Comparative study between generations of transgenic and non-transgenic soybeans seeds using a proteomic and metabolomic approach

Orientador: Marco Aurélio Zezzi ArrudaTese (doutorado) - Universidade Estadual de Campinas, Instituto de QuímicaResumo: Este trabalho baseou-se no estudo comparativo entre sementes de soja transgênica (T) e não-transgênica (NT) no que diz respeito a suas proteínas e metabólitos em diferentes gerações da planta, pois acredita-se que o gene inserido possa alterar seu perfil em nível molecular, uma vez que seu genoma foi alterado. Para tal avaliação, foi necessário o plantio das sementes de soja em casa de vegetação sob as mesmas condições até a obtenção da terceira geração. Além disso, foram realizadas diversas extrações das proteínas das sementes de soja para posterior separação por eletroforese. Os géis de eletroforese obtidos mostram um perfil bastante semelhante, mostrando que a separação foi adequada, e que existem proteínas diferentes e também diferenciais entre as mesmas. A quantificação relativa das proteínas foi realizada por meio da técnica de 2-D DIGE, revelando um total de 89 proteínas diferenciais com critério de seleção que partiu de p<0,1 e fator de regulação maior que 2,0 entre os grupos estudados. Um menor número de proteínas diferenciais foi observado nos grupos de sementes T. Após identificação por espectrometria de massas, 29,2% das proteínas corresponderam a classe de armazenamento. Com relação aos metabólitos, os resultados indicaram que o solvente composto por metanol/água (70/30% v/v) foi considerado o mais adequado. Com isso, as duas gerações de sementes tiveram sua análise realizada por LC-MS com analisador QTOF. Os dados obtidos foram avaliados por uma análise estatística para melhor compreensão dos resultados. Os resultados mostraram uma diferença entre os perfis T e NT e também entre suas gerações. Esses resultados indicam, que as sementes de soja T e NT estão sofrendo uma alteração em seu proteoma e metaboloma. A soja T se mostrou mais robusta, mas não é possível afirmar se é devido a transgenia. O fato das menores diferenças terem sido observadas entre os grupos transgênicos, com um menor número de proteínas diferenciais e menores variações estatísticas com relação aos metabólitos, nos levam a concluir que a soja transgênica se adapta melhor as mudanças que estão ocorrendo em seu metabolismo, sejam elas causadas por fatores biológicos, químicos ou físicosAbstract: This work was based on the comparative study between transgenic (T) and non-transgenic (NT) soybean seeds with respect to their proteins and metabolites in different generations of the plant, since it is believed that the inserted gene can change its profile at the molecular level, once their genome was altered. For this evaluation, it was necessary to plant the soybean seeds in a greenhouse under the same conditions until obtaining the third generation. In addition, several extractions of the proteins of the soybean seeds were carried out for subsequent separation by electrophoresis. The obtained electrophoresis gels show a very similar profile, showing that the separation was adequate, and that there are different proteins and also differentials between them. The relative quantification of the proteins was performed using the 2-D DIGE technique, revealing a total of 89 differential proteins with a selection criterion starting from p<0.1 and a regulation factor greater than 2.0 between the groups studied. A lower number of differential proteins was observed in the T seed groups. After identification by mass spectrometry, 29.2% of the proteins corresponded to the storage class. Regarding the metabolites, the results indicated that the solvent composed of methanol/water (30/70% v/v) was considered the most adequate. Thus, the two seed generations were analyzed by LC-MS with Q-TOF analyser. The obtained data were evaluated by a statistical analysis to better understand the results. The results showed a difference between the T and NT profiles and also between their generations. These results indicate that soybean seeds T and NT are undergoing a change in their proteome and metabolome. The T soybean was more robust, but it is not possible to say if it is due to transgenic. The fact that the smaller differences were observed between the transgenic groups, with a lower number of differential proteins and lower statistical variations with respect to the metabolites, lead us to conclude that transgenic soya is better adapted to the changes that are occurring in its metabolism, whether they are caused by biological, chemical or physical factorsDoutoradoQuimica AnaliticaDoutora em Ciências2013/17514-7, 02015/24668-6FAPES

Biomonitoramento de Cr e Pb em peixes de água doce

Trace metal content of three fish species harvested from a tank located in the City of Guarapuava-PR, Brazil were determined by digestion and voltametry. Trace metal content in fish samples were 0.10-1.30 µg/g (muscle) and 0.45-15.90 µg/g (liver) for chromium, 0.35-2.56 µg/g (muscle) and 2.32-34.30 µg/g (liver) for lead. The levels of lead and chromium in fish samples were higher than the recommended legal limits for human consumption. To assess the dietary intake of Cr and Pb by fish, weekly intake was calculated based on average metals contents and weekly consumption of fish and it compared with PTWI (Provisional Tolerable Weekly Intake). The Cr and Pb have achieved up to 25 and 45%, respectively, of PTWI recommended by WHO/FAO

Bipolar Disorder: Recent Advances And Future Trends In Bioanalytical Developments For Biomarker Discovery.

In this manuscript we briefly describe bipolar disorder (a depressive and manic mental disease), its classification, its effects on the patient, which sometimes include suicidal tendencies, and the drugs used for treatment. We also address the status quo with regard to diagnosis of bipolar disorder and recent advances in bioanalytical approaches for biomarker discovery. These approaches focus on blood samples (serum and plasma) and proteins as the main biomarker targets, and use various strategies for protein depletion. Strategies include use of commercially available kits or other homemade strategies and use of classical proteomics methods for protein identification based on bottom-up or top-down approaches, which used SELDI, ESI, or MALDI as sources for mass spectrometry, and up-to-date mass analyzers, for example Orbitrap. We also discuss some future objectives for treatment of this disorder and possible directions for the correct diagnosis of this still-unclear mental illness.407661-

Avaliação da labilidade de alumínio em infusões de erva-mate empregando voltametria adsortiva de redissolução catódica

Infusions of yerba mate obtained at different stages of industrialization were evaluated to determine the bioavailable fraction of Al. Adsorptive Cathodic Stripping Voltammetry using DASA (complexing agent) was applied to determine the labile fraction of Al at pH 5.0 and pH 8.0 for the total fraction of dissolved Al. The results indicate that on average 60% of Al is complexed with organic compounds, minimizing their bioavailability; however, the labile fraction exceeds by up to 4 times the maximum weekly intake recommended by the World Health Organization