Orientador: Hércules Menezes / Banca: Jonas Contiero / Banca: Wilma Aparecida Spinosa / Banca: Derlene Attili de Angelis / Banca: Patricia de Miranda Brusantin / Resumo: O principal objetivo deste trabalho foi o de avaliar uma mistura de frutooligossacarídeos (FOS), sacarose residual, glicose e frutose, produzida por Aspergullus japonicus - 119, quanto à capacidade de produzir alterações nas populações de bactérias láticas e coliformes nos intestinos de ratos machos Wistar. Foram constituídos 13 grupos de animais, sendo um controle e 12 experimentais, cada grupo contendo 7 animais. Durante 3 meses os animais dos grupos experimentais foram suplementados com 350 mL diários de FOS I (49,7% de FOS) ou FOS II (34,3% de FOS), utilizando como excipiente de diluição: água, iogurte convencional e um extrato de soja (2,5% p:v). Estes constituíram os seguintes grupos: Ga; Gi e Gs. As bebidas formuladas com iogurte e extrato de soja foram submetidas a análises sensoriais para verificação de aceitabilidade, visando seu emprego em humanos, no futuro. Para efeito de comparação foram constituídos também 3 grupos de animais que receberam um FOS comercial purificado (ORAFTI) dissolvido nos mesmos excipientes. As médias de UFCs/g de fezes foram submetidas à análise de variância pelos testes de Wilcoxon e/ou Mann-Whitney. Em todos os subgrupos constituídos o FOS I foi mais eficiente que o FOS II para a desejável alteração da microbiota intestinal: aumento das bactérias láticas e diminuição dos coliformes. Os resultados apontam para a seguinte ordem de eficiência para a diminuição de coliformes: ORAFTI+Iogurte > FOS I+Iogurte > ORAFT+soja = ORAFTI+Água. Para o crescimento de bactérias láticas, a mistura de FOS I mostrou-se mais eficiente que o ORAFTI em todos os excipientes utilizados. Os resultados sugerem a possibilidade do emprego comercial da FOS I de Aspergillus japonicus - 119, independentemente de sua purificação / Abstract: The main objective of this work was to evaluate a mixture of fructooligosaccharide (FOS), residual sucrose, glucose and fructose, from Aspergullus japonicus - 119, considering their ability to produce changes in populations of lactic acid bacteria and coliform bacteria in the intestines of male Wistar rats. The animals (91 at all) were divided in 13 groups with 7 rat in each group: one control group and 12 experimental groups. During three months the animals of experimental groups were daily supplemented with 350 mL of FOS I (49.7% FOS) or FOS II (31.3% FOS), using as a vehicle for dilution water, conventional yogurt and soybean extract ( 2.5% w: v). These were designed as Ga, Gi and Gs groups. The drinks made with yogurt and soy extract were subjected to sensory analysis for verification of acceptability, aiming at their employment in humans beings. For comparison were also formed three subgroups of animals that received a purified commercial FOS (Orafti) dissolved in the same dilution vehicle. The averages of CFU/g of feces were submitted to variance analysis by Wilcoxon's test and/or Mann-Whitney test. In all constituted subgroupes, the FOS I was more efficient than the FOS II for desirable change in intestinal microbiologic populations: increase of lactic acid bacteria and coliforms decreased. Overall, the results indicate the following order of efficiency for coliforms decreasing: ORAFTI+Yogurt > FOS I+Yogurt > ORAFT+soybean extract = ORAFTI+Água. For the increasing of lactic bacteria population the mixture of FOS I in all dissolution vehicle showed more efficient then ORAFTI I. The results suggest the possibility of commercial using of FOS I from Aspergillus japonicus - 119, even without purification / Mestre

Introdução: Isoflavonóides são compostos encontrados em vários vegetais e apresentam diversos efeitos farmacológicos. Dentre estes compostos, encontramos os fitoestrógenos, assim chamados por possuírem ações que mimetizam o efeito do estrógeno natural sobre as células. A soja (Glycine max), um dos vegetais ricos nos fitoestrógenos daidzeína e genisteína, tem sido indicada pela literatura como terapia alternativa para a menopausa pela atividade estrogênica que apresenta, visto que a terapia estroprogestiva para tratar os sintomas desta fase aponta um aumento da incidência de câncer de mama. Objetivo: Avaliar a promoção de apoptose e/ou necrose por um Extrato de Soja (Glycine max) Biotransformado pelo fungo Aspergillus awamori (ESBF) em células de linhagem de adenocarcinoma mamário estrógeno-dependentes (MCF-7) e estrógeno-independentes (SK-BR-3). Materiais e métodos: o ESBF foi produzido na Faculdade de Ciências Farmacêuticas de Ribeirão Preto (FCFRP/USP), com concentração determinada de daidzeína (D) e genisteína (G) por CLAE e avaliado em dois modelos de células de adenocarcinoma mamário: estrógeno-dependentes (MCF-7) e estrógeno-independentes (SK-BR-3). Nestes modelos experimentais, foram avaliados também, o Extrato de Soja (ES) e os padrões comerciais de daidzeína (D) e genisteína (G) isoladas ou em combinação (D+G). Neste estudo avaliamos estes compostos perante os parâmetros: a) citotoxicidade pelo método de MTT; b) necrose e apoptose celular pelo ensaio de marcação por iodeto de propídio (IP) e anexina-V e IP; c) a atividade da caspase-3 por western blotting. Resultados: o ESBF nas linhagem MCF-7 e SK-BR-3 apresentou citotoxicidade dose-dependente a partir de 2,184 mg/mL; o ES apresentou aumento na viabilidade celular em todas as concentrações estudadas; os padrões D e G nas concentrações 1,3 e 1,5 µM respectivamente aumentou a viabilidade celular apenas para a linhagem MCF-7; resultado este não observado nas células SK-BR-3. Quanto aos ensaios de necrose e apoptose, encontramos que as duas linhagens celulares apresentaram marcação pelo IP a partir da concentração de 1,638 mg/mL do ESBF, enquanto que o ES e D+G não apresentaram marcação nas concentrações testadas. Somente para a linhagem MCF-7 encontramos no teste de anexina-V + IP apoptose precoce a partir da concentração 0,819 mg/mL e apoptose tardia/necrose a partir da concentração de 2,717 mg/mL frente ao ESBF, enquanto que frente ao ES e aos padrões D+G este resultado não foi observado. Utilizando apenas a linhagem MCF-7, com relação a detecção da caspase-3 íntegra, não foi possível visualizar sua presença a partir da concentração de 1,638 mg/mL do ESBF. Conclusão: Com este estudo verificamos que o ESBF favorece a indução a morte celular das linhagens MCF-7 e SK-BR-3, não acontecendo o mesmo com o ES e os padrões D+G. Nossos achados sugerem que componentes do fungo são os responsáveis por este efeito biológico, e não os metabólitos da soja, visto que os padrões de daidzeína e genisteína, bem como o ES, não apresentaram os resultados de morte celular evidenciados aqui. / Introduction: Isoflavones are compounds found in various vegetables and have different pharmacological effects. Among these compounds there are phytoestrogens, so called because they have actions that mimic the effects of natural estrogen on cells. Soybean (Glycine max), a plant rich in phytoestrogens genistein and daidzein, have been cited in the literature as an alternative therapy for menopause because this plant has estrogen activity. Since oestroprogestative therapy to treat the symptoms of this phase, has many collateral effects, like increased incidence of breast cancer. Objective: To evaluate the promotion of apoptosis and/or necrosis caused by an extract of soybean (Glycine max) biotransformed by the fungus Aspergillus awamori (ESBF) by cell lineage of estrogen-dependent (MCF-7) and estrogen-independent (SK- BR-3) breast adenocarcinoma. Materials and methods: ESBF was produced at the Faculty of Pharmaceutical Sciences of Ribeirão Preto (FCFRP / USP), with known concentration of daidzein (D) and genistein (G) by HPLC and subjected to two models of breast adenocarcinoma cells: estrogen- dependent (MCF-7) and estrogen-independent (SK-BR-3). In these experimental models were also evaluated, Soy Extract (ES) and the commercial standards of daidzein (D) and genistein (G) alone or in combination (D+G). In this study we evaluated all these compounds the following parameters: a) cytotoxicity by MTT method; b) necrosis and apoptosis assay by dialing propidium iodide (PI) and annexin-V + PI; c) the activity of caspase-3 by western blotting. Results: ESBF in cell line MCF-7 and SK-BR-3 showed dose-dependent cytotoxicity starting from 2.184 mg/mL, the ES showed an increase in cell viability at all concentrations studed, D and G standards at concentrations of 1, 3 and 1.5 mM respectively increased cell viability only in line MCF-7, this result not observed in SK-BR-3. For the tests of necrosis and apoptosis, we found that that two cell lines presented labeling IP from the concentration of 1.638 mg/mL of ESBF, while the ES and D + G showed no labeling at all concentrations tested. Only line MCF-7 in the test of annexin-V + PI early apoptosis from the concentration 0.819 mg / mL and late apoptosis or necrosis from the concentration of 2.717 mg / mL against the ESBF, while facing the ES and D+G standards this result was not observed. Using only the cell line MCF-7 in assay to detection of caspase-3 intact, we could not see his presence from the concentration of 1.638 mg/mL ESBF. Conclusion: This study verified that the ESBF favors the induction of cell death in cell line MCF-7 and SK-BR-3, the same not happening with the ES and D+G standards. Our findings suggest that components of the fungus are responsible for this biological effect and not the soy metabolites, since the standards of daidzein and genistein, as well as the ES, the results showed no cell death.

A soja é uma leguminosa amplamente cultivada mundialmente, sendo o Brasil o segundo maior produtor mundial. Seu alto conteúdo proteico e baixo custo são fatores potenciais para o desenvolvimento de produtos tendo como base o isolado proteico de soja ou seus derivados. Neste sentido, a partir do farelo de soja (após extração do óleo) e métodos convencionais de extração foi obtido o concentrado proteico, substrato para o desenvolvimento de hidrolisados enzimáticos parciais de proteína. Para tanto, foram avaliadas endopeptidases (Neutrase® 0,8L, Alcalase® 2,4L e papaína) e exopeptidase (Flavourzyme® 1000L). A partir do hidrolisado foram preparados complexos/quelatos de metal-peptídeo. Em cada etapa foi avaliada a viabilidade econômica do produto gerado. A condição de extração proteica que proporcionou o melhor resultado foi a relação sólido/solvente de 1:30 (m/v), pH 9,0 ajustado com NaOH 4,0 M, com tempo de extração de 45 minutos, seguido de filtração e ajuste do pH para 4,5 com HCl 2,0 M para a precipitação de proteínas. Nestas condições foi obtido rendimento aproximado de 68,6 % de extrato com teor proteico de 84 %. O processo de hidrólise que proporcionou melhor perfil de peptídeos foi obtido com a Alcalase® 2,4L, cuja relação proteína/enzima foi de 7,5 mg:10 ?L, com tempo de incubação de 30 minutos em solução de tampão fosfato de sódio 30 mM a 55 ºC. Porém, não foi possível a secagem do hidrolisado devido ao teor de glicerol oriundo da enzima. Este inconveniente foi superado com a purificação parcial da mistura enzimática em coluna de Sephadex G25, eluída com tampão acetato de sódio (50 mM, pH 5,0), obtendo assim o concentrado enzimático sem prejuízo para a atividade da enzima. O hidrolisado assim obtido representa a proteína em seu conteúdo de aminoácido tanto qualitativamente quanto quantitativamente. Na preparação dos complexos metálicos de cobre, ferro, zinco e manganês, o ponto de equivalência metal/ligante foi determinado com a utilização de métodos eletroquímicos (voltametria cíclica ou titulação potenciométrica) e a quantificação do metal por absorção atômica revelou uma quantidade de metal ligado de 15,19; 5,55; 3,13 e 2,94 % de manganês, ferro, cobre e zinco respectivamente. A análise econômica mostrou a viabilidade para a produção de complexo de zinco, porém não se descartou a viabilidade dos outros produtos mediante ao ajuste da escala produtiva. / The soybean is a legume widely cultivated worldwide, with Brazil being the second largest world producer. Its high protein content and low cost are potential factors for the development of products based on isolated soybean protein or its derived products. In this way, from the soybean meal ( after oil extraction ) and conventional extraction methods, the protein concentrate was obtained, which is a substrate for the development of partial hydrolysates of protein. For this, were evaluated endopeptidases (Neutrase® 0.8L, Alcalase® 2.4L and papain) and a exopeptidase (Flavourzyme® 1000L). From the hydrolysate were prepared metal-peptide complexes / chelates. At each stage were evaluated the economic feasibility of the generated product. The protein extraction condition which provided the best result was the relationship solid/solvent 1:30 (w/v), pH 9.0 adjusted with 4.0 M NaOH, with extraction time of 45 minutes, followed by filtration and pH adjustment to 4.5 with 2.0 M HCl for proteins precipitation. In these conditions was obtained an income of about 68.6 % of extract with 84% of protein content. The hydrolysis process which provided the best peptides profile was obtained with Alcalase® 2.4L, whose ratio of protein / enzyme was 7.5 mg:10 ?L, with incubation time of 30 minutes in a buffer solution of sodium phosphate 30 mM at 55 º C. However, the drying of the hydrolyzed was not possible due to the glycerol content coming from the enzyme. This drawback was overcome by partial purification of the enzyme mixture on a column of Sephadex G25, eluted with sodium acetate buffer (50 mM, pH 5.0), thus obtaining the enzymatic concentrate without any loss to the enzyme activity. The thus obtained hydrolysate represents the protein in its amino acid content qualitatively and quantitatively. In the preparation of metal complexes of copper, iron, zinc and manganese, the equivalence point metal / ligand was determined using electrochemical methods (cyclic voltammetry or potentiometric titration) and metal quantification by atomic absorption revealed an amount of bounded metal of the 15.19; 5.55; 3.13 e 2.94 % of manganese, iron, copper and zinc respectively. The economic analysis showed the feasibility for the production of zinc complex, but not dismissed the feasibility of using the other products adjusting the scale of production.

Made available in DSpace on 2016-04-28T20:12:49Z (GMT). No. of bitstreams: 1 2006- Biancca Scarpeline de Castro.pdf: 1185624 bytes, checksum: 75c4a07321c0f473fd6f7fdc7f3bcf57 (MD5) Previous issue date: 2006-09-26 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior / This master's degree dissertation analyzes the process of institutionalization of transgenic soy in Brazil, starting from the perspective of social networks, specifically in the years of 2003 and 2005. For such an intention, a methodological chapter was accomplished, in which the focus of nets, its appearance in the Economical Sociology, its different types and properties and its uses in the present research were explained. Initially, the context that the transgenic soy was diffused at the country was expose, as in relation to the general panorama of its conventional cultivation, as in the historical, political, juridical and social context which those discussions and disputes about that genetically modified organism have begun. The analyses of social nets for the years of 2003 (chapter III) and 2005 (chapter IV), have been filled with detailed reports of the average current situation of the subject and, starting from these, the actors that have been part of the process were defined and pointed, and being grouped according to their common characteristics. Having the definition of actors and their relationships, it was possible to accomplish the construction and the analysis of the social net illustration and some of their properties could be applied. By its turn, the analyses of the transgenic soy institutionalization were accomplished based on the institutionalist theory, starting from the application of the social nets methodology. Two hypotheses were confirmed with the research: The first has ratified the usefulness and relevance of social nets methodology to analyze institutionalization processes, since it does not privilege agency to structure. The second has pointed that the deliberations about the use and diffusion of the transgenic soy were economical and political deliberations, in spite of involving studies and scientific decisions. In the research, it was concluded that the institutionalization studied in this dissertation has not finished in the formal and informal processes observed, due to the lack of government structure to accomplish the legislation about the subject, to its ignorance by a great part of the population and to the refusal of another portion of this population, who are against its production and commercialization. This way, the process of transgenic soy institutionalization, in spite of its legalization has happened, is still in course. / Esta disserta??o de mestrado analisa o processo de institucionaliza??o da soja transg?nica no Brasil, a partir da perspectiva das redes sociais, especificamente nos anos de 2003 e 2005. Para tal foi realizado um cap?tulo metodol?gico onde o enfoque de redes, seu surgimento na sociologia econ?mica, seus diferentes tipos e propriedades e, suas utiliza??es na presente pesquisa foram explicadas. Inicialmente, foi apresentado o contexto em que a soja transg?nica se difundiu no pa?s, tanto no que se refere ao panorama geral do seu cultivo convencional, quanto no contexto hist?rico, pol?tico, jur?dico e social em que as discuss?es e disputas a respeito deste organismo geneticamente modificado se iniciaram. As an?lises de redes sociais para os anos de 2003 (cap?tulo III) e 2005 (cap?tulo IV), contaram com hist?ricos detalhados da situa??o corrente sobre o assunto e, a partir destes, foram definidos e apontados os atores que fizeram parte do processo, sendo agrupados de acordo com suas caracter?sticas comuns. Com a defini??o dos atores e suas rela??es, foi poss?vel realizar a constru??o e a an?lise da figura da rede social e puderam ser aplicadas algumas de suas propriedades. J? as an?lises da institucionaliza??o da soja transg?nica foram realizadas com base na teoria institucionalista, a partir da aplica??o da metodologia das redes sociais. Duas hip?teses foram confirmadas com a pesquisa: A primeira ratificou a utilidade e relev?ncia da metodologia de redes sociais para analisar processos de institucionaliza??o j? que n?o privilegia ag?ncia sobre estrutura. A segunda apontou que as delibera??es a respeito da utiliza??o e difus?o da soja transg?nica eram pol?ticas e econ?micas, a despeito de envolverem estudos e decis?es cient?ficas. Na pesquisa foi conclu?do que a institucionaliza??o estudada nesta disserta??o n?o se encerrou nos processos formais e informais observados, devido ? falta de estrutura governamental para cumprir a legisla??o sobre o tema, ao seu desconhecimento por grande parte da popula??o e ao recha?o destes de outra parcela, contr?ria ? sua produ??o e comercializa??o. Assim sendo, o processo de institucionaliza??o da soja transg?nica, a despeito de sua legaliza??o ter ocorrido, encontra-se ainda em curso.

This research investigates the protein interactions that occur when soy protein is added to milk and subjected to renneting or heating. Milk was fortified with 20% soy protein and enzymic coagulation studied at 35°C at various pH's and CaCl2 levels. The first part deals with the interaction between milk and soy proteins during rennet-induced milk coagulation. The first goal was to determine how soy proteins affected milk coagulation. The effects of native versus heat-denatured soy proteins on rennet coagulation time and curd firmness were compared. lmmunogold labeling along with transmission electron microscopy was used to identify and localfze soy proteins in coagulated milk. Partitioning of ß-conglycinin and glycinin, the two main soy protein fractions, between cheese and whey was determined by electrophoresis. Soy proteins affected milk coagulation to the greatest extent at pH 6.6. Both heat-denatured and native soy proteins increased rennet coagulation time. Only heat-denatured soy proteins affected final curd firmness. Most of ß-conglycinin was lost in whey, whereas glycinin was retained in curd. Soy proteins existed in the curd as aggregates that were less electron dense than casein micelles. At pH 6.6, heat-denatured soy proteins were fibrous and adhered to the surfaces of casein micelle, preventing direct micelle-micelle contact. This would delay aggregation rate and decrease curd firmness by decreasing the number and strength of links between casein micelles. Native soy proteins did not bind to the casein micelles but rather were physically trapped within curd. Their effect of delaying aggregation is thought to be a function of their binding of calcium. Adding CaCl2 or lowering the pH to 6.3 or 6.0 helped restore coagulation properties. The second goal was to determine what heat-induced interaction occurs between milk and soy proteins, specifically between κ-casein and glycinin. Both κ-casein and glycinin are heat labile and form insoluble aggregates when heated. When glycinin and κ-casein were heated together, some acidic polypeptides of glycinin crosslinked with κ-casein via disulfide linkages. However, when disulfide linkage was prevented by adding ß-mercaptoethanol , non-covalent interactions between κ-casein and both acidic and basic polypeptides of glycinin occurred that prevented the heat precipitation of glycinin. This non-covalent interaction between glycinin polypeptides and κ-casein may explain why the heat-treated soy proteins became attached to the surfaces of casein micelles during rennet coagulation of milk.

No description available.

Antioxidants are molecules capable of stabilizing and preventing oxidation. Certain peptides, protein hydrolysates, have shown antioxidant capacities, which are obtained once liberated from the native protein structure. Soy protein isolates (SPI) were enzymatically hydrolyzed by pepsin and pancreatin mixtures. The soy protein hydrolysates (SPH) were fractionated with sequential ultrafiltration (UF) and nanofiltration (NF) membrane steps. Heat pre-treatment of SPI at 95 degrees celsius (C) for 5 min prior to enzymatic hydrolysis was investigated for its effect on peptide distribution and antioxidant capacity. SPH were subjected to UF with a 10 kDa molecular weight cut off (MWCO) polysulfone membrane. UF permeate fractions (lower molecular weight than 10 kDa) were fractionated by NF with a thin film composite membrane (2.5 kDa MWCO) at pH 4 and 8. Similar peptide content and antioxidant capacity (α=0.05) were obtained in control and pre-heated SPH when comparing the respective UF and NF permeate and retentate fractions produced. FCR antioxidant capacities of the SPH fractions were significantly lower than their ORAC antioxidant capacities, and the distribution among the UF and NF fractions was generally different. Most UF and NF fractions displayed higher antioxidant capacities when compared to the crude SPI hydrolysates, showing the importance of molecular weight on antioxidant capacity of peptides. The permeate fractions produced by NF at pH 8 displayed the highest antioxidant capacity, expressed in terms of Trolox equivalents (TE) per total solids (TS): 5562 μmol TE/g TS for control SPH, and 5187 μmol TE/g TS for pre-heated SPH. Due to the improvement in antioxidant capacity of peptides by NF at pH 8, the potential for NF as a viable industrial fractionation process was demonstrated. Principal component analysis (PCA) of fluorescence excitation-emission matrix (EEM) data for UF and NF peptide fractions, followed by multi-linear regression analysis, was assessed for its potential to monitor and identify the contributions to ORAC and FCR, two in vitro antioxidant capacity assays, of SPH during membrane fractionation. Two statistically significant principal components (PCs) were obtained for UF and NF peptide fractions. Multi-linear regression models (MLRM) were developed to estimate their fluorescence and PCA-captured ORAC (ORAC-FPCA) and FCR (FCR-FPCA) antioxidant capacities. The ORAC-FPCA and FCR-FPCA antioxidant capacities for NF samples displayed strong, linear relationships at different pH conditions (R-squared>0.99). Such relationships are believed to reflect the individual and relative combined contributions of tryptophan and tyrosine residues present in the SPH fractions to ORAC and FCR antioxidant capacities. Therefore, the proposed method provides a tool for the assessment of fundamental parameters of antioxidant capacities captured by ORAC and FCR assays.

A novel approach for the production of soy protein isolates was investigated integrating electroacidification and membrane ultrafiltration. The effect of electroacidification on the ultrafiltration performance and physicochemical properties of the soy protein extracts was obtained by comparing an electroacidified (pH 6) and a non-electroacidified (pH 9) soy protein extract. The effect of membrane fouling on the permeate flux decline was studied in a hollow fiber and a dead end ultrafiltration system. Due to more significant membrane fouling, the permeate flux was always lower for the electroacidified extract, resulting in at least 1.5-fold increase in the total fouling resistance compared to the non-electroacidified extract. The total amount of protein deposited on the membrane surface during unstirred dead-end ultrafiltration was comparable (about 7 mg/cm2) for both soy protein extracts. The discrepancy between the total fouling resistance and the protein deposition estimates was attributed to the formation of denser (less permeable) fouling deposit for the electroacidified extract, which was supported by scanning electron microscopy studies of fouled membranes. The removal of carbohydrates and minerals was evaluated for direct ultrafiltration and two-stage discontinuous diafiltration using a hollow fiber system. The carbohydrate removal results were always consistent with the theoretical predictions, indicating that the carbohydrates were freely permeable across the membrane. In contrast, the minerals were partially retained by the membrane, but to a higher extent for the non-electroacidified extract, which demonstrated that the electroacidification pretreatment enhanced the mineral removal during the ultrafiltration. Incorporation of the diafiltration step improved the ash (mineral) and carbohydrate removal. Stronger electrostatic interactions between soy proteins, calcium/magnesium, and phytic acid (antinutrient) at alkaline pH resulted in less efficient removal of calcium, magnesium, and phytic acid during the ultrafiltration of the non-electroacidified extract compared to the electroacidified extract. Consequently, the soy protein isolates produced by electroacidification and the hollow fiber ultrafiltration had a lower mineral and phytic acid content. The protein content was at least 88 % (dry basis), with or without the electroacidification pretreatment. The study of the viscosity revealed that the electroacidification pretreatment reduced the viscosity of the soy protein extract, which resulted in a lower axial pressure drop increase during the ultrafiltration of the electroacidified extract compared to the non-electroacidified extract. Adjusting the pH of the electroacidified extract to 9 and the pH of the non-electroacidified extract to 6 had a great impact on the particle size distribution but only a marginal effect on the viscosity of the pH adjusted extracts. This indicated that the pH and the particle size distribution were not responsible for the viscosity difference between the electroacidified and the non-electroacidified soy protein extracts. It was proposed that the electroacidification pretreatment had some impact on the water hydration capacity of the soy proteins, which consequently affected the viscosity.

A novel approach for the production of soy protein isolates was investigated integrating electroacidification and membrane ultrafiltration. The effect of electroacidification on the ultrafiltration performance and physicochemical properties of the soy protein extracts was obtained by comparing an electroacidified (pH 6) and a non-electroacidified (pH 9) soy protein extract. The effect of membrane fouling on the permeate flux decline was studied in a hollow fiber and a dead end ultrafiltration system. Due to more significant membrane fouling, the permeate flux was always lower for the electroacidified extract, resulting in at least 1.5-fold increase in the total fouling resistance compared to the non-electroacidified extract. The total amount of protein deposited on the membrane surface during unstirred dead-end ultrafiltration was comparable (about 7 mg/cm2) for both soy protein extracts. The discrepancy between the total fouling resistance and the protein deposition estimates was attributed to the formation of denser (less permeable) fouling deposit for the electroacidified extract, which was supported by scanning electron microscopy studies of fouled membranes. The removal of carbohydrates and minerals was evaluated for direct ultrafiltration and two-stage discontinuous diafiltration using a hollow fiber system. The carbohydrate removal results were always consistent with the theoretical predictions, indicating that the carbohydrates were freely permeable across the membrane. In contrast, the minerals were partially retained by the membrane, but to a higher extent for the non-electroacidified extract, which demonstrated that the electroacidification pretreatment enhanced the mineral removal during the ultrafiltration. Incorporation of the diafiltration step improved the ash (mineral) and carbohydrate removal. Stronger electrostatic interactions between soy proteins, calcium/magnesium, and phytic acid (antinutrient) at alkaline pH resulted in less efficient removal of calcium, magnesium, and phytic acid during the ultrafiltration of the non-electroacidified extract compared to the electroacidified extract. Consequently, the soy protein isolates produced by electroacidification and the hollow fiber ultrafiltration had a lower mineral and phytic acid content. The protein content was at least 88 % (dry basis), with or without the electroacidification pretreatment. The study of the viscosity revealed that the electroacidification pretreatment reduced the viscosity of the soy protein extract, which resulted in a lower axial pressure drop increase during the ultrafiltration of the electroacidified extract compared to the non-electroacidified extract. Adjusting the pH of the electroacidified extract to 9 and the pH of the non-electroacidified extract to 6 had a great impact on the particle size distribution but only a marginal effect on the viscosity of the pH adjusted extracts. This indicated that the pH and the particle size distribution were not responsible for the viscosity difference between the electroacidified and the non-electroacidified soy protein extracts. It was proposed that the electroacidification pretreatment had some impact on the water hydration capacity of the soy proteins, which consequently affected the viscosity.

With the aim to enhance the efficiency of utilization of crystalline methionine supplemented in the high plant-protein diet for grouper (Epinephelus coioides), this study used Bacillus subtitlis spore as a probiotic additive in the diet to shorten the absorption time difference between protein-bound amino acid and crystalline methionine. The study was conducted in two parts. In the first part, juvenile groupers were fed for 14 weeks with 5 experimental diets: fish meal diet, high plant-protein diet with/without crystalline methionine, as well as with/without B. subtitlis spore separately. Growth performance, PER, protein digestibility, amino acid digestibility, non-specific immune responses, and free amino acid concentration in both muscle and serum were assessed. The second part was a time-series study on serum free amino acids concentration after a force-feeding experiment. The results showed that crystalline methionine supplementation in the high plant-protein diet effectively improved the growth of E. coioides (P<0.05). However, B. subtitlis spore supplementation did not affect fish growth performance significantly (P>0.05). A delay in the appearance of peak serum amino acid concentration was observed when fishmeal was partially replaced by soy protein. On the other hand, the force-feeding experiment showed that serum essential amino acid (include methionine) concentrations droped drastically after they reached the peak concentrations from being forced-fed with the B. subtilis containing diet. Supplementation of crystalline methionine seemed to ease the drop of serum methionine concentration. Based on these results, it is concluded that addition of B. subtitlis spore in high plant-protein diet for the grouper does not enhance the utilization of crystalline methionine, but supplementation of crystalline methionine significantly improve the growth performance of the grouper.