O principal crescimento na indústria de alimentos funcionais corresponde ao dos produtos probióticos e prebióticos. A literatura mostra efeitos imunomoduladores de certas cepas probióticas, contudo, os resultados são às vezes controversos e os mecanismos implicados ainda são pouco elucidados. Sabe-se, no entanto que algumas cepas de probióticos aumentam significantemente a liberação de IL-10 e γ-INF modulando a resposta imune, além destas respostas serem de forma mais branda relacionada às bactérias Gram-positivas probióticas do que às Gram-positivas patogênicas. O presente trabalho teve como objetivo geral estudar o efeito do leite probiótico fermentado na resposta imune celular em cólon de camundongos BALB/c. Os objetivos específicos foram: (i) determinar o efeito imunomodulador do leite adicionado de probiótico em camundongos normais, (ii) identificar os tipos celulares implicados na resposta imune específica por citometria de fluxo e, (iii) colocalizá-los nos cortes histológicos. Simultaneamente, a análise e a comparação da resistência do probiótico à digestão gastrintestinal in vitro e a produção de metabólitos bioativos de acordo com os deferentes produtos foi realizada. Foram preparados leites nos quais as variáveis estudadas foram a tecnologia empregada para a produção das formulações (a) leite; (b) água, (c) leite não fermentado; (d) leite fermentado; (e) leite fermentado seguido de pasteurização, usando a mesma concentração da cepa comercial Bifidobacterium animalis subsp. lactis HOWARU HN019. O leite desnatado e a água foram usados como controles. / Functional food industry is in expansion mainly due to probiotic and prebiotic products. Studies have shown some probiotic strains develop immune modulation effect, however, these results are controversial and the mechanisms are not been well understood. Although, some probiotic strains increase IL-10 and γ-INF release modulating immune response, this response is weaker in probiotic strains when compared to pathogenic Gram-positive bacteria. The major aim of the present study was to assess the effect of probiotic fermented milk in cellular immune response of Balb/c mice colon. The specific objectives were: (i) to determine the immunomodulation of the milk added of probiotic in normal mice; (ii) to identify the cellular types implied in immune specific response and, (iii) to colocalize them in histological sections. Besides, the analyze and comparation of the probiotic resistance upon in vitro gastrointestinal and bioactive metabolites release in fermented or unfermented bifido milk using the same matrix, probiotic strain and probiotic dose in CFU. mL-1 were conducted. Dairy products were prepared in which variable form of technological appliance were: (i) milk, (ii) water, (iii) unfermented milk, (iv) fermented milk, and (v) fermented and heat treatment milk, all using Bifidobacterium subsp. lactis HOWARU HN019 strain in the same concentration. The skimmed milk and water were used as controls. The immune effects were evaluated by histological sections and the lymphocytic infiltrated was analyzed by flow citometry and histology.

O principal crescimento na indústria de alimentos funcionais corresponde ao dos produtos probióticos e prebióticos. A literatura mostra efeitos imunomoduladores de certas cepas probióticas, contudo, os resultados são às vezes controversos e os mecanismos implicados ainda são pouco elucidados. Sabe-se, no entanto que algumas cepas de probióticos aumentam significantemente a liberação de IL-10 e γ-INF modulando a resposta imune, além destas respostas serem de forma mais branda relacionada às bactérias Gram-positivas probióticas do que às Gram-positivas patogênicas. O presente trabalho teve como objetivo geral estudar o efeito do leite probiótico fermentado na resposta imune celular em cólon de camundongos BALB/c. Os objetivos específicos foram: (i) determinar o efeito imunomodulador do leite adicionado de probiótico em camundongos normais, (ii) identificar os tipos celulares implicados na resposta imune específica por citometria de fluxo e, (iii) colocalizá-los nos cortes histológicos. Simultaneamente, a análise e a comparação da resistência do probiótico à digestão gastrintestinal in vitro e a produção de metabólitos bioativos de acordo com os deferentes produtos foi realizada. Foram preparados leites nos quais as variáveis estudadas foram a tecnologia empregada para a produção das formulações (a) leite; (b) água, (c) leite não fermentado; (d) leite fermentado; (e) leite fermentado seguido de pasteurização, usando a mesma concentração da cepa comercial Bifidobacterium animalis subsp. lactis HOWARU HN019. O leite desnatado e a água foram usados como controles. / Functional food industry is in expansion mainly due to probiotic and prebiotic products. Studies have shown some probiotic strains develop immune modulation effect, however, these results are controversial and the mechanisms are not been well understood. Although, some probiotic strains increase IL-10 and γ-INF release modulating immune response, this response is weaker in probiotic strains when compared to pathogenic Gram-positive bacteria. The major aim of the present study was to assess the effect of probiotic fermented milk in cellular immune response of Balb/c mice colon. The specific objectives were: (i) to determine the immunomodulation of the milk added of probiotic in normal mice; (ii) to identify the cellular types implied in immune specific response and, (iii) to colocalize them in histological sections. Besides, the analyze and comparation of the probiotic resistance upon in vitro gastrointestinal and bioactive metabolites release in fermented or unfermented bifido milk using the same matrix, probiotic strain and probiotic dose in CFU. mL-1 were conducted. Dairy products were prepared in which variable form of technological appliance were: (i) milk, (ii) water, (iii) unfermented milk, (iv) fermented milk, and (v) fermented and heat treatment milk, all using Bifidobacterium subsp. lactis HOWARU HN019 strain in the same concentration. The skimmed milk and water were used as controls. The immune effects were evaluated by histological sections and the lymphocytic infiltrated was analyzed by flow citometry and histology.

Foram conduzidos três experimentos no confinamento experimental do Departamento de Zootecnia da ESALQ/USP com o objetivo de estudar os efeitos de diferentes aditivos em rações para bovinos terminados em confinamento. No experimento 1 foram utilizados 100 bovinos machos Nelore castrados (392 kg), distribuídos em 20 baias, por 60 dias. As rações continham 41% de sorgo moído, 40% de polpa cítrica peletizada e 15% de silagem de cana-de-açúcar. Os tratamentos foram: (1) controle, (2) monesina sódica Rumensin (MON1), (3) monensina sódica Rumenfort (MON2), levedura Saccharomyces cerevisiae Yea-Sacc 1026 (LEV1) e levedura Saccharomyces cerevisiae Biosaf SC47 (LEV2). A IMS foi reduzida pelo tratamento MON1 (P<0,05), quando comparada ao tratamento controle. O GPD dos animais não foi afetado pelos tratamentos (P>0,05). A EA dos animais não foi afetada pelos tratamentos (P>0,05). O tratamento MON2 apresentou um menor rendimento de carcaça (P<0,05) e o tratamento MON 1 apresentou uma maior AOL (P<0,05). No experimento 2 foram utilizados 96 tourinhos Nelore não castrados (396 kg), distribuídos em 16 baias por 95 dias. Os tratamentos foram: (1) Controle, (2) levedura Saccharomyces cerevisiae Biosaf SC47 (LEV), (3) combinação de levedura Saccharomyces cerevisiae e bactérias probióticas na dose de 1g /bovino/dia (PROB1) e (4) combinação de levedura Saccharomyces cerevisiae e bactérias probióticas na dose de 3g/bovino/dia (PROB2). As rações continham 59% de polpa cítrica peletizada, 35% de farelo de glúten de milho úmido e 5% de feno de tifton 65. A adição dos aditivos levedura (Saccharomyces cerevisae) e a combinação de leveduras e bactérias probióticas não afetou a IMS o GPD e a EA dos animais (P>0,05). A energia líquida de manutenção e de ganho das rações também não foi afetada pelos tratamentos (P>0,05), assim como os dados de carcaça. No experimento 3, que avaliou a digestibilidade das rações, foram utilizados 20 tourinhos Nelore, alocados em 20 baias individuais durante 15 dias, sendo 10 dias de adaptação ao marcador e 5 dias de coleta de fezes. A ração foi a mesma utilizada no experimento 2, e o marcador utilizado foi o óxido de cromo. Os tramentos utilizados foram: (1) Controle, (2) monensina sódica Rumenfort (MON), (3) levedura Saccharomyces cerevisiae Biosaf SC47 (LEV), (4) combinação de levedura Saccharomyces cerevisiae e bactérias probióticas na dose de 1g /bovino/dia (PROB1) e (5) combinação de levedura Saccharomyces cerevisiae e bactérias probióticas na dose de 3g/bovino/dia (PROB2). Os aditivos testados não afetaram as digestibilidades da MS, da MO, da FDN e da PB das rações (P>0,05). Com os resultados obtidos é possível afirmar que bovinos Nelore, castrados ou não, confinados com rações com altos teores de concentrado ricos em co-produtos como polpa cítrica e farelo de glúten de milho úmido não apresentam melhor desempenho nem melhor digestibilidade dos nutrientes quando suplementados com monensina sódica ou com micorganismos probióticos. / Three trials were conducted at the ESALQ/USP Animal Sciences Department experimental feedlot to evaluate the effects of different feed additives in feedlot finished cattle. On trial 1 100 Nellore steers (392kg) were allocated to 20 pens for 60 days. Experimental rations had 41% ground milo, 40% dried citrus pulp and 15% sugarcane silage. Treatments were: (1) control, (2) sodium monensin Rumensin (MON1), (3) sodium monensin Rumenfort (MON2), (4) yeast culture Saccharomyces cerevisiae Yea-Sacc 1026 (LEV1) and (5) yeast culture Saccharomyces cerevisiae Biosaf SC47 (LEV2). DMI was reduced by MON1 trestment (P<0,05) in relation to control. WDG was not affected by treatments (P>0,05). Treatments did not affect FE (P>0,05). Animals on treatment MON2 showed the lowest dressing percentage (DP) and those on MON1 showed the highest rib eye area (REA) (P<0,05). Trial 2 used 96 young Nellore bulls (396kg), allocated to 16 pens for 95 days. Treatments were: (1) control, (2) Saccharomyces cerevisiae Biosaf SC47 (LEV), (3) Saccharomyces cerevisiae and probiotic bacteria mix at 1g /animal/day dose (PROB1) and (4) Saccharomyces cerevisiae probiotic bacteria mix at 3g/animal/day dose (PROB2). Rations contained 59% dried citrus pulp, 35% wet corn gluten feed and 5% Tifton 65 hay. Treatments didnt affect DMI, WDG and FE (P>0,05). Rations net energy for maintenance and gain were also not affect by treatments (P>0,05), well as carcass data. Trial 3 utilized 20 young Nellore bulls allocated to individual pens for 15 days (10 days for adaptation to marker and 5 days for data collection) to evaluate rations digestibility. Experimental ration was the same utilized on trial 2, with chromium oxide as an external marker. Treatments were (1) control, (2) sodium monensin Rumenfort (MON1), (3) yeast culture Saccharomyces cerevisiae Biosaf SC47 (LEV), (4) Saccharomyces cerevisiae and probiotic bacteria mix at 1g /animal/day dose (PROB1) and (5) Saccharomyces cerevisiae probiotic bacteria mix at 3g/animal/day dose (PROB2). Treatments did not affect rations DM, OM, NDF and CP digestibilities (P>0,05). Results show that Nellore cattle, castrated or not, feedlot finished receiving rations with high levels of byproducts such as dried citrus pulp and wet corn gluten feed dont have higher performance nor better nutrients digestibility when supplemented with sodium monensin or probiotic microorganisms.

Atopic allergy is the most common chronic disease among children in the developed world. This high prevalence could be associated with low microbial exposure. The early gut microbiota appears to be important for immune maturation. Immunomodulatory components in human milk might differ between mothers and could therefore explain the contradictory results seen regarding breastfeeding and allergy development. The aim of this thesis was to investigate whether early colonization with certain gut microbiota species influences childhood immune responses and allergy development up to age five. Also, as human milk oligosaccharides (HMOs) might stimulate the growth of certain gut microbiota species, the consumption of neutral colostrum HMOs was investigated for their role in allergy development up to 18 months. The concentrations of neutral colostrum HMOs varied considerably between women; however this variation could not be explained by their allergic status. Neither was the consumption of neutral colostrum HMOs related to allergy development in their children up to 18 months. Infants who harboured lactobacilli group I and Bifidobacterium adolescentis one week after birth developed allergic disease less frequently during their first five years than infants who did not harbour these bacteria at the same time. Also, colonization with several Bifidobacterium species was associated with higher levels of house dust endotoxin and larger family size. The early Bifidobacterium flora influenced levels of salivary secretory IgA at six and 12 months but not during later childhood. Moreover, the intensity of early Bacteroides fragilis colonization was inversely associated with spontaneous Toll-like receptor 4 mRNA expression in peripheral blood cells collected 12 months after birth. In conclusion, these results indicate that the early infant gut microbiota influences systemic and mucosal immune maturation during infancy, and that it might be altered in infants developing allergic disease.

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No. of bitstreams: 1 zordao_op_me_arafc.pdf: 1351578 bytes, checksum: bc36b449347fea51a7c4bc4e2a6d703b (MD5) Previous issue date: 2016-07-08 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Objetivos: A utilização de microrganismos probióticos na redução do risco de doenças inflamatórias intestinais (DIIs), incluindo a colite ulcerativa (CU), tem se mostrado promissora, sendo que o efeito benéfico depende da cepa utilizada e da fase da patologia. O objetivo do presente estudo foi averiguar o efeito de um produto probiótico à base de soja, fermentado com Enterococcus faecium CRL 183 e Lactobacillus helveticus 416 e com adição de Bifidobacterium longum ATCC 15707, na fase aguda da colite quimicamente induzida em camundongos. Métodos: O protocolo experimental teve duração total de 14 dias e os animais foram distribuídos aleatoriamente em quatro grupos (n=10): Grupo C - Animais sadios que não receberam os produtos em estudo; Grupo CL - Animais com colite quimicamente induzida e que não receberam os produtos em estudo; Grupo CLF - Animais com colite quimicamente induzida e que receberam o produto fermentado probiótico; Grupo CLP - Animais com colite quimicamente induzida e que receberam o produto não fermentado (placebo). A indução da colite foi feita pela administração de dextran sulfato de sódio (DSS) a 3% dissolvido na água fornecida diariamente aos animais, por um período de sete dias, e os produtos foram administrados ao longo dos 14 dias de estudo. Durante o período experimental foram monitorados os seguintes parâmetros: peso corpóreo dos animais, ingestão hídrica e de ração, consistência e presença de sangue oculto ou aparente nas fezes e composição da microbiota fecal. Ao final do protocolo, os animais foram eutanasiados e o intestino grosso foi removido para realização das análises macroscópica e histológica. Resultados: De acordo com os resultados do índice de atividade da doença (IAD), os animais que consumiram o produto probiótico (CLF) apresentaram redução dos sintomas associados à colite durante o período de indução em comparação aos animais do grupo CL. As análises histológicas evidenciaram que o cólon dos camundongos dos grupos CL e CLP apresentou áreas com infiltrados de células, além de alterações nas criptas, representadas por degeneração ou desaparecimento de tais estruturas. O colón dos animais pertencentes ao grupo CLF exibiu infiltrado de células inflamatórias, sem alteração nas criptas. Os resultados da análise da composição da microbiota fecal mostraram que a ingestão do produto fermentado probiótico resultou no maior aumento na população de Lactobacillus spp. (1,5 log10 UFC/g) e menor aumento de enterobactérias (1,34 log10 UFC/g) ao longo do protocolo. Conclusão: Os resultados obtidos no presente estudo indicam que a ingestão regular do produto fermentado probiótico avaliado pode auxiliar na redução dos sintomas e na preservação da integridade do cólon durante a fase aguda da colite induzida em camundongos / Objectives: The use of probiotic microorganisms in the reduction of inflammatory bowel diseases (IBD) risks, including ulcerative colitis (UC), has been shown promising, and the beneficial effect depends on the strain used and the stage of disease. The objective of this study was to investigate the effect of a probiotic soy product fermented with Enterococcus faecium CRL 183 and Lactobacillus helveticus 416 with addition of Bifidobacterium longum ATCC 15707 in the acute phase of colitis chemically induced in mice. Methods: The experimental protocol had total duration of 14 days and the animals were randomized into four groups (n = 10): Group C - healthy animals that have not received the products under study; CL Group - Animals with chemically induced colitis and who have not received the products under study; CLF Group - Animals with chemically induced colitis and receiving the probiotic fermented product; Group CLP - animals with chemically induced colitis and receiving the unfermented product (placebo). The induction of colitis was made by the administration of dextran sodium sulfate (DSS) 3% dissolved in the drinking water for a period of seven days, and the products were administered over the 14 days of the study. During the trial period the following parameters were monitored: body weight, water and food intake, consistency and presence of hidden or apparent blood in stool and composition of the fecal microbiota. At the end of the protocol, the animals were euthanized and the large intestine was removed to perform the macroscopic and histological analysis. Results: According to the results of disease activity index (DAI), the animals that consumed the probiotic product (CLF) showed a reduction of symptoms associated with colitis during the induction period, compared to the CL group. Histological analysis showed that the colon of mice of CL and CLP groups showed areas with cell infiltrates, as well as changes in the crypts, represented by degeneration or disappearance of such structures. The colon of animals belonging to the CLF group exhibited infiltration of inflammatory cells, without changing in the crypts. The results of the analysis of fecal microbiota composition revealed that the ingestion of the probiotic fermented product resulted in the greatest increase in the population of Lactobacillus spp. (1.5 log10 CFU / g) and less increase of enterobacteria (1.34 log10 CFU / g) over the protocol. Conclusion: The results obtained in this study indicate that regular intake of probiotic fermented product evaluated can help in the reduction of the symptoms and preservation of colon health during the acute colitis induced in mice.

Foram conduzidos três experimentos no confinamento experimental do Departamento de Zootecnia da ESALQ/USP com o objetivo de estudar os efeitos de diferentes aditivos em rações para bovinos terminados em confinamento. No experimento 1 foram utilizados 100 bovinos machos Nelore castrados (392 kg), distribuídos em 20 baias, por 60 dias. As rações continham 41% de sorgo moído, 40% de polpa cítrica peletizada e 15% de silagem de cana-de-açúcar. Os tratamentos foram: (1) controle, (2) monesina sódica Rumensin (MON1), (3) monensina sódica Rumenfort (MON2), levedura Saccharomyces cerevisiae Yea-Sacc 1026 (LEV1) e levedura Saccharomyces cerevisiae Biosaf SC47 (LEV2). A IMS foi reduzida pelo tratamento MON1 (P<0,05), quando comparada ao tratamento controle. O GPD dos animais não foi afetado pelos tratamentos (P>0,05). A EA dos animais não foi afetada pelos tratamentos (P>0,05). O tratamento MON2 apresentou um menor rendimento de carcaça (P<0,05) e o tratamento MON 1 apresentou uma maior AOL (P<0,05). No experimento 2 foram utilizados 96 tourinhos Nelore não castrados (396 kg), distribuídos em 16 baias por 95 dias. Os tratamentos foram: (1) Controle, (2) levedura Saccharomyces cerevisiae Biosaf SC47 (LEV), (3) combinação de levedura Saccharomyces cerevisiae e bactérias probióticas na dose de 1g /bovino/dia (PROB1) e (4) combinação de levedura Saccharomyces cerevisiae e bactérias probióticas na dose de 3g/bovino/dia (PROB2). As rações continham 59% de polpa cítrica peletizada, 35% de farelo de glúten de milho úmido e 5% de feno de tifton 65. A adição dos aditivos levedura (Saccharomyces cerevisae) e a combinação de leveduras e bactérias probióticas não afetou a IMS o GPD e a EA dos animais (P>0,05). A energia líquida de manutenção e de ganho das rações também não foi afetada pelos tratamentos (P>0,05), assim como os dados de carcaça. No experimento 3, que avaliou a digestibilidade das rações, foram utilizados 20 tourinhos Nelore, alocados em 20 baias individuais durante 15 dias, sendo 10 dias de adaptação ao marcador e 5 dias de coleta de fezes. A ração foi a mesma utilizada no experimento 2, e o marcador utilizado foi o óxido de cromo. Os tramentos utilizados foram: (1) Controle, (2) monensina sódica Rumenfort (MON), (3) levedura Saccharomyces cerevisiae Biosaf SC47 (LEV), (4) combinação de levedura Saccharomyces cerevisiae e bactérias probióticas na dose de 1g /bovino/dia (PROB1) e (5) combinação de levedura Saccharomyces cerevisiae e bactérias probióticas na dose de 3g/bovino/dia (PROB2). Os aditivos testados não afetaram as digestibilidades da MS, da MO, da FDN e da PB das rações (P>0,05). Com os resultados obtidos é possível afirmar que bovinos Nelore, castrados ou não, confinados com rações com altos teores de concentrado ricos em co-produtos como polpa cítrica e farelo de glúten de milho úmido não apresentam melhor desempenho nem melhor digestibilidade dos nutrientes quando suplementados com monensina sódica ou com micorganismos probióticos. / Three trials were conducted at the ESALQ/USP Animal Sciences Department experimental feedlot to evaluate the effects of different feed additives in feedlot finished cattle. On trial 1 100 Nellore steers (392kg) were allocated to 20 pens for 60 days. Experimental rations had 41% ground milo, 40% dried citrus pulp and 15% sugarcane silage. Treatments were: (1) control, (2) sodium monensin Rumensin (MON1), (3) sodium monensin Rumenfort (MON2), (4) yeast culture Saccharomyces cerevisiae Yea-Sacc 1026 (LEV1) and (5) yeast culture Saccharomyces cerevisiae Biosaf SC47 (LEV2). DMI was reduced by MON1 trestment (P<0,05) in relation to control. WDG was not affected by treatments (P>0,05). Treatments did not affect FE (P>0,05). Animals on treatment MON2 showed the lowest dressing percentage (DP) and those on MON1 showed the highest rib eye area (REA) (P<0,05). Trial 2 used 96 young Nellore bulls (396kg), allocated to 16 pens for 95 days. Treatments were: (1) control, (2) Saccharomyces cerevisiae Biosaf SC47 (LEV), (3) Saccharomyces cerevisiae and probiotic bacteria mix at 1g /animal/day dose (PROB1) and (4) Saccharomyces cerevisiae probiotic bacteria mix at 3g/animal/day dose (PROB2). Rations contained 59% dried citrus pulp, 35% wet corn gluten feed and 5% Tifton 65 hay. Treatments didnt affect DMI, WDG and FE (P>0,05). Rations net energy for maintenance and gain were also not affect by treatments (P>0,05), well as carcass data. Trial 3 utilized 20 young Nellore bulls allocated to individual pens for 15 days (10 days for adaptation to marker and 5 days for data collection) to evaluate rations digestibility. Experimental ration was the same utilized on trial 2, with chromium oxide as an external marker. Treatments were (1) control, (2) sodium monensin Rumenfort (MON1), (3) yeast culture Saccharomyces cerevisiae Biosaf SC47 (LEV), (4) Saccharomyces cerevisiae and probiotic bacteria mix at 1g /animal/day dose (PROB1) and (5) Saccharomyces cerevisiae probiotic bacteria mix at 3g/animal/day dose (PROB2). Treatments did not affect rations DM, OM, NDF and CP digestibilities (P>0,05). Results show that Nellore cattle, castrated or not, feedlot finished receiving rations with high levels of byproducts such as dried citrus pulp and wet corn gluten feed dont have higher performance nor better nutrients digestibility when supplemented with sodium monensin or probiotic microorganisms.

Probiotic lactic acid bacteria (LAB) are very often used in food procesing industry, such as milk products, cheese and fermentsd salami production in nova days. In diploma thesis were tested symbiotic food supplements from different producers. Bacterial DNA was isolated from crude cell lysates of six food suplements by magnetic particles P(HEMA-co-GMA). PCR-ready DNAs were isolated. from all products The detection of Bifidobacterium bacteria identified by PCR was in agreement with those declared by the manufacturers. Magnetic particles with immobilized antibodies against Bifidobacterium were used in the next part of thesis. These particles were used for the isolation of target cells from two products with cell identification by genus specific PCR.

The aim of this diploma thesis was to introduce real-time PCR with high-resolution melting analysis for Bifidobacterium species. Currently a small number of publication, dealing with identification of Bifidobacterium species using high-resolution melting analysis, is available. According to publications dealing with identification of lactic acid bacteria were selected primers P1V1 and P2V1, LAC1 and LAC2, LsppUPF and LsppUPR, V3F and V3R, V6F and V6R. Using this primers bacterial DNA was amplified by real-time PCR with high-resolution melting analysis. After evaluation of the measured results efficiency of selected primers was verified on DNA izolated from complex sample of probiotic product. After further optimisation real-time PCR with high-resolution melting analysis could be suitable using selected primers for Bifidobacterium species.

In the food industry, it is important to correctly identify the species of bacteria and thier properties so that they can be used as a probiotic in dietary supplements. This is performed using DNA diagnostics. In the experimental part, the DNA from four probiotic dietary supplements for children was isolated. Magnetic particles P(HEMA-co-GMA) were tested for isolation. Isolated DNA was amplified by PCR and the presence of DNA of genus Lactobacillus, Bifidobacterium and Bacillus was demonstrated in the products according to the data declared by the manufacturer. The presence of species L.acidophilus, B.animalis in accordance with the data on the product has been demonstrated by PCR with species specific primers. Using PCR, the presence of L.casei, which was declared by the manufacturer, has not been proven in one product at given experimental conditions.

Presented diploma thesis is focused on testing encapsulation methods of enzymes and probiotic bacteria. In the theoretical part a summary of different encapsulation techniques used in food industry is given. Further, materials for encapsulation, above all polysaccharides are presented. Next, some procedures of encapsulation of biopolymers and microorganisms – mainly enzymes and probiotic cultures are discussed. In the experimental part methods for preparation of several types of particles based on polysaccharides and liposomes are introduced. Particles were used for encapsulation of selected hydrolytic enzymes and probiotic strains Bifidobacterium breve a Lactobacillus acidophilus. The encapsulation effectiveness was evaluated by analysis of total proteins and enzyme activities. Particles sizes and their stability in water, in selected model foods and model body fluids were observed, too. According to results obtained in this work it was found that encapsulation of enzymes into polysaccharide particles were succesfull in all types of particles (encapsulation effectivness was more than 50 %). Polysaccharide particles showed a very good stability in body fluids as well as in model foods. As the most suitable materials for enzymes encapsulation chitosan and liposomes were found. Polysaccharide particles were used also for the encapsulation of microorganisms. The stability of particles with lactic acid bacteria was similar to particles containig enzymes, very good stability was verified aslo in model foods and model body fluids. Encapsulation enables long-term stabilization of biologically active compounds as well as posibility of their transport and controlled releasing in gastrointestinal tract. Encapsulation of probiotic bacteria could preserve their viability and long-term survival until the product expiration date. Thus, encapsulation is one of the most promissing procedures for production of foods and food suplements of great quality and high additional value.