Chemical Composition of Magonia pubescens Essential Oils and Gamma-Radiation Effects on Its Constituents and Cytotoxic Activity in Leukemia and Breast Cancer Model.

Magonia pubescens A. St.-Hil. is a Brazilian species often used in ethnopharmacology for wound and pain healing and seborrhea treatment. For the first time, essential oils (EOs) obtained from M. pubescens inflorescences were studied. The plant materials (Montes Claros, Brazil, 2018) were submitted to different gamma-radiation doses and their chemical compositions were analyzed by GC/MS and GC-FID. The cytotoxic activity of the EOs was evaluated against K562 and MDA-MB-231 cancer cell lines. A total of 30 components were identified, being 24 compounds detected for the first time in M. pubescens. The main obtained components were hotrienol (35.9 %), cis-linalool oxide (17.0 %) and trans-linalool oxide (10.2 %). The chemical composition of the EO was slightly affected by the applied radiation doses. Irradiated and non-irradiated EOs showed cytotoxic activity against both cell lines and the non-irradiated EO sample was the most active against the K562 cell lines (IC50 =22.10±1.98).

Larvicidal activity of essential oil of Peumus boldus Molina and its ascaridole-enriched fraction against Culex quinquefasciatus.

Culex quinquefasciatus (Say, 1823), known as the domestic mosquito, is a common and abundant species throughout the world, and a cosmopolitan species. The adults of this mosquito are important in terms of public and animal health since they display adaptability to different hosts. In humans, they are responsible for the transmission of various diseases. One manner of control of this vector is the use of insecticidal or larvicidal products, which may have the drawback of toxicity to mammals and can be harmful to the environment. The present work evaluated the larvicidal potential of the essential oil (EO) and ascaridole-enriched fraction (EF4-5) obtained from the leaves of Peumus boldus Molina (boldo). The EO, obtained by steam distillation, was analyzed by GC/MS and fractionated on silica gel. EO and EF4-5, containing 31.4% and 89.5% ascaridole, respectively, were evaluated against C. quinquefasciatus at concentrations of 100, 50 and 25 µg/ml on the third and fourth instars. They showed lethal concentrations (LC50) of 82.14 and 41.85 µg/ml, respectively. Larvae treated with the EF4-5 showed morphological changes in the midgut, with cells possessing a cytoplasm that contained small vacuole-like structures, as well as a nucleus with decondensed chromatin and a cell apex with a short brush border. The cells of the fat body showed larger protein granules, which were acidophilic relative to the larvae of the control group. Moreover, the enriched fraction at a dose of 50 µg/ml showed a residual larvicidal effect according to exposure time on C. quinquefasciatus. This residual effect deserves consideration, since a long-term larvicidal product may be a useful tool for vector control.

Comparative presynaptic effects of the volatile anesthetics sevoflurane and isoflurane at the mouse neuromuscular junction.

INTRODUCTION: Sevoflurane and isoflurane are anesthetics that cause muscle relaxation and potentiate the effects of neuromuscular blocking agents. Their presynaptic mechanisms of action are not understood completely, especially at the motor nerve terminal. METHODS: We compared the presynaptic effects of these anesthetics on the exocytosis of synaptic vesicles labeled with the dye FM1-43 at the mouse neuromuscular junction. RESULTS: Neither anesthetic evoked spontaneous exocytosis of synaptic vesicles, but both significantly inhibited the depolarization evoked by 4-aminopyridine and veratridine, suggesting a putative action on sodium channels. Exocytosis evoked by veratridine was inhibited by tetrodotoxin alone or in conjunction with sevoflurane or isoflurane, indicating that both agents may target voltage-gated sodium channels. CONCLUSIONS: We suggest that sevoflurane and isoflurane inhibit exocytosis evoked by sodium-dependent depolarization and might act on tetrodotoxin-sensitive sodium channels. These findings contribute to a better understanding of some clinical neuromuscular effects induced by these anesthetics.

Phytotoxic and antioxidant effects of dichloromethane fraction of Smilax Brasiliensis Sprengel.

Dichloromethane (DCM) fraction and sub-fractions obtained from Smilax brasiliensis leaves were examined in order to determine their phytotoxic and antioxidant effects. The dichloromethane fraction was submitted to a preparative layer chromatography leading to seven sub-fractions (DCM1-DCM7). Gas chromatography-mass spectrometry (GC-MS) was performed on the dichloromethane sub-fractions. The DCM sub-fractions presented phytotoxic potential; at a concentration of 125 µg per plate, DCM6 and DCM4 showed the strongest results on Lactuca sativa and Allium cepa, respectively. The DCM fraction and DCM4 sub-fraction were more effective than 2,6-di-tert-butyl-4-methylphenol (BHT) at scavenging the 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical. Analysis by GC-MS showed the presence of methyl palmitate (33.05%) in DCM4 and methyl palmitate (17.29%) and methyl oleate (50.96%) in DCM6, suggesting that the activities exhibited by the sub-fractions may be attributed, at least partially, to these major compounds. These results indicate that the DCM sub-fractions of S. brasiliensis could be used as natural herbicides and antioxidants.

Impact on cholinesterase-inhibition and in silico investigations of sesquiterpenoids from Amazonian Siparuna guianensis Aubl.

The plant popularly known as "negramina" (Siparuna guianensis Aubl.), member of the family Siparunaceae produces an essential oil that presents several biological activities reported in literature. Here, the essential oil was obtained by hydrodistillation from fresh leaves collected in the state of Roraima, far north of the Amazon. Chemical composition of the essential oil was characterized by gas chromatography coupled to mass spectrometry (GC-MS) and flame ionization detector (GC-FID). The sesquiterpenoid shyobunone and its derivatives were identified as major compounds in the oil (>40%). The effect of S. guianensis essential oil on the acetylcholinesterase (AChE) activity from Crassostrea rhizophorae, Litopenaeus vannamei and Electrophorus electricus was tested by spectrophotometric assays. The essential oil has been identified as an AChE inhibitor. The mechanism of inhibition was investigated as well as spectrofluorimetric interactions between the essential oil and the enzyme. 1H NMR titration and molecular docking were also investigated. The spectrophotometric results revealed that shyobunone and its derivatives strongly interact with AChE with a kind of non-competitive inhibition. Interaction studies support the results of enzyme inhibition. Molecular coupling predicted that iso-shyobunone is the strongest ligand, corroborated by fluorescence suppression and 1H NMR titration results. In conclusion, Siparuna guianensis essential oil can be a new source of shyobunone and derivatives capable to reversibly inhibit AChE showing potential neuroprotective properties to be applied in the treatment of Alzheimer's disease.

Effect of copaíba essential oil (Copaifera officinalis L.) as a natural preservative on the oxidation and shelf life of sheep burgers.

We evaluated the effects of the inclusion of copaíba (Copaifera officinalis L.) essential oil at 0.05 and 0.1% as a possible replacement of synthetic additive butylated hydroxytoluene (BHT) in sheep burgers during 14 days of storage in a refrigerated display case (4°C). During the shelf life days, analyzes of antioxidant activity, lipid oxidation, pH, color, cooking loss, texture and consumer acceptability were carried out on refrigerated burgers. The addition of copaíba essential oil showed an antioxidant effect in sheep burgers (P > 0.05), reducing lipid oxidation. Copaiba essential oil added at 0.05% showed the highest antioxidant activity, decreased cooking losses and delaying discoloration (loss of redness) during storage; it further improved, the tenderness of sheep burgers (P < 0.05). The treatments had no effect (P > 0.05) on consumer acceptability. Copaiba essential oil is a promising natural antioxidant to increase the shelf life of meat products, as well as being a viable solution to replace synthetic antioxidant BHT, thereby promoting the fresh-like quality appeal of sheep burgers.

The sensitivity of Demodex canis (Acari: Demodicidae) to the essential oil of Melaleuca alternifolia - an in vitro study.

The essential oil of the Melaleuca alternifolia (Maiden & Betche) (tea tree oil) has been effective in previous studies, in the treatment of infestation by Demodex mites in humans. The present study aimed at evaluating the in vitro acaricidal effect of this herbal medicine on Demodex canis. For the parasitological examination, samples were collected from a dog's skin and examined using optical microscopy. Only samples with intact mites and with evident movement of chelicerae and tarsi were selected. Twenty-one samples were tested with the oil, in seven different concentrations: 100%; 50%; 25%; 12.5%; 6.25%; 5.0% and 3.13%. Three samples were tested with the positive control amitraz, and three with the negative control neutral shampoo. The interval between the time the solution was added and the moment the movement of the last mite ceased defined the survival time in the sample. By comparing the times at different concentrations and controls, the results showed that the higher the concentration of the oil, the more quickly its lethal effect occurred, and that the survival times in the controls were longer than in the different concentrations of melaleuca oil.

Improved high-performance liquid chromatographic method for GABA and glutamate determination in regions of the rodent brain.

A C18 reversed-phase column and isocratic fluorescence HPLC method for the simultaneous detection of glutamate and gamma-aminobutyric acid (GABA) is described. In this article a fast and more efficient method for the extraction of these neurotransmitters in rat brain tissue is also presented. The supernatant was derivatized with o-phthalaldehyde (OPA) and analyzed by HPLC with fluorescence detection. Intraday reproducibility was 97.0% and 96.7% and interday reproducibility was 97.1% and 93.7% for GABA and glutamate, respectively. Recovery assays indicate that the accuracy of the method for GABA is 99.6+/-2.3% and for glutamate is 101.9+/-1.8%. In addition, the time consumed to run a sample is lower than that described by other authors. Mean elution time was 3.10 min and 8.22 min for glutamate and GABA, respectively. Thus, in a total runtime of less than 9 min both neurotransmitters were detected. Moreover, when compared to the current methods, the extraction solution used here allowed a high drawing out of the neurotransmitters, glutamate and GABA, from the hippocampus, thalamus and prefrontal cortex of the rat brain.

Coffee oil as a potential feedstock for biodiesel production.

A preliminary evaluation of the feasibility of producing biodiesel using oil extracted from defective coffee beans was conducted as an alternative means of utilizing these beans instead of roasting for consumption of beverage with depreciated quality. Direct transesterifications of triglycerides from refined soybean oil (reference) and from oils extracted from healthy and defective coffee beans were performed. Type of alcohol employed and time were the reaction parameters studied. Sodium methoxide was used as alkaline catalyst. There was optimal phase separation after reactions using both soybean and healthy coffee beans oils when methanol was used. This was not observed when using the oil from defective beans which required further processing to obtain purified alkyl esters. Nevertheless, coffee oil was demonstrated to be a potential feedstock for biodiesel production, both from healthy and defective beans, since the corresponding oils were successfully converted to fatty acid methyl and ethyl esters.

Production of omega 3, 6, and 9 fatty acids from hydrolysis of vegetable oils and animal fat with Colletotrichum gloeosporioides lipase.

Hydrolysis of vegetable oils (Olive, corn, peanut, sesame, flaxseed, soy, canola, garlic, sunflower, almond, castor bean oils) and beef marrow bone oil by Colletotrichum gloeosporioides lipase was studied. The enzyme was capable of generating free fatty acids from all oils tested. The higher hydrolytic activity of the enzyme was towards olive (18.0 IU) and soybean (17.8 IU) oils. The average percentage of essential fatty acids generated from hydrolysis of the oils was 32.92% of omega 9 (as oleic acid C18:1), 26.24% of omega 6 (linoleic C18:2), and 5.86% of omega 3 (such as α-linolenic acid C18:3). Comparison between chromatographic profile of the oils and its enzymatic hydrolysate showed a good equivalence, stressing the applicability of these vegetable substrates under the action of lipase from C. gloeosporioides produce essential fatty acids, being more efficient production of α-linolenic acid from flaxseed oil, linoleic acid from sunflower oil, and oleic acid from olive.

Chemical composition, antioxidant and cytotoxic activities of extracts from the leaves of Smilax brasiliensis Sprengel (Smilacaceae).

The antioxidant and cytotoxic activities of petroleum ether and methanol extracts, fatty acids and methyl esters from leaves of Smilax brasiliensis were evaluated, and the composition of the extracts was determined. Palmitic, linoleic and linolenic acids were major components of the extracts. For antioxidant activity, all samples exhibited IC50 values lower than BHT (2,6-di-tert-butyl-4-methylphenol). The extracts, fatty acids and methyl esters from S. brasiliensis presented no toxicity to larvae of the brine shrimp, Artemia salina. Among the purified substances, only methyl linolenate showed toxicity (LD50 = 21.47 µg/mL). This study showed, for the first time, the composition of petroleum ether and methanol extracts from S. brasiliensis leaves, as well as the antioxidant and cytotoxic activities of extracts, fatty acids and methyl esters.

Correlation between phosphorylation level of a hippocampal 86kDa protein and extinction of a behaviour in a model of Wernicke-Korsakoff syndrome.

The effects of chronic ethanol and thiamine deficiency, alone or associated, on hippocampal protein phosphorylation profiles ranging in molecular weight from 30 to 250kDa molecular weight, in stimulated (high K(+) concentration) and unstimulated (basal) conditions were investigated. These treatments significantly changed the phosphorylation level of an 86kDa phosphoprotein. Thiamine deficiency, but not chronic ethanol, induced a decrease in a behavioural extinction index, which is significantly correlated to the phosphorylation level of the p86 protein. These data add to and extend previous findings by our laboratory implicating the involvement of hippocampal neurotransmission components in extinction of a behaviour which involves learning of environmental spatial cues.

Age-related deficit in behavioural extinction is counteracted by long-term ethanol consumption: correlation between 5-HIAA/5HT ratio in dorsal raphe nucleus and cognitive parameters.

We investigated age-related changes in learning and memory performance and behavioural extinction in the water maze; and in endogenous levels of serotonin (5-HT) and 5-hydroxyindole acetic acid (5-HIAA) in the neocortex, hippocampus, thalamus and dorsal raphe nucleus of Wistar rats. Another aim was to assess the correlation between behavioural and biochemical parameters, which were measured in rodents of two different ages: 5 months (adults) and 16 months (middle-aged). The middle-aged subjects succeeded in learning the behavioural task, albeit with significantly worse performance when compared to adult animals. Aging also had significant main effects on memory and extinction. An age-dependent decrease in 5-HIAA levels was observed in both hippocampus and dorsal raphe nucleus (DRN). The decrease in DRN 5-HIAA was paralleled by a decrease in 5-HIAA/5-HT ratio in this brain area, which was significantly correlated to the animals' spatial memory performance and behavioural extinction. In addition, using middle-aged rats, a 2x2 factorial study was carried out to examine the effects of food restriction and chronic ethanol consumption on rat's performance in a spatial behavioural task and on central serotonergic parameters. None of these two treatments had a significant effect on the behavioural and biochemical parameters assessed, with the exception of extinction index, which was significantly affected by ethanol consumption. Long-term ethanol ameliorated the impairment in behavioural flexibility caused by aging. In conclusion, long-term ethanol consumption may have a role in protecting against age-related deficit in behavioural extinction. Moreover, the present results also indicate that DRN serotonergic system is involved in spatial memory and behavioural extinction.

Thiamine deficiency decreases glutamate uptake in the prefrontal cortex and impairs spatial memory performance in a water maze test.

Using an animal model of Wernicke-Korsakoff syndrome, in which rats were submitted to a chronic ethanol treatment with or without a thiamine deficiency episode, the glutamate uptake in the prefrontal cortex and spatial memory aspects were studied. It was found that (i) thiamine deficiency, but not chronic ethanol consumption, induced a significant decrease of glutamate uptake; (ii) thiamine-deficient subjects showed an impaired performance in the water maze spatial memory test though these animals were able to learn the task during the acquisition. In spite of the fact that thiamine deficiency affects both glutamate uptake and spatial reference memory, there was no significant correlation between these two data. The present results show that, although prefrontal cortex is considered by some authors a not vulnerable area to lesions caused by thiamine deficiency, this vitamin deficiency does cause a neurochemistry dysfunction in that region.

Production of the radioactive antitumoral cisplatin.

This work presents the preparation of radiolabelled cis-dichlorodiammineplatinum (II), CDDP*, sealed in a cadmium capsule. The irradiation of CDDP covered by cadmium, employing exposure times longer than 2 h, demonstrated good chemical purity and high specific activity. This finding allowed a better detection of in vivo CDDP* and suggests that it may be a good tool for studies of long-term biodistribution of pharmaceutical formulations containing this drug.

Analysis of fatty acids released by crotoxin in rat brain synaptosomes.

Crotoxin, the main toxin of Crotalus durissus terrificus venom, exerts its lethal effect by blocking neurotransmission at the neuromuscular junction level through a triphasic mechanism. This effect seems to depend on its phospholipasic activity, suggesting that the mechanism of neurotransmission blockage may be related to fatty acids release in specific sites of the nervous terminal. In this work, we purified the fatty acids released by crotoxin's activity and this outline was compared with other phospholipases A(2), including CB, a subunit of crotoxin. Our results show a higher release of palmitate and arachidonate by crotoxin when compared to other phospholipases A(2). Since palmitate has a role in protein acylation processes and arachidonate participates in signal transduction events, these mechanisms may be related to the neurotoxic actions of crotoxin.

Chemical characterization and antiherpes activity of sulfated polysaccharides from Lithothamnion muelleri.

We report herein the chemical characterization and antiherpes activity of polysaccharides from the red alga Lithothamnion muelleri (Hapalidiaceae). The polysaccharide-rich fractions B1 and B2 were obtained by extraction with Na2CO3 and were purified by size exclusion chromatography to afford Fra-B1 and Fra-B2. The polysaccharides were characterized by FT-IR and chemical analysis (total contents of carbohydrates, proteins, sulfate and uronic acid), whereas their average molecular weights were estimated by high performance gel permeation chromatography. The monosaccharide analysis detected galactose, glucose, xylose, mannose, rhamnose and arabinose in the four polysaccharide samples. Antiherpetic in vitro assays showed that B1 and B2 inhibited Herpes Simplex Virus types 1 and 2 (HSV-1 and HSV-2) when added simultaneously to viral infection affording selectivity indices (SI=CC50/EC50) higher than 20. Investigation of the mechanism of action indicated that B1 and B2 act on the initial steps of HSV replication, mainly inhibiting viral adsorption but also viral penetration into the cells.

Composition in fatty acids of mature milk of nursing mothers / Composição em ácidos graxos do leite maduro de nutrizes

Abstract Objectives: to determine the fatty acid composition of mature milk of nursing mothers and its distribution according to some maternal variables. Methods: this is a cross-sectional observational epidemiological study based on the eva-luation of the fatty acid profile of mature human milk. Samples of mature breast milk were taken from 106 nursing mothers, by manual milking and who were after the 5th postpartum week. The milk fat extraction was carried out by using the Bligh and Dyer method and methy-lated with 0.25 mol/L sodium methoxide in methanol diethyl ether. The fatty acid of the milk profile was determined by a Gas Chromatograph equipped with a flame ionization detector. Results: among the saturated fatty acids, the highest values were observed for palmitic (C16:0), stearic (C18:0), myristic (C14:0) and lauric (C12:0) fatty acids, respectively. Among the monounsaturated fatty acids, there was a higher contribution of oleic (C18:1) and palmi-toleic (C16:1) fatty acids, respectively. The total essential fatty acids (linoleic and α-linolenic) was 14.94%. Conclusions: a low content of essential fatty acids in the breast milk of the nursing mothers was observed in the present study, which are important for infant growth and deve-lopment. We suggest the need to implement nutrition education strategies aimed for pregnant women and nursing mothers who should be advised to eat healthier foods.

Resumo Objetivos: determinar a composição em ácidos graxos do leite maduro de nutrizes e sua distribuição segundo algumas variáveis maternas. Métodos: trata-se de um estudo epidemiológico observacional, transversal realizado a partir da avaliação do perfil de ácidos graxos do leite humano maduro. Amostras de leite materno maduro foram obtidas de 106 nutrizes, a partir da 5ª semana pós-parto, por meio de ordenha manual. A extração da gordura do leite foi realizada através do método de Bligh e Dyer, e metiladas com metóxido de sódio 0,25 mol/L em metanol dietil - éter. O perfil de ácidos graxos do leite foi determinado por um Cromatógrafo a Gás equipado com detector por ionização de chamas. Resultados: dentre os ácidos graxos saturados, foram observados valores mais elevados para os ácidos graxos palmítico (C16:0), esteárico (C18:0), mirístico (C14:0) e láurico (C12:0), respectivamente. Entre os ácidos graxos mono-insaturados, verificou-se maior contribuição dos ácidos graxos oleico (C18:1) e palmitoleico (C16:1), respectivamente. O total de ácidos graxos essenciais (linoleico e α- linolênico) foi de 14,94%. Conclusões: foi observado baixo teor de ácidos graxos essenciais no leite materno das nutrizes do presente estudo, que são importantes para o crescimento e desenvolvimento do lactente, sugerindo-se a necessidade de implementação de estratégias de educação nutricional direcionadas a gestantes e nutrizes que devem ser orientadas a consumir alimentos mais saudáveis.

Different leaf extracts from Brunfelsia uniflora in mice / Diferentes extratos das folhas de Brunfelsia uniflora em camundongos

ABSTRACT: In Brazil, at least 16 poisonous plant species can affect the central nervous system of livestock. Recently in the state of Piauí, Northeastern Brazil, Brunfelsia uniflora was reported as a cause of nervous signs in donkeys, cattle, and small ruminants. In order to assess the toxicity of B. uniflora extracts, 20 Swiss mice were distributed into four groups (n=5) receiving by gavage alkaloids, flavonoids, saponins, or saline (control group). After administration of the extracts in a single dose (5g kg-1), all mice were observed daily for clinical signs. Mice that received the extracts showed moderate to severe clinical signs, including piloerection, vocalization, and seizures. All mice dosed with saponins died between 10 and 20min after administration. Serum biochemical evaluation of animals that received saponins revealed slight increases in total protein levels and decreased magnesium and chlorite levels. In conclusion, saponins of B. uniflora leaves induced acute toxic neurological effects and death in mice.

RESUMO: No Brasil, pelo menos 16 espécies de plantas tóxicas podem afetar o sistema nervoso central dos bovinos. Recentemente no estado do Piauí, no Nordeste do Brasil, Brunfelsia uniflora foi relatada como causadora de sinais nervosos em jumentos, bovinos e pequenos ruminantes. Para avaliar a toxicidade de B. uniflora, 20 camundongos foram distribuídos em quatro grupos de cinco animais recebendo diferentes extratos da planta. Os extratos de alcalóides, flavonóides, saponinas ou solução salina (grupo controle) foram administrados por gavagem. Após a administração dos extratos, em dose única (5g kg-1), todos os animais foram observados diariamente. Os camundongos que receberam os extratos de B. uniflora apresentaram sinais clínicos moderados a graves, incluindo piloeração, vocalização e convulsões. Todos os camundongos que receberam extrato de saponinas morreram entre 10 e 20 minutos após. A avaliação bioquímica sérica dos animais que receberam saponinas, revelou discretos aumentos nos níveis de proteína total e diminuição nos níveis de magnésio e cloro. Conclui-se que, as saponinas presentes nas folhas de B. uniflora induzem efeitos neurológicos tóxicos agudos e, morte em camundongos.

Changes in the essential oil composition of leaves of Echinodorus macrophyllus exposed to

Leaves of Echinodorus macrophyllus (Kunth) Micheli, Alismataceae, were exposed to different doses of γ-radiation (0.00, 1.00, 3.00, 5.00, 10.00, and 20.00 kGy) and the chemical composition of their essential oils was investigated. The extractive process of the essential oil was more favored when the leaves were irradiated. The essential oil components were identified by correlation between GC-FID data and retention parameters obtained from the Kováts method. Moreover, GC-MS analyses of the essential oils were correlated with fragmentation profiles in the NIST standard mass fragmentation data bank. The essential oil of E. macrophyllus contains biologically active constituents of different chemical classes. Acyclic monoterpenes and sesquiterpenes showed increase in concentration when the leaves were exposed to γ-radiation. On the other hand, the component concentrations of some chemical classes were lightly decreased, i.e., for bicyclic monoterpenes, diterpenes, triterpenes, carboxylic esters, and carotenoid derivatives.