The stepwise selection for ketoconazole resistance induces upregulation of C14-demethylase (CYP51) in Leishmania amazonensis

Ketoconazole is a clinically safe antifungal agent that also inhibits the growth of Leishmania spp. A study was undertaken to determine whether Leishmania parasites are prone to becoming resistant to ketoconazole by upregulating C14-demethylase after stepwise pharmacological pressure. Leishmania amazonensis promastigotes [inhibitory concentration (IC)50 = 2 µM] were subjected to stepwise selection with ketoconazole and two resistant lines were obtained, La8 (IC50 = 8 µM) and La10 (IC50 = 10 µM). As a result, we found that the resistance level was directly proportional to the C14-demethylase mRNA expression level; we also observed that expression levels were six and 12 times higher in La8 and La10, respectively. This is the first demonstration that L. amazonensis can up-regulate C14-demethylase in response to drug pressure and this report contributes to the understanding of the mechanisms of parasite resistance.

Modelling Amazonian forest eddy covariance data: a comparison of big leaf versus sun/shade models for the C-14 tower at Manaus I. Canopy photosynthesis

In this study, we concentrate on modelling gross primary productivity using two simple approaches to simulate canopy photosynthesis: "big leaf" and "sun/shade" models. Two approaches for calibration are used: scaling up of canopy photosynthetic parameters from the leaf to the canopy level and fitting canopy biochemistry to eddy covariance fluxes. Validation of the models is achieved by using eddy covariance data from the LBA site C14. Comparing the performance of both models we conclude that numerically (in terms of goodness of fit) and qualitatively, (in terms of residual response to different environmental variables) sun/shade does a better job. Compared to the sun/shade model, the big leaf model shows a lower goodness of fit and fails to respond to variations in the diffuse fraction, also having skewed responses to temperature and VPD. The separate treatment of sun and shade leaves in combination with the separation of the incoming light into direct beam and diffuse make sun/shade a strong modelling tool that catches more of the observed variability in canopy fluxes as measured by eddy covariance. In conclusion, the sun/shade approach is a relatively simple and effective tool for modelling photosynthetic carbon uptake that could be easily included in many terrestrial carbon models.

Tracer studies in the coffee plant (Coffea arabica L.)

Due to the great importance of coffee to the Brazilian economy, a good deal of the work carried out in the "Laboratório de Isótopos", E. E. A. "Luiz de Queiroz", Piracicaba, S. Paulo, Brazil, was dedicated to the study of some problems involving that plant. The first one was designed to verify a few aspects of the control of zinc deficiency which is common in many types of soils in Brazil. An experiment conducted in nutrient solution showed that the leaf absorption of the radiozinc was eight times as high as the root uptake; the lower surface of the leaves is particularly suited for this kind of absorption. Among the heavy metal micronutrients, only iron did not affect the absorption of the radiozinc; manganese, copper, and molybdenum brought about a decrease of fifty per cent in total uptake. In another pot experiment in which two soils typical of the coffee growing regions were used, namely, a sandy soil called "arenito de Bauru" and a heavy one, "terra roxa", only O.l and 0.2 per cent of the activity supplied to the roots was recovered", respectively. This indicates that under field conditions the farmer should not attempt to correct zinc deficiency by applying zinc salts to the soil: leaf sprays should be used wherever necessary. In order to find out the most suitable way to supply phosphatic fertilizers to the coffee plant, under normal farm conditions, an experiment with tagged superphosphate was carried out with the following methods of distribution of this material: (1) topdressed in a circular area around the trees; (2) placed in the bottom of a 15 cm deep furrow made around the plant; (3) placed in a semicircular furrow, as in the previous treatment; (4) sprayed directly to the leaves. It was verified that in the first case, circa 10 per cent of the phosphorus in the leaves came from the superphosphate; for the other treatments, the results ware, respectively: 2.4, 1.7, and 38.0 per cent. It is interesting to mention that the first and the last methods of distribution were those less used by the farmers; now they are being introduced in many coffee plantations. In a previous trial it was demonstrated that urea sprays were an adequate way to correct nitrogen deficiency under field conditions. An experiment was then set up in which urea-C14 was used to study the metabolism of this fertilizer in coffee leaves. In was verified that in a 9 hours period circa 95 per cent of the urea supplied to the leaves had been absorbed. The distribution of the nitrogen of the urea was followed by standard chemical procedures. On the other hand the fate of the carbonic moiety was studied with the aid of the radiochromatographic technique. Thus, the incorporation of C14 in aminoacids, sugars and organic acids was ascertained. Data obtained in this work gave a definite support to the idea that in coffee leaves, as in a few other higher plants, a mechanism similar to the urea cycle of animals does exist.

Ergosterol biosynthesis and drug development for Chagas disease

This article presents an overview of the currently available drugs nifurtimox (NFX) and benznidazole (BZN) used against Trypanosoma cruzi, the aetiological agent of Chagas disease; herein we discuss their limitations along with potential alternatives with a focus on ergosterol biosynthesis inhibitors (EBI). These compounds are currently the most advanced candidates for new anti-T. cruzi agents given that they block de novo production of 24-alkyl-sterols, which are essential for parasite survival and cannot be replaced by a host's own cholesterol. Among these compounds, new triazole derivatives that inhibit the parasite's C14± sterol demethylase are the most promising, as they have been shown to have curative activity in murine models of acute and chronic Chagas disease and are active against NFX and BZN-resistant T. cruzi strains; among this class of compounds, posaconazole (Schering-Plough Research Institute) and ravuconazole (Eisai Company) are poised for clinical trials in Chagas disease patients in the short term. Other T. cruzi-specific EBI, with in vitro and in vivo potency, include squalene synthase, lanosterol synthase and squalene epoxidase-inhibitors as well as compounds with dual mechanisms of action (ergosterol biosynthesis inhibition and free radical generation), but they are less advanced in their development process. The main putative advantages of EBI over currently available therapies include their higher potency and selectivity in both acute and chronic infections, activity against NFX and BZN-resistant T. cruzi strains, and much better tolerability and safety profiles. Limitations may include complexity and cost of manufacture of the new compounds. As for any new drug, such compounds will require extensive clinical testing before being introduced for clinical use, and the complexity of such studies, particularly in chronic patients, will be compounded by the current limitations in the verification of true parasitological cures for T. cruzi infections.

Perfil de ácidos graxos da carne de novilhos europeus e zebuínos alimentados com milheto

O objetivo deste trabalho foi avaliar o perfil de ácidos graxos da carne de novilhos mestiços alimentados, em confinamento, com dietas contendo níveis crescentes de grão de milheto moído em substituição ao grão de milho moído. Foram utilizados 24 tourinhos mestiços europeus e 24 mestiços zebuínos, abatidos aos 24 meses de idade, após 96 dias de confinamento. Utilizou-se o delineamento experimental inteiramente casualizado, com os tratamentos em arranjo fatorial 4x2 (quatro dietas com 0, 33, 66 e 100% de milheto no concetrado e dois grupos genéticos) com seis repetições. O aumento dos níveis de grão de milheto na dieta elevou linearmente a participação dos ácidos graxos, araquídico (C20:0), heneicosanoico (C21:0), α-linolênico (C18:3 n-3) e dihomo-γ-linolênico (C20:3 n-6). Tourinhos europeus apresentaram carne com menor teor dos ácidos graxos, mirístico (C14:0), heneicosanoico (C21:0) e γ-linolênico (C18:3 n-6) do que tourinhos zebuínos. A concentração total de ácidos graxos saturados (45,2%), monoinsaturados (41,2%) e poli-insaturados (8,7%), e a relação monoinsaturados/saturados (1,09) e poli-insaturados/saturados (0,18) não foi alterada pelos grupos genéticos e pelas dietas. O aumento da percentagem de grão de milheto na dieta de tourinhos europeus ou zebuínos melhora a relação entre os ácidos graxos ω-6/ω-3.

Estudo microcalorimétrico da interação de tensoativos n-alquil-sulfato de sódio com tripsina a 298 k

Systematic study of the interactions of ionic surfactants with protein trypsin in buffer solution pH 3.5, 7.0 and 9.0, ionic strength 10 mM at 298 K was done using the microcalorimetric technique. In this study, anionic surfactant solutions of the sodium n-alkyl sulfates series (C8, C10, C12 and C14) were used. The enthalpy of interaction (ΔintHº) shows that the interaction of the surfactants C8, C10, C12 and C14 with trypsin in the solution pH 3.5 is an endothermic process with the value of ΔintHº decreasing linearly with increasing carbon chain length, which is attributed to the unfolding of the polypeptide chain. In the solution pH 7.0, we observed the same trend except for C14. In the solution pH 9.0, from C10 the enthapy of interaction didn't change with the increasing of the carbon chain length due to unfolding of the polypeptide. We concluded that when trypsin is folded, the enthalpy of interaction shows a linear relationship with the surfactant's hydrophobicity, in agreement with Traube's rule.

FAST GC-FID METHOD FOR MONITORING ACIDIC AND BASIC CATALYTIC TRANSESTERIFICATION REACTIONS IN VEGETABLE OILS TO METHYL ESTER BIODIESEL PREPARATION

A fast gas chromatography with a flame ionisation detector (GC-FID) method for the simultaneous analysis of methyl palmitate (C16:0), stearate (C18:0), oleate (C18:1), linoleate (C18:2) and linolenate (C18:3) in biodiesel samples was proposed. The analysis was conducted in a customised ionic-liquid stationary-phase capillary, SLB-IL 111, with a length of 14 m, an internal diameter of 0.10 mm, a film thickness of 0.08 µm and operated isothermally at 160 °C using hydrogen as the carrier gas at a rate of 50 cm s-1 in run time about 3 min. Once methyl myristate (C14:0) is present lower than 0.5% m/m in real samples it was used as an internal standard. The method was successful applied to monitoring basic and acidic catalysis transesterification reactions of vegetable oils such as soybean, canola, corn, sunflower and those used in frying process.

AVALIAÇÃO DA QUALIDADE DE MANTEIGAS TIPO EXTRA COMERCIALIZADAS NO ESTADO DO RIO DE JANEIRO

A manteiga é um produto gorduroso onde a fase aquosa está dispersa na fase oleosa formando emulsão do tipo água/óleo. As análises físicas e químicas revelam se a fase oleosa apresenta alterações decorrentes de hidrólise, rancificação ou adulteração. A adição de outras gorduras na manteiga provoca alterações em seus índices físicos e químicos. Para avaliar a qualidade de manteigas tipo extra com sal comercializadas no Estado do Rio de Janeiro, cinco marcas nacionais e duas importadas, totalizando 66 amostras, foram submetidas às análises de acidez, índice de peróxidos, índice de iodo, teor de umidade, teor de cloreto de sódio e composição em ácidos graxos por cromatografia gasosa de alta resolução. A acidez das amostras variou de 0,76 a 2,03 ml SN%; o índice de peróxidos de 0,15 a 3,82 meq peróxidos/kg gordura; o índice de iodo de 20,76 a 53,46 g I2/100g gordura; o teor de umidade de 11,3 a 20,3%; o teor de cloreto de sódio de 0,14 a 2,21%; a composição em % dos principais ácidos graxos foi C14:0 (9,7-12,1%); C16:0 (22,0-26,1%; C18:0 (9,6-11,7%) e C18:1 (21,9-24,2%. Os resultados das amostras foram comparados com o RIISPOA (1952) e com a Portaria n0. 146/96 do MAARA. Todas as amostras analisadas estavam com a acidez dentro dos padrões; 85,5% estavam com índice de peróxidos acima do permitido; 15,2% estavam fora dos limites de índice de iodo; 34,8% estavam com excesso de umidade e 7,5% estavam com o teor de cloreto de sódio acima do permitido. A composição em ácidos graxos estava dentro dos padrões.

Efeitos dos métodos de abate e sexo na composição centesimal, perfil de ácidos graxos e colesterol da carne de capivaras

O objetivo desse trabalho foi avaliar os efeitos de métodos de abate (MA) (humanitário (MH) e por tiro (MT)) e sexo na composição centesimal, perfil de ácidos graxos (AG) e colesterol da carne de capivara (Hydrochaeris hydrochaeris L. 1766). Foram abatidos 20 animais (13 machos (M) e 7 fêmeas (F)) com peso médio de 45,71kg. No músculo longissimus dorsi (LD) foram determinados: umidade, lipídios totais (LT), proteína e cinzas e no semimembranosus (SM), o colesterol por colorimetria e o perfil de AG por cromatografia gasosa. O LD apresentou: 75,87% de umidade, 1,37% de LT, 22,11% de proteínas e 1,09% de cinzas. Houve diferença (p<0,01) nos teores de LT de M (1,75%) e F (0,98%). Os fatores sexo e MA não influenciaram: umidade, proteína e cinzas. Os AG encontrados em maior proporção (%) foram: C16:0 (29,57); C18:1w9 (27,87); C18:2w6 (19,19); C18:0 (6,57); C18:3w3 (4,97); C14:0 (3,64); C20:4w6 (3,45); C18:1w7 (3,31) e C16:1w7 (1,90). A relação AG poliinsaturados/saturados foi 0,82. As médias de AG w6 e w3 foram 23,41 e 5,63%. Os fatores sexo e MA não influenciaram os poliinsaturados e colesterol (28,11mg/100g). A carne de capivara apresentou teores elevados de proteína e valores reduzidos de lipídios totais e de colesterol e relação w6/w3 nutricionalmente adequada.

Reflexos da utilização de farelo de coco sobre o valor nutricional do filé de tilápia-do-nilo (Oreochromis niloticus Linnaeus, 1857)

O uso do farelo de coco pode representar uma fonte alternativa na alimentação de peixe. Por esse motivo avaliaram-se os reflexos de sua utilização sobre o valor nutricional do filé de tilápia-do-nilo. O experimento foi realizado em um delineamento inteiramente ao acaso com 4 tratamentos (T1 - 0%, T2 - 10%, T3 - 20% e T4 - 30% de inclusão de farelo de coco) e cinco repetições. Foram determinados a composição centesimal, o teor de colesterol e os teores de ácidos graxos. Observou-se diferença (p < 0,05) entre T1 e T3, T2 e T3 para os teores de umidade; entre T2 e T3 para os lipídios; T1 apresentou diferença com relação aos demais tratamentos referentes aos carboidratos; e quanto aos teores de cinzas foi observada diferença de T1 e T2 entre T3 e T4. Os teores de colesterol apresentaram um aumento gradual significativo. Com relação aos ácidos graxos, observou-se diferença (p < 0,05) entre os tratamentos referentes aos C10:0; C12:0; C14:0; C18:1; C18:3 (gama); C20:3; e C22:6 numa proporção tal que não determinou diferença na qualidade nutricional do produto.