Isolation and characterization of a serine proteinase with thrombin-like activity from the venom of the snake Bothrops asper

A serine proteinase with thrombin-like activity was isolated from the venom of the Central American pit viper Bothrops asper. Isolation was performed by a combination of affinity chromatography on aminobenzamidine-Sepharose and ion-exchange chromatography on DEAE-Sepharose. The enzyme accounts for approximately 0.13% of the venom dry weight and has a molecular mass of 32 kDa as determined by SDS-PAGE, and of 27 kDa as determined by MALDI-TOF mass spectrometry. Its partial amino acid sequence shows high identity with snake venom serine proteinases and a complete identity with a cDNA clone previously sequenced from this species. The N-terminal sequence of the enzyme is VIGGDECNINEHRSLVVLFXSSGFL CAGTLVQDEWVLTAANCDSKNFQ. The enzyme induces clotting of plasma (minimum coagulant dose = 4.1 µg) and fibrinogen (minimum coagulant dose = 4.2 µg) in vitro, and promotes defibrin(ogen)ation in vivo (minimum defibrin(ogen)ating dose = 1.0 µg). In addition, when injected intravenously in mice at doses of 5 and 10 µg, it induces a series of behavioral changes, i.e., loss of the righting reflex, opisthotonus, and intermittent rotations over the long axis of the body, which closely resemble the `gyroxin-like' effect induced by other thrombin-like enzymes from snake venoms.

In vitro evaluation of the cytotoxic and trypanocidal activities of Ampelozizyphus amazonicus (Rhamnaceae)

Ampelozizyphus amazonicus Ducke is a tree commonly found in the Amazon region and an extract of its stem bark is popularly used as an antimalarial and anti-inflammatory agent and as an antidote to snake venom. Ursolic acid; five lupane type triterpenes: betulin, betulinic acid, lupenone, 3ß-hydroxylup-20(29)-ene-27,28-dioic acid, and 2a,3ß-dihydroxylup-20(29)-ene-27,28-dioic acid, and three phytosteroids: stigmasterol, sitosterol and campesterol, have been isolated from stem extracts of A. amazonicus Ducke. Their structures were characterized by spectral data including COSY and HMQC. In an in vitro biological screening of the isolated compounds, 3ß-hydroxylup-20(29)-ene-27,28-dioic acid was cytotoxic against the SKBR-3 human adenocarcinoma cell line (1 to 10 mg/mL), while 2a,3ß-dihydroxylup-20(29)-ene-27,28-dioic acid exhibited cytotoxicity against both SKBR-3 human adenocarcinoma and C-8161 human melanoma tumor cell lines (>0.1 mg/mL). In the present study, different extracts and some fractions of this plant were also investigated for trypanocidal activity due to the presence of pentacyclic triterpenes. The triterpene classes are potent against Trypanosoma cruzi. The bioassays were carried out using blood collected from Swiss albino mice by cardiac puncture during the parasitemic peak (7th day) after infection with the Y strain of T. cruzi. The results obtained showed that A. amazonicus is a potential source of bioactive compounds since its extracts and fractions isolated from it exhibited in vitro parasite lysis against trypomastigote forms of T. cruzi at concentrations >100 µg/mL. Fractions containing mainly betulin, lupenone, 3ß-hydroxylup-20(29)-ene-27,28-dioic acid, and 2a,3ß-dihydroxylup-20(29)-ene-27,28-dioic acid showed more activity than crude extracts.

Randomized clinical trial comparing the efficacy of the vaginal use of metronidazole with a Brazilian pepper tree (Schinus) extract for the treatment of bacterial vaginosis

A 7.4% vaginal extract of the Brazilian pepper tree (Schinus terebinthifolius Raddi) was compared with 0.75% vaginal metronidazole, both manufactured by the Hebron Laboratory, for the treatment of bacterial vaginosis, used at bedtime for 7 nights. The condition was diagnosed using the combined criteria of Amsel and Nugent in two groups of 140 and 137 women, aged between 18 and 40 years. Intention-to-treat analysis was performed. Women were excluded from the study if they presented delayed menstruation, were pregnant, were using or had used any topical or systemic medication, presented any other vaginal infections, presented hymen integrity, or if they reported any history suggestive of acute pelvic inflammatory disease. According to Amsel’s criteria separately, 29 patients (21.2%) treated with the extract and 87 (62.1%) treated with metronidazole were considered to be cured (P < 0.001). According to Nugent’s score separately, 19 women (13.9%) treated with the extract and 79 (56.4%) treated with metronidazole were considered to be cured (P < 0.001). Using the two criteria together, the so-called total cure was observed in 17 women (12.4%) treated with the extract and in 79 women (56.4%) treated with metronidazole (P < 0.001). In conclusion, the cure rate for bacterial vaginosis using a vaginal gel from a pepper tree extract was lower than the rate obtained with metronidazole gel, while side effects were infrequent and non-severe in both groups.

Participation of the GABAergic system in the anesthetic effect of Lippia alba (Mill.) N.E. Brown essential oil

The objective of this study was to identify the possible involvement of the GABAergic system in the anesthetic effect of Lippia alba essential oil (EO). We propose a new animal model using silver catfish (Rhamdia quelen) exposed to an anesthetic bath to study the mechanism of action of EO. To observe the induction and potentiation of the anesthetic effect of EO, juvenile silver catfish (9.30 ± 1.85 g; 10.15 ± 0.95 cm; N = 6) were exposed to various concentrations of L. alba EO in the presence or absence of diazepam [an agonist of high-affinity binding sites for benzodiazepinic (BDZ) sites coupled to the GABA A receptor complex]. In another experiment, fish (N = 6) were initially anesthetized with the EO and then transferred to an anesthetic-free aquarium containing flumazenil (a selective antagonist of binding sites for BDZ coupled to the GABA A receptor complex) or water to assess recovery time from the anesthesia. In this case, flumazenil was used to observe the involvement of the GABA-BDZ receptor in the EO mechanism of action. The results showed that diazepam potentiates the anesthetic effect of EO at all concentrations tested. Fish exposed to diazepam and EO showed faster recovery from anesthesia when flumazenil was added to the recovery bath (12.0 ± 0.3 and 7.2 ± 0.7, respectively) than those exposed to water (9.2 ± 0.2 and 3.5 ± 0.3, respectively). In conclusion, the results demonstrated the involvement of the GABAergic system in the anesthetic effect of L. alba EO on silver catfish.

Bioactive compounds and antioxidant activity in fruits of clone and ungrafted genotypes of yellow mombin tree

Yellow mombin is a fruit tree that grows spontaneously in the Semi-Arid Northeastern Brazil. Its fruits are still extractively exploited. The pulp of yellow mombin fruit stands out regarding the commercial aspect due to the characteristic flavor and aroma felt when consumed in diverse ways. This study aimed to evaluate the presence of bioactive compounds, total extractable polyphenols, and antioxidant activity of yellow mombin fruits (Spondias mombin, L.), from clone and ungrafted genotypes. The fruits were harvested at commercial maturity from twelve yellow mombin tree genotypes from an experimental orchard located at the municipality of Joao Pessoa, Paraíba, Brazil, and evaluated for chlorophyll, carotenoids, yellow flavonoids, total extractable polyphenols, and antioxidant activity, which was measured by the β-carotene/linoleic acid method. The antioxidant activity showed a percentage of inhibition of oxidation higher than 75% for all genotypes evaluated at the time of 120 minutes. The fruits from clone genotypes showed a higher percentage of antioxidant activity.

Green and brown propolis: efficient natural biocides for the control of bacterial contamination of alcoholic fermentation of distilled beverage

This study aimed to evaluate the efficiency of natural biocides, brown and green propolis, for the control of bacterial contamination in the production of sugarcane spirit. The treatments consisted of brown and green propolis extracts, ampicillin, and a control and were assessed at the beginning and end of harvest season in ten fermentation cycles. In the microbiological analyses, the lactic acid bacteria were quantified in the inoculum before and after the treatment with biocides, and the viability of yeast cells during fermentation was evaluated. The levels of acids, glycerol, total residual reducing sugars, and ethanol were analyzed for the wine resulting from each fermentation cycle. A reduction in the number of bacterial contaminants in the inoculum in the treatments with the natural biocides was observed, but it did not affect the viability of yeast cells. The control of the contaminants led to the production of higher levels of ethanol and reduced acidity in the wine produced. The results of the use of brown and green propolis to control the growth microorganisms in the fermentation of sugarcane spirit can be of great importance for using alternative strategies to synthetic antibacterials in fermentation processes including other distilled beverage or spirits.

Physicochemical and antioxidative properties of black, brown and red rice varieties of northern Thailand

Rice, the seed of Oryza species, is the major cereal crop in most of the developing countries. Nearly 95% of global rice production is done in Asian countries, and about half of the world’s population consumes it. Some speciality rices are not commonly consumed. Colored rice is one of such variety. In these varieties, high amounts of anthocyanin pigment are deposited in the rice coat to form its black (also known as purple), brown and red colors. Minimum studies are there to explain the properties of these rice varieties of Thailand. Thus, the current study was aimed to assess the physicochemical and antioxidative properties of three rice varieties (Chiang Mai Black rice, Mali Red rice and Suphanburi-1 Brown rice) of different cultivars of northern Thailand. Rice bran extracts of these three cultivars were prepared with different solvents (polar and non-polar) for the evaluation of total phytochemical content and anti-oxidant free-radical-scavenging properties. Chiang Mai Black rice contained higher concentration of phenolic acid, flavonoids, and anthocyanins (Cyanidin 3-glucoside, peonidin 3-glucoside, cyanidin chloride). Chiang Mai Black rice is richer in free-radical-scavenging compounds and activities than the other tested varieties. Polar extractions of rice bran are high in anti-oxidative compounds and activities than non-polar extractions.

Avaliação de métodos de extração do arilo e tratamento com ethephon em sementes de Passiflora giberti N.E. Brown pelos testes de germinação e de tetrazólio

Com o objetivo de avaliar o efeito de dois métodos de extração de arilo e cinco concentrações de ethephon em sementes de Passiflora giberti N.E.Brown, empregou-se o teste de germinação e de tetrazólio para a realização deste experimento, no Laboratório de Tecnologia de Sementes da Universidade Estadual do Oeste do Paraná - UNIOESTE, campus de Marechal Cândido Rondon, no período de 01/06 a 03/07/01. O delineamento foi o inteiramente casualizado com cinco repetições de 25 sementes por parcela. Os frutos foram coletados em Marechal Cândido Rondon e os tratamentos foram constituídos pela combinação de dois tipos de extração do arilo e embebição das sementes em cinco concentrações de ethephon por cinco horas. A semeadura para o teste de germinação foi realizada em rolos de papel germitest mantidos em câmara de germinação com temperatura alternada (25-30ºC). O teste de tetrazólio foi feito no lote de sementes antes da instalação do experimento e no final do teste de germinação, 30 dias após a semeadura. Os resultados demonstram que ocorerram interações significativas entre os métodos de extração do arilo e as concentrações de ethephon para a vitalidade de sementes. Pode-se observar que somente na extração por fermentação com a concentração de 600mg.l-1 verificou-se diminuição significativa na vitalidade das sementes, diferindo dos demais tratamentos. Na extração do arilo por fricção com pano todas as concentrações demonstraram-se semelhantes. Os métodos de extração do arilo não afetaram a vitalidade das sementes de Passiflora giberti N.E. Brown com presença de dormência, exceto quando se empregou 600mg.l-1 de ethephon. Tratamentos com ethephon não foram adequados para proporcionar a germinação das sementes.

Quantitative characteristics of fruits and seeds of Pouteria pachycarpa Pires - Sapotaceae

Pouteria pachycarpa is a tree species, found in the Brazilian Amazon and Bolivia whose wood has been exploited from the native forest. The present research describes the quantitative characteristics of fruits and seeds and quantifies the seed germination of this species. The fruit and seed color were characterized and measurements taken of the mass, length, diameter and number of seeds per fruit, the seed length, width and thickness, the germination percentage, abnormal seedlings and dead seeds. Sowing was carried out on a substrate containing sand and sawdust (1:1), in four replications of 50 seeds. The predominant fruit and seed colors were vivid yellowish orange (9YR) and dark grayish brown (6YR), respectively. Fruit mass, length and diameter ranged from 37.7 to 192.4g, 41.3 to 87.3mm and 39.7 to 71.7mm, respectively. Fruits had from two to seven seeds, and 42.6% were damaged by insects. Seed length, width and thickness ranged from 22.4 to 35.2mm, 9.7 to 15.5mm and 5.5 to 10.8mm, respectively. Seedling emergence began 18 days after sowing. Maximum germination, 86%, was recorded 33 days after sowing. The germination curve was sigmoid, similar to the majority of species. The percentage of abnormal seedlings and dead seeds were 3% and 11%, respectively. Both fruits and seeds show great variation in quantitative characteristics and the germination is slow and non-uniform.

Trophic cascades in boreal landscapes: top predator protection on tree seedlings and forest grouse

Changes in the abundance of top predators have brought about notable, cascading effects in ecosystems around the world. In this thesis, I examined several potential trophic cascades in boreal ecosystems, and their separate interspecific interactions. The main aim of the thesis was to investigate whether predators in the boreal forests have direct or indirect cascading effects on the lower trophic levels. First, I compared the browsing effects of different mammalian herbivores by excluding varying combinations of voles, hares and cervids from accessing the seedlings of silver birch (Betula pendula), Scots pine (Pinus sylvestris) and Norway spruce (Picea abies). Additionally, I studied the effect of simulated predation risk on vole browsing by using auditory cues of owls. Moving upwards on the trophic levels, I examined the intraguild interactions between the golden eagle (Aquila chrysaetos), and its mesopredator prey, the red fox (Vulpes vulpes) and the pine marten (Martes martes). To look at an entire potential trophic cascade, I further studied the combined impacts of eagles and mesopredators on the black grouse (Tetrao tetrix) and the hazel grouse (Tetrastes bonasia), predicting that the shared forest grouse prey would benefit from eagle presence. From the tree species studied, birch appears to be the most palatable one for the mammalian herbivores. I observed growth reductions in the presences of cervids and low survival associated with hares and voles, which suggests that they all weaken regeneration in birch stands. Furthermore, the simulated owl predation risk appeared to reduce vole browsing on birches in late summer, although the preferred grass forage is then old and less palatable. Browsing by voles and hares had a negative effect on the condition and survival of Scots pine, but in contrast, the impact of mammalian herbivores on spruce was found to be small, at least when more preferred food is available. I observed that the presence of golden eagles had a negative effect on the abundance of adult black grouse but a positive, protective effect on the proportion of juveniles in both black grouse and hazel grouse. Yet, this positive effect was not dependent on the abundance foxes or martens, nor did eagles seem to effectively decrease the abundance of these mesopredators. Conversely, the protection effect on grouse could arise from fear effects and also be mediated by other mesopredators. The results of this thesis provide important new information about trophic interactions in the boreal food webs. They highlight how different groups of mammalian herbivores vary in their effects on the growth and condition of different tree seedlings. Lowered cervid abundances could improve birch regeneration, which indirectly supports the idea that the key predators of cervids could cause cascading effects also in Fennoscandian forests. Owls seem to reduce vole browsing through an intimidation effect, which is a novel result of the cascading effects of owl vocalisation and could even have applications for protecting birch seedlings. In the third cascade examined in this thesis, I found the golden eagle to have a protective effect on the reproducing forest grouse, but it remains unclear through which smaller predators this effect is mediated. Overall, the results of this thesis further support the idea that there are cascading effects in the forests of Northern Europe, and that they are triggered by both direct and non‐lethal effects of predation.

Between Dugin’s Eurasianism and Mahan’s Sea Power Concept: Russia’s Geopolitical Discourse within Foucault’s ‘Discursive Tree’ before and After the Incorporation of the Crimea

The fall of 2013 could be characterized as a crossroad in the geopolitics of Eastern Europe, namely Ukraine. Two rivalry geopolitical projects have been developing throughout the post-Cold War years, and it seems that they reached a collision point in Ukraine; a country whose authorities have been for long switching sides between the European Union and the Russian Federation in their foreign policy commitments. The refusal/postponing to sign the Association Agreement with Brussels, an expected event by a large category of the Ukrainian society, by Yanukovich’s government led to the outset of the latter; and brought a pro-Western, anti-Russian government in Kyiv. It seems that Ukraine, after those events, has embarked definitively on the path of integration into the West (European Union and possibly NATO). The Russian Federation, who has been throughout Putin’s years engaged into the re-integration of post-Soviet space, reacted to these developments in an assertive manner by violating borders, agreements and the territorial integrity of Ukraine. Thus, the incorporation of the Crimea into the Russian Federation is the first in its kind in the post-Soviet space, despite the existence of various other conflicts that broke out in the region after the Soviet Union broke up. I will investigate in this thesis the nature of what will be labelled, in this work, the Crimean issue. I argue that the incorporation of the Crimean peninsula into the Russian Federation marks a new era in Russian geopolitical thinking that shapes, to a far extent, Russian foreign policy. Discourse analysis will be the methodological basis for this study, with a special focus on Michel Foucault’s Archaeology of Knowledge. The innovation that this research brings is the fact that it discusses Russian geopolitical discourse within the scope of Foucault’s ‘discursive tree’, with a reference to the Crimean issue. A wide range of primary sources will be consulted in this study such as presidential addresses to the Federal Assembly (2000-2014), Foreign Policy Concepts of the Russian Federation (2000, 2008), Russian maritime doctrines, as wells as Dugin’s Osnovy Geopolitiki (Foundations of Geopolitics), Mahan’s (The Influence of Sea Power Upon History, 1660–1783) and other Eurasianism related literature.

The tree we heard fall : analyzing the societal processes following focusing events

Fokuserande händelser är plötsliga, ovanliga och för med sig negativa konsekvenser för ett stort antal människor. Det handlar om det vi i vardagligt tal kallar kriser och katastrofer. En intensiv medierapportering i kombination med oroade medborgare gör att kriser och katastrofer är viktiga samhällspåfrestningar som beslutsfattare måste hantera. Syftet med avhandlingen är att öka förståelsen för hur fokuserande händelser påverkar samhälleliga processer. Avhandlingens två övergripande forskningsfrågor tar fasta på jämförelsen av olika typer av fokuserande händelser. För det första, skillnader i hur naturkatastrofer och katastrofer som tillkommit genom mänskligt handlade påverkar samhället, och för det andra, hur händelser med olika stark fokuseringskraft påverkar samhället. Medborgarreaktioner studeras i en experimentell laboratoriestudie, den mediala rapporteringen analyseras genom kvantitativ innehållsanalys av texter och bilder, och på den parlamentariska arenan analyseras riksdagsdebatten genom kvantitativ innehållsanalys. De temaområden som identifieras i den teoretiska referensramen ligger till grund för analysen och tar fasta på de känslor som händelsen ger upphov till, hur händelsen kontextualiseras (framing), skuldbeläggning samt möjliga lösningar som förs fram. Resultatet från studierna visar att skillnader mellan olika typer av händelser syns tydligast hos medborgare, uttryckt genom starkare emotionella reaktioner för händelser med stark fokusering, och på den mediala arenan, genom mer utrymme för dessa händelser. Då kriser och katastrofer med olika stark fokuseringskraft når riksdagen får de däremot ett relativt likadant mottagande. I jämförelsen mellan naturkatastrofer och människoskapade händelser visar resultaten att naturkatastrofer ofta diskuteras som specifika händelser, medan människoskapade händelser placeras i en bredare samhällelig kontext och diskuteras under längre tid. Avhandlingen bidrar till diskussionen om hur våra känslor och vår kognition samspelar i krissituationer. De omedvetna signaler som vår kropp ger oss gällande hur vi ska reagera på en farlig situation kan påverka om vi är rädda, oroliga eller till och med positivt inställda. I ljuset av de tre studierna är det tydligt att de emotionella reaktionerna minskar ju högre upp i den politiska processen vi kommer. Trots att medborgare och medier reagerar starkt är de emotionella reaktionerna på den politiska arenan relativt milda och skuldbeläggningen näst intill icke-existerande.