Objectives- The purpose of this study was to determine if differences in breast milk (BM) short chain fatty acid (SCFA) and microbiome profiles are correlated to maternal BMI. Our hypothesis is that BM SCFA are a reflection of colonic SCFA distribution and concentration and may reflect microbiome diversity in the maternal gut. Methods- Study design-This was a cohort study in which forty-six Caucasian participants were recruited from BABE Breastfeeding Coalition of Tri-Cities, divided into two groups, one group with normal pre-gravid BMI between 18.5 and 24.9 kg/m2 (n=23) and the other group with overweight or obese pre-gravid BMI greater than 25.0 kg/m2 (n=23). Each participant completed a demographic and health survey and provided 4 ounces of expressed BM. This study was approved by the ETSU IRB (0915.8s-ETSU). 16s rRNA Isolation & Quantification- Microbiome analysis was performed on thirty-four samples (n=13 for overweight/obese, and n=21 for normal weight). Qiagen QIAmp PowerFecal Pro DNA Kit was utilized for isolation of microbiome DNA; Amplicon sequencing of the 16S rRNA region was performed at the University of Tennessee Genomics Core Laboratory utilizing a modified Klindworth et al method. Microbiome Analysis- Operational Taxonomic Unit (OUT) clustering and taxonomic analysis were performed using CLC Genomics Workbench. Alpha diversity indexes were calculated using the Abundance Analysis tool, and the weighted Unifrac metric was used to calculate Beta diversity. Fatty Acid Profile- BM samples were subjected to SCFA extraction and analysis using a modified Schwiertz et al. method. The resulting SCFA profiles were then utilized to determine if there were any significant differences between groups. Results- No significance was observed in BM microbiome between the normal weight and overweight/obese groups for alpha or beta diversity. Significance was detected between the groups for valeric (p=0.02) and isocaproic acids (p=0.05) with the normal weight group higher than the overweight/obese group. No significance was observed for any of the other SCFAs. Conclusions- Although these results are not significant due to low sample size and lack of diversity, they potentially offer insights into the impact of maternal BMI on microbiome and SCFA profiles, which can have implications for infant health and development. Funding Sources- ETSU Small RDC Grant

Bisphenols are used in the process of polymerization of polycarbonate plastics and epoxy resins. Bisphenols can easily migrate out of plastic products and enter the gastrointestinal system. By increasing colonic inflammation in mice, disrupting the intestinal bacterial community structure and altering the microbial membrane transport system in zebrafish, bisphenols seem to interfere with the gut microbiome. The highly abundant human commensal bacterium Bacteroides thetaiotaomicron was exposed to bisphenols (Bisphenol A (BPA), Bisphenol F (BPF), Bisphenol S (BPS)), to examine the mode of action, in particular of BPF. All chemicals caused a concentration-dependent growth inhibition and the half-maximal effective concentration (EC50) corresponded to their individual logP values, a measure of their hydrophobicity. B. thetaiotaomicron exposed to BPF decreased membrane fluidity with increasing BPF concentrations. Physiological changes including an increase of acetate concentrations were observed. On the proteome level, a higher abundance of several ATP synthase subunits and multidrug efflux pumps suggested an increased energy demand for adaptive mechanisms after BPF exposure. Defense mechanisms were also implicated by a pathway analysis that identified a higher abundance of members of resistance pathways/strategies to cope with xenobiotics (i.e., antibiotics). Here, we present further insights into the mode of action of bisphenols in a human commensal gut bacterium regarding growth inhibition, and the physiological and functional state of the cell. These results, combined with microbiota-directed effects, could lead to a better understanding of host health disturbances and disease development based on xenobiotic uptake.

Chronic infection with hepatitis B virus (HBV) is a primary risk factor for the development of hepatocellular carcinoma (HCC). HCC is the fifth most common cancer type worldwide with few treatment options. The hepatitis B encoded x antigen (HBx) plays a crucial role in the pathogenesis of HCC through several mechanisms. HBx alters signaling pathways shown to promote carcinogenesis and mediates epigenetic changes that silence tumor suppressor genes and activate host oncogenes. Short chain fatty acids (SCFAs) are made by selected gut bacteria with largely anti-inflammatory properties. They alter gene expression by functioning as histone deacetylase inhibitors (HDACi) and can bind to G protein coupled receptors (GCPR) to stimulate signaling pathways. Due to the documented anti-cancer properties of SCFAs, experiments were designed to test the hypothesis that SCFAs delay the development of HCC in HBx transgenic (HBxTg) mice. A diet of SCFAs was fed to HBxTg for three months prior to the expected appearance of dysplastic nodules and HCC. The results showed a statistically significant reduction in the number of dysplastic nodules as well as the presence and frequency of HCC. The effect of SCFAs on tumor growth was also evaluated in nude mice subcutaneously injected with human HCC cells. Tumor size in SCFA-treated mice was statistically smaller compared to the controls. The effect of SCFAs on cell viability of cancer and primary human hepatocytes was evaluated. SCFAs were shown to reduce cell viability in cancer cells only, with no effect on primary hepatocytes. Proteomics was performed on SCFA-treated compared to control livers from HBxTg to investigate changes on the molecular level that are associated with reduced preneoplastic and neoplastic nodule formation. Pathway analysis showed a decrease in important cancer-promoting pathways altered by HBx in HCC, including inflammation, oxidative stress, PI3K, VEGF, EGF, and Ras. These pathways are involved in biological processes central to carcinogenesis such as cell proliferation, survival, and angiogenesis. The ability of SCFAs to decrease these pathways has never been demonstrated. Further investigation confirmed that Ras activity was decreased in 12-month old livers treated with SCFAs. Taken together, these results show that SCFAs are capable of delaying the rate of tumor growth and tumor frequency in two mouse models of HBV-associated HCC, as well as reduce cell viability in cancer cells specifically. This data suggests that SCFAs may be a novel treatment option for HBV-associated HCC. / Biology

Introduction: Microbial fermentation of carbohydrates in forestomach of ruminants produces large amounts of short-chain fatty acids (SCFA, mainly acetic acid, propionic acid, and n-butyric acid). The majority of these substrates is taken up directly across the ruminal wall. After luminal uptake into the epithelial cells, SCFA mainly occur in the dissociated form due to the intracellular pH of ~7.4. Moreover, a big portion of SCFA is metabolised within the cytosol. Main end products of epithelial SCFA metabolism are ketone bodies (D-3-hydroxybutyric acid and acetoacetic acid) and lactic acid. Both intact SCFA and ketone bodies and lactate need to be efficiently extruded from the ruminal epithelial cells to prevent a lethal drop of intracellular pH and counteract osmotic load of the cytosol. All these substances are less lipophilic in comparison to the undissociated form of SCFA. Thus, dissociated SCFA (SCFA-) and their metabolites need Protein mediated mechanisms for the extrusion across the basolateral side of ruminal epithelium. One mechanism suggested to be involved in the extrusion of SCFA- across basolateral membrane of the ruminal epithelium is the monocarboxylate transporter 1 (MCT1). Functionally, MCT1 was first assumed to operate as proton-coupled transporter for monocarboxylates including SCFA. Nonetheless, a recent study found a bicarbonate dependent anion exchange mechanism which turned out to be sensitive to MCT1 Inhibitors at the basolateral side of the ruminal epithelium pointing to the ability of MCT1 to act as an anion exchanger. However, in these experiments the inhibition of MCT1 abolished bicarbonate dependent transport only by half. This suggests the involvement of further anion exchanger(s) in the transport of SCFA across the basolateral membrane of ruminal epithelium. Promising candidates to underlie this exchange are the putative Anion exchanger 1 (PAT1) and a transport protein designated „down-regulated in adenoma“ (DRA). Materials and Methods: Sheep rumen epithelium was mounted in Ussing Chambers under short-circuit conditions. Radioactively labelled acetate (ac) was added to the serosal side. Serosal to mucosal flux of ac (Jsm ac) was measured with or without anion Exchange inhibitors (50 mM NO3- or 1 mM DIDS) or the MCT1 inhibitor p-hydroxy mercuribenzoic acid (pHMB; 1.5 mM) in the serosal buffer solution. The inhibitors were added alone or in combination with each other. Furthermore, mucosal to serosal flux of radioactivelly labelled ac or butyrate (bu) (Jms ac, bu) was measured in the presence or absence of SO42-, Cl- or NO3- (50 mM respectively) as exchange substrate in the serosal buffer solution. Immunohistochemical staining was conducted to locate PAT1 and DRA by use of commercially available antibodies. Results: NO3- and pHMB significantly reduced Jsm ac by 57 % and 51 %, respectively. When pHMB was applied after pre-incubation with NO3- an additional inhibition of Jsm ac was observed. Vice versa, NO3- further inhibited Jsm ac when epithelia were pre-incubated with pHMB before. DIDS had no inhibitory effect on SCFA flux. Serosal presence of SO42- or Cl- enhanced Jms ac significantly. Regarding bu, Cl- or SO4 2- also enhanced Jms bu significantly. The different anions available in the serosal buffer solution numerically enhanced Jms in the order of SO4 2- > Cl- for both ac and bu, which corresponds to the known affinity sequence of PAT1 and DRA. Immunohistochemistry revealed localization of PAT 1 in the stratum basale, whereas DRA was not detectable using this method. Conclusions: Basically, this study supports the suggestion that MCT1 works as an Anion exchanger in ruminal epithelium. In addition, it clearly shows that there is at least one further anion exchanger involved in the basolateral extrusion of SCFA and their metabolites. The functional and immunohistochemical findings suggest that PAT1 holds a significant role in this respect.

As leveduras Saccharomyces cerevisiae são consideradas importantes matérias primas na nutrição animal pela sua capacidade prebiótica. Os prebióticos são compostos não digeridos pelo organismo animal, mas que são fermentados pelos microrganismos do trato gastrintestinal, cujos produtos são capazes de prover benefícios ao hospedeiro. A fermentação depende de fatores como: substrato utilizado para o seu crescimento, método de fermentação, modo e condição de secagem e idade das células. Assim, os processos produtivos modernos têm como intuito a produção de leveduras com elevado potencial prebiótico. Este estudo objetivou avaliar os efeitos de teores crescentes de leveduras com metabólitos ativos (LSC; baseada na fermentação de substratos específicos) na digestibilidade aparente dos nutrientes da dieta, microbiota e produtos da fermentação fecal e parâmetros imunológicos de gatos adultos. Foram utilizados 27 gatos adultos, idade média de 9,44±5,35 anos, machos e fêmeas, sem raça definida e saudáveis. Os animais foram distribuídos em delineamento em blocos casualizados (idade), constituído de três tratamentos experimentais, denominados: DC (dieta controle), LSC 0,3 (dieta controle com 0,3% de leveduras com metabólitos ativos) e LSC 0,6 (dieta controle com 0,6% de leveduras com metabólitos ativos). Os resultados obtidos foram analisados através do programa computacional Statistical Analysis System (SAS Institute Inc., 2004), sendo considerados significativos valores de p<0,05 e as médias comparadas pelo teste de Tukey. Verificou-se que a inclusão do aditivo alterou apenas a digestibilidade aparente da fibra bruta, da matéria mineral e energia metabolizável (p<0,05). Já em relação aos produtos da fermentação e microbiota das fezes, observou-se redução em ácido lático (p=0,0040) e Clostridium perfringens (p=0,0226) com a inclusão do prebiótico e diminuição do ácido isovalérico (p=0,0144) no tratamento LSC 0,3. Pode-se concluir que o aditivo, nos teores de inclusão avaliados, parece apresentar potencial prebiótico em relação aos produtos da fermentação e microbiota fecal / Saccharomyces cerevisiae yeast are considered important raw materials in animal nutrition due their prebiotic capacity. Prebiotics are compounds not digested by animal organism, but are fermented by microorganisms in the gastrointestinal tract, which products are capable of providing benefits to the host. The fermentation depends on factors such as: substrate used for growth, fermentation method, way and drying condition and age of the cells. Thus, modern production processes have the objective of producing yeast with high prebiotic potential. This study aimed to evaluate the effects of increasing levels of yeast with active metabolites (LSC) based on the fermentation of specific substrates on apparent digestibility of diet nutrients, microbiota and fecal fermentation products and immunological parameters in adult cats. Twenty seven male or female cats with mean weight of 4.19 ± 0.83kg and mean age of 9.44 ± 5.35 years were used and distributed in an unbalanced randomized block design (age), consisting of three experimental treatments, DC (control diet), LSC 0.3 (control diet with 0.3% yeast with active metabolites) and LSC 0.6 (control diet with 0.6% yeast with active metabolites). The results were analyzed using the computer program Statistical Analysis System (SAS Institute Inc., 2004), with significance level of p<0.05 and the averages compared by Tukey test. The inclusion of the additive only changed the apparent digestibility of crude fiber and mineral content (p<0.05). Regarding the fermentation products and microbiota of feces, there was a reduction in lactic acid (p=0,0040) and Clostridium perfringens (p=0,0226) with inclusion of prebiotic and decreased of isovalerate (p=0,0144) in the LSC 0.3 treatment. It can be concluded that the additive, in the levels of inclusion assessed, seems to have prebiotic potential on fecal fermentation products and microbiota

Os grãos de linhaça e amaranto recebem grande atenção da população em decorrência de suas alegações nutricionais e funcionais à saúde. Uma vez que estes grãos apresentam significativos teores de fibras alimentares (28 e 10%, respectivamente), os efeitos fisiológicos benéficos associados a estas frações foram investigados por meio de parâmetros do metabolismo colônico (fermentativos) e lipídico em ensaio biológico com ratos (Wistar), divididos em quatro grupos (n=12) durante 28 dias. As dietas dos grupos Referência e Controle apresentaram 7% (p/p) de celulose, e as dietas dos grupos experimentais foram formuladas com: farinha desengordurada de linhaça; amaranto extrusado, de modo que apresentassem 7% de fibra alimentar (FA). Com exceção da dieta do grupo Referência, as demais apresentaram 0,5% de colesterol (p/p). Foram observados para os grupos Linhaça e Amaranto: aumento do peso do ceco e conteúdo; aumento da massa fecal úmida; aumento dos ácidos graxos de cadeia curta (AGCC) e diminuição do pH no conteúdo do ceco, indicando fermentação colônica. Os animais do grupo Linhaça apresentaram menor peso e colesterol hepático, porém, nenhuma alteração no colesterol plasmático e na excreção de colesterol e ácidos biliares nas fezes. Já os animais do grupo Amaranto apresentaram menor peso hepático, porém, nenhuma alteração do colesterol total do órgão. Neste grupo também não foi observada redução do colesterol plasmático, acompanhada do aumento da excreção do colesterol e redução dos ácidos biliares nas fezes. Sugere-se que o trofismo da parede do ceco está relacionado à produção de ácido butírico, principal substrato energético dos colonócitos, que contribui para uma mucosa mais resistente a patógenos e carcinógenos. O mecanismo envolvido no efeito hepatoprotetor observado no grupo Linhaça pode estar relacionado à produção de ácido propiônico no cólon, uma vez que a literatura sugere que o mesmo pode estar envolvido na inibição da HMG-CoA redutase. Desconhecem-se, até o momento, os mecanismos que promoveram a menor excreção de ácidos biliares nas fezes no grupo Amaranto. De qualquer modo, as FAs do grão de linhaça e amaranto não foram capazes de promover efeito hipercolesterolemiante nas condições em que foi realizado este estudo. Conclui-se que as frações indigeríveis da linhaça e amaranto são benéficas para a saúde da mucosa intestinal. / The flaxseed and amaranth grains are recognized by their nutritional and functional health attributes. These grains have significant contents of dietary fiber (28 and 10%, respectively), and the physiological effects related to these fractions were investigated by colonic fermentative and lipid metabolism parameters with rats (Wistar) distributed in five groups (n=12) during twenty-eight days. The diets of the Reference and Control groups contained 7% (w/w) of cellulose and the diets of the experimental groups were formulated with: defatted flaxseed flour and defatted extruded amaranth, in order to provide 7% dietary fiber (DF). With the exception of the Reference, the other diets received 0,5% cholesterol. The Flaxseed and Amaranth groups compared to the Control showed: greater weight of caecum and its contents; greater weight of feces (wet mass); increase of short chain fatty acids (SCFA) concentration and decrease of pH of cecal content, indicating colonic fermentation. The Flaxseed group showed lower liver hepatic weight and cholesterol, but no changes in plasmatic cholesterol, cholesterol fecal excretion, or bile acids fecal excretion. The Amaranth group exhibited lesser hepatic weight, along with no changes on hepatic cholesterol. Also, no plasma cholesterol reduction was observed, with the increase of fecal cholesterol excretion and decrease in fecal bile acid excretion. It is suggest that the caecum wall trophism was related to butiric acid production, main energy colonic cell substrate that fosters a more resistant mucosa to pathogens and carcinogens. The mechanism involved in liver protecting effect, observed in the Flaxseed group, could be related to propionic acid production, since that it is suggest in the literature that this fatty acid could be envolved in the HMG-CoA reductase inhibition. Until this moment, the mechanisms that promoted the lesser bile acid excretion by the Amaranth group were not kwon. The DFs of flaxseed and amaranth grains were unable to promote hypocholestelolemic effects under the present assay conditions. It could be concluded that DFs of flaxseed and amaranth grains are beneficial to the health of the intestinal mucosa.

Os objetivos destes trabalhos foram avaliar a inclusão de altos níveis de monensina em suplementos proteico de baixo consumo para bovinos sobre (i) desempenho; (ii) fermentação e degradabilidade ruminal e (iii) produção de metano. Para tanto, foram realizados três experimentos distintos. Os experimentos realizados são apresentados na forma de capítulos. Nos Capítulos I e II são apresentados a introdução e a revisão de literatura. No Capítulo III (desempenho), foram utilizados 64 novilhos, distribuídos aleatoriamente em quatro tratamentos: 0, 400, 800 e 1200 mg de monensina sódica/ kg de suplemento proteico. Os animais permaneceram em quatro piquetes de Brachiaria decumbens onde eram submetidos a rodízios semanais. Amostras de pasto foram colhidas para determinação das variáveis qualitativas e quantitativas da forragem. Foram estimados o consumo médio de suplemento, consumo individual de suplemento e ganho de peso médio diário. A inclusão de monensina nos níveis estudados diminuiu o consumo de suplemento e piorou o desempenho animal. No Capítulo IV (fermentação e degradabilidade ruminal), foram utilizados quatro bovinos da raça nelore, canulados no rúmen, em experimento com delineamento quadrado latino 4x4 por 84 dias. Os animais foram arraçoados com dieta composta de feno de Brachiaria decumbens (à vontade), 500 g/cabeça/dia de suplemento proteinado de baixo consumo e monensina sódica conforme os tratamentos: 0, 200, 400 e 600 mg/dia. Os parâmetros ruminais avaliados foram: produções de ácidos graxos de cadeia curta (AGCC); concentração de nitrogênio amoniacal; pH; consumo de matéria seca; degradabilidade in situ da MS e FDA do feno de B. decumbens. Os valores encontrados para a fermentação ruminal (pH, N-NH3 e AGCC) e degradabilidade in situ da MS não foram afetados pelos tratamentos. O aumento dos níveis de monensina diminuíram linearmente o consumo de matéria seca e a degradabilidade potencial da fibra. No Capítulo V (produção de metano), quatro níveis de monensina foram testados em ensaio in vitro de produção de gases com o objetivo de avaliar a cinética fermentativa, produção de AGCC, degradabilidade da matéria seca, e produção do gás metano. Para tanto, feno de Brachiaria decumbens e suplemento proteico de baixo consumo foram utilizados como substrato. Foi simulado um consumo médio de suplemento de 500g/animal/dia acrescido dos níveis de monensina: 0, 200, 400 e 600 mg. Foram coletados amostras dos gases para mensuração da produção de metano em 24 hrs de incubação. A degradabilidade da matéria seca e a produção de ácidos graxos de cadeia curta foram determinados às 24 e 96 horas de incubação. A produção potencial de gases diminuiu conforme a inclusão monensina. O tempo de colonização das amostras aumentou com a inclusão de monensina. Não foram observadas diferenças na degradabilidade da matéria seca e na produção de metano e AGCC entre os tratamentos. / The aim of this research was to evaluate the inclusion of high monensin levels on protein supplements for cattle on (i) performance, (ii) fermentation and ruminal degradability and (iii) production of methane. To this end, three different experiments were performed. The experiments are presented in the chapters form. The introduction and literature review are presented in Chapter I and II. In Chapter III (performance): 64 cattle were randomly assigned to four treatments: control, containing protein supplement and control plus three monensin levels (400, 800 or 1200 mg/kg of supplement). The animals remained in four paddocks of Brachiaria decumbens where they were undergo weekly casters. Pasture samples were collected to determination of qualitative and quantitative variables. The average supplement intake, individual supplement intake and daily weight gain were estimated. The increased monensin levels decreased the supplement consumption and the animal performance. In Chapter IV (fermentation and ruminal degradability), four male cattle with rumen cannula were utilized in a Latin Square assay design 4x4, during 84 days. The animals were fed with Brachiaria decumbens, protein supplement of low consumption (500g/animal /day) and monensina. The treatments were 0, 200, 400 and 600 mg of monensin/day. The ruminal parameters evaluated were: production of short chain fatty acids (SCFA), ammonia concentration, pH, dry matter intake, in situ degradability of DM and ADF. The values found for ruminal fermentation and degradability of DM were not affected by treatments. Increased levels of monensin decreases linearly dry matter intake and the ADF degradability. In Chapter V (production of methane), four monensin levels were tested in vitro gas production. The objectives were to evaluate the fermentation kinetics, short chain fatty acids production, dry matter degradation and methane production. For this purpose, B. decumbens hay and protein supplement low consumption were used as substrate. An average supplement intake 500g/animal/day with levels of monensin (0, 200, 400 and 600 mg) was simulated. Gas samples were collected for measurement of methane production in 24hrs of incubation. The dry matter degradability and production of short chain fatty acids were determined after 24 and 96 hours of incubation. The potential gas production decreased as increased of monensin. The colonization time of the samples increased with monensin. No differences were observed in the degradability of dry matter and in the methane and SCFA production between treatments.

Small proline-rich protein-2 (SPRR2) functions as a determinant of flexibility and permeability in the mature cornified envelope of the skin. SPRR2 is strongly upregulated by the commensal flora and may mediate signaling to differentiated epithelia of the small intestine and colon. Yet, SPRR2 function in the GI tract is largely unexplored. Using the Caco-2 model of intestinal epithelial differentiation along the crypt-villus axis, we hypothesized that SPRR2 would be preferentially expressed in post-confluent differentiated Caco-2 cells and examined SPRR2 regulation by the protein kinase A pathway (PKA) and short chain fatty acids (SCFAs). Differentiation-dependent SPRR2 expression was examined in cytoskeletal-, membrane-, and nuclear-enriched fractions by immunoblotting and confocal immunofluorescence. We studied the effect of SCFAs, known inducers of differentiation, on SPRR2 expression in pre-confluent undifferentiated Caco-2 cells and explored potential mechanisms involved in this induction using MAP kinase inhibitors. SPRR2 expression was also compared between HIEC crypt cells and 16 to 20 week primary fetal villus cells as well as in different segments in mouse small intestine and colon. We determined if SPRR2 is increased by gram negative bacteria such as S. typhimurium. SPRR2 expression increased in a differentiation-dependent manner in Caco-2 cells and was present in human fetal epithelial villus cells but absent in HIEC crypt cells. Differentiation-induced SPRR2 was down-regulated by 8-Br-cAMP as well as by forskolin/IBMX co-treatment. SPRR2 was predominantly cytoplasmic and did not accumulate in Triton X-100-insoluble cytoskeletal fractions. SPRR2 was present in the membrane- and nuclear-enriched fractions and demonstrated co-localization with F-actin at the apical actin ring. No induction was seen with the specific HDAC inhibitor trichostatin A, while SCFAs and the HDAC inhibitor SBHA all induced SPRR2. SCFA responses were inhibited by MAP kinase inhibitors SB203580 and U0126, thus suggesting that the SCFA effect may be mediated by orphan G-protein receptors GPR41 and GPR43. S. typhimurium induced SPRR2 in undifferentiated cells. We conclude that SPRR2 protein expression is associated with differentiated epithelia and is regulated by PKA signaling and by by-products of the bowel flora. This is the first report to establish an in vitro model to study the physiology and regulation of SPRR2. / Thesis (Master, Anatomy & Cell Biology) -- Queen's University, 2008-04-25 12:39:06.427 / This work was funded by the CIHR GIDRU Training Grant and Aid in Research from Crohn's and Colitis Foundation of Canada

Os objetivos destes trabalhos foram avaliar a inclusão de altos níveis de monensina em suplementos proteico de baixo consumo para bovinos sobre (i) desempenho; (ii) fermentação e degradabilidade ruminal e (iii) produção de metano. Para tanto, foram realizados três experimentos distintos. Os experimentos realizados são apresentados na forma de capítulos. Nos Capítulos I e II são apresentados a introdução e a revisão de literatura. No Capítulo III (desempenho), foram utilizados 64 novilhos, distribuídos aleatoriamente em quatro tratamentos: 0, 400, 800 e 1200 mg de monensina sódica/ kg de suplemento proteico. Os animais permaneceram em quatro piquetes de Brachiaria decumbens onde eram submetidos a rodízios semanais. Amostras de pasto foram colhidas para determinação das variáveis qualitativas e quantitativas da forragem. Foram estimados o consumo médio de suplemento, consumo individual de suplemento e ganho de peso médio diário. A inclusão de monensina nos níveis estudados diminuiu o consumo de suplemento e piorou o desempenho animal. No Capítulo IV (fermentação e degradabilidade ruminal), foram utilizados quatro bovinos da raça nelore, canulados no rúmen, em experimento com delineamento quadrado latino 4x4 por 84 dias. Os animais foram arraçoados com dieta composta de feno de Brachiaria decumbens (à vontade), 500 g/cabeça/dia de suplemento proteinado de baixo consumo e monensina sódica conforme os tratamentos: 0, 200, 400 e 600 mg/dia. Os parâmetros ruminais avaliados foram: produções de ácidos graxos de cadeia curta (AGCC); concentração de nitrogênio amoniacal; pH; consumo de matéria seca; degradabilidade in situ da MS e FDA do feno de B. decumbens. Os valores encontrados para a fermentação ruminal (pH, N-NH3 e AGCC) e degradabilidade in situ da MS não foram afetados pelos tratamentos. O aumento dos níveis de monensina diminuíram linearmente o consumo de matéria seca e a degradabilidade potencial da fibra. No Capítulo V (produção de metano), quatro níveis de monensina foram testados em ensaio in vitro de produção de gases com o objetivo de avaliar a cinética fermentativa, produção de AGCC, degradabilidade da matéria seca, e produção do gás metano. Para tanto, feno de Brachiaria decumbens e suplemento proteico de baixo consumo foram utilizados como substrato. Foi simulado um consumo médio de suplemento de 500g/animal/dia acrescido dos níveis de monensina: 0, 200, 400 e 600 mg. Foram coletados amostras dos gases para mensuração da produção de metano em 24 hrs de incubação. A degradabilidade da matéria seca e a produção de ácidos graxos de cadeia curta foram determinados às 24 e 96 horas de incubação. A produção potencial de gases diminuiu conforme a inclusão monensina. O tempo de colonização das amostras aumentou com a inclusão de monensina. Não foram observadas diferenças na degradabilidade da matéria seca e na produção de metano e AGCC entre os tratamentos. / The aim of this research was to evaluate the inclusion of high monensin levels on protein supplements for cattle on (i) performance, (ii) fermentation and ruminal degradability and (iii) production of methane. To this end, three different experiments were performed. The experiments are presented in the chapters form. The introduction and literature review are presented in Chapter I and II. In Chapter III (performance): 64 cattle were randomly assigned to four treatments: control, containing protein supplement and control plus three monensin levels (400, 800 or 1200 mg/kg of supplement). The animals remained in four paddocks of Brachiaria decumbens where they were undergo weekly casters. Pasture samples were collected to determination of qualitative and quantitative variables. The average supplement intake, individual supplement intake and daily weight gain were estimated. The increased monensin levels decreased the supplement consumption and the animal performance. In Chapter IV (fermentation and ruminal degradability), four male cattle with rumen cannula were utilized in a Latin Square assay design 4x4, during 84 days. The animals were fed with Brachiaria decumbens, protein supplement of low consumption (500g/animal /day) and monensina. The treatments were 0, 200, 400 and 600 mg of monensin/day. The ruminal parameters evaluated were: production of short chain fatty acids (SCFA), ammonia concentration, pH, dry matter intake, in situ degradability of DM and ADF. The values found for ruminal fermentation and degradability of DM were not affected by treatments. Increased levels of monensin decreases linearly dry matter intake and the ADF degradability. In Chapter V (production of methane), four monensin levels were tested in vitro gas production. The objectives were to evaluate the fermentation kinetics, short chain fatty acids production, dry matter degradation and methane production. For this purpose, B. decumbens hay and protein supplement low consumption were used as substrate. An average supplement intake 500g/animal/day with levels of monensin (0, 200, 400 and 600 mg) was simulated. Gas samples were collected for measurement of methane production in 24hrs of incubation. The dry matter degradability and production of short chain fatty acids were determined after 24 and 96 hours of incubation. The potential gas production decreased as increased of monensin. The colonization time of the samples increased with monensin. No differences were observed in the degradability of dry matter and in the methane and SCFA production between treatments.

As leveduras Saccharomyces cerevisiae são consideradas importantes matérias primas na nutrição animal pela sua capacidade prebiótica. Os prebióticos são compostos não digeridos pelo organismo animal, mas que são fermentados pelos microrganismos do trato gastrintestinal, cujos produtos são capazes de prover benefícios ao hospedeiro. A fermentação depende de fatores como: substrato utilizado para o seu crescimento, método de fermentação, modo e condição de secagem e idade das células. Assim, os processos produtivos modernos têm como intuito a produção de leveduras com elevado potencial prebiótico. Este estudo objetivou avaliar os efeitos de teores crescentes de leveduras com metabólitos ativos (LSC; baseada na fermentação de substratos específicos) na digestibilidade aparente dos nutrientes da dieta, microbiota e produtos da fermentação fecal e parâmetros imunológicos de gatos adultos. Foram utilizados 27 gatos adultos, idade média de 9,44±5,35 anos, machos e fêmeas, sem raça definida e saudáveis. Os animais foram distribuídos em delineamento em blocos casualizados (idade), constituído de três tratamentos experimentais, denominados: DC (dieta controle), LSC 0,3 (dieta controle com 0,3% de leveduras com metabólitos ativos) e LSC 0,6 (dieta controle com 0,6% de leveduras com metabólitos ativos). Os resultados obtidos foram analisados através do programa computacional Statistical Analysis System (SAS Institute Inc., 2004), sendo considerados significativos valores de p<0,05 e as médias comparadas pelo teste de Tukey. Verificou-se que a inclusão do aditivo alterou apenas a digestibilidade aparente da fibra bruta, da matéria mineral e energia metabolizável (p<0,05). Já em relação aos produtos da fermentação e microbiota das fezes, observou-se redução em ácido lático (p=0,0040) e Clostridium perfringens (p=0,0226) com a inclusão do prebiótico e diminuição do ácido isovalérico (p=0,0144) no tratamento LSC 0,3. Pode-se concluir que o aditivo, nos teores de inclusão avaliados, parece apresentar potencial prebiótico em relação aos produtos da fermentação e microbiota fecal / Saccharomyces cerevisiae yeast are considered important raw materials in animal nutrition due their prebiotic capacity. Prebiotics are compounds not digested by animal organism, but are fermented by microorganisms in the gastrointestinal tract, which products are capable of providing benefits to the host. The fermentation depends on factors such as: substrate used for growth, fermentation method, way and drying condition and age of the cells. Thus, modern production processes have the objective of producing yeast with high prebiotic potential. This study aimed to evaluate the effects of increasing levels of yeast with active metabolites (LSC) based on the fermentation of specific substrates on apparent digestibility of diet nutrients, microbiota and fecal fermentation products and immunological parameters in adult cats. Twenty seven male or female cats with mean weight of 4.19 ± 0.83kg and mean age of 9.44 ± 5.35 years were used and distributed in an unbalanced randomized block design (age), consisting of three experimental treatments, DC (control diet), LSC 0.3 (control diet with 0.3% yeast with active metabolites) and LSC 0.6 (control diet with 0.6% yeast with active metabolites). The results were analyzed using the computer program Statistical Analysis System (SAS Institute Inc., 2004), with significance level of p<0.05 and the averages compared by Tukey test. The inclusion of the additive only changed the apparent digestibility of crude fiber and mineral content (p<0.05). Regarding the fermentation products and microbiota of feces, there was a reduction in lactic acid (p=0,0040) and Clostridium perfringens (p=0,0226) with inclusion of prebiotic and decreased of isovalerate (p=0,0144) in the LSC 0.3 treatment. It can be concluded that the additive, in the levels of inclusion assessed, seems to have prebiotic potential on fecal fermentation products and microbiota