Involvement of NO in the inhibitory effect of Calotropis procera latex protein fractions on leukocyte rolling, adhesion and infiltration in rat peritonitis model

Aim of the study: The latex of Calotropis procera has been used in the traditional medicinal system for the treatment of leprosy, ulcers, tumors, piles and diseases of liver, spleen, abdomen and toothache. it comprises of a non-dialyzable protein fraction (LP) that exhibits anti-inflammatory properties and a dialyzable fraction (DF) exhibiting pro-inflammatory properties. The present study was carried out to evaluate the effect of LP sub-fractions on neutrophil functions and nociception in rodent models and to elucidate the mediatory role of nitric oxide (NO). Material and methods: The LP was subjected to ion exchange chromatography and the effect of its three sub-fractions (LP(PI), LP(PII), and LP(PIII)) thus obtained was evaluated on leukocyte functions in the rat peritonitis model and on nociception in the mouse model. Results: LP sub-fractions exhibit distinct protein profile and produce a significant decrease in the carrageenan and DF induced neutrophil influx and exhibit anti-nociceptive property. The LP and its sub-fractions produced a marked reduction in the number of rolling and adherent leukocytes in the mesenteric microvasculature as revealed by intravital microscopy. The anti-inflammatory effect of LP(PI), the most potent anti-inflammatory fraction of LP, was accompanied by an increase in the serum levels of NO. Further, our study shows that NO is also involved in the inhibitory effect of LP(PI) on neutrophil influx. Conclusions: Our study shows that LP fraction of Calotropis procera comprises of three distinct sets of proteins exhibiting anti-inflammatory and anti-nociceptive properties of which LP(PI) was most potent in inhibiting neutrophil functions and its effects are mediated through NO production. (C) 2009 Elsevier Ireland Ltd. All rights reserved.

Latex constituents from Calotropis procera (R. Br.) display toxicity upon egg hatching and larvae of Aedes aegypti (Linn.)

Calotropis procera R. Br. (Asclepiadaceae) is a well-known medicinal plant with leaves, roots, and bark being exploited by popular medicine to fight many human and animal diseases. This work deals with the fractionation of the crude latex produced by the green parts of the plant and aims to evaluate its toxic effects upon egg hatching and larval development of Aedes aegypti. The whole latex was shown to cause 100% mortality of 3rd instars within 5 min. It was fractionated into water-soluble dialyzable (DF) and non-dialyzable (NDF) rubber-free materials. Both fractions were partially effective to prevent egg hatching and most of individuals growing under experimental conditions died before reaching 2nd instars or stayed in 1st instars. Besides, the fractions were very toxic to 3rd instars causing 100% mortality within 24 h. When both fractions were submitted to heat-treatment the toxic effects were diminished considerably suggesting low thermostability of the toxic compounds. Polyacrylamide gel electrophoresis of both fractions and their newly fractionated peaks obtained through ion exchange chromatography or desalting attested the presence of proteins in both materials. When submitted to protease digestion prior to larvicidal assays NDF lost most of its toxicity but DF was still strongly active. It may be possible that the highly toxic effects of the whole latex from C. procera upon egg hatching and larvae development should be at least in part due to its protein content found in NDF. However the toxicity seems also to involve non protein molecules present in DF.

CARDINAL TEMPERATURES FOR GERMINATION OF THE MEDICINAL ANDDESERT PLANT, Calotropis procera

ABSTRACT Calotropis procera, Apocynaceae, is a wild perennial shrub that originated in the Persian deserts. It is known to provide key resources in degraded ecosystems to about 80 animal species. C. procera is regenerated by seed and produces lots of small seeds that are dispersed by wind; nonetheless, its density is very low. The purpose of this study is to estimate the cardinal temperatures including the base, optimum, and maximum temperatures of Calotropis procera looking at two different ecotypes in the Iranian desert. The germination behavior of C. procera seeds was tested at temperature regimens of 0, 5, 10, 15, 20, 25, 30, 35 and 40oC and was analyzed using linear regression models. The rate of germination increased between base and optimum thermal conditions, and decreased between optimum and maximum thermal conditions. The base, optimum and maximum temperatures for germination of C. procera seeds were estimated at 19.10, 30.75 and 47.80 oC for the Fars and 20.00, 31.82 and 49.69oC for the Zahedan desert, respectively. Temperature and germination were rated to determine the seeding dates of the C. procera. Overall, cardinal temperatures for germination were dependent on local climate characteristics for the range of adaptations in plant growth of the given species.

Osmotin from Calotropis procera latex: New insights into structure and antifungal properties

This study aimed at investigating the structural properties and mechanisms of the antifungal action of CpOsm, a purified osmotin from Calotropis procera latex. Fluorescence and CD assays revealed that the CpOsm structure is highly stable, regardless of pH levels. Accordingly, CpOsm inhibited the spore germination of Fusarium solani in all pH ranges tested. The content of the secondary structure of CpOsm was estimated as follows: alpha-helix (20%), beta-sheet (33%), turned (19%) and unordered (28%). RMSD 1%. CpOsm was stable at up to 75 degrees C, and thermal denaturation (T(m)) was calculated to be 77.8 degrees C. This osmotin interacted with the negatively charged large unilamellar vesicles (LUVs) of 1-palmitoyl-2-oleoyl-sn-glycero-3-phospho-rac-1-glycerol (POPG), inducing vesicle permeabilization by the leakage of calcein. CpOsm induced the membrane permeabilization of spores and hyphae from Fusarium solani, allowing for propidium iodide uptake. These results show that CpOsm is a stable protein, and its antifungal activity involves membrane permeabilization, as property reported earlier for other osmotins and thaumatin-like proteins. (C) 2011 Elsevier B.V. All rights reserved.

Flavonóides de Calotropis procera R. Br. (Asclepiadaceae)

In this work we report the identification of two flavonol glycosides isolated from the leaves of Calotropis procera R. Br. (Asclepiadaceae), a plant species with large occurrence in Northwest of Brazil with some applications in folk medicine. Some proved pharmacological activities in this species could be attributed to the presence of flavonol glycosides. The extraction and isolation of flavonol glycosides was carried out firstly by a liquid-liquid partition, and then by elution of n-BuOH fraction with MeOH over a Sephadex LH-20 column. The identification of flavonol glycosides isorhamnetin-3-O-rutinoside (1), and isorhamnetin-3-O-robinobioside (2), was obtained by 1H and 13C NMR, one- and two-dimensional techniques.

Avaliação do potencial das fibras de Ceiba Pentandra (Kapok) e Calotropis Procera como adsorventes de petróleo

On the basis of human evolution and the population increase was necessary, the emergence of new sources of energy, the development of new products and technologies. One such product, object of the industry revolution and of great importance to the development of humanity is the oil, a substance composed primarily of hydrocarbons which give rise to several other products as fuels, lubricants, polymers, solvents, cooking gas, asphalt for roads, fertilizers, medicines, paints, among other. However, mishandling this product may cause leaks and spills that generate huge damages to the environment and the economy. Soon, with the purpose of contributing to decrease is problematic, in this master's work was carried out an intensive search of the possible potential of the fibers of Ceiba pentandra (L.) "Kapok" and Calotropis Procera as bioadsorbents of petroleum in water. The choice of these fibers is due to surface properties such as oleophylics and hydrophobic, their buoyancy and yet, being biodegradable natural polymers derived from the Brazilian Northeast. This research was used experimental planning with response surface methodology (RSM) with the software Design Expert. The results were statistically efficient, obtaining a R2= 0.9995 for Calotropis Procera and a R2= 0.9993 for Kapok. And that, both fibers showed adsorption efficiency, removing more than 80% petroleum in water static and dynamic state.

Vascular permeability, neutrophil migration and edematogenic effects induced by the latex of Cryptostegia grandiflora

Inflammatory responses have been described as occurring after exposure to some latex materials. In this study pro-inflammatory activity in the latex of Cryptostegia grandiflora was investigated. The soluble proteins of the latex (CgLP) were isolated from the whole latex and evaluated by in vivo assays. CgLP induced strong inflammatory activity mediated by neutrophil migration, enlarging vascular permeability and increasing myeloperoxidase activity locally in rats. CgLP-induced inflammation was observed in peritonitis, paw edema and air push models. In addition, CgLP caused hyperemia in a healing model. The peritonitis effect was lost when CgLP was previously boiled suggesting the involvement of proinflammatory proteins. Thioglycollate increased the neutrophil migration induced by CgLP, but not by fMLP Mast cell depletion provoked by 40/80 compound did not modify the course of inflammation triggered by CgLP, being similar to fMLP, which suggested that neutrophil migration was induced by direct mechanism mediated by macrophages. Neutrophil migration stimulated by CgLP was strongly inhibited by Dexamethasone and to a lesser extent by Thalidomide, indicating the involvement of cytokines in mediating neutrophil infiltration. Celecoxib and Indomethacin were inhibitory suggesting the involvement of prostaglandins. Cimetidine was effective only in the initial phase of edema. PCA 4248 was ineffective. It is concluded that the latex of C. grandiflora is a potent inflammatory fluid, and also that laticifer proteins may be implicated in this process. (c) 2008 Elsevier Ltd. All rights reserved.

Potential of laticifer fluids for inhibiting Aedes aegypti larval development: evidence for the involvement of proteolytic activity

It has been shown previously that the laticifer fluid of Calotropis procera (Ait.) R.Br. is highly toxic to the egg hatching and larval development of Aedes aegypti L. In the present study, the larvicidal potential of other laticifer fluids obtained from Cryptostegia grandiflora R.Br., Plumeria rubra L. and Euphorbia tirucalli L. was evaluated. We attempted to correlate larvicidal activity with the presence of endogenous proteolytic activity in the protein fraction of the fluids. After collection, the fluids were processed by centrifugation and dialysis to obtain the soluble laticifer protein (LP) fractions and eliminate water insoluble and low molecular mass molecules. LP did not visibly affect egg hatching at the doses assayed. LP from Cr. grandiflora exhibited the highest larval toxicity, while P. rubra was almost inactive. E. tirucalli was slightly active, but its activity could not be correlated to proteins since no protein was detected in the fluid. The larvicidal effects of LP from C. procera and Cr. grandiflora showed a significant relationship with the proteolytic activity of cysteine proteinases, which are present in both materials. A purified cysteine proteinase (papain) from the latex of Carica papaya (obtained from Sigma) was similarly effective, whereas trypsin and chymotrypsin (both serine proteinases) were ineffective. The results provide evidence for the involvement of cysteine proteinase activity in the larvicidal action of some laticifer fluids. C. procera is an invasive species found in areas infested with Ae. aegypti and thus could prove useful for combating mosquito proliferation. This is the first report to present evidence for the use of proteolytic enzymes as chemical agents to destroy Ae. aegypti larvae.

First record of Passalora calotropidis in Australia and its generic position

Passalora calotropidis has been found for the first time in Australia on rubber bush (Calotropis procera) in northern Queensland where it was associated with a damaging leaf spot disease. Analysis of sequence data of the ITS region indicated that P. calotropidis belonged to a group that consisted of species of Pseudocercospora. The generic position of P. calotropidis and its potential for biological control are discussed.

Natural parasitism of lepidopteran eggs by Trichogramma species (Hymenoptera: Trichogrammatidae) in agricultural crops in Minas Gerais, Brazil.

The genus Trichogramma Westwood (Hymenoptera: Trichogrammatidae) includes insect egg parasitoids that are widely used throughout the world as control agents of pest insects. The aim of this study was to identify the species of Trichogramma naturally associated with the eggs of lepidopteran pests of the following agricultural and horticultural crops: collards, Brassica oleracea L. (Brassicales: Brassicaceae); papaya, Carica papaya L. (Capparales: Caricaceae); tomato, Lycopersicon esculentum Mill. (Solanales: Solanaceae); cassava, Manihot esculenta Crantz (Malpighiales: Euphorbiaceae); banana, Musa sp. L. (Zingiberales: Musaceae); passion fruit, Passiflora sp. Degener (Malpighiales: Passifloraceae); sugarcane, Saccharum sp. L. (Poales: Poaceae); and corn (maize), Zea mays L. (Poales: Poaceae); and an invasive species (Sodom?s apple milkweed, Calotropis procera Aiton; Gentianales: Apocynaceae) in the semiarid region of Minas Gerais, Brazil. We report natural parasitism by Trichogramma in eggs of Agraulis vanillae vanillae (L.) (Lepidoptera: Nymphalidae), Antichloris eriphia F. (Lepidoptera: Arctiidae), Danaus sp. (L.) (Lepidoptera: Nymphalidae), Diatraea saccharalis F. (Lepidoptera: Crambidae), Erinnyis ello L. (Lepidopera: Sphingidae), and Protambulyx strigilis L. (Lepidopera: Sphingidae). In total, 2,242 specimens of Trichogramma were obtained, belonging to the species T. pretiosum Riley, T. manicobai Brun, Moraes & Soares, T. marandobai Brun, Moraes & Soares, and T. galloi Zucchi. These species of Trichogramma may be candidates for biological control programs of lepidopteran pests in the semiarid region of Minas Gerais and in other semiarid regions.

DISTINGUISHING Campa guianensis Aubl. FROM Campa procera D.C. (MELIACEAE) BY MORPHOLOGY OF YOUNG SEEDLINGS

Seedling morphology was studied in Campa guianensisAubl. and Carapa proceraD. C. from germination to 90 days age. In both species germination is hypogeal and cryptocotylar. Both have rare albino seedlings. Though both species have compound leaves when adult, C, proceraputs out an average total of six simple leaves at germination, while leaves of C. guianensisare compound at all stages. This is the best diagnostic character for separation of the two species at the young seedling stage. Not diagnostic, but none the less useful, is the fact, that polyembryonic seeds are often found in C. procera,but not observed in C guianensis.

Sementes e plântulas de andiroba (Carapa guianensis Aubl. e Carapa procera D. C.): aspectos botânicos, ecológicos e tecnológicos

Na Amazônia, encontram-se ambas espécies do gênero Carapa (Meliaceae), conhecidas pela qualidade da madeira e pelo óleo extraído das sementes. O objetivo deste trabalho foi apresentar informações sobre sementes e plântulas de uma maneira abrangente para subsidiar interesses científicos e aplicados à propagação e manejo das espécies. Para tanto são apresentadas a bio-morfologia dos frutos, sementes e plântulas e uma descrição botânica comparativa, permitindo a distinção das duas espécies. A distribuição natural, abundância e fenologia são apresentados; como também fatores que afetam a regeneração natural, relacionados com as altas taxas de predação de sementes e o ataque da broca-do-ponteiro (Hypsipyla grandella). Andiroba produz sementes grandes e regularmente (em média com peso de 25 g - C. guianensis el6 g -C. procera), porém a produção de mudas enfrenta dificuldades devido a intolerância das sementes ao dessecamento e resfriamento. A espessura do tegumento das sementes de C. guianensis pode causar dormência e prolongar a germinação em até seis meses. O tegumento das sementes de C. procera é normalmente mais delgado e elas necessitam em média apenas um mês para germinar. Práticas de viveiro e plantio, assim como o uso destas espécies são abordados.

Predação de sementes de andiroba [Carapa guianensis Aubl. e Carapa procera DC. (Meliaceae)] por insetos na Amazônias

Os objetivos deste trabalho foram identificar os insetos associados à predação de sementes de Carapa guianensis e Carapa procera e avaliar o potencial de dano nas sementes por insetos e a ocorrência de estratificação vertical na predação de sementes de andiroba. O estudo foi realiado em plantios de C. guianensis e C. procera na Reserva Florestal Ducke, Manaus, Estado do Amazonas, Brasil. As coletas foram realizadas semanalmente no chão da floresta e mensalmente em três diferentes alturas (terços) da copa das árvores. Para avaliar o efeito da predação na germinação, 30 sementes não predadas e 30 sementes predadas coletadas mensalmente do chão da floresta foram colocadas para germinar por um período de um mês. Os resultados indicaram que Hypsipyla grandella e H. ferrealis (Lepidoptera, Pyralidae) foram as principais espécies de insetos associadas à predação das sementes de C. procera e C. guianensis, resultando em taxas de predação média de 39% a 61,96%, respectivamente. Observou-se estratificação vertical na predação dos frutos e sementes na copa das árvores de ambas as espécies de Carapa. A predação das sementes de C. procera e C. guianensis por Hypsipyla spp. reduziu o processo de germinação. Este estudo produziu informações sobre a associação entre C. procera e C. guianensis e as espécies de Hypsipyla.

Influência do ácido peracético sobre a resistência flexural e rugosidade das cerâmicas do sistema Procera AllCeram

O objetivo deste trabalho foi avaliar a influência da imersão em ácido peracético sobre a resistência flexural e rugosidade das cerâmicas do sistema Procera AllCeram. Para cada ensaio foram analisados, o grupo controle e experimental cuja variável foi a imersão em ácido peracético (0,2%), por 50 minutos. A resistência flexural biaxial foi avaliada de acordo com a especificação 6872, da International Organization for Standardization (ISO), que requer um valor mínimo de 50MPa para a cerâmica de cobertura (AllCeram) e 100MPa para a cerâmica de subestrutura (Procera). Para todos os ensaios foram confeccionados 10 corpos de prova, de cada material para compor os grupos experimental (imerso em desinfetante) e controle, com dimensões de (16±0,2)mm de diâmetro por (1,6±0,1)mm de espessura para o AllCeram e, (12±0,2)mm de diâmetro e (1,2±0,2)mm de espessura para o Procera, totalizando 60 corpos de prova. A rugosidade foi avaliada com um rugosímetro utilizando o parâmetro Ra. Os valores médios de resistência flexural biaxial e o desvio padrão dos grupos controle e experimental do Procera e AllCeram foram respectivamente, (440,7±114,77)MPa, (411,6±45,02)MPa, (76,5±3,59)MPa e (86,5±15,41)MPa. Os valores médios da rugosidade e o desvio padrão dos grupos controle e experimental do Procera e AllCeram foram respectivamente: (0,090±0,014)µm, (0,086±0,009) µm, (0,022±0,004)µm e (0,0210±0,006) µm. Quando comparados com a ISO 6872, todos os corpos de prova foram aprovados quanto à resistência flexural. Quando comparados entre si, através do teste “t” de Student não houve diferença significativa entre os grupos experimental e controle de cada um dos ensaios, mostrando que o ácido peracético não interferiu significativamente nas propriedades de resistência flexural e rugosidade dos materiais. Sendo assim, pode-se prever que tal procedimento não trará prejuízos ao desempenho clínico das próteses confeccionadas com as cerâmicas do sistema Procera AllCeram quando consideradas as propriedades analisadas, podendo ser um procedimento clinicamente recomendável. Unitermos: cerâmica odontológica, resistência flexural, rugosidade, ácido peracético.