols, their structural analogs and semisynthetic derivatives will be discussed, with particular emphasis on the structural features essential for antimycobacterial activity.

Chondropsin D, a new 37-membered-ring macrolide lactam from the marine sponge Chondropsis species.

Chondropsin D (2), a new 37-membered-ring macrolide lactam, was isolated as a minor constituent of an aqueous extract of the marine sponge Chondropsis sp. This sponge sample had previously been the source of chondropsins A (1) and B, two novel polyketide-derived macrolides with potent cytotoxic activity. The structure of 2 was initially deduced from analysis of spectral data. This assignment was supported by the observation that chondropsin A (1), which contains a 35-membered macrocyclic ring, could be converted to chondropsin D (2) by a base-catalyzed intramolecular transesterification reaction. Rearrangement of the methylated derivative of chondropsin A (3) to the corresponding methylated analogue of chondropsin D (4) confirmed the structure of 2.

Phase I dose escalation study of topotecan combined with alternating schedules of paclitaxel and carboplatin in advanced solid tumors.

BACKGROUND: Combining topotecan with other cytotoxics has been problematic due to marrow suppression. A phase I trial was initiated to identify the optimal sequence and maximum-tolerated dose of topotecan in combination with paclitaxel and carboplatin. PATIENTS AND METHODS: Patients with advanced cancer and performance status ECOG < or = 2. The starting dose was paclitaxel 175 mg/m2 day 1, carboplatin AUC 6.0 day 1, and topotecan 0.5 mg/m2 daily day 1-5 (early sequence). The next course of paclitaxel and carboplatin administration was delayed to day 5 (late sequence). Treatment was repeated every three weeks. After determining maximum-tolerated dose without cytokines, granulocyte colony-stimulating factor (G-CSF) was added and further dose escalation was pursued. RESULTS: Fifty-one patients were entered; men: women ratio 30:21. Dose-limiting toxicity (DLT) for the early sequence was neutropenia at doses paclitaxel mg/m2/carboplatin AUC 5/topotecan mg/m2 (PCT) 175/5/0.75 for four to five days. DLT for the late sequence was neutropenia at PCT doses of 175/5/ 1.0 for four days. G-CSF 5 microg/kg subcutaneously starting day 6 permitted further topotecan dose escalation. After adding G-CSF, late sequence DLT was neutropenia at doses 175/5/1.25 for four days. Forty-six patients were evaluable for response and of those, there were thirteen partial responses. CONCLUSIONS: The late sequence resulted in less toxicity and was better tolerated. The early sequence maximum-tolerated dose (MTD) was 175/6/0.5 for five days. The late sequence MTD was PCT 175/5/0.75 for five days. The late sequence MTD with G-CSF was 175/5/1.0 for four days. The recommended phase II PCT dose is the late sequence 175/5/1.0 for four days with G-CSF.

Stolonic acids A and B, new cytotoxic cyclic peroxides from an Indian Ocean ascidian Stolonica species.

Two new 3,6-epidioxy-7,10-tetrahydrofurano C(26) unsaturated fatty acids, stolonic acids A (1) and B (2), were isolated from a previously undescribed ascidian species, Stolonica sp. collected off the Maldive Islands in the Indian Ocean. The structures and relative stereochemistry of 1 and 2 were determined using conventional spectroscopic methods. Both compounds exhibited antiproliferative activity against selected human melanoma and ovarian tumor cell lines, with IC(50) values of approximately 0.05-0.1 microg/mL.

Antimycobacterial eudesmanolides from Inula helenium and Rudbeckia subtomentosa.

In a bioassay guided search for antimycobacterial compounds from higher plants, the root extracts of Elecampane (Inula helenium L.; Asteraceae) and Sweet Coneflower (Rudbeckia subtomentosa Pursh.; Asteraceae) were chemically investigated for their active constituents. Chromatographic fractions of root extracts of l. helenium, which exhibited significant activity against Mycobacterium tuberculosis, provided the known eudesmanolides alantolactone, isoalantolactone, and 11 alpha H, 13-dihydroisoalantolactone. Peracid epoxidation of alantolactone and isoalantolactone provided 5 alpha-epoxyalantolactone and 4(15) alpha-epoxyisoalantolactone, respectively and oxidation of alantolactone with OsO4 gave 11,13-dihydroxyalantolactone. Active fractions from R subtomentosa contained the known alloalantolactone and 3-oxoalloalantolactone. The structures of the above compounds were established by spectroscopic methods including 1D and 2D NMR techniques as well as spectral comparison with previously reported data. The molecular structure of 5 alpha-epoxyalantolactone was determined by single crystal X-ray diffraction. Eleven natural and semisynthetic eudesmanolides were tested in a radiorespirometric bioassay for activity against M. tuberculosis. 5 alpha-Epoxyalantolactone and encelin from Montanoa speciosa showed minimum inhibitory concentrations (MICs) of 8 and 16 micrograms ml-1, respectively. Alantolactone, isoalantolactone and its 4 alpha, 15-epoxide, 1,2-dehydro-3-epi-isotelekin and alloalantolactone gave MICs of 32 micrograms ml-1. All other compounds showed MIC values of 128 micrograms ml-1 or higher.

Antimycobacterial triterpenes from Melia volkensii.

In a bioassay-guided search for antimycobacterial compounds from higher plants, we have chemically investigated methanolic extracts of seeds of Melia volkensii. Chromatographic fractions provided two new euphane (20R)-type triterpenoids. The structures of the new compounds, 12beta-hydroxykulactone (1) and 6beta-hydroxykulactone (2), were elucidated by 1D and 2D NMR (13C, 1H, 1H-1H COSY, HMQC, HMBC, and NOESY spectra) and FABMS studies and shown to be hydroxyl derivatives of kulactone (3). Also isolated was the known kulonate (4). In a radiorespirometric bioassay against Mycobacterium tuberculosis, compounds 1, 2, and 4 exhibited minimum inhibitory concentrations of 16, 4, and 16 microg/mL, respectively.

Antimycobacterial ergosterol-5,8-endoperoxide from Ajuga remota.

In a bioassay-guided search for antimycobacterial natural products from higher plants, we have chemically investigated the methanol extract of aerial parts of Ajuga remota Benth. (Labiatae) for its active constituent(s). Bioactive chromatographic fractions of the crude extract provided the known triterpene ergosterol-5,8-endoperoxide plus the diterpenes clerodin, ajugarin-I, and ajugarin-II, which had been previously isolated from A. remota. This is the first report on the isolation of ergosterol-5,8-endoperoxide from this plant. The above compounds were tested in a radiorespirometric bioassay for activity against Mycobacterium tuberculosis. Ergosterol-5,8-endoperoxide showed a minimum inhibitory concentration (MIC) of 1 microgram/ml, while ergosterol-5,8-endoperoxide acetate, ergosterol, and ergosta-5,7,9(11),22-tetraen-3 beta-ol gave MICs of 8 micrograms/ml, > 128 micrograms/ml, and 128 micrograms/ml, respectively. Clerodin, ajugarin-I, and ajugarin-II were inactive with MICs of > 128 micrograms/ml.

Antimycobacterial matricaria esters and lactones from Astereae species.

Six matricaria esters (MEs) and two matricaria lactones (MLs), isolated from members of the tribe Astereae (Asteraceae), were tested against Mycobacterium tuberculosis and M. avium, using a radiorespirometric bioassay. (2Z,8Z)-ME and (2E-8Z)-ME gave minimum inhibitory concentrations (MICs) of 50 micrograms ml-1 against M. tuberculosis and respective MICs of 25 and 50 micrograms ml-1 against M. avium. The (4Z,8Z)-ML, (2Z)-8-dehydro-ME and (2Z,8Z)-10-angeloyloxy-(2Z,8Z)-ME showed respective MICs of 12.5, 25, 25 micrograms ml-1 against M. tuberculosis and MICs of 50, 25, 25 micrograms ml-1 against M. avium, respectively. The MICs of (2Z,8Z)-10-tigloyloxy-ME and (2E,8Z)-10-angeloyloxy-ME and (4E,8Z)-ML ranged from 50 to > 100 micrograms ml-1 against both pathogenic mycobacteria.

Antimycobacterial activities of dehydrocostus lactone and its oxidation products.

In an attempt to study the structural dependence of antimycobacterial activity of the guaianolide dehydrocostus lactone and its derivatives, m-chloroperoxybenzoic acid oxidations of dehydrocostus lactone (1a) were performed. Three new monoepoxides, one previously synthesized diepoxide, and two new diepoxides were obtained. Two of the monoepoxides are C-10 epimers (3a, 3b), while the 4(15)-monoepoxide (2) has the 4alpha-O-configuration. The known diepoxide (4a) contains a C-10 alpha-epoxide and a beta-epoxide at C-4. The diepoxides 4b and 4c, each with a C-4 alpha-epoxy group, differ in the configuration of the epoxide ring at C-10. Allylic oxidation of dehydrocostus lactone (1a) with selenium dioxide/tert-butyl hydroperoxide afforded the known 3-epizaluzanin C (1b). The relative configurations of compounds 1b-4c were established by 1D and 2D NMR techniques (1H, 13C, COSY, NOESY, HMQC, and HMBC) as well as comparison with literature data. The molecular structures of lactones 1b, 4a, and 4c were determined by single-crystal X-ray diffraction. In radiorespirometric bioassays against Mycobacterium tuberculosis and Mycobacterium avium, dehydrocostus lactone (1a) exhibited minimum inhibitory concentrations of 2 and 16 microgram/mL, respectively. In contrast, its monoepoxides (2, 3a, and 3b) and diepoxides (4a-c), as well as its hydrogenated derivatives and other analogues (1b, 1c, 5, and 6), showed significantly lower activities against M. tuberculosis.

Antimycobacterial evaluation of germacranolides.

The minimum inhibitory concentrations (MIC) against Mycobacterium tuberculosis and M. avium of parthenolide, costunolide, 1 (10)-epoxycostunolide and other germacranolide-type sesquiterpene lactones and derivatives were determined by use of a radiorespirometric bioassay. Structure-activity relationship studies with natural and semisynthetic sesquiterpene lactones suggested that the alpha-methylene-gamma-lactone moiety is an essential, but not sufficient, structural requirement for significant in vitro activity against M. tuberculosis and M. avium.

Antimycobacterial activity of (E)-phytol and derivatives: a preliminary structure-activity study.

The crude methanol extract of the Kenyan shrub Leucas volkensii Gürke (Labiatae) displayed in a radiorespirometric bioassay antimycobacterial activity against Mycobacterium tuberculosis. Bioassay-guided fractionation of the crude extract led to the identification of (E)-phytol as the principal active component with a minimum inhibitory concentration (MIC) of 2 micrograms/ml, a value also observed for (3R,S,7R,11R)-phytanol, (Z)-phytol, and a commercially available 2:1 mixture of (E)- and (Z)-phytol. The derivatives (E)-phytol acetate, a mixture of the (2S,3S)- and (2R,3R)-isomers of (E)-phytol epoxide and (3R,S,7R,11R)-phytanic acid displayed lower activities with MICs of 8, 16, and > 128 micrograms/ml, respectively. Geraniol and farnesol, displayed MICs of 64 and 8 micrograms/ml, respectively. The activities of (E)-phytol, (Z)-phytol and (3R,S,7R,11R)-phytanol were found to be in the same range as ethambutol, a clinically useful drug with an MIC in the range 0.95-3.8 micrograms/ml.

Antimycobacterial crude plant extracts from South, Central, and North America.

Two-hundred and thirty crude organic extracts from 118 plant species distributed among 10 families were evaluated for anti-mycobacterial activity. Activity was determined against Mycobacterium tuberculosis H(37)Rv and Mycobacterium avium using the BACTEC 460 radiorespirometric assay. At 100 µg/ml, twenty-four and ten of the extracts caused more than 95% inhibition of growth of M. tuberculosis and M. avium, respectively. The most active plant species (100% inhibition) were Borrichia frutescens, Solidago arguta, and Inula helenium against M. tuberculosis, Euthamia leptocephala against M. avium, and Erigeron strigosus and Magnolia acuminata against both mycobacteria.

Antimycobacterial cycloartanes from Borrichia frutescens.

In a bioassay-guided search for antimycobacterial compounds from higher plants of the southeastern United States, we have chemically investigated the sea daisy (Borrichia frutescens) from coastal marshes of Louisiana for their active constituents. Bioactive chromatographic fractions provided two new triterpenes, (24R)-24,25-epoxycycloartan-3-one (1) and (23R)-3-oxolanosta-8,24-dien-23-ol (4), and (3 alpha H, 24R)-24,25-epoxycycloartan-3-ol (3a). Compound 3a had been previously isolated as a mixture of C-24 epimers. The structures of 1, 3a, and 4 were established by spectroscopic methods and chemical transformations, and the molecular structures of 1 and 4 were determined by single-crystal X-ray diffraction. In a radiorespirometric bioassay against Mycobacterium tuberculosis, the epoxycycloartanes 1 and 3a exhibited minimum inhibitory concentrations of 8 micrograms/mL. In contrast, the lanostadiene-type triterpene 4 showed no significant inhibition at 128 micrograms/mL, as did the acetate 3b. Cytotoxicity for Vero cells gave IC50 values of 71.8, 39.8, and 103.6 micrograms/mL for triterpenes 1, 3a, and 4, respectively.