The effect of germination on the levels of certain antinutritional factors, nutritional composition and in vitro protein digestibility (IVPD) of soybean seeds were determined. Raw soybeans contain antinutritional factors, such as trypsin inhibitors (83.05 mg/g), total polyphenols (10.83%) and condensed tannins (1.24%). These factors limit the use of soybeans as a source of protein, carbohydrates and minerals in the poultry industry. In general, soybeans are subjected to heat treatment to reduce the amount of antinutritional factors within the seeds, but it is costly and needs high technology equipment. In order to assist small-scale chicken farmers or people living in developing countries, traditional or domestic processing methods are implemented to address the problem of antinutritional factors (ANFs) in soybeans. This study focused on germination as a versatile and low cost practice. Different lengths of germination time were assessed and compared to each another. Soybeans were soaked for 24 hours and allowed to germinate from one day up to six days. Changes within seeds were noted for nutritional and antinutritional factors during germination. Condensed tannins and trypsin inhibitors decreased significantly (P<0.05) while total polyphenols increased from 5 to 6 days. Protein and fat content increased significantly (P<0.05), but starch content decreased with germination. There were no significant changes for IVPD during germination of soybeans. Germination was also compared to other traditional methods such as roasting, soaking and dehulling. Results showed that dehulled soybeans had the highest proximate composition which was significantly different from other treatments (P<0.05), but the total polyphenols increased. It was concluded that germination for a period of three days effectively improved the nutritional value of soybeans and can be considered as an alternative treatment of soybeans in situations where heat treatment is impossible or impractical. Copyright / Dissertation (MSc(Agric))--University of Pretoria, 2011. / Animal and Wildlife Sciences / unrestricted

Doctor of Philosophy / Department of Grain Science and Industry / Sajid Alavi / The feasibility of using a wheat flour mill to refine corn, sorghum and cowpea was studied. Milling of white sorghum grain resulted in decrease in fiber content from 1.89% to 0.38% and 0.45% in raw, finely milled and coarsely milled sorghum respectively. Similarly, there was a reduction in fat (3.17% to 1.75% and 0.51%) content from raw to fine and coarse milled fractions. Starch content increased from 61.85% in raw to 69.80% in fine and 72.30% in coarse fractions. Protein content was almost unchanged at about 7.40% in all the fractions. In de-hulling and milling of cowpeas, starch and protein content increased whereas fiber, fat and ash content decreased. There was a significant difference in expansion characteristics between whole and decorticated binary blends on account of different levels of inherent starch content. Sorghum cowpea (SC) blends had the highest specific mechanical energy (SME) range (285.74 – 361.52 kJ/kg), followed by corn soy (CS) (138.73 – 370.99 kJ/kg) and the least SME was found in sorghum soy (SS) blends (66.56 – 332.93 kJ/kg). SME was found to be positively correlated to starch content in the blends. SC blends had the most stable process followed by SSB and CSB in that order. The milling of expanded extrudates was found to be dependent on bulk density and low bulk density extrudates had bigger particle size and vice-versa. The water absorption index (WAI) for SC was 4.17 g/g to 5.97 g/g, SS ranged from 2.85 g/g to 5.91 g/g and CS ranged from 2.63 g/g to 5.40 g/g. Starch gelatinization ranged from 85.42 – 98.83% for SC, 90.70 – 96.27% for SS, and 72.57 – 95.49% for CS. The starch digestibility increased after extrusion and cooking but there was no significant change in protein digestibility. There was a significant reduction in anti-nutritional factors – phytic acid (26.06 – 44.03%), tannins (18.69 – 26.67%) and trypsin inhibitor (16.55 – 50.85%) after extrusion. Thus, the study showed that high protein blends with superior nutrition density needed for preparation of FBFs could be produced by using existing/traditional milling capabilities and extrusion process.

The use of full fat soybeans (FFSB) in animal feeds has, to date, been limited due to the presence of anti-nutritional factors (ANF). It is, therefore, increasingly recognized that, if the full potential of full fat soybeans for the use in diets is to be realized, reliable analytical procedures must be available for the quality control of processed FFSB which would enable feed manufactures to determine the degree of soybean processing. Results of currently available analytical techniques vary widely between laboratories, causing uncertainty and confusion among soybean processors, feed manufacturers and end users. A collaborative study was conducted to standardize a number of existing analytical procedures used for determining the effects of heat treatment on FFSB and to generate South African ranges for standardized laboratory procedures. Raw soybeans, in a mixture of cultivars, were processed by dry extrusion at eight different temperatures (110ºC, 120ºC, 127ºC, 136ºC, 140ºC, 145ºC, 151ºC and 164ºC). In vivo trials were conducted on broiler chicks which were fed the eight extruded FFBB. Their performance was monitored with regard to average daily gain (ADG) and feed conversion ratio (FCR). The Protein Solubility in KOH (PSKOH) and the Protein Dispersibility index (PDI) procedures were used for standardization in an inter-laboratory study with the participation of ten South African analytical laboratories. Statistical analysis of the in vivo trials with broilers showed no significant difference (P>0.05) between broilers fed FFSB processed at 136 ºC, 140 ºC and 145 ºC. In addition, samples processed at those temperatures (136 ºC, 140 ºC and 145 ºC) showed the best chick performance with regard to average daily gain (ADG) and the feed conversion ratio (FCR). There was a significant difference (P<0.05) between broilers fed FFSB processed at 110 ºC, 120 ºC and 120 ºC as well as those fed FFSB processed at 151 ºC and 164 ºC. In vitro results showed that the PSKOH and PDI values corresponding to temperatures which showed the best chicken performance were between 67-77% and 10.3-8.5 index units respectively. Therefore, the South African ranges for describing the degree of soybean processing using the PSKOH method are 66-77% with repeatability and reproducibility limits of 3.5 and 10.9 respectively and, when using the PDI method, are 8.5-10.3 index units with repeatability and reproducibility limits of 2.1 and 7.7 respectively. A very good correlation was established between the animal production parameters and the PSKOH values, while a poor correlation between animal production parameters (ADWG and FCR) and PDI values was established. The PSKOH method was found to be the most reliable method for FFSB quality control under standardized South African conditions. / Dissertation (MSc)--University of Pretoria, 2011. / Animal and Wildlife Sciences / unrestricted

Made available in DSpace on 2016-08-15T20:27:59Z (GMT). No. of bitstreams: 1 IvanaCNGL_TESE.pdf: 14299009 bytes, checksum: 5bf2d46500f8d157d958429f1dc7dad5 (MD5) Previous issue date: 2015-04-16 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / Gossypol is a phenolic compound produced by pigment glands in cotton stems, leaves, seeds and flower buds (Gossypium spp). Cottonseed meal is a by-product of cotton that is used for animal feeding because it is rich in oil and proteins. However, gossypol toxicity limits cottonseed use in animal feed. High concentrations of free gossypol may be responsible for acute clinical signs of gossypol poisoning, the most common toxic effect is a reduction in reproduction of males and females. The objective of this study was to evaluate the effects of gossypol in the estrous cycle, folliculogenesis in vivo and in vitro and in developing embryos in vitro rodent and chickens. For the first experiment were used 24 Wistar rats divided into two groups: a control group (saline injection, subcutaneously) and the other treated with gossypol (25 mg/kg/day subcutaneously) for 15 days. The rats were euthanized after the last day of treatment for blood and fragments of ovaries, uterus and thyroid. Blood was collected for hormone analysis by ELISA and the ovaries were collected for histological study for qualitative and quantitative evaluation of ovarian follicles. For the second experiment, it took 32 ovaries of female rats and mice and four chickens. The ovaries of rats and mice, chickens and fragments ovary cells were grown in 24-well culture plates containing 1 ml of culture medium specific. We tested four concentrations of gossypol: 0 (control), 5, 10 and 20 µg / ml. All plates were incubated at 39 ° C and 5% CO2 for 24 hours and seven days. After incubation, the ovaries were fixed and processed for histological analysis. For the third experiment, nulliparous female Balb-C mice were superovulated and then allocated to males. Next day, harvest embryos was performed in 2-8 cell stage. Sequential washes the tubes were performed to remove the embryos on modified HTF. Lots 5 - 8 are selected embryos collected and transferred to new culture HTF medium without HEPES supplemented with 10% FBS. Subsequently they were collected and deposited into microdroplets (30 µ L) medium supplemented in 35x10 mm plates. The embryos were cultured in medium without addition of gossypol (control group) or with concentrations of 5, 10 and 20 µg / ml of gossypol added to each drop. These droplets were immersed in mineral oil and grown in a glasshouse with 5% CO2 at 37 ° C for 24 hours. After the cultivation, the embryos were stained for the detection of ruptured membranes and cells in cell death. It can be seen that gossypol promotes degeneration of ovarian follicles and embryolethality and this toxicity occurs by direct action of gossypol and not a product of its biotransformation / Gossipol é um composto fenólico produzido pelas glândulas de pigmento presente nas raízes, caules, folhas, sementes e botões de flores do algodão (Gossypium spp). O farelo e a torta de algodão são subprodutos do algodão utilizados para a alimentação animal, pois são ricos em óleo e proteínas. No entanto, o gossipol limita a utilização do algodão na alimentação animal, pois altas concentrações desta substância podem ser responsáveis por sinais clínicos agudos de intoxicação, sendo o efeito tóxico mais comum é a redução na reprodução de machos e fêmeas. O objetivo deste estudo foi avaliar os efeitos do gossipol no ciclo estral, na foliculogênese in vivo e in vitro e no desenvolvimento de embriões in vitro de roedores e galinhas. Para o primeiro experimento, foram utilizadas 24 ratas Wistar, distribuídas em dois grupos iguais: um grupo controle (injeção de solução salina, por via subcutânea) e o outro tratado com gossipol (25 mg/kg/dia, por via subcutânea) durante 15 dias. As ratas sofreram eutanásia após o último dia do tratamento para coleta de sangue e de fragmentos de ovários, útero e tireoide. Foi coletado sangue para análise hormonal por ELISA e os ovários foram coletados para estudo histológico para avaliação qualitativa e quantitativa dos folículos ovarianos. Para o segundo experimento, foram necessários 32 ovários de ratas e camundongas e quatro de galinhas. Os ovários das ratas e de camundongas, e fragmentos dos ovários de galinhas foram cultivadas em placas de 24 poços de cultura de células contendo 1 ml de meio de cultura específico. Foram testadas quatro concentrações de gossipol: 0 (controle), 5, 10 e 20 ug/mL. Todas as placas foram incubadas a 39 °C e 5% de CO2 durante 24 horas e sete dias. Após a incubação, os ovários foram fixados e processados para análise histológica. Para o terceiro experimento, fêmeas nulíparas de camundongos Balb-C foram superovuladas e em seguida, alocadas com machos. No dia seguinte, foi realizada a colheita de embriões na fase de 2-8 células. Foram realizadas lavagens sequenciais das tubas para remoção dos embriões com meio HTF modificado. Lotes de 5 8 embriões coletados e selecionados foram transferidos para novo meio de cultivo HTF sem HEPES suplementado com 10% de SFB. Posteriormente foram recolhidos e depositados em microgotas (30 µl) de meio suplementado em placas de 35x10 mm. Os embriões foram cultivados em meio sem a adição de gossipol (grupo controle) ou com concentrações de 5, 10 e 20 µg/ml de gossipol, adicionadas em cada gota. Estas gotas foram imersas em óleo mineral e cultivadas em estufa com 5% de CO2 a 37 °C, por 24 horas. Após o cultivo, os embriões foram corados para a detecção de membranas rompidas e de células em processo de morte celular. Pode-se observar que o gossipol promove degeneração de folículos ovarianos e embrioletalidade, e esta toxicidade ocorre por meio de ação direta do gossipol e não por algum produto de sua biotransformação

The giant freshwater prawn, Macrobrachium rosenbergii (De Man, 1879), is gaining popularity as a key aquaculture species; global production currently exceed 220,000 tonnes, however, industry expansion is limited by high operational costs, with the feed accounting for between 40 to 60% of these. Attention, therefore, has been devoted to increasing the inclusion of plant proteins into the formulation of aqua feeds as a consequence of the limited, unpredictable supply and increasing price of fishmeal. The concomitant introductions of anti-nutritional factors (ANFs), such as phytic acid (PA) with the plant protein fraction, however, are major impediments in the efforts toward the increased use of plant protein ingredients in aqua feeds. Phytic acid is an anti-nutrient that can curtail the development of this as PA has been reported to suppress growth impairing proper nutrient intake, diminishing the availability of minerals, and causing damage to the body tissues and organs which can result in mortality. Although the anti-nutritive effects of PA have been studied extensively in terrestrial agriculture farm species, as well as in a variety of fish species, there is almost no information regarding the effects of PA in crustaceans, including the freshwater prawn, M. rosenbergii. The aims of this present thesis were, therefore, to gain a greater understanding of dietary PA and microbial phytase and their effect on growth performance, feed utilisation, nutrient utilisation and digestibility, mineral availability and whole body proximate composition in juvenile M. rosenbergii. Specifically, the first major experiment set out to investigate the effect of including increasing amounts of PA in the diets presented to M. rosenbergii on growth. The dose-response relationships between PA and growth performance, feed utilisation, nutrient digestibility and utilisation and whole proximate composition were investigated. Four replicate groups of M. rosenbergii with a mean initial carapace length of 6.03 ± 0.30 mm and mean initial weight of 0.29 ± 0.02 g were fed graded levels of PA for 140 days. The basal diet, to which different levels of PA were added to obtain 0.26 (control), 6.48, 11.28, 16.53, 21.45 and 26.16 g PA kg-1, contained fishmeal, soy protein concentrate, wheat meal and corn starch. The results indicated that growth performance, feed utilisation and survival did not differ significantly between the groups receiving the different inclusions of PA within their diets. The apparent protein, lipid and energy utilisations responded negatively, decreasing significantly (p<0.05) with an increasing inclusion of PA, particularly within the groups of prawns fed the diet with the highest inclusions of PA, i.e. the 21.45–26.16 g PA kg-1 diets. The digestibility of protein and lipid were also reduced as the inclusion of PA increased. The whole body composition of protein (p<0.04), lipid (p<0.01) and gross energy (p<0.05) decreased significantly with an increasing supplementation of PA, while the ash content significantly increased (p<0.01), most notably in the groups of prawns receiving the highest levels of dietary PA. The second major experimental trial investigated the effect of microbial phytase on the growth of juvenile M. rosenbergii, when fed diets supplemented with various doses for a period of 80 days. The study set out to improve the growth performance, feed utilisation, nutrient digestibility and utilisation and body composition of M. rosenbergii when fed diets high in plant protein ingredients. To investigate this, four plant protein based diets, which included soybean meal, wheat gluten and wheat meal, were formulated and supplemented with microbial phytase at levels of 0, 500, 1000 and 2000 FTU kg-1 (one phytase unit per kg) and fed to sixty juvenile M. rosenbergii (mean initial carapace length of 8.51 ± 0.52 mm; mean initial weight of 0.40 ± 0.07 g) for 80 days. High levels of plant protein in the diets supplemented with 0–2000 FTU kg-1 did not result in any negative effect on growth performance, feed utilisation nor on the survival of M. rosenbergii. Noticeable moderate growth improvements in line with increasing microbial phytase supplement levels were observed and the highest growth performance was seen in the group fed 2000 FTU kg-1. Supplementation of the diets with 500–2000 FTU kg-1 were found to affect the nutrient utilisation, resulting in a significant (p<0.05) increase in the protein and lipid utilisation when compared to the prawns analysed from the control group. In addition to this latter finding, an increasing supplement of phytase in the diet also resulted in an increase in the dry matter fraction as well as improvements in the digestibility of protein and lipid. There were, however, no significant differences in moisture, protein, lipid, gross energy and the ash content of the whole body of the M. rosenbergii among the groups. This thesis, in a third major trial, explored the impact of other potential ANFs associated to PA, the binding effect of PA with mineral. The effects of graded levels of PA (i.e. 0.26–control, 6.48, 11.28, 16.53, 21.45 and 26.16 g PA kg-1) on the moult frequency and mineral availability in juvenile M. rosenbergii fed over a period of 140 days were determined. The levels of PA assessed in this feed trial had no major detrimental effects on moult frequency. Negative effects (p<0.05) of high PA levels (i.e. 21.45–26.16 g PA kg-1), however, were found on the whole body P concentration. An inverse trend was recorded for the Ca content in the whole body (p<0.005) and carapace (p<0.004) with increasing PA inclusion. These results are consistent with findings for marine shrimp species such as Marsupenaeus japonicus and Litopenaeus vannamei. The graded inclusion of PA in the experimental diets also resulted in a significant reduced (p<0.03) P content in the carapace. Significant changes (p<0.05) were observed in the carapace Zn, Cu, K and Na compositions, particularly in the prawns fed the diet containing 11.28 g PA kg-1, which suggests that the specific minerals were either selectively utilised or retained in the carapace. Accordingly, this thesis investigates the potential of adding dietary supplements of microbial phytase in order to improve mineral availability as proven in several fish species. To explore this, four experimental diets were formulated - three incorporating different levels of microbial phytase (i.e. 0 FTU kg-1, 1000 FTU kg-1 and 2000 FTU kg-1) and a fourth, a control. An aliquot of 15 g PA kg-1 was also added to each treatment. The mineral premix was omitted from the test group diets except the control diet was identical to the 0 FTU kg-1 diet but included a mineral premix and an aliquot of 8 g kg-1 monosodium phosphate which replaced an equal amount of wheat meal fraction. The trial found no significant differences in growth, feed utilisation and moult frequency with the microbial phytase level within the diet, however, survival was compromised. The proximate composition of the prawn whole body was in most cases unaffected by the level of phytase. Supplementation of the diets with microbial phytase did, however, result in significant higher (p<0.05) concentrations of minerals including: 1) Ca, Mg, K and Na in the whole body; 2) Ca and Zn in muscle tissue; and, 3) Ca and Mg in the carapace. The Zn content of the carapace, however, was negatively affected by the inclusion of microbial phytase suggesting the necessity of this mineral within the diet of M. rosenbergii. This thesis contributes to current understanding surrounding the inclusion of dietary PA and the benefits of microbial phytase within the experimental diets consumed by juvenile M. rosenbergii. The knowledge gained from this work provides a means to optimise the use of plant protein ingredients and with the potential to decrease the dependability of fishmeal without compromising M. rosenbergii production and profitability, thus ultimately promoting the sustainable expansion of M. rosenbergii aquaculture.

Sementes de feijão são freqüentemente infestadas por diferentes insetos-praga incluindo Zabrotes subfasciatus, Callosobruchus maculatus e Acanthocelides obtectus. Estratégias visando o controle do ataque desses insetos têm sido desenvolvidas através do estudo de substâncias tóxicas e/ou proteínas antinutricionais que atuem sobre o desenvolvimento destes insetos. Os inibidores protéicos de &#945;-amilases presentes em leguminosas como o feijão comum, bem como em sementes de cereais como o trigo, têm a capacidade de inibir a &#945;-amilase de Z. subfasciatus. O estudo do mecanismo de interação entre os inibidores e as enzimas digestivas de insetos pode propiciar o delineamento de inibidores potentes que atuariam especificamente contra essas pragas. O estudo da interação do inibidor de trigo 0.19 e com a &#945;-amilase de Z. subfasciatus desenvolvido neste trabalho demonstra a atuação de vários componentes de força na superfície de interação entre a enzima e o inibidor, garantindo a estabilidade do complexo. A presença de pontes de hidrogênio, interações hidrofóbicas e ligações iônicas foram evidenciadas. Através da análise da seqüência primaria de outros inibidores de cereais, foi verificado que o inibidor de &#945;-amilase de trigo denominado 0.53 e o inibidor de cevada possivelmente atuam da mesma maneira sobre a &#945;- amilase do inseto. Estes resultados tornam esses inibidores protéicos potenciais candidatos à transformação de plantas de feijão visando o controle do desenvolvimento de Z. subfasciatus durante seu estágio larval. / Several insects included Callosobruchus maculatus, Acanthocelides obtectus and Zabrotes subfasciatus have frequently infest bean seeds. Strategies for the control of these predators have been developed through the study of toxic substances and antinutritional factors, which act on the development of these insects. Proteinaceus Inhibitors of &#945;-amylases present in varieties of beans, as well as in wheat seeds have been demonstrated to be active to the &#945;-amylase of Z. subfasciatus. The study of the mechanism of interaction between these inhibitors and the digestive enzymes can promote a powerful inhibitor that acts specifically against these insect-pest. Studies on the interaction of the inhibitor of wheat kernels 0.19 and Z. subfasciatus &#945;-amylase demonstrated the performance of some intermolecular forces on the surface of the enzyme-inhibitor interaction giving the stability to the complex. The presence of hydrogen bond, hydrophobic interactions and saline bridges had been showed . The analysis of the primary sequences of inhibitors from other cereals allowed to verify that the wheat amylase inhibitor 0.53 and the barley inhibitor possibly act by the same way on the insect a-amylase. These results make these proteinaceus inhibitors potential candidates to be used on the plant transformation for the control of the development of Z. subfasciatus during of larval stage.

As leguminosas constituem uma importante fonte de alimentos para os ruminantes e podem ser exploradas para pastejo direto ou, se conservadas, para fornecimento na forma de feno ou silagem. Algumas leguminosas possuem compostos fenólicos em sua composição. A caracterização química dessas as plantas possibilita melhor uso das mesmas na alimentação animal assim como um melhor entendimento dos efeitos positivos e negativos dos compostos fenólicos na nutrição dos animais. Objetivou-se com o presente estudo: (i) avaliar a composição química de cinco leguminosas de interesse para a alimentação de ruminantes; (ii) estudar os efeitos dos taninos de diferentes leguminosas na produção de gases in vitro; e (iii) estudar os efeitos de dietas constituídas com estas leguminosas no consumo voluntário e digestibilidade aparente dos nutrientes em ovinos. O primeiro trabalho (Capítulo 3) refere-se à caracterização in vitro das leguminosas forrageiras (Leucaena leucocephala (Leucena), Arachis pintoi (Amendoim forrageiro), Stylosanthes guianenses cv mineirão (Estilosantes mineirão), Stylosanthes guianenses cv Campo Grande (Estilosantes Campo Grande e Calopogonio sp. (Calopogônio). Foram avaliadas a composição química, a quantificação de taninos, a fermentabilidade ruminal e a síntese microbiana e os resultados obtidos demonstraram valores de proteína bruta compatíveis com a literatura, exceto para o Calopogônio e Estilosantes Campo Grande (< 60 g kg-1 MS). Os teores de taninos variaram significativamente dentre as plantas estudadas, entretanto o teor de tanino condensado pode ser considerado seguro para os animais (entre 30 a 40 eq-g leucocianidina kg-1 MS). As cinco leguminosas apresentaram boa fermentabilidade in vitro, com baixo tempo de colonização (~ 4 h) e T ½ (tempo gasto para atingir metade do valor da Produção Potencial de Gases) inferior a 25 h. A técnica in vitro de incorporação de radiofósforo mostrou efeito significativo da adição de polietileno glicol (PEG) na avaliação da síntese microbiana, demonstrando o efeito dos taninos presentes nas leguminosas estudadas sobra a síntese de proteína no rúmen. O segundo trabalho (Capítulo 4) refere-se à caracterização e avaliação nutricional in vitro de Medicago sativa (Alfafa), Cajanus cajan (Feijão guandu), Mucuna aterrina (Mucuna preta) e Mucuna pluriens (Mucuna cinza). A composição química, a quantificação de taninos e a cinética de fermentação destas leguminosas e dietas experimentais constituídas de fenos de Tifton-85 (Cynodon sp), milho triturado, sal mineralizado na proporção de 30:18:2 com a adição (50%) de cada uma das leguminosas foram estudadas (tratamento ALF, GND, MCZ, MPT respectivamente para Alfafa, Feijão guandu, Mucuna cinza e Mucuna preta). Alfafa apresentou o maior conteúdo de proteína bruta (185 g kg-1 MS) e os teores de taninos variaram significativamente entre as plantas estudadas. Mucuna cinza apresentou maior e Alfafa menor valores de tanino condensado (50 e 0,2 eg. G leucocianidina kg-1 MS respectivamente). Com exceção da Alfafa, todas as outras leguminosas apresentaram incremento de gases quando incubadas in vitro na presença de PEG; o que reflete a atividade biológica dos taninos presente nestas plantas. Em relação às dietas experimentais, apenas as dietas GND e MCZ apresentaram incrementos de gases na presença de PEG. O terceiro trabalho (Capítulo 5) descreve a avaliação nutricional in vivo das dietas experimentais utilizadas no Capítulo 4 (ALF, GND, MCZ, e MPT). Foram utilizados ovinos da raça Santa Inês, machos, castrados com peso vivo médio de 53 ± 5,1 kg. As dietas foram oferecidas de acordo com o peso vivo de cada animal (3%) durante o ensaio de consumo voluntário, enquanto que durante o ensaio de digestibilidade, foi oferecida em cerca de 90% do consumo voluntário determinado anteriormente. Nenhuma dieta apresentou quantidade de taninos condensados considerado prejudicial aos animais. Todas as dietas estudadas apresentaram consumo voluntário e consumo de nutrientes semelhantes. Apenas ALF apresentou maior digestibilidade de PB, enquanto que a MCZ apresentou menor digestibilidade da mesma fração. É concluído com esses trabalhos que as leguminosas estudadas constituem uma importante fonte de nutrientes para os ruminantes, contudo, os teores de taninos condensados devem ser monitorados a fim de que sejam preparadas dietas que não alterem o consumo voluntário nem a fermentação ruminal, e consequentemente disponibilização dos nutrientes aos animais / Leguminous plants constitute an important source of food for the ruminants and they can be explored as direct grazing or, if conserved, for supply in the form of hay or silage. Some of those plants possess phenolic compounds in its composition. The chemical characterization of such plants make possible better use of them in the animal feeding as well as a better understanding of the positive and negative effects of the phenolic compounds in the nutrition of the animals. It was aimed with the present study: (i) to evaluate the chemical composition of five leguminous plants of interest for the feeding of ruminant; (ii) to study the effects of the tannins of different leguminous plants in the in vitro gas production assay; and (iii) to study the effects of diets constituted with these leguminous plants in the voluntary intake and apparent digestibility of the nutrients in sheep. The first work (Chapter 3) refers to the characterization in vitro of the leguminous foragers (Leucaena leucocephala (Leucena), Arachis pintoi (Amendoim forrageiro), Stylosanthes guianenses cv mineirão (Estilosantes mineirão), Stylosanthes guianenses cv Campo Grande (Estilosantes Campo Grande) and Calopogonio sp. (Calopogônio). They were appraised for the chemical composition, the quantification of tannins, the ruminal fermentability and the microbial synthesis. The obtained results demonstrated compatible values for crude protein content, except for Calopogônio and Estilosantes Campo Grande (< 60 g kg-1 DM). The tannin content varied significantly among the studied plants, however the condensed tannin content can be considered safe for the animals (between 30 to 40 eq-g leucocianidin kg-1 DM). The five leguminous plants presented good in vitro fermentability, with low time of colonization (~ 4 h) and T ½ (time spent to reach half of the value of the potential gas production) less than 25 h. The in vitro technique of radio labeled phosphorus incorporation showed significant effect of the addition of polyethylene glycol (PEG) in the evaluation of the microbial synthesis, demonstrating the effect of the presence of tannins in the studied leguminous plants. The second work (Chapter 4) refers to the characterization and in vitro nutritional evaluation of Medicago sativa (Alfalfa), Cajanus cajan (Feijão guandu), Mucuna aterrina (Mucuna preta) and Mucuna pluriens (Mucuna cinza). The chemical composition, the quantification of tannins and the kinetics of fermentation of these leguminous plants and four constituted experimental diets with Tifton-85 hay (Cynodon sp), corn grain, mineralized salt (30:18:2) with the addition (50%) of each one of the leguminous plants were studied (treatment ALF, GND, MCZ, MPT respectively for Alfalfa, Feijão guandu, Mucuna cinza and Mucuna preta). Alfalfa presented the highest content of crude protein (185 g kg-1 DM) and the tannin content varied significantly among the studied plants. Mucuna cinza presented greater and Alfalfa lower values of condensed tannin (50 and 0.2 eg g leucocianidin kg-1 DM respectively). Except for the Alfalfa, all the other leguminous plants presented increment of gases when incubated in vitro in the presence of PEG; what reflects the biological activity of the tannins present in these plants. In relation to the experimental diets, just the diets GND and MCZ presented increments of gases in the presence of PEG. The third work (Chapter 5) describes the in vivo nutritional evaluation of the experimental diets used in the Chapter 4 (ALF, GND, MCZ, and MPT). Santa Inês males, castrated with live weight of 53 ± 5.1 kg sheep were used. The diets were offered in agreement with the liveweight of each animal (3%) during the voluntary intake assay, while during the digestibility trial, it was offered in about 90% of the voluntary intake determined previously. No diet presented amount of condensed tannins considered harmful to the animals. All of the studied diets presented similar voluntary intake and nutrients consumption. Just ALF presented higher crude protein digestibility, while MCZ presented lower digestibility of the same fraction. It is concluded with those works that the studied leguminous plants constitutes an important source of nutrients for the ruminants, however, the condensed tannins content should be monitored so that it can prepare diets to neither alter the voluntary intake nor the ruminal fermentation, and consequently guaranteeing the supply of nutrients to the animals

A soja tolerante ao herbicida glifosato é o vegetal geneticamente modificado mais cultivado. Entretanto, questões sobre a biossegurança dos alimentos GM são ainda levantadas, como as incertezas sobre a expressão das novas proteínas, mutações indesejadas, alterações de perfis nutricionais e aparecimento de compostos tóxicos. Apesar dos comprovados efeitos benéficos à saúde, a soja apresenta naturalmente, entre suas proteínas, fatores antinutricionais, que podem: provocar efeitos fisiológicos adversos; diminuir a biodisponibilidade de nutrientes; e induzir reações de hipersensibilidade. Paralelamente, a soja possui sabor e aroma desagradáveis por ação de enzimas lipoxigenases. Os fatores antinutricionais estão relacionados às aglutininas (lectinas) e aos inibidores de proteases (inibidor de tripsina, tipo Kunitz, e inibidor de tripsina e quimotripsina, tipo Bowman-Brik), enquanto as globulinas da soja respondem pelas reações de hipersensibilidade. Objetivou-se neste trabalho fazer a comparação dos mapas protéicos de soja GM, suas isolinhas convencionais e sojas orgânicas visando a detecção de alterações nos perfis protéicos destes diferentes tipos de cultivo, e também a análise da expressão dos fatores antinutricionais, como os inibidores de proteases e aglutininas, considerando a extensão das variações naturais existentes nas amostras. Para tanto, foram comparadas seis amostras de sementes de variedades comerciais de soja cultivadas em paralelo, sob as mesmas condições ambientais e de solo, compostas por três isolinhas genitoras e suas três correspondentes GMs e duas amostras orgânicas, sendo que uma delas é comercial e a outra ainda esta em campos de pesquisa, fornecidas pela Embrapa Soja. Foram analisados extratos protéicos de todas as amostras, após extração com ácido tricloroacético (TCA) e acetona, através de eletroforese unidimensional (1D) e bidimensional (2D). Nestas foi empregado gradiente de pH de 3-10 e as imagens avaliadas pelo software ImageMaster 2D Platinum. Diversos spots selecionados foram identificados por espectrometria de massas. Nas imagens dos géis 2D, foi possível identificar e quantificar os spots correspondentes às proteínas isoladas e não houve diferença estatística ao nível de significância de 5% entre os diferentes tipos de cultivo. Na verificação da sobreposição dos géis, obtivemos porcentagens de matchings superiores a 70% entre as amostras GMs e não GMs. As amostras orgânicas apresentaram % matchings menores que entre convencionais e GMs. Nos resultados da espectrometria de massas foi possível reconhecer os principais grupos protéicos da soja, como as frações e subunidades de &#946;-conglicinina e de glicinina, bem como os inibidores de proteases, aglutininas e lipoxigenases e não foi possível perceber alterações na expressão dos peptídeos identificados e analisados. Podemos concluir que as variações encontradas entre as três amostras convencionais e entre as amostras dos grupos convencionais e orgânicas foram maiores que a comparação das amostras GMs com suas genitoras correspondentes. / The glyphosate tolerant soybean is the most cultivated GM plant. However, questions about the bio-safety of GM foods are still rising, as there are uncertainty about the expression of new proteins, undesired mutations, changes in nutritional profile and the production of toxic compounds. Despite of the beneficial effects to human health, the soybean has anti-nutritional factors that can cause adverse physiological effects, reduce the bioavailability of nutrients, and induce hypersensitivity reactions. At the same time the soybean develop undesirable flavor due to the action of lipoxygenases. The anti-nutritional factors are related to agglutinins (lectins) and to the proteases inhibitors (Kunitz\'s Trypsin Inhibitor and Bowman-Birk\'s Inhibitor of Trypsin and Chymotrypsin) while the soybean globulins are responsible for hypersensitivity reactions. The objective of this work was to compare proteic maps of GM soybean, conventional isolines and organic soybean aiming to detect changes in protein profiles of the different types of cultivation and also analyze the expression of the anti-nutritional factors, such as protease inhibitors and agglutinins, taking into consideration the natural variation existing in the samples. For this, it was performed the comparisons between six seed samples of commercial varieties of soybeans, grown in parallel under the same environmental conditions and soil, composed of three parental isolines and their three corresponding GM and two organic sample, one of them is commercial and the other is still in search fields. The samples were provided by Embrapa Soja. Protein extracts were analyzed from the samples after extraction with trichloroacetic acid (TCA) and acetone using regular monodimensional electrophoresis (1D) and two-dimensional (2D) ones. For 2D were used strips of pH gradient 3-10 and the final images analyzed by ImageMaster 2D Platinum software. Several selected spots were identified by mass spectrometry. In the images of the 2D gels, we could identify and quantify the spots corresponding to proteins isolated and there was no statistical difference at 5% of significance between the different types of cultivation. Checking the overlap of the gels, we obtained matchings above 70% between GM and non GM sample. The organic sample had lower matching index between conventional and GM. It was possible to recognize the major groups of soy protein as a result of mass spectrometry such as¨&#946;-conglycinin fractions and glycinin as well as protease inhibitors, lipoxygenase, and agglutinins. We concluded that the variations found among the tree conventional samples and between samples of conventional and organic groups were higher than the comparison of sample GMs with their corresponding parentals.

Submitted by Franciele Moreira (francielemoreyra@gmail.com) on 2018-06-08T13:46:39Z No. of bitstreams: 2 Dissertação - Juliana Aparecida Correia Bento - 2018.pdf: 3915998 bytes, checksum: 02825a5c8f16825aad6955538d1369a8 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Approved for entry into archive by Luciana Ferreira (lucgeral@gmail.com) on 2018-06-08T15:06:59Z (GMT) No. of bitstreams: 2 Dissertação - Juliana Aparecida Correia Bento - 2018.pdf: 3915998 bytes, checksum: 02825a5c8f16825aad6955538d1369a8 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Made available in DSpace on 2018-06-08T15:06:59Z (GMT). No. of bitstreams: 2 Dissertação - Juliana Aparecida Correia Bento - 2018.pdf: 3915998 bytes, checksum: 02825a5c8f16825aad6955538d1369a8 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Previous issue date: 2018-02-28 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / Cochlospermum regium, a native plant of the Brazilian Cerrado, known as “Algodãozinho-do- campo”, is used in folk medicine for the treatment of infections and inflammation. Jatropha elliptica (Pohl) Muell Arg is a plant belonging to the Euphorbiaceae family, native to the Brazilian Cerrado, and popularly known as "Batata-de-teiú". There is a lack of scientific knowledge about the physico-chemical and technological aspects of Jatropha elliptic and its possible applications, in foods and or medicines. Hedychium coronarium Koen, native to Nepal, is considered an invasive plant in Brazil, as it presents rapid and negative growth for plant biodiversity. Rhizomes are man-made due to their medicinal properties, such as the treatment of headache, cancer and inflammation, but it can also be a potential source for starch extraction. Algodãozinho-do-campo, Batata-de-teiú and lírio-do-brejo are rhizomatous plants with the potential to obtain starch and flour. The objective of this work was to contribute with unpublished data on the characterization of flour and starch with a probable functional value, extracted from Hedychium coronarium Koen, Cochlospermum regium and Jatropha elliptica (Pohl) Muell Arg, as well as the application of these products as substrate in the semi-solid fermentation, aiming the production of enzymatic extracts, and to evaluate the effect of the ultrasound on the lírio-do-brejo starch. The extracted starches present peculiar technological characteristics, being: the algodãozinho-do-campo starch under cooking temperature absorbs less water and solubilizes less than the corn starch, maintaining more the structure of the granules in the final product, being able to contribute to the improvement of final product texture; the batata-de-teiú starch had a low tendency to retrograde; the lírio- do-brejo starch presented high amylose content (about 59.16%), crystallinity of 19.30%, and when it was submitted to ultrasound for 15 min at 62% intensity and 12.5 g 100g-1 an increase in viscosity peak (from 1917 to 2257 cP) and reduction in tendency to retrograde (from 2079 to 1295 cP) was observed. The flours presented peculiar nutritional characteristics, antioxidant capacity, presence of phenolic compounds, low toxicity in Artêmia salina, presence of antinutritional factors, and were not efficient for the production of enzymatic extracts by semi-solid fermentation. It is concluded that they are plants with potential to obtain starch and flour. / Cochlospermum regium, planta nativa do cerrado brasileiro, conhecida como algodãozinho- do-campo, é utilizada na medicina popular para o tratamento de infecções e inflamações. Jatropha elliptica (Pohl) Muell Arg é uma planta pertencente à família das Euphorbiaceae, nativa do cerrado brasileiro, e conhecida popularmente como “batata-de-teiú”, sendo que existe carência de conhecimentos científicos sobre os aspectos fisíco-quimicos e tecnológicos da Jatropha elliptica e suas possíveis aplicações, em alimentos e ou medicamentos. O lírio-do- brejo (Hedychium coronarium Koen), natural do Nepal, é considerado planta invasora no Brasil, pois apresenta crescimento rápido e negativo para a biodiversidade vegetal. Os rizomas são aproveitados pelo homem, devido suas propriedades medicinais, como o tratamento da cefaleia, câncer e inflamações, mas também pode ser uma fonte potencial para a extração de amido. Algodãozinho-do-campo, batata-de-teiú e lírio-do-brejo são plantas rizomatosas com potencial para a obtenção de amido e farinha. Com isso, este trabalho tem por objetivo contribuir com dados inéditos sobre a caracterização das farinhas e féculas, com provável valor funcional, extraídas do Hedychium coronarium Koen, Cochlospermum regium e Jatropha elliptica (Pohl) Muell Arg, bem como a aplicação destes produtos como substrato na fermentação semissólida, visando a produção de extratos enzimáticos, e avaliar o efeito do ultrassom sobre o amido de lírio-do-brejo. Os amidos extraídos apresentam características tecnológicas peculiares, sendo que: o amido de algodãozinho-do-campo sob temperatura de cozimento absorve menos água e se solubiliza menos que o amido de milho, mantendo mais a estrutura dos gânulos no produto final, podendo contribuir para a melhoria da textura do produto final; o amido de batata-de-teiú apresentou baixa tendência à retrogradação; o amido de lírio-do-brejo apresentou alto teor de amilose (cerca de 59.16%), cristalinidade de 19.30%, sendo que quando este foi submetido ao ultrassom, por 15 min, a 62% de intensidade e 12.5 g 100g-1 de concentração, observou-se um aumento do pico de viscosidade (de 1917 para 2257 cP) e a redução da tendência à retrogradação (de 2079 para 1295 cP). As farinhas apresentaram caracteristicas nutricionais peculiares, capacidade antioxidante, presença de compostos fenólicos, baixa toxicidade em ensaio com Artêmia salina, presença de fatores antinutricionais, e não foram eficientes para a produção de extratos enzimáticos por fermentação semissólida. Conclui-se que são plantas com potencial para a obtenção de amido e farinhas.

As leguminosas constituem uma importante fonte de alimentos para os ruminantes e podem ser exploradas para pastejo direto ou, se conservadas, para fornecimento na forma de feno ou silagem. Algumas leguminosas possuem compostos fenólicos em sua composição. A caracterização química dessas as plantas possibilita melhor uso das mesmas na alimentação animal assim como um melhor entendimento dos efeitos positivos e negativos dos compostos fenólicos na nutrição dos animais. Objetivou-se com o presente estudo: (i) avaliar a composição química de cinco leguminosas de interesse para a alimentação de ruminantes; (ii) estudar os efeitos dos taninos de diferentes leguminosas na produção de gases in vitro; e (iii) estudar os efeitos de dietas constituídas com estas leguminosas no consumo voluntário e digestibilidade aparente dos nutrientes em ovinos. O primeiro trabalho (Capítulo 3) refere-se à caracterização in vitro das leguminosas forrageiras (Leucaena leucocephala (Leucena), Arachis pintoi (Amendoim forrageiro), Stylosanthes guianenses cv mineirão (Estilosantes mineirão), Stylosanthes guianenses cv Campo Grande (Estilosantes Campo Grande e Calopogonio sp. (Calopogônio). Foram avaliadas a composição química, a quantificação de taninos, a fermentabilidade ruminal e a síntese microbiana e os resultados obtidos demonstraram valores de proteína bruta compatíveis com a literatura, exceto para o Calopogônio e Estilosantes Campo Grande (< 60 g kg-1 MS). Os teores de taninos variaram significativamente dentre as plantas estudadas, entretanto o teor de tanino condensado pode ser considerado seguro para os animais (entre 30 a 40 eq-g leucocianidina kg-1 MS). As cinco leguminosas apresentaram boa fermentabilidade in vitro, com baixo tempo de colonização (~ 4 h) e T ½ (tempo gasto para atingir metade do valor da Produção Potencial de Gases) inferior a 25 h. A técnica in vitro de incorporação de radiofósforo mostrou efeito significativo da adição de polietileno glicol (PEG) na avaliação da síntese microbiana, demonstrando o efeito dos taninos presentes nas leguminosas estudadas sobra a síntese de proteína no rúmen. O segundo trabalho (Capítulo 4) refere-se à caracterização e avaliação nutricional in vitro de Medicago sativa (Alfafa), Cajanus cajan (Feijão guandu), Mucuna aterrina (Mucuna preta) e Mucuna pluriens (Mucuna cinza). A composição química, a quantificação de taninos e a cinética de fermentação destas leguminosas e dietas experimentais constituídas de fenos de Tifton-85 (Cynodon sp), milho triturado, sal mineralizado na proporção de 30:18:2 com a adição (50%) de cada uma das leguminosas foram estudadas (tratamento ALF, GND, MCZ, MPT respectivamente para Alfafa, Feijão guandu, Mucuna cinza e Mucuna preta). Alfafa apresentou o maior conteúdo de proteína bruta (185 g kg-1 MS) e os teores de taninos variaram significativamente entre as plantas estudadas. Mucuna cinza apresentou maior e Alfafa menor valores de tanino condensado (50 e 0,2 eg. G leucocianidina kg-1 MS respectivamente). Com exceção da Alfafa, todas as outras leguminosas apresentaram incremento de gases quando incubadas in vitro na presença de PEG; o que reflete a atividade biológica dos taninos presente nestas plantas. Em relação às dietas experimentais, apenas as dietas GND e MCZ apresentaram incrementos de gases na presença de PEG. O terceiro trabalho (Capítulo 5) descreve a avaliação nutricional in vivo das dietas experimentais utilizadas no Capítulo 4 (ALF, GND, MCZ, e MPT). Foram utilizados ovinos da raça Santa Inês, machos, castrados com peso vivo médio de 53 ± 5,1 kg. As dietas foram oferecidas de acordo com o peso vivo de cada animal (3%) durante o ensaio de consumo voluntário, enquanto que durante o ensaio de digestibilidade, foi oferecida em cerca de 90% do consumo voluntário determinado anteriormente. Nenhuma dieta apresentou quantidade de taninos condensados considerado prejudicial aos animais. Todas as dietas estudadas apresentaram consumo voluntário e consumo de nutrientes semelhantes. Apenas ALF apresentou maior digestibilidade de PB, enquanto que a MCZ apresentou menor digestibilidade da mesma fração. É concluído com esses trabalhos que as leguminosas estudadas constituem uma importante fonte de nutrientes para os ruminantes, contudo, os teores de taninos condensados devem ser monitorados a fim de que sejam preparadas dietas que não alterem o consumo voluntário nem a fermentação ruminal, e consequentemente disponibilização dos nutrientes aos animais / Leguminous plants constitute an important source of food for the ruminants and they can be explored as direct grazing or, if conserved, for supply in the form of hay or silage. Some of those plants possess phenolic compounds in its composition. The chemical characterization of such plants make possible better use of them in the animal feeding as well as a better understanding of the positive and negative effects of the phenolic compounds in the nutrition of the animals. It was aimed with the present study: (i) to evaluate the chemical composition of five leguminous plants of interest for the feeding of ruminant; (ii) to study the effects of the tannins of different leguminous plants in the in vitro gas production assay; and (iii) to study the effects of diets constituted with these leguminous plants in the voluntary intake and apparent digestibility of the nutrients in sheep. The first work (Chapter 3) refers to the characterization in vitro of the leguminous foragers (Leucaena leucocephala (Leucena), Arachis pintoi (Amendoim forrageiro), Stylosanthes guianenses cv mineirão (Estilosantes mineirão), Stylosanthes guianenses cv Campo Grande (Estilosantes Campo Grande) and Calopogonio sp. (Calopogônio). They were appraised for the chemical composition, the quantification of tannins, the ruminal fermentability and the microbial synthesis. The obtained results demonstrated compatible values for crude protein content, except for Calopogônio and Estilosantes Campo Grande (< 60 g kg-1 DM). The tannin content varied significantly among the studied plants, however the condensed tannin content can be considered safe for the animals (between 30 to 40 eq-g leucocianidin kg-1 DM). The five leguminous plants presented good in vitro fermentability, with low time of colonization (~ 4 h) and T ½ (time spent to reach half of the value of the potential gas production) less than 25 h. The in vitro technique of radio labeled phosphorus incorporation showed significant effect of the addition of polyethylene glycol (PEG) in the evaluation of the microbial synthesis, demonstrating the effect of the presence of tannins in the studied leguminous plants. The second work (Chapter 4) refers to the characterization and in vitro nutritional evaluation of Medicago sativa (Alfalfa), Cajanus cajan (Feijão guandu), Mucuna aterrina (Mucuna preta) and Mucuna pluriens (Mucuna cinza). The chemical composition, the quantification of tannins and the kinetics of fermentation of these leguminous plants and four constituted experimental diets with Tifton-85 hay (Cynodon sp), corn grain, mineralized salt (30:18:2) with the addition (50%) of each one of the leguminous plants were studied (treatment ALF, GND, MCZ, MPT respectively for Alfalfa, Feijão guandu, Mucuna cinza and Mucuna preta). Alfalfa presented the highest content of crude protein (185 g kg-1 DM) and the tannin content varied significantly among the studied plants. Mucuna cinza presented greater and Alfalfa lower values of condensed tannin (50 and 0.2 eg g leucocianidin kg-1 DM respectively). Except for the Alfalfa, all the other leguminous plants presented increment of gases when incubated in vitro in the presence of PEG; what reflects the biological activity of the tannins present in these plants. In relation to the experimental diets, just the diets GND and MCZ presented increments of gases in the presence of PEG. The third work (Chapter 5) describes the in vivo nutritional evaluation of the experimental diets used in the Chapter 4 (ALF, GND, MCZ, and MPT). Santa Inês males, castrated with live weight of 53 ± 5.1 kg sheep were used. The diets were offered in agreement with the liveweight of each animal (3%) during the voluntary intake assay, while during the digestibility trial, it was offered in about 90% of the voluntary intake determined previously. No diet presented amount of condensed tannins considered harmful to the animals. All of the studied diets presented similar voluntary intake and nutrients consumption. Just ALF presented higher crude protein digestibility, while MCZ presented lower digestibility of the same fraction. It is concluded with those works that the studied leguminous plants constitutes an important source of nutrients for the ruminants, however, the condensed tannins content should be monitored so that it can prepare diets to neither alter the voluntary intake nor the ruminal fermentation, and consequently guaranteeing the supply of nutrients to the animals