O objetivo do presente estudo clínico randomizado foi avaliar in vivo, por meio da técnica de biologia molecular Checkerboard DNA-DNA Hybridization: 1) A contaminação de bráquetes metálicos por 4 espécies bacterianas de microrganismos cariogênicos (Streptococcus mutans, Streptococcus sobrinus, Lactobacillus casei e Lactobacillus acidophilus); e 2) A eficácia da utilização do gluconato de clorexidina a 0,12% (Periogard&reg;) sob a forma de bochechos, sobre esses microrganismos. Participaram do estudo 39 pacientes de 11 a 33 anos de idade, em tratamento com aparelho ortodôntico fixo, nos quais foram colados 2 bráquetes metálicos novos, nos pré-molares. Os pacientes do Grupo Controle (n=20) foram orientados a fazer 2 bochechos semanais com solução placebo, durante 30 dias. Os pacientes do Grupo Experimental (n=19) foram orientados a fazer bochechos com solução à base de gluconato de clorexidina a 0,12% (Periogard&reg;), da mesma forma que o grupo Controle. Decorridos 30 dias, os 2 bráquetes foram removidos de cada paciente e processados para detecção das 4 cepas bacterianas, pela técnica Checkerboard DNADNA Hybridization. Os resultados obtidos foram analisados por meio do teste nãoparamétrico de Kruskal-Wallis, utilizando o software SAS (Statistical Analysis System). O nível de significância adotado foi de 5%. De acordo com os resultados obtidos, observou-se que S. mutans, S. sobrinus, L. casei e L. acidophilus foram detectados em 100% das amostras dos bráquetes de ambos os grupos. No entanto, os bráquetes do grupo Controle encontravam-se mais densamente contaminados por S. mutans e S. sobrinus (p<0,01). No grupo Experimental, embora as quantidades de S. mutans, S. sobrinus, L. casei e L. acidophilus tenham sofrido redução numérica após o uso dos bochechos com solução de gluconato de clorexidina a 0,12%, a comparação com os valores observados no grupo Controle evidenciou diferença estatisticamente significante apenas para S. mutans (p=0,03). Concluiu-se que os bochechos com solução de gluconato de clorexidina a 0,12% podem ser úteis, na prática clínica, com a finalidade de reduzir os níveis de microrganismos cariogênicos, em pacientes portadores de aparelhos ortodônticos fixos. / The purpose of this randomized clinical study was to evaluate in vivo, by the checkerboard DNA-DNA hybridization biomolecular technique: 1) The contamination of metallic brackets by four cariogenic bacterial strains (Streptococcus mutans, Streptococcus sobrinus, Lactobacillus casei and Lactobacillus acidophilus); and 2) The efficacy of 0.12% chlorhexidine gluconate mouthrinses (Periogard&reg;) against these microorganisms. Thirty-nine 11-33-year-old patients under treatment with fixed orthodontic appliances were enrolled in the study and had 2 new metallic brackets bonded to the premolars. For the patients of the control group (n=20), mouthrinses with a placebo solution were prescribed twice a week during 30 days. The patients randomized to the experimental group (n=19) were instructed to use a 0.12% chlorhexidine gluconate solution (Periogard&reg;) as an oral rinse twice a week during 30 days, in the same way as in control group. After this period, the 2 brackets were removed from each patient and processed for detection of the 4 bacterial strains by checkerboard DNA-DNA hybridization. The obtained data were analyzed statistically by the non-parametric Kruskal-Wallis test using the SAS (Statistical Analysis System) software. A level of significance of 5% was set for all analysis. S. mutans, S. sobrinus, L. casei and L. acidophilus were detected in 100% of the samples from brackets of both groups. However, the brackets of the control group were more heavily contaminated by S. mutans and S. sobrinus (p<0.01). In the experimental group, although S. mutans, S. sobrinus, L. casei and L. acidophilus counts decreased after rinsing with the 0.12% chlorhexidine gluconate solution, the comparison with the values obtained in the control group showed statistically significant difference only for S. mutans (p=0.03). In conclusion, the use of 0.12% chlorhexidine gluconate mouthrinses can be useful in clinical practice to reduce the levels of cariogenic microorganisms in patients under treatment with fixed orthodontic appliances.

A possibilidade de se obter um produto com grande aceitabilidade nacional, principalmente voltado ao público infantil, com propriedades microbiológicas, nutricionais, físico-químicas e organolépticas ótimas associadas aos benefícios creditados aos probióticos é bastante promissora. Assim, o presente trabalho teve como objetivo verificar a influência do emprego de culturas probióticas compostas pelos microrganismos Lactobacillus acidophilus La-5 e Bifidobacterium animalis subsp. lactis BL04 isolados e em co-cultura, em queijo tipo petit-suisse processado com a adição de Streptococcus thermophilus como cultura starter, sobre as características do produto, ao longo do seu armazenamento refrigerado. Quatro tratamentos de queijo petit-suisse foram estudados (em triplicata): T1 (controle - Streptococcus thermophilus ST), T2 (ST + Lactobacillus acidophilus La-5), T3 (ST + B. animalis subsp. lactis BL04) e T4 (ST + La-5 + BL04) e armazenados a 4±1ºC. Foram avaliados parâmetros microbiológicos (população de probióticos, starter e contaminantes), de textura, físico-químicos e sensoriais (aceitabilidade). As análises sensoriais foram realizadas após 7 e 14 dias e as demais análises, após 1, 7, 14, 21 e 28 dias de armazenamento dos produtos a 4±1oC. As populações dos probióticos estiveram sempre superiores ao mínimo recomendado para um alimento probiótico, tendo variado de 7,22 a 7,60 log UFC/g e de 8,56 a 8,72 log UFC/g durante o armazenamento, para L. acidophilus e B. animalis subsp. lactis, respectivamente. A cultura starter apresentou populações entre 9,20 e 9,61 log UFC/g no mesmo período. As populações máximas para coliformes totais, bolores e leveduras foram de 2,79 e 3,60 log UFC/g, respectivamente. Os valores de pH diminuíram ao longo do armazenamento, devido à atividade dos microrganismos acidificantes presentes. A umidade variou entre 67,16% e 70,26% no mesmo período. Todos os queijos apresentaram queda na dureza e adesividade ao longo do armazenamento. Na análise sensorial, o queijo T4, com a co-cultura, revelou uma aceitabilidade significativamente maior em todos os atributos avaliados (p<0,05). Após 14 dias de armazenamento, o queijo T3 apresentou uma aceitabilidade significativamente inferior aos demais (p<0,05). A suplementação de queijo petit-suisse T4 com Lactobacillus acidophilus La-5 e B. animalis subsp. lactis BL04 em co-cultura com a cultura starter resultou em um produto com excelente potencial comercial, tendo apresentado características sensoriais, microbiológicas e de textura bastante vantajosas. / The possibility of obtaining a product with wide acceptability in Brazil, and which is mainly targeted at children and has excellent microbiological, nutritional, physical-chemical and organoleptic properties associated with the benefits ascribed to probiotic microorganisms is rather promising. Thus, the aim of the present work was to examine the influence of the probiotic cultures of Lactobacillus acidophilus La-5 and Bifidobacterium animalis subsp. lactis BL04, isolated and in co-culture, on the characteristics of petit-suisse cheese produced with the addition of Streptococcus thermophilus as a starter culture, throughout the period of refrigerated storage. Four petit-suisse cheese trials were studied (in triplicates): T1 (control - Streptococcus thermophilus ST), T2 (ST + Lactobacillus acidophilus La-5), T3 (ST + Bifidobacterium animalis subsp. lactis BL04) and T4 (ST + La-5 + BL04) and stored at 4±1ºC. Microbiological (population of probiotic bacteria, of the starter and of contaminants), instrumental texture, physical-chemical and sensory (acceptability test) parameters were evaluated. The sensory evaluation was carried out after 7 and 14 days of storage of the products at 4±1ºC, and further analyses were carried out after 1, 7, 14, 21 and 28 days of storage. The populations of the probiotics always remained above the minimum level recommended for a probiotic food, having varied between 7.22 and 7.60 log CFU/g, and between 8.72 and 8.56 log CFU/g during storage, respectively, for L. acidophilus and B. animalis subsp. lactis. The starter culture presented populations between 9.20 and 9.61 log CFU/g in the same period. Coliforms, yeasts and molds presented maximum populations of, respectively, 2.79 and 3.60 log CFU/g. The pH values decreased throughout storage, due to the activity of the acidifying microorganisms present. The moisture ranged from 67.16% up to 70.26% in the same period. All the cheeses studied presented a decrease in hardness and adhesiveness throughout storage. Both after 7 and 14 days, cheese T4 presented the highest sensory acceptance for all attributes evaluated and differed significantly from the other cheeses (p<0.05). After 14 days of storage, cheese T3 presented the lowest acceptance and differed significantly from the other cheeses (p<0.05). The supplementation of petit-suisse cheese T4 with both L. acidophilus and B. animalis subsp. lactis in coculture with a starter culture resulted in a product with excellent market potential, as it presented favorable sensory, microbiological and texture features.

A possibilidade de se obter um produto com grande aceitabilidade nacional, principalmente voltado ao público infantil, com propriedades microbiológicas, nutricionais, físico-químicas e organolépticas ótimas associadas aos benefícios creditados aos probióticos é bastante promissora. Assim, o presente trabalho teve como objetivo verificar a influência do emprego de culturas probióticas compostas pelos microrganismos Lactobacillus acidophilus La-5 e Bifidobacterium animalis subsp. lactis BL04 isolados e em co-cultura, em queijo tipo petit-suisse processado com a adição de Streptococcus thermophilus como cultura starter, sobre as características do produto, ao longo do seu armazenamento refrigerado. Quatro tratamentos de queijo petit-suisse foram estudados (em triplicata): T1 (controle - Streptococcus thermophilus ST), T2 (ST + Lactobacillus acidophilus La-5), T3 (ST + B. animalis subsp. lactis BL04) e T4 (ST + La-5 + BL04) e armazenados a 4±1ºC. Foram avaliados parâmetros microbiológicos (população de probióticos, starter e contaminantes), de textura, físico-químicos e sensoriais (aceitabilidade). As análises sensoriais foram realizadas após 7 e 14 dias e as demais análises, após 1, 7, 14, 21 e 28 dias de armazenamento dos produtos a 4±1oC. As populações dos probióticos estiveram sempre superiores ao mínimo recomendado para um alimento probiótico, tendo variado de 7,22 a 7,60 log UFC/g e de 8,56 a 8,72 log UFC/g durante o armazenamento, para L. acidophilus e B. animalis subsp. lactis, respectivamente. A cultura starter apresentou populações entre 9,20 e 9,61 log UFC/g no mesmo período. As populações máximas para coliformes totais, bolores e leveduras foram de 2,79 e 3,60 log UFC/g, respectivamente. Os valores de pH diminuíram ao longo do armazenamento, devido à atividade dos microrganismos acidificantes presentes. A umidade variou entre 67,16% e 70,26% no mesmo período. Todos os queijos apresentaram queda na dureza e adesividade ao longo do armazenamento. Na análise sensorial, o queijo T4, com a co-cultura, revelou uma aceitabilidade significativamente maior em todos os atributos avaliados (p<0,05). Após 14 dias de armazenamento, o queijo T3 apresentou uma aceitabilidade significativamente inferior aos demais (p<0,05). A suplementação de queijo petit-suisse T4 com Lactobacillus acidophilus La-5 e B. animalis subsp. lactis BL04 em co-cultura com a cultura starter resultou em um produto com excelente potencial comercial, tendo apresentado características sensoriais, microbiológicas e de textura bastante vantajosas. / The possibility of obtaining a product with wide acceptability in Brazil, and which is mainly targeted at children and has excellent microbiological, nutritional, physical-chemical and organoleptic properties associated with the benefits ascribed to probiotic microorganisms is rather promising. Thus, the aim of the present work was to examine the influence of the probiotic cultures of Lactobacillus acidophilus La-5 and Bifidobacterium animalis subsp. lactis BL04, isolated and in co-culture, on the characteristics of petit-suisse cheese produced with the addition of Streptococcus thermophilus as a starter culture, throughout the period of refrigerated storage. Four petit-suisse cheese trials were studied (in triplicates): T1 (control - Streptococcus thermophilus ST), T2 (ST + Lactobacillus acidophilus La-5), T3 (ST + Bifidobacterium animalis subsp. lactis BL04) and T4 (ST + La-5 + BL04) and stored at 4±1ºC. Microbiological (population of probiotic bacteria, of the starter and of contaminants), instrumental texture, physical-chemical and sensory (acceptability test) parameters were evaluated. The sensory evaluation was carried out after 7 and 14 days of storage of the products at 4±1ºC, and further analyses were carried out after 1, 7, 14, 21 and 28 days of storage. The populations of the probiotics always remained above the minimum level recommended for a probiotic food, having varied between 7.22 and 7.60 log CFU/g, and between 8.72 and 8.56 log CFU/g during storage, respectively, for L. acidophilus and B. animalis subsp. lactis. The starter culture presented populations between 9.20 and 9.61 log CFU/g in the same period. Coliforms, yeasts and molds presented maximum populations of, respectively, 2.79 and 3.60 log CFU/g. The pH values decreased throughout storage, due to the activity of the acidifying microorganisms present. The moisture ranged from 67.16% up to 70.26% in the same period. All the cheeses studied presented a decrease in hardness and adhesiveness throughout storage. Both after 7 and 14 days, cheese T4 presented the highest sensory acceptance for all attributes evaluated and differed significantly from the other cheeses (p<0.05). After 14 days of storage, cheese T3 presented the lowest acceptance and differed significantly from the other cheeses (p<0.05). The supplementation of petit-suisse cheese T4 with both L. acidophilus and B. animalis subsp. lactis in coculture with a starter culture resulted in a product with excellent market potential, as it presented favorable sensory, microbiological and texture features.

O objetivo do presente estudo clínico randomizado foi avaliar in vivo, por meio da técnica de biologia molecular Checkerboard DNA-DNA Hybridization: 1) A contaminação de bráquetes metálicos por 4 espécies bacterianas de microrganismos cariogênicos (Streptococcus mutans, Streptococcus sobrinus, Lactobacillus casei e Lactobacillus acidophilus); e 2) A eficácia da utilização do gluconato de clorexidina a 0,12% (Periogard&reg;) sob a forma de bochechos, sobre esses microrganismos. Participaram do estudo 39 pacientes de 11 a 33 anos de idade, em tratamento com aparelho ortodôntico fixo, nos quais foram colados 2 bráquetes metálicos novos, nos pré-molares. Os pacientes do Grupo Controle (n=20) foram orientados a fazer 2 bochechos semanais com solução placebo, durante 30 dias. Os pacientes do Grupo Experimental (n=19) foram orientados a fazer bochechos com solução à base de gluconato de clorexidina a 0,12% (Periogard&reg;), da mesma forma que o grupo Controle. Decorridos 30 dias, os 2 bráquetes foram removidos de cada paciente e processados para detecção das 4 cepas bacterianas, pela técnica Checkerboard DNADNA Hybridization. Os resultados obtidos foram analisados por meio do teste nãoparamétrico de Kruskal-Wallis, utilizando o software SAS (Statistical Analysis System). O nível de significância adotado foi de 5%. De acordo com os resultados obtidos, observou-se que S. mutans, S. sobrinus, L. casei e L. acidophilus foram detectados em 100% das amostras dos bráquetes de ambos os grupos. No entanto, os bráquetes do grupo Controle encontravam-se mais densamente contaminados por S. mutans e S. sobrinus (p<0,01). No grupo Experimental, embora as quantidades de S. mutans, S. sobrinus, L. casei e L. acidophilus tenham sofrido redução numérica após o uso dos bochechos com solução de gluconato de clorexidina a 0,12%, a comparação com os valores observados no grupo Controle evidenciou diferença estatisticamente significante apenas para S. mutans (p=0,03). Concluiu-se que os bochechos com solução de gluconato de clorexidina a 0,12% podem ser úteis, na prática clínica, com a finalidade de reduzir os níveis de microrganismos cariogênicos, em pacientes portadores de aparelhos ortodônticos fixos. / The purpose of this randomized clinical study was to evaluate in vivo, by the checkerboard DNA-DNA hybridization biomolecular technique: 1) The contamination of metallic brackets by four cariogenic bacterial strains (Streptococcus mutans, Streptococcus sobrinus, Lactobacillus casei and Lactobacillus acidophilus); and 2) The efficacy of 0.12% chlorhexidine gluconate mouthrinses (Periogard&reg;) against these microorganisms. Thirty-nine 11-33-year-old patients under treatment with fixed orthodontic appliances were enrolled in the study and had 2 new metallic brackets bonded to the premolars. For the patients of the control group (n=20), mouthrinses with a placebo solution were prescribed twice a week during 30 days. The patients randomized to the experimental group (n=19) were instructed to use a 0.12% chlorhexidine gluconate solution (Periogard&reg;) as an oral rinse twice a week during 30 days, in the same way as in control group. After this period, the 2 brackets were removed from each patient and processed for detection of the 4 bacterial strains by checkerboard DNA-DNA hybridization. The obtained data were analyzed statistically by the non-parametric Kruskal-Wallis test using the SAS (Statistical Analysis System) software. A level of significance of 5% was set for all analysis. S. mutans, S. sobrinus, L. casei and L. acidophilus were detected in 100% of the samples from brackets of both groups. However, the brackets of the control group were more heavily contaminated by S. mutans and S. sobrinus (p<0.01). In the experimental group, although S. mutans, S. sobrinus, L. casei and L. acidophilus counts decreased after rinsing with the 0.12% chlorhexidine gluconate solution, the comparison with the values obtained in the control group showed statistically significant difference only for S. mutans (p=0.03). In conclusion, the use of 0.12% chlorhexidine gluconate mouthrinses can be useful in clinical practice to reduce the levels of cariogenic microorganisms in patients under treatment with fixed orthodontic appliances.

Lactic acid bacteria were isolated from 10 samples of the traditionally fermented foods (5 samples of Vietnamese fermented pork roll and 5 samples of the salted field cabbage) and 5 samples of fresh cow milks collected from households in Vietnam. 22 strains of lactic acid bacteria were isolated for inhibition to Lactobacillus plantarum JCM 1149. Of these, only 2 strains including DC1.8 and NC1.2 have rod shape, the others have coccus shape. 7 strains showing higher antibacterial activity were selected for checking spectrum of antibacteria with indicator bacteria consistting of Bacillus subtilis ATCC 6633, Enterococcus faecium JCM 5804 and Staphylococcus aureus TLU. By which, 3 strains including NC3.5 (from Vietnamese fermented pork roll), DC1.8 (from salted field cabbage) and MC3.19 (from fresh cow milk) were selected because of their higher antibacterial ability. However, the antibacterial activity of the lactic acid bacteria can be based on their disposable compounds and some other antibacterial compounds produced during their growth (such as lactic acid, H2O2, bacteriocins, etc.). For seeking lactic acid bacteria with capability of producing bacteriocins, antibacterial compounds with protein nature, 3 above strains were checked sensitiveness to proteases (including protease K, papain, α – chymotrypsin and trypsin). Because bacteriocins are proteinaceous antibacterial compounds, so their antibacterial activity will be reduced if proteases are added. The result showed DC1.8 and MC3.19 were capable of producing bacteriocin during culture process. They were identified as Lactobacillus acidophilus and Lactococcus lactis and classified, respectively, based on analysis chemical characterisitcs by standard API 50 CHL kit and phylogeny relationship by 16s rRNA sequences. / Các chủng vi khuẩn lactic được phân lập từ 10 mẫu thực phẩm lên men truyền thống (5 mẫu nem chua, 5 mẫu dưa cải bẹ muối) và 5 mẫu sữa bò tươi được thu thập từ các hộ gia đình ở Việt Nam. 22 chủng vi khuẩn lactic đã được phân lập với tiêu chí có khả năng kháng lại vi khuẩn kiểm định Lactobacillus plantarum JCM 1149. Trong số đó, 2 chủng DC1.8 và NC1.2 có tế bào hình que, các chủng còn lại có tế bào hình cầu. 7 chủng thể hiện hoạt tính kháng khuẩn cao được lựa chọn để xác định phổ kháng khuẩn rộng hơn với ba loài vi khuẩn kiểm định Bacillus subtilis ATCC 6633, Enterococcus faecium JCM 5804 và Staphylococcus aureus TLU. Từ đó lựa chọn được 3 chủng có hoạt tính kháng khuẩn cao hơn hẳn. Các chủng này gồm NC3.5 phân lập từ nem chua, DC1.8 phân lập từ dưa cải bẹ muối và MC3.19 phân lập từ sữa bò tươi. Tuy nhiên, hoạt tính kháng khuẩn của vi khuẩn lactic bao gồm những hợp chất nội tại có trong nó và cả những hợp chất được sinh ra trong quá trình phát triển của nó (như axit lactic, H2O2, bacteriocin, …). Với định hướng tìm chủng vi khuẩn lactic có khả năng sinh bacteriocin, chất kháng khuẩn có bản chất protein, 3 chủng trên được kiểm tra độ nhạy cảm với các protease (gồm protease K, papain, α – chymotrypsin và trypsin). Do bacteriocin là chất kháng khuẩn có bản chất protein nên hoạt tính kháng khuẩn của chúng sẽ bị giảm nếu protease được bổ xung vào. Kết quả lựa chọn được chủng DC1.8 và MC3.19 có khả năng sinh bacteriocin. Hai chủng này được phân loại đến loài nhờ vào phân tích đặc điểm sinh hóa bằng kit API 50 CHL và mối quan hệ di truyền thông qua trình tự gen 16s rRNA. Kết quả phân loại đã xác định chủng DC1.8 thuộc loài Lactobacillus acidophilus và chủng MC3.19 thuộc loài Lactococcus lactis.

The Diploma thesis deals with designing of probiotic dietary supplement for children with strains Lactobacillus acidophilus and Bifidobacterium breve and with prebiotics. Used prebiotics were Inulin, Chia fiber, Bamboo fiber, Chlorella + Spirulina and Yakon syrup. The theoretical part is focused on probiotics, prebiotics and their biological influence. In experimental part the possibilities of encapsulation into alginate particle and lyophilisation of probiotic cells were observed to find their good form to final nutritional product for children. Several types of probiotic with addition of prebiotics were tested in model conditions of gastrointestinal tract. It was found that addition of prebiotic highly increases viability of probiotic cells and their resistance to model conditions of gastrointestinal tract. In this case, the best prebiotic was found in Yakon syrup. The prebiotics were also characterised in terms of nutritional composition, amount of total and reducing sugars, oligosaccharides, proteins, lipids, polyphenols and chlorophyll were obtained. Finally, Chia fiber, Chlorella + Spirulina and Yakon syrup were chosen as prebiotics with best characterisation/properties. In conclusion, a dietary supplement with lyophilized alginate particles containing probiotic cells and with the most appropriate prebiotics were designed. Forms of the product were powder and gummy-bear.

The presented diploma thesis is focused on the preparation and the monitoring of the biological stability of beer enriched with probiotics. Probiotic bacterial strains of Lactobacillus acidophilus, Bifidobacterium breve and Bifidobacterium bifidum were used in this study. The theoretical part is divided into two sections. In the first section, probiotics are generally characterized, and their role within a gut microbiota is described. Next, the microbiology of particular probiotic microorganisms including the genera of Lactobacillus and Bifidobacterium is described. Then, some factors influencing the viability and the growth of probiotics are stated. In this section, biological effects of probiotics on the human organism and their potential clinical applications are described. The second section of the theoretical part deals with the technology of brewing, the chemical composition of beer and particular beer styles. In the experimental part, the methods of probiotic bacterial cell concentration and viability determination were optimized. Several techniques for determination of these parameters were selected, particularly the cultivation method, flow cytometry and the spectrophotometric measurement of turbidity. Then, the growth curves of the probiotic strains were measured in MRS medium. Probiotic bacteria were cultivated in model beer samples, i.e. in MRS media with several different concentrations of ethanol. It is possible to say that ethanol did not have significant effect on probiotics growth. Next, experimental cultivations of individual probiotic bacteria and their mixtures in nine real beer samples were conducted. No increase of viable cells concentrations was detected in the samples. On the contrary, a decrease of the concentrations was observed, mainly in the samples with individual bacterial strains. However, certain values of viable cells concentrations were determined at the end of the cultivations in all cases. A pale, top-fermented beer was brewed and supplemented with probiotics, and the concentrations of viable probiotic cells were monitored during 37 days of fermentation. A decrease of concentrations by two orders of magnitude of CFU/ml was observed in almost all samples. Yet, viable cells of probiotic bacteria were detected in all samples of beer at the end of the fermentation. Maximal concentration of viable probiotic cells in the brewed beer was determined with the cultivation method at (3,80 ± 0,14)10^5 CFU/ml. Chosen samples were analyzed with HPLC-RI method that quantified the common beer concentrations of ethanol in all chosen samples, lactic acid was not detected. Sensory analysis was conducted as well. Based on the results of the experimental part and the bibliography, an optimal technology of the preparation of beer enriched with probiotics is discussed in this study.

Probiotic lactic acid bacteria (LAB) are an important part of fermented dairy products, pharmaceuticals and food supplements. At present, rapid and accurate identification of bacteria is carried out using molecular biological methods based on DNA amplification. The aim of the thesis was to identify by non-cultivation bacteria of genus Lactobacillus and bacteria of species Lactobacillus acidophilus in complex matrices at total of seven different food supplements. Total DNA was isolated from crude cell lysates using magnetic carrier P(HEMA-co-GMA). Amplificability of DNA was verified by PCR using primers specific for the domain Bacteria. In next step isolated DNA was amplified using primers specific for the genus Lactobacillus and species Lactobacillus acidophilus to demonstrate the presence of this bacterial genus and species declared by the producers. The results of bacteria identification obtained by PCR were compared with declared specification given by the producers.

This diploma thesis is focused on encapsulation probiotics and co-encapsulation with some types of prebiotics. In theoretical part is aimed to probiotics, their general characteristics and application of probiotics in food industry. There are described prebiotics and their classification, there is described principles of encapsulation and encapsulation techniques. Methods, which are used for analysis of particles and encapsulation components were introduced too. The experimental part describes methods of prebiotics characterization by high performance liquid chromatography, thin layer chromatography and spectrophotometric methods. Cultivation of probiotics with prebiotics - hydrolyzed and non-hydrolyzed wad tested. Using flow cytometry cell viability was measured too. Finally probiotics and prebiotics were encapsulated, mainly by encapsulator machine. Long-term stability of particles during 6 week storage was observed. The particles were exposed to effect of artificial intestinal, gastric and bile juices.

The Diploma thesis deals with use of selected probiotic strains Lactobacillus acidophilus and Bifidobacterium breve in different forms in food for infants. The theoretical part is focused on describing probiotics, encapsulation methods and intestinal gut microbiota of infants. Further, characterization of individual periods of infant feeding and food for infants were introduced. In experimental part the possibilities of encapsulation and lyophilisation of probiotic cells were observed. Probiotic cells were encapsulated into alginate particles. The encapsulator was used for preparation of particles and the most appropriate particles were prepared by encapsulation nozzle with size of 300 µm. Moreover, probiotics viability was monitored by Flow Cytometry, Fluorescence Microscopy and by cultivation (CFU method). Viability of probiotics was monitored during long-term storage in selected food for infants. The appropriate shelf life of non-lyophilized alginate particles in real food have been set at 1 to 2 months. Lyophilized alginate particles could be stored for more than 3 months. Finally, the stability of the particles and viability of encapsulated and non-encapsulated cells in the gastrointestinal tract conditions were also examined. The viabilities of lyophilized cells and cells encapsulated in lyophilized particles were also compared. From the results obtained, non-encapsulated probiotic bacteria cells are more susceptible to negative effects of digestive juices, the percentage of dead probiotic cells after digestion was approximately 80 %. On the other hand, alginate particles showed cell protection from digestive juices, after incomplete cell releasing from particles the percentage of dead probiotic cells did not exceed 20 %. After adequate rehydration, similar results were gained with lyophilized alginate particles. Lyophilized alginate particles have been determined to be the most suitable application form for infants’ food.