Percepción de la erosión agraria

Esta investigación analiza mediante la metodología Delphi varios indicadores de calidad del suelo del olivar español, en particular, la textura, la fertilidad, el color, la profundidad, la compactación, la aireación, la materia orgánica y la infiltración del agua. El objetivo consiste en describir la percepción de los olivicultores acerca del grado de erosión en la zona de estudio, teniendo en cuenta su propia autovaloración referida a su nivel de adopción de innovaciones y a su actitud frente al riesgo de aplicar nuevas tecnologías. El panel de expertos seleccionados está formado por 63 olivicultores de la cuenca granadina del Alto Genil en el año 2005; esta zona se caracteriza por el cultivo del olivar en ladera y sus consiguientes niveles de erosión medio-altos, por lo que la actividad agraria se encuentra fuertemente condicionada por el grado de implementación de prácticas de conservación del suelo, específicamente por el laboreo según curvas de nivel, el no laboreo con herbicidas y el mantenimiento de restos de poda sobre la capa edáfica (según una encuesta preliminar del año 2004), así como por su promoción a través de la reciente legislación europea en materia agroambiental.

Manejo de suelo mediante coberturas vegetales establecidas

El objetivo de esta investigación fue determinar los efectos de las coberturas vegetales en el microclima de la planta de vid. Se compararon cinco coberturas de diferente ciclo vegetativo con respecto al manejo de suelo sin labranza mediante aplicación de herbicidas. El estudio se desarrolló en un viñedo cv. Malbec conducido en espaldera alta, situado en Agrelo, Luján de Cuyo, Mendoza, Argentina. Se determinaron parámetros microclimáticos, temperatura, humedad relativa y radiación a nivel de racimos, temperatura del suelo, cantidad y calidad de la radiación reflejada por la cobertura. También se midió la expresión vegetativa y de uvas y el potencial enológico. Se verificó una significativa disminución de la radiación fotosintéticamente activa (RFA) reflejada por las coberturas con una menor relación “Rojo/Rojo lejano" comparada con el suelo descubierto. Sin embargo, el efecto no se percibió dentro de la canopia debido a que las coberturas permanentes de trébol rojo (Trifolium pratensis) y agropiro alargado (Agropyron elongatum) restringieron el vigor de las cepas, disminuyendo el crecimiento de brotes y el tamaño de hojas, lo cual se tradujo en una mayor recepción directa de la RFA a nivel de racimos. No hubo una significativa variación en cuanto a temperatura máxima, mínima y amplitud térmica a nivel de racimos. No obstante ello, los tratamientos con mayor cobertura de suelo tendieron a reducir levemente la temperatura mínima a nivel de racimos. La humedad relativa en la canopia no fue significativamente afectada. El trébol rojo, el agropiro alargado, la mezcla centeno-cebadilla (Secale cereale-Bromus catharticus) y el sorgo del Sudán (Sorghum sudanensis) redujeron notablemente la amplitud térmica del suelo. El efecto fue determinado principalmente por la disminución de la temperatura máxima. Las coberturas vegetales con alguna dificultad para desarrollarse durante su ciclo vegetativo tuvieron un comportamiento intermedio o uno muy similar al de un suelo descubierto. La introducción de una cobertura permanente con buena invasión del sitio interfilar permitió modificar indirectamente las características microclimáticas de la canopia, a través del control del crecimiento vegetativo y de los rendimientos de la planta de vid, modificando el equilibrio vigor / producción del viñedo, y por lo tanto la composición de las uvas y del vino elaborado.

Agricultural pesticide in sediment cores from the Lac de Saint-André, France

Agricultural pesticide use has increased worldwide during the last several decades, but the long-term fate, storage, and transfer dynamics of pesticides in a changing environment are poorly understood. Many pesticides have been progressively banned, but in numerous cases, these molecules are stable and may persist in soils, sediments, and ice. Many studies have addressed the question of their possible remobilization as a result of global change. In this article, we present a retro-observation approach based on lake sediment records to monitor micropollutants and to evaluate the long-term succession and diffuse transfer of herbicides, fungicides, and insecticide treatments in a vineyard catchment in France. The sediment allows for a reliable reconstruction of past pesticide use through time, validated by the historical introduction, use, and banning of these organic and inorganic pesticides in local vineyards. Our results also revealed how changes in these practices affect storage conditions and, consequently, the pesticides' transfer dynamics. For example, the use of postemergence herbicides (glyphosate), which induce an increase in soil erosion, led to a release of a banned remnant pesticide (dichlorodiphenyltrichloroethane, DDT), which had been previously stored in vineyard soil, back into the environment. Management strategies of ecotoxicological risk would be well served by recognition of the diversity of compounds stored in various environmental sinks, such as agriculture soil, and their capability to become sources when environmental conditions change.

Even-sowing pattern strategies for a low-input agricultural system in forage maize

The objective of this study was to verify the effectiveness of new patterns of sowing and to achieve a low-input organic system in two different environments (northern and southern Europe). The study was motivated by the hypothesis that more even sowing patterns (triangular and square) would significantly enhance the growth and yield of forage maize under widely varying conditions, compared with traditional mechanised rectangular seed patterns. An experiment was conducted in Madrid and duplicated in Copenhagen during 2010. A random block design was used with a 2 × 2 factorial arrangement based on two seed-sowing patterns: traditional (rectangular) and new (even) and two weed-management conditions (herbicide use and a low-input system). In both weed-management conditions and locations, the production of aerial maize biomass was greater for the new square seed patterns. In addition, the new pattern showed a greater effectiveness in the control of weeds, both at the initial crop stages (36 and 33% fewer weeds m-2 at the 4- and 8-leaf stages, respectively, in the Copenhagen field experiment) and at the final stage. The final weed biomass for the new pattern was 568 kg ha-1 lower for the Copenhagen experiment and 277 kg ha-1 lower in Madrid field experiments. In the light of these results, the new pattern could potentially reduce the use of herbicides. The results of the experiments support the hypothesis formulated at the beginning of this study that even-sowing patterns would be relatively favourable for the growth and yield of the maize crop. In the near future, new machinery could be used to achieve new seed patterns for the optimisation of biomass yield under low-input systems. This approach is effective because it promotes natural crop-weed competition.

Effects of soil management in vineyard on soil physical and chemical characteristics

Cover crops in Mediterranean vineyards are scarcely used due to water competition between the cover crop and the grapevine; however, bare soil management through tillage or herbicides tends to have negative effects on the soil over time (organic matter decrease, soil structure and soil fertility degradation, compaction, etc). The objective of this study was to understand how soil management affects soil fertility, compaction and infiltration over time. To this end, two bare soil techniques were compared, tillage (TT) and total herbicide (HT) with two cover crops; annual cereal (CT) and annual grass (AGT), established for 8 years. CT treatment showed the highest organic matter content, having the biggest amount of biomass incorporated into the soil. The annual adventitious vegetation in TT treatment (568 kg dry matter ha-1) that was incorporated into the soil, kept the organic matter content higher than HT levels and close to AGT level, in spite of the greater aboveground annual biomass production of this treatment (3632 kg dry matter ha-1) whereas only its roots were incorporated into the soil. TT presented the highest bulk density under the tractor track lines and a greatest resistance to penetration (at 0.2 m depth). AGT presented bulk density values (upper 0.4 m) lower than TT and penetration resistance in CT lower (at 0.20 m depth) than TT too. Effects of soil management in vineyard on soil physical and chemical characteristics - ResearchGate. Available from: http://www.researchgate.net/publication/268520480_Effects_of_soil_management_in_vineyard_on_soil_physical_and_chemical_characteristics [accessed May 20, 2015].

Evaluación de tecnologías de manejo de arvenses en el cultivo de la caña de azúcar (Saccharum spp. híbrido)

En Cuba, las arvenses que afectan al cultivo de la caña de azúcar son una de las causas fundamentales de los bajos rendimientos agrícolas y su control constituye unas de las principales partidas de gastos. En general, se aplican los herbicidas, así como otros métodos de control, sin tener en cuenta el tipo de suelo y las características de estas plantas. Sobre el manejo de arvenses no existen trabajos de investigación que aborden aspectos de eficiencia energética de las producciones y daños al ambiente. Por lo antes señalado, el objetivo de esta investigación fue evaluar diversas tecnologías de manejo de arvenses en el cultivo de la caña de azúcar (Saccharum spp. híbrido), en cepas de primavera y retoño, en tres tipos de suelos, con el propósito de obtener producciones sustentables. El área de estudio se localizó en los campos de la Empresa Azucarera del municipio “Majibacoa”, provincia de Las Tunas (oriente de Cuba), que posee condiciones edafoclimáticas que abundan a lo largo del país. Los tres tipos de suelos más representativos son Fersialítico Pardo Rojizo ócrico, Pardo Mullido y Vertisol Crómico gléyco. En dicha área se han identificado 31 especies de arvenses, 16 de la clase Liliopsida y 15 de la Magnoliopsida. En un primer grupo de experimentos, se desarrollaron nueve ensayos de campo para evaluar la efectividad de herbicidas y mezclas de estos en el manejo de arvenses en el cultivo de la caña de azúcar, tanto en cepas de primavera como de retoño, en los tres tipos de suelos. Se establecieron parcelas de 80 m2 distribuidas en bloques al azar con cuatro réplicas. La efectividad se evaluó por medio del porcentaje de cobertura por arvenses y la fitotoxicidad provocada a las plantas de caña, teniendo en cuenta el coste asociado a cada tratamiento. En aplicaciones preemergentes en caña planta de primavera, el herbicida más eficiente fue el Isoxaflutole con dosis de 0,15; 0,20 y 0,25 kg.ha-1 de producto comercial (pc) en los suelos Fersialítico, Pardo y Vertisol respectivamente. En aplicaciones postemergentes tempranas la mezcla más eficiente fue la de Isoxaflutole + Ametrina +2,4-D con las dosis de Isoxaflutole citadas anteriormente. En aplicaciones preemergentes en cepa de retoño, el herbicida más eficiente fue el Isoxaflutole a dosis de 0,20 kg.ha-1 pc para el suelo Fersialítico y a 0,25 kg.ha-1 pc para los suelos Pardo y Vertisol. En un segundo grupo, se realizaron seis ensayos de campo distribuidos en dos fases. En la primera fase, se desarrollaron tres experimentos, uno por cada tipo de suelo, para evaluar la eficiencia de nueve tecnologías de manejo de arvenses (químicas y físicas combinadas) en cepa de primavera de caña de azúcar. En la siguiente fase, los tres ensayos restantes (uno por tipo de suelo) evaluaron tecnologías de manejo de arvenses durante dos ciclos de producción de caña de azúcar (etapa de primavera y retoño). En la etapa de primavera se aplicó la tecnología más eficiente de los tres experimentos anteriores y durante la etapa de retoño se evaluaron otras nueve tecnologías propias de este tipo de cepa. En estos experimentos los diferentes tratamientos se aplicaron en franjas distribuidas al azar con cuatro réplicas. En las tecnologías evaluadas se emplearon los herbicidas y mezclas que resultaron más eficientes en el primer grupo de experimentos. En cada caso, se evaluaron la eficiencia energética de la producción de azúcar y otros derivados, la resistencia a la penetración de los suelos, la carga contaminante hacia la atmósfera producto de la combustión del diésel y los beneficios al aplicar las diferentes tecnologías. En la primera fase (cepa de primavera), la tecnología con mejor resultado fue la aplicación preemergente de Isoxaflutole inmediatamente después de la plantación, seguida de descepe químico con Glufosinato de amonio, más labor con grada múltiple aproximadamente a los 80 días de la plantación y aplicación pre-cierre con Glufosinato de amonio. En la segunda fase (dos ciclos del cultivo), el mejor resultado se obtuvo cuando en la etapa de retoño se realizó una aplicación preemergente de Isoxaflutole, descepe químico con Glufosinato de amonio y aplicación pre-cierre con este mismo herbicida. En los tres tipos de suelos durante los dos ciclos, la eficiencia energética tuvo valores de 7,2 - 7,5, la resistencia a la penetración 1,2 - 1,5 MPa, la carga contaminante hacia la atmósfera fue de 63,3 - 64,9 kg.t-1 de caña cosechada y beneficios de 8.324 - 8.455 pesos cubanos por hectárea. Este estudio demuestra que un control eficiente de las arvenses debe tener en cuenta necesariamente el tipo de suelo. Así, en los Vertisoles, con mayor contenido en arcilla, se requieren mayores dosis de Isoxaflutole y la eficiencia energética de la producción es menor. La persistencia de ciertas arvenses, especialmente de la clase Liliopsida, requiere de un manejo integrado que incluya diferentes tipos de herbicidas. ABSTRACT In Cuba, weeds affecting the sugarcane are one of the main causes of low agricultural yields, and their control constitutes some of the main items of expenditure. In general, herbicides are applied, as well as other control methods, without keeping in mind the soil type and the characteristics of these plants. Moreover, weed control research approaching aspects about energy efficiency of the crop production, and environmental damages are missing. Hence, the objective of this investigation was to evaluate diverse technologies of weed handling in sugarcane (Saccharum spp. hybrid), both in spring cane plant and ratoon, in three types of soils, with the purpose of obtaining sustainable productions. The study area was located in the fields of the Sugar Enterprise of the Municipality "Majibacoa”, Las Tunas province (east of Cuba) that possesses ecological conditions that are plentiful along the country. The three more representative types of soils are Fersialitic, Brown, and Vertisol. In this area 31 weeds species have been identified, 16 of the Class Liliopsida and 15 of the Magnoliopsida. In a first group of experiments, nine field rehearsals were developed to evaluate the effectiveness of herbicides and mixtures of these for weed handling in sugarcane, in spring cane plants as well as in ratoon, in the three types of soils. Plots of 80 m-2 were distributed at random blocks with four replications. The effectiveness was evaluated by means of the covering percentage by weeds and the provoked toxicity to the cane plants, keeping in mind the cost associated to each treatment. In preemergence applications in spring cane plant, the most efficient herbicide was the Isoxaflutole with dose of 0.15; 0.20 and 0.25 kg.ha-1 of commercial product (pc) in the soils Fersialítico, Brown and Vertisol respectively. In early postemergence applications the most efficient mixture was that of Isoxaflutole + Ametrina + 2,4-D with the doses of Isoxaflutole mentioned previously. In preemergence applications in ratoon, the most efficient herbicide was the Isoxaflutole at dose of 0.20 kg.ha-1 pc for the soil Fersialític and to 0.25 kg. ha-1 pc for the Brown soil and Vertisol. In a second group, six field rehearsals distributed in two phases were carried out. In the first phase, three experiments were developed, one for each soil type, to evaluate the efficiency of nine technologies of weed handling in spring cane plant. In the following phase, the three remaining rehearsals (one for each soil type) diverse technologies of weed handling were evaluated during two cycles of sugarcane production (spring stage and ratoon). In the spring stage the most efficient technology in the three previous experiments was applied and during ratoon stage other nine technologies were evaluated. In these experiments the different treatments were applied in fringes distributed at random with four replicas. In the evaluated technologies the herbicides and mixtures were used selecting those that were more efficient in the first group of experiments. In each case, the energy efficiency of the sugar production and other derivatives, the soil penetration resistance, the polluting load toward the atmosphere product of the combustion, and the benefits when applying the different technologies were all evaluated. In the first phase (spring cane plant), the technology with better result was the preemergence application of Isoxaflutole immediately after the plantation, followed by chemical eradication with Ammonia Glufosinate, hoeing work with multiple tier approximately to the 80 days of the plantation and pre-closing application with Ammonia Glufosinate. In the second phase (two cycles of the cultivation), the best result was obtained when a preemergence application of Isoxaflutole was carried out in sprout's stage, chemical eradication with Ammonia Glufosinate and pre-closing application with this same herbicide. In the three types of soils during the two cycles, the energy efficiency achieved values of 7.2 to 7.5, the resistance to the penetration 1.2 - 1.5 MPa, the polluting load toward the atmosphere was of 63.3 - 64.9 kg.t-1 of the harvested cane and the obtained benefits of 8,324 - 8,455 Cuban pesos per hectare. This study demonstrates that an efficient control of the weeds should necessarily keep in mind the soil type. This way, in the Vertisols, with more clay content, bigger dose of Isoxaflutole is required and the energy efficiency of the production is smaller. The persistence of certain weeds, especially of the class Liliopsida, requires of an integrated handling him to include different types of herbicides.

Wheat cytosolic acetyl-CoA carboxylase complements an ACC1 null mutation in yeast

Spores harboring an ACC1 deletion derived from a diploid Saccharomyces cerevisiae strain, in which one copy of the entire ACC1 gene is replaced with a LEU2 cassette, fail to grow. A chimeric gene consisting of the yeast GAL10 promoter, yeast ACC1 leader, wheat cytosolic acetyl-CoA carboxylase (ACCase) cDNA, and yeast ACC1 3′ tail was used to complement a yeast ACC1 mutation. The complementation demonstrates that active wheat ACCase can be produced in yeast. At low concentrations of galactose, the activity of the “wheat gene” driven by the GAL10 promoter is low and ACCase becomes limiting for growth, a condition expected to enhance transgenic yeast sensitivity to wheat ACCase-specific inhibitors. An aryloxyphenoxypropionate and two cyclohexanediones do not inhibit growth of haploid yeast strains containing the yeast ACC1 gene, but one cyclohexanedione inhibits growth of the gene-replacement strains at concentrations below 0.2 mM. In vitro, the activity of wheat cytosolic ACCase produced by the gene-replacement yeast strain is inhibited by haloxyfop and cethoxydim at concentrations above 0.02 mM. The activity of yeast ACCase is less affected. The wheat plastid ACCase in wheat germ extract is inhibited by all three herbicides at concentrations below 0.02 mM. Yeast gene-replacement strains will provide a convenient system for the study of plant ACCases.

Herbicide sensitivity determinant of wheat plastid acetyl-CoA carboxylase is located in a 400-amino acid fragment of the carboxyltransferase domain

A series of chimeral genes, consisting of the yeast GAL10 promoter, yeast ACC1 leader, wheat acetyl-CoA carboxylase (ACCase; EC 6.4.1.2) cDNA, and yeast ACC1 3′-tail, was used to complement a yeast ACC1 mutation. These genes encode a full-length plastid enzyme, with and without the putative chloroplast transit peptide, as well as five chimeric cytosolic/plastid proteins. Four of the genes, all containing at least half of the wheat cytosolic ACCase coding region at the 5′-end, complement the yeast mutation. Aryloxyphenoxypropionate and cyclohexanedione herbicides, at concentrations below 10 μM, inhibit the growth of haploid yeast strains that express two of the chimeric ACCases. This inhibition resembles the inhibition of wheat plastid ACCase observed in vitro and in vivo. The differential response to herbicides localizes the sensitivity determinant to the third quarter of the multidomain plastid ACCase. Sequence comparisons of different multidomain and multisubunit ACCases suggest that this region includes part of the carboxyltransferase domain, and therefore that the carboxyltransferase activity of ACCase (second half-reaction) is the target of the inhibitors. The highly sensitive yeast gene-replacement strains described here provide a convenient system to study herbicide interaction with the enzyme and a powerful screening system for new inhibitors.

Interaction of the herbicide glyphosate with its target enzyme 5-enolpyruvylshikimate 3-phosphate synthase in atomic detail

Biosynthesis of aromatic amino acids in plants, many bacteria, and microbes relies on the enzyme 5-enolpyruvylshikimate 3-phosphate (EPSP) synthase, a prime target for drugs and herbicides. We have identified the interaction of EPSP synthase with one of its two substrates (shikimate 3-phosphate) and with the widely used herbicide glyphosate by x-ray crystallography. The two-domain enzyme closes on ligand binding, thereby forming the active site in the interdomain cleft. Glyphosate appears to occupy the binding site of the second substrate of EPSP synthase (phosphoenol pyruvate), mimicking an intermediate state of the ternary enzyme⋅substrates complex. The elucidation of the active site of EPSP synthase and especially of the binding pattern of glyphosate provides a valuable roadmap for engineering new herbicides and herbicide-resistant crops, as well as new antibiotic and antiparasitic drugs.

Subcellular localization of acetyl-CoA carboxylase in the apicomplexan parasite Toxoplasma gondii

Apicomplexan parasites such as Toxoplasma gondii contain a primitive plastid, the apicoplast, whose genome consists of a 35-kb circular DNA related to the plastid DNA of plants. Plants synthesize fatty acids in their plastids. The first committed step in fatty acid synthesis is catalyzed by acetyl-CoA carboxylase (ACC). This enzyme is encoded in the nucleus, synthesized in the cytosol, and transported into the plastid. In the present work, two genes encoding ACC from T. gondii were cloned and the gene structure was determined. Both ORFs encode multidomain proteins, each with an N-terminal extension, compared with the cytosolic ACCs from plants. The N-terminal extension of one isozyme, ACC1, was shown to target green fluorescent protein to the apicoplast of T. gondii. In addition, the apicoplast contains a biotinylated protein, consistent with the assertion that ACC1 is localized there. The second ACC in T. gondii appears to be cytosolic. T. gondii mitochondria also contain a biotinylated protein, probably pyruvate carboxylase. These results confirm the essential nature of the apicoplast and explain the inhibition of parasite growth in cultured cells by herbicides targeting ACC.

An isoleucine/leucine residue in the carboxyltransferase domain of acetyl-CoA carboxylase is critical for interaction with aryloxyphenoxypropionate and cyclohexanedione inhibitors

cDNA fragments encoding the carboxyltransferase domain of the multidomain plastid acetyl-CoA carboxylase (ACCase) from herbicide-resistant maize and from herbicide-sensitive and herbicide-resistant Lolium rigidum were cloned and sequenced. A Leu residue was found in ACCases from herbicide-resistant plants at a position occupied by Ile in all ACCases from sensitive grasses studied so far. Leu is present at the equivalent position in herbicide-resistant ACCases from other eukaryotes. Chimeric ACCases containing a 1000-aa fragment of two ACCase isozymes found in a herbicide-resistant maize were expressed in a yeast ACC1 null mutant to test herbicide sensitivity of the enzyme in vivo and in vitro. One of the enzymes was resistant/tolerant, and one was sensitive to haloxyfop and sethoxydim, rendering the gene-replacement yeast strains resistant and sensitive to these compounds, respectively. The sensitive enzyme has an Ile residue, and the resistant one has a Leu residue at the putative herbicide-binding site. Additionally, a single Ile to Leu replacement at an equivalent position changes the wheat plastid ACCase from sensitive to resistant. The effect of the opposite substitution, Leu to Ile, makes Toxoplasma gondii apicoplast ACCase resistant to haloxyfop and clodinafop. In this case, inhibition of the carboxyltransferase activity of ACCase (second half-reaction) of a large fragment of the Toxoplasma enzyme expressed in Escherichia coli was tested. The critical amino acid residue is located close to a highly conserved motif of the carboxyltransferase domain, which is probably a part of the enzyme active site, providing the basis for the activity of fop and dim herbicides.

The Chemically Inducible Plant Cytochrome P450 CYP76B1 Actively Metabolizes Phenylureas and Other Xenobiotics1

Cytochrome P450s (P450s) constitute one of the major classes of enzymes that are responsible for detoxification of exogenous molecules both in animals and plants. On the basis of its inducibility by exogenous chemicals, we recently isolated a new plant P450, CYP76B1, from Jerusalem artichoke (Helianthus tuberosus) and showed that it was capable of dealkylating a model xenobiotic compound, 7-ethoxycoumarin. In the present paper we show that CYP76B1 is more strongly induced by foreign compounds than other P450s isolated from the same plant, and metabolizes with high efficiency a wide range of xenobiotics, including alkoxycoumarins, alkoxyresorufins, and several herbicides of the class of phenylureas. CYP76B1 catalyzes the double N-dealkylation of phenylureas with turnover rates comparable to those reported for physiological substrates and produces nonphytotoxic compounds. Potential uses for CYP76B1 thus include control of herbicide tolerance and selectivity, as well as soil and groundwater bioremediation.

Molecular Characterization of Photomixotrophic Tobacco Cells Resistant to Protoporphyrinogen Oxidase-Inhibiting Herbicides1

Peroxidizing herbicides inhibit protoporphyrinogen oxidase (Protox), the last enzyme of the common branch of the chlorophyll- and heme-synthesis pathways. There are two isoenzymes of Protox, one of which is located in the plastid and the other in the mitochondria. Sequence analysis of the cloned Protox cDNAs showed that the deduced amino acid sequences of plastidial and mitochondrial Protox in wild-type cells and in herbicide-resistant YZI-1S cells are the same. The level of plastidial Protox mRNA was the same in both wild-type and YZI-1S cells, whereas the level of mitochondrial Protox mRNA YZI-1S cells was up to 10 times the level of wild-type cells. Wild-type cells were observed by fluorescence microscopy to emit strong autofluorescence from chlorophyll. Only a weak fluorescence signal was observed from chlorophyll in YZI-1S cells grown in the Protox inhibitor N-(4-chloro-2-fluoro-5-propagyloxy)-phenyl-3,4,5,6-tetrahydrophthalimide. Staining with DiOC6 showed no visible difference in the number or strength of fluorescence between wild-type and YZI-1S mitochondria. Electron micrography of YZI-1S cells showed that, in contrast to wild-type cells, the chloroplasts of YZI-1S cells grown in the presence of N-(4-chloro-2-fluoro-5-propagyloxy)-phenyl-3,4,5,6-tetrahydrophthalimide exhibited no grana stacking. These results suggest that the herbicide resistance of YZI-1S cells is due to the overproduction of mitochondrial Protox.

Herbicide Safener-Binding Protein of Maize1 : Purification, Cloning, and Expression of an Encoding cDNA

Dichloroacetamide safeners protect maize (Zea mays L.) against injury from chloroacetanilide and thiocarbamate herbicides. Etiolated maize seedlings have a high-affinity cytosolic-binding site for the safener [3H](R,S)-3-dichloroacetyl-2,2,5-trimethyl-1,3-oxazol-idine ([3H]Saf), and this safener-binding activity (SafBA) is competitively inhibited by the herbicides. The safener-binding protein (SafBP), purified to homogeneity, has a relative molecular weight of 39,000, as shown by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and an isoelectric point of 5.5. Antiserum raised against purified SafBP specifically recognizes a 39-kD protein in etiolated maize and sorghum (Sorghum bicolor L.), which have SafBA, but not in etiolated wheat (Triticum aestivum L.), oat (Avena sativa L.), barley (Hordeum vulgare L.), tobacco (Nicotiana tabacum L.), or Arabidopsis, which lack SafBA. SafBP is most abundant in the coleoptile and scarcest in the leaves, consistent with the distribution of SafBA. SBP1, a cDNA encoding SafBP, was cloned using polymerase chain reaction primers based on purified proteolytic peptides. Extracts of Escherichia coli cells expressing SBP1 have strong [3H]Saf binding, which, like binding to the native maize protein, is competitively inhibited by the safener dichlormid and the herbicides S-ethyl dipropylthiocarbamate, alachlor, and metolachlor. SBP1 is predicted to encode a phenolic O-methyltransferase, but SafBP does not O-methylate catechol or caffeic acid. The acquisition of its encoding gene opens experimental approaches for the evaluation of the role of SafBP in response to the relevant safeners and herbicides.

Desenvolvimento e avaliação de métodos de extração e separação cromatográfica em colunas monolíticas e superficialmente porosas para determinação de herbicidas triazínicos em solos e águas

O uso de pesticidas levou ao aumento da produtividade e qualidade dos produtos agrícolas, porém o seu uso acarreta na intoxicação dos seres vivos pela ingestão gradativa de seus resíduos que contaminam o solo, a água e os alimentos. Dessa forma, há a necessidade do monitoramento constante de suas concentrações nos compartimentos ambientais. Para isto, busca-se o desenvolvimento de métodos de extração e enriquecimento de forma rápida, com baixo custo, gerando um baixo volume de resíduos, contribuindo com a química verde. Dentre estes métodos destacam-se a extração por banho de ultrassom e a extração por ponto nuvem. Após o procedimento de extração, o extrato obtido pode ser analisado por técnicas de Cromatografia a Líquido de Alta Eficiência (HPLC) e a Cromatografia por Injeção Sequencial (SIC), empregando fases estacionárias modernas, tais como as monolíticas e as partículas superficialmente porosas. O emprego de SIC com coluna monolítica (C18, 50 x 4,6 mm) e empacotada com partículas superficialmente porosas (C18, 30 x 4,6 mm, tamanho de partícula 2,7 µm) foi estudado para separação de simazina (SIM) e atrazina (ATR), e seus metabólitos, desetilatrazina (DEA), desisopropilatrazina (DIA) e hidroxiatrazina (HAT). A separação foi obtida por eluição passo-a-passo, com fases móveis compostas de acetonitrila (ACN) e tampão Acetato de Amônio/Ácido acético (NH4Ac/HAc) 2,5 mM pH 4,2. A separação na coluna monolítica foi realizada com duas fases móveis: MP1= 15:85 (v v-1) ACN:NH4Ac/HAc e MP2= 35:65 (v v-1) ACN:NH4Ac/HAc a uma vazão de 35 µL s-1. A separação na coluna com partículas superficialmente porosas foi efetivada com as fases móveis MP1= 13:87 (v v-1) ACN: NH4Ac/HAc e MP2= 35:65 (v v-1) ACN:NH4Ac/HAc à vazão de 8 µL s-1. A extração por banho de ultrassom em solo fortificado com os herbicidas (100 e 1000 µg kg-1) resultou em recuperações entre 42 e 160%. A separação de DEA, DIA, HAT, SIM e ATR empregando HPLC foi obtida por um gradiente linear de 13 a 35% para a coluna monolítica e de 10 a 35% ACN na coluna com partículas superficialmente porosas, sendo a fase aquosa constituída por tampão NH4Ac/HAc 2,5 mM pH 4,2. Em ambas as colunas a vazão foi de 1,5 mL min-1 e o tempo de análise 15 min. A extração por banho de ultrassom das amostras de solo com presença de ATR, fortificadas com concentrações de 250 a 1000 µg kg-1, proporcionou recuperações entre 40 e 86%. A presença de ATR foi confirmada por espectrometria de massas. Foram realizados estudos de fortificação com ATR e SIM em amostras de água empregando a extração por ponto nuvem com o surfactante Triton-X114. A separação empregando HPLC foi obtida por um gradiente linear de 13 a 90% de ACN para a coluna monolítica e de 10 a 90% de ACN para a coluna empacotada, sempre em tampão NH4Ac/HAc 2,5 mM pH 4,2. Em ambas as colunas a vazão foi de 1,5 mL min-1 e o tempo de análise 16 min. Fortificações entre 1 e 50 µg L-1 resultaram em recuperações entre 65 e 132%.