Desenvolvimento de método enzimático para determinação de sacarose e correlações entre o conteúdo de açúcares solúveis e proteína em soja

• Autor(a) principal: Teixeira, Arlindo Inês
• Data de Publicação: 2008
• Tipo de documento: Tese
• Idioma: por
• Título da fonte: LOCUS Repositório Institucional da UFV
• Texto Completo: http://locus.ufv.br/handle/123456789/334
• topic: Sacarose; Quantificação; Correlação; Rafinose; Estaquiose; Sucrose; Quantification; Correlation; Raffinose; Stachyose; CNPQ::CIENCIAS AGRARIAS::AGRONOMIA::FITOTECNIA::MELHORAMENTO VEGETAL

The sucrose content in the soybean grain is one of the most important characteristics that accounts for quality improvement, especially concerning flavor and aroma, and it is a critical factor in elaborating soy-based products. The objectives of this study were first to develop a simple and cheap spectrophotometric methodology to quantify sucrose in soybean grains. Next, parents were selected with high sucrose contents and analyze the sucrose, raphinose and staquiose contents, total soluble sugars and total proteins in a population derived from the selected parents, and the correlation among these characteristics was estimated at the end. To develop the methodology, the action of the glucose-oxydase enzyme, which is widely used in glucose quantification, was combined with invertase, a hydrolytic enzyme that cleaves the sucrose glucosidic link forming fructose and glucose. This system was adapted for Elisa plates, that permitted low cost, large scale analyses, with great potential for application in routine analyses in soybean breeding. To validate this new methodology, the sucrose content was also quantified in the grain of 14 soybean genotypes by high performance liquid chromotography (HPLC) and by an enzymatic method already validated, and the correlation coefficient between the methodologies was calculated at the end. These correlations were highly significant, 13 especially between the HPLC quantification and the methodology we developed (r=0.9766), showing that this was efficient in quantifying sucrose. To continue the study, the genetic material used consisted of a population of 19 F2:3 progenies derived from the cross between the CD2013PTWA4-1-1 line, derived from a variety with high sucrose contents, with the CD2013PTA106B-4 isoline that is derived from a high protein content line. This material, together with the parents, was analyzed for the following characteristics: sucrose concentration, analyzed by HPLC and by the GOD/invertase method, the concentration of raphinose, staquiose, total soluble sugars, total protein and weight of 100 seeds. Normality tests were carried out for each characteristic and the heritability was determined for each one and their phenotypic correlations. The broad sense heritabilities of the characteristics sucrose and raphinose contents, total soluble sugars and protein content were high, meaning that most of the variation observed in these characteristics was due to genetic causes. The sucrose concentrations determined by HPLC and the GOD/invertase methods had relatively high Pearson and Spearman correlation coefficients, showing the applicability of the enzymatic method developed. The sucrose concentration determined by HPLC had high and positive correlation coefficients with the raphinose, staquiose and total soluble sugar concentrations, indicating that indirect selection of individuals with low raphinose and staquiose contents in this population would be impossible. The sucrose concentration determined by HPLC correlated negatively with the total protein concentration, indicating that direct selection could not be made of individuals with high contents of both characteristics. The correlation between the raphinose and staquiose concentrations and the protein concentration was not significant. Correlation between the sucrose concentration determined by HPLC with the weight of 100 seeds was not significant. The same performance was observed for the concentrations of raphinose, staquiose and total soluble sugars. The correlation between the sucrose concentration determined by the GOD/invertase method with the other characteristics were smaller than those determined between the sucrose concentration determined by HPLC with the other characteristics. A possible explanation may be the fact that the correlation between the two methodologies was not very high (54.8%). This suggested the need for similar analyses in other populations to increase the range of these results. Only the estaquiose concentration, by the Lilliefors test, and the protein concentration, by the 14 curtose test, was not significant, showing an approximately normal distribution. The other characteristics did not present this distribution.