Abstract Nutritional status affects every aspect of a child?s health. Thorough nutritional assessment is hampered by the lack of a single comprehensive tool, which can cover all aspects of nutritional assessment. In three distinct studies, this thesis investigated the nutritional status of hospitalised children, children with coeliac disease and children with inflammatory bowel disease. Study 1 The objectives of this study were to assess prevalence of malnutrition and nutritional risk, and define demographic and anthropometric factors associated with nutritional risk among hospitalized children. In this cross sectional study, 157 hospitalised children were assessed for nutritional status using nutritional risk score (NRS) and anthropometric measurements. We found that 4.5%, 8.9%, 15.1% and 10.4% children were wasted, stunted, overweight and obese respectively. However, with the NRS, 47.8% of the children were at high nutritional risk. These children at high risk had lower weight for age (p=0.02), lower BMI percentiles for age (p=0.001) and longer hospitalization (p=0.001) than children at no risk. Study 2 The objectives of this study were to determine nutritional parameters in children with coeliac disease. Twenty-five children with coeliac disease and an equal number of age and gender matched controls were enrolled and anthropometric measurements, BIA and leptin levels were analysed in all. No significant differences were found between the children with coeliac disease and controls in these parameters. BMI percentile correlated with leptin levels in children with coeliac disease. Study 3 The objectives of this study were to determine anthropometric parameters and leptin levels in children with IBD and ascertain if BMI correlates with leptin levels in these children. Thirty children with IBD and 60 age and gender matched controls were enrolled. Anthropometric measurements and leptin levels were analysed and compared with controls. IBD children had significantly low weight for age (p=0.002), BMI percentiles (p=0.001) and leptin levels (p=0.009) compared to controls. There was a correlation between BMI and leptin levels in IBD children. In conclusion, this thesis has shown that one quarter of hospitalized children were overweight or obese, and further, that half of the hospitalised children were at high risk of nutritional deterioration and these children had longer hospital stay than children at no risk. Children with coeliac disease had similar anthropometric measurements, body compartments and leptin levels to controls. However, children with IBD had lower anthropometric measurements and leptin levels, indicating under-nutrition. Nutritional assessment should be a mandatory part of clinical management with nutritional status assessed by various tools including NRS, anthropometry, BIA and leptin levels.

Abstract Nutritional status affects every aspect of a child?s health. Thorough nutritional assessment is hampered by the lack of a single comprehensive tool, which can cover all aspects of nutritional assessment. In three distinct studies, this thesis investigated the nutritional status of hospitalised children, children with coeliac disease and children with inflammatory bowel disease. Study 1 The objectives of this study were to assess prevalence of malnutrition and nutritional risk, and define demographic and anthropometric factors associated with nutritional risk among hospitalized children. In this cross sectional study, 157 hospitalised children were assessed for nutritional status using nutritional risk score (NRS) and anthropometric measurements. We found that 4.5%, 8.9%, 15.1% and 10.4% children were wasted, stunted, overweight and obese respectively. However, with the NRS, 47.8% of the children were at high nutritional risk. These children at high risk had lower weight for age (p=0.02), lower BMI percentiles for age (p=0.001) and longer hospitalization (p=0.001) than children at no risk. Study 2 The objectives of this study were to determine nutritional parameters in children with coeliac disease. Twenty-five children with coeliac disease and an equal number of age and gender matched controls were enrolled and anthropometric measurements, BIA and leptin levels were analysed in all. No significant differences were found between the children with coeliac disease and controls in these parameters. BMI percentile correlated with leptin levels in children with coeliac disease. Study 3 The objectives of this study were to determine anthropometric parameters and leptin levels in children with IBD and ascertain if BMI correlates with leptin levels in these children. Thirty children with IBD and 60 age and gender matched controls were enrolled. Anthropometric measurements and leptin levels were analysed and compared with controls. IBD children had significantly low weight for age (p=0.002), BMI percentiles (p=0.001) and leptin levels (p=0.009) compared to controls. There was a correlation between BMI and leptin levels in IBD children. In conclusion, this thesis has shown that one quarter of hospitalized children were overweight or obese, and further, that half of the hospitalised children were at high risk of nutritional deterioration and these children had longer hospital stay than children at no risk. Children with coeliac disease had similar anthropometric measurements, body compartments and leptin levels to controls. However, children with IBD had lower anthropometric measurements and leptin levels, indicating under-nutrition. Nutritional assessment should be a mandatory part of clinical management with nutritional status assessed by various tools including NRS, anthropometry, BIA and leptin levels.

Abstract Nutritional status affects every aspect of a child?s health. Thorough nutritional assessment is hampered by the lack of a single comprehensive tool, which can cover all aspects of nutritional assessment. In three distinct studies, this thesis investigated the nutritional status of hospitalised children, children with coeliac disease and children with inflammatory bowel disease. Study 1 The objectives of this study were to assess prevalence of malnutrition and nutritional risk, and define demographic and anthropometric factors associated with nutritional risk among hospitalized children. In this cross sectional study, 157 hospitalised children were assessed for nutritional status using nutritional risk score (NRS) and anthropometric measurements. We found that 4.5%, 8.9%, 15.1% and 10.4% children were wasted, stunted, overweight and obese respectively. However, with the NRS, 47.8% of the children were at high nutritional risk. These children at high risk had lower weight for age (p=0.02), lower BMI percentiles for age (p=0.001) and longer hospitalization (p=0.001) than children at no risk. Study 2 The objectives of this study were to determine nutritional parameters in children with coeliac disease. Twenty-five children with coeliac disease and an equal number of age and gender matched controls were enrolled and anthropometric measurements, BIA and leptin levels were analysed in all. No significant differences were found between the children with coeliac disease and controls in these parameters. BMI percentile correlated with leptin levels in children with coeliac disease. Study 3 The objectives of this study were to determine anthropometric parameters and leptin levels in children with IBD and ascertain if BMI correlates with leptin levels in these children. Thirty children with IBD and 60 age and gender matched controls were enrolled. Anthropometric measurements and leptin levels were analysed and compared with controls. IBD children had significantly low weight for age (p=0.002), BMI percentiles (p=0.001) and leptin levels (p=0.009) compared to controls. There was a correlation between BMI and leptin levels in IBD children. In conclusion, this thesis has shown that one quarter of hospitalized children were overweight or obese, and further, that half of the hospitalised children were at high risk of nutritional deterioration and these children had longer hospital stay than children at no risk. Children with coeliac disease had similar anthropometric measurements, body compartments and leptin levels to controls. However, children with IBD had lower anthropometric measurements and leptin levels, indicating under-nutrition. Nutritional assessment should be a mandatory part of clinical management with nutritional status assessed by various tools including NRS, anthropometry, BIA and leptin levels.

SAMMANFATTNING Bakgrund: Celiaki är en autoimmun sjukdom med en prevalens på 2/100 i Sverige. Enda behandlingen är att äta en kost fri från gluten. Gluten förekommer i spannmålen vete, råg, korn och att diagnostiseras med celiaki innebär ofta att en förändring av matvanor blir nödvändig. Att leva tillsammans med någon med celiaki har visat sig kunna vara problematiskt då familjelivet kan påverkas negativt och föräldrar till barn med celiaki upplever ofta oro och ängslan för sitt barn. Syfte: Syftet med studien var att undersöka hur mödrar till barn med celiaki upplever och hanterar familjens vardag i relation till barnets specialkost. Metod: Ett målinriktat urval användes och kvalitativa, semi-strukturerade intervjuer genomfördes med nio mödrar till barn med celiaki. Materialet spelades in och transkriberades ordagrant för att sedan analyseras med kvalitativ innehållsanalys. Resultat: Samtliga mödrar upplevde att vardagen hade anpassats efter barnet med celiaki och det fanns olika sätt att hantera den glutenfria kosten. Vissa valde att ha helt glutenfritt hemma för att undvika stress över rädslan att ge barnet fel mat. Andra valde att ha både gluteninnehållande och glutenfria livsmedel hemma p.g.a. ekonomiska skäl samt preferenser från övriga familjemedlemmar. Kunskapsnivån kring celiaki i samhället ansågs låg, vilket upplevdes försvåra vardagen för mödrar till ett barn med celiaki. Majoriteten av mödrarna uttryckte att bästa stödet fanns att tillgå via internet och sociala medier. Slutsats: Enligt mödrar till barn med celiaki innebar sjukdomen en del dilemman och vardagen upplevdes stundvis som orofylld, vilket kan påverka livskvaliteten. Internet och sociala medier sågs som ett bra stöd i vardagen. Ökad kunskap samt större förståelse från omgivningen skulle kunna förbättra familjernas vardag. / ABSTRACT                          Background: Celiac disease (CD) is an autoimmune disorder with a prevalence of 2/100 in Sweden. The only treatment is a diet free from gluten. Gluten occurs in grains of wheat, rye, barley and being diagnosed with CD often means that a change in eating habits is necessary. Living together with someone with CD has been shown to be problematic. Family life may be negatively affected and parents of children with CD experience worry and anxiety for their child. Objective: The aim of the study was to examine how mothers of children with CD experienced and handled everyday life in relation to their child's gluten-free diet (GFD). Method: A targeted selection was used and qualitative, semi-structured interviews were conducted with nine mothers of children with CD. The interviews were recorded and transcribed verbatim and then analyzed with qualitative content analysis. Results: All participants felt that everyday life had been adapted to the child with CD and there were different ways to deal with the GFD. Some chose to have the home as a gluten-free zone to avoid the stress of risking to give the child wrong food. Others chose to have both gluten-containing and gluten-free foods at home because of economic aspects and preferences from other family members. The level of knowledge about CD in the community was considered low, which was perceived to complicate everyday life for mothers of a child with CD. The majority of participants expressed that best support was found through Internet and social media. Conclusion: According to mothers of children with CD, the disease involved some dilemmas and daily life was at times perceived as worrisome, which can affect quality of life. Internet and social media were considered as good support in everyday life. Increased knowledge in society and greater understanding from the environment could improve families everyday life.

Coeliac disease (CD) is a chronic gastrointestinal condition, the only treatment for which is a gluten-free diet (GFD). Following a GFD is restrictive, burdensome, and can impact health-related quality of life (HRQOL). People with CD can experience long delays to diagnosis and evidence suggests large variations in follow-up care, but the relationship between health care experiences and HRQOL is unknown. The main aim of this research was to develop a patient-reported outcome measure and patient experience questionnaire, and use these to investigate the relationship between adults' experiences of health care services and HRQOL in CD. The questionnaires, named the Coeliac Disease Assessment Questionnaire (CDAQ) and the Coeliac Disease Patient Experience Questionnaire (CD-PEQ), were developed following qualitative interviews with adults with CD, and refined with input from experts, and cognitive interviews. The CDAQ was also subject to a translatability assessment to assess its linguistic and cultural translatability, and a cross-sectional survey to assist with item reduction and scale generation. Members of Coeliac UK (n=267) completed the CDAQ and CD-PEQ, together with the SF-36v2 and demographic questions as part of a postal survey. Psychological health, vitality, general health, and dietary burden were found to have the greatest impact on HRQOL, with physical health and social isolation the least affected. HRQOL was found to have a strong correlation with patients' experiences of health care services. Aspects most strongly related were: the provision of information; communication with HCPs; difficulty obtaining prescriptions; and GPs' knowledge. This research has identified aspects of health care services that are strongly related to HRQOL in CD. Health care providers are recommended to focus service improvement efforts on these areas. A reliable and valid disease-specific patient-reported outcome measure and patient experience questionnaire have been developed as part of this study. The CDAQ is suitable for use in research studies, including clinical trials, to assess HRQOL in CD.

Orientador: Yoon Kil Chang / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos / Made available in DSpace on 2018-08-18T02:02:28Z (GMT). No. of bitstreams: 1 Almeida_OsmarPatriciode_D.pdf: 100022462 bytes, checksum: 269e201898013558151a84656469d230 (MD5) Previous issue date: 2011 / Resumo: Disponibilizar aos celíacos a variedade de alimentos que não lhes é permitida consumir tem sido um desafio aos pesquisadores na área de alimentos. O pão tem sido objeto de estudo profundo pois nesse produto, em especial, a substituição da funcionalidade tecnológica do glúten que é particularmente não trivial. O objetivo do presente trabalho foi o de desenvolver um pão de forma sem glúten a base de farinha de arroz, com características similares a um pão de forma tradicional. O intento tecnológico de pesquisa foi dividido em cinco etapas, a saber: revisão bibliográfica, mapeamento geral e desenvolvimento de formulação-base, estudando-se variáveis significativas via planejamento de Plackett e Burman, a partir da determinação das variáveis significativas o delineamento composto central rotacional 24 incluindo 8 ensaios axiais e 4 repetições no ponto central, seguido de um delineamento composto central rotacional 22 incluindo 4 ensaios axiais e 4 repetições no ponto central. Na etapa inicial estudaram-se 50 formulações, envolvendo 32 ingredientes e 12 variáveis de processo. Ao final desenvolveu-se um pão de forma sem glúten, cuja formulação pode ser obtida mediante a substituição da farinha de trigo pela mistura de farinha de arroz, fécula de mandioca modificada e albumina de ovo acidificada e desidratada. A farinha de arroz mostrou-se uma boa base de trabalho, atuando como agente de corpo. A fécula de mandioca modificada proveu consistência e coesividade à massa. A albumina de ovo acidificada e desidratada substituiu o glúten, sendo a responsável pelo crescimento da massa. Adicionou-se ainda o hidrocoloide carboximetilcelulose que, atuando como espessante, contribuiu para a viscosidade da massa e absorção de água pela mesma. Os demais ingredientes atuaram de forma análoga a uma formulação tradicional de pão de forma produzidos com farinha de trigo. O processo de produção mostrou que o uso de batedeira com agitador tipo raquete, relação volume de forma/peso de massa de 3,71 mL/g em uma forma de 21 x 10 x 10 cm (c x l x a), tempo de fermentação de 60 minutos a 40°C realizada a 85% de umidade relativa, seguida de forneamento a temperatura de 180°C por 23 minutos e resfriamento de 60 minutos promoveu os melhores resultados. Todas as amostras após 24 horas do seu preparo apresentaram-se excessivamente firmes. O delineamento experimental de Plackett e Burman foi empregado em sequência para determinar quais variáveis possuíam efeito significativo (p<0,10) sobre a altura média da fatia central e a firmeza do miolo nos dias 01, 03, 07 e 10 após a produção de pães de forma sem glúten a base de farinha de arroz. A firmeza foi influenciada pela fécula de mandioca modificada, amilase maltogênica e água, enquanto a protease fúngica, albumina de ovo acidificada e desidratada e água apresentaram efeito sobre a altura média da fatia central. A falta de preenchimento da forma pela massa batida afetou a cor da crosta dos pães, devido a sua falta de exposição ao calor. A consistência da massa influenciou a característica da casca. Massas com excesso de água e consequentemente menos estrutura colapsaram comprometendo a simetria. A cor do miolo apresentou-se mais clara à medida que a altura média da fatia central das peças estava acima de 100 mm. A estrutura do miolo seguiu a tendência da cor, com exceção de PB11, onde os níveis de albumina de ovo acidificada e desidratada e água eram máximos e o nível de fécula de mandioca modificada era mínimo, que além do bom crescimento possuía uma estrutura do miolo mais fechada. A formulação PB11 apresentou uma mastigação similar a um pão de forma tradicional. O aroma e o sabor variaram entre o leve de farinha de arroz a neutro, com uma ressalva para as fórmulas PB02 e PB05, onde os níveis de fécula de mandioca modificada e albumina de ovo acidificada e desidratada eram máximos e o nível de água era mínimo, cujos sabores foram desagradáveis. Uma vez definidas as variáveis significativas empregou-se o delineamento composto central rotacional 24 incluindo 8 ensaios axiais e 4 repetições no ponto central para determinar quais variáveis independentes - albumina de ovo desidratada e acidificada, água, enzima amilase maltogênica, fécula de mandioca modificada - possuíam efeito significativo sobre a altura média da fatia central e a firmeza do miolo nos dias 01, 03, 07 e 10 de pães de forma sem glúten a base de farinha de arroz. Como resultado deste delineamento, verificou-se que a altura média da fatia central foi positiva e significativamente impactada (p<0,05) pela água na forma em um contorno quadrático com um máximo no nível 1,1. Com relação à firmeza, esta foi significativa, linear e positivamente afetada (p<0,10 dia 01 e p<0,05 demais dias) pela fécula de mandioca modificada, albumina de ovo acidificada e desidratada. Os termos quadrático e linear da água sobre a firmeza foram significativamente positivo e negativo, respectivamente (p<0,10 dia 01 e p<0,05 demais dias). Ao contrário do que se esperava, a enzima amilase maltogênica não apresentou efeito sobre a firmeza. Todos os pães apresentaram bom crescimento e características visuais, carecendo de sabor que variou entre o leve de farinha de arroz ao amargo. Como a etapa anterior não contemplou a enzima protease fúngica, uma das variáveis significativas determinada pela planejamento de Plackett e Burman, realizou-se o delineamento composto central rotacional 22 incluindo 4 ensaios axiais e 4 repetições no ponto central foi empregado para determinar quais variáveis independentes ¿ carboximetilcelulose (cmc) e enzima protease fúngica (eprt) - possuíam efeito significativo sobre a altura média da fatia central, a firmeza do miolo nos dias 01, 03, 07 e 10, volume específico e tempo de fermentação de pães de forma sem glúten a base de farinha de arroz. Como resultado deste trabalho, verificou-se que a altura média da fatia central foi positiva e significativamente impactada (p<0,05) pela eprt na forma em um contorno quadrático, linearmente pela cmc e pela relação dos termos lineares da eprt e da cmc. Com relação à firmeza esta foi significativa e positivamente impactada pela cmc em termos linear e quadrático para os dias 01, 03 e 07 após a produção, (p<0,05, p<0,05 e p<0,10), enquanto a firmeza do dia 10 foi afetada negativamente apenas pelo termo linear da cmc (p<0,10). O volume específico foi positivamente afetado (p<0,10) pela cmc de forma linear, enquanto o tempo de fermentação o foi positivamente pelo termo quadrático apenas (p<0,10). Todos os pães apresentaram bom crescimento e boa mastigabilidade. O sabor manteve-se neutro, com exceção do experimento que possuía o nível mínimo de cmc que colapsou e apresentou amargor. A partir destes resultados criaram-se modelos matemáticos para a firmeza nos dias 01, 03, 07 e 10 de vida de prateleira, altura média da fatia central, volume específico e tempo de fermentação. Os modelos foram testados e validados no ponto ótimo de firmeza no dia 01 de vida de prateleira e altura média da fatia central, cmc = 0,8 e eprt = 1,41 (variáveis codificadas). A análise sensorial mostrou que o pão sem glúten apresentou características muito próximas de um pão de forma tradicional (p<0,01). Concluiu-se que é possível produzir pães de forma com formulação a base de farinha de arroz com características similares de um pão de forma tradicional, havendo ainda oportunidade de melhoria para a textura e o sabor / Abstract: It is being a challenge for researchers in the food field to make available to the celiac the variety of foods that they are not allowed to consume. The bread has been studied as deep in it, especially the replacement of the technological functionality of gluten, which is particularly nontrivial. The objective of this work was to develop gluten-free bread based on rice flour, with similar characteristics of a traditional loaf. The technological intent of research was divided in five parts: literature review, general mapping and basic formulation development, screening of significant variables via Plackett and Burman design, once determined the significant variables, central composite rotational design 24 including eight star points and four replicates at the central point, followed by central composite rotational design 22 including four star points and four replicates at the central point. In the initial stage 50 formulations were examined, involving 32 ingredients and 12 process variables. At the end, gluten-free pan bread was developed, whose formulation can be obtained by replacing the wheat flour by a mix of rice flour, tapioca starch and modified egg albumen acidified and dehydrated. The rice flour was found to be a good working basis, acting as agent of the body. The modified tapioca starch provided consistency and cohesiveness to the dough. The egg albumen acidified and dehydrated replaced the gluten being responsible for the growth of the dough. Hydrocolloid (carboxymethyl) cellulose was also added, acting as a thickener, contributing to the dough viscosity and water absorption. The other ingredients acted similarly to a traditional formulation of pan bread prepared with wheat flour. The production process has shown that using an mixer with paddle, dough ratio of 3.71 mL/g in a pan 21 x 10 x 10 cm (l x w x h), fermentation of 60 minutes at 40°C held at 85% of relative humidity, followed by baking temperature of 180°C during 23 minutes and cooling during 60 minutes promoted the best results. All samples after 24 hours of its preparation presented an excessive firmness. The experimental design of Plackett and Burman was applied in sequence to determine which variables had significant effect (p <0.10) on the average height of the central slice and firmness on days 01, 03, 07 and 10 after produced of loaves of gluten-free bread based on rice flour. The firmness was influenced by modified cassava starch, maltogenic amylase and water, while the protease, powdered and acidified egg albumen and water had effect on the average height of the center slice The lack of pan flow affected the bread¿s crust color, due to lower heat contact on the surface. The dough consistency influenced the characteristic of the crust. Dough with water in excess and consequently with less structure, had collapsed compromising the symmetry. The crumb color was whiter as the average height of the central slice of the pieces was above 100 mm. The crumb structure followed the trend of color, except PB11, where the levels of powdered and acidified egg albumen and water were maximum and modified cassava starch was minimum, that aside from presenting good height, had a more closed crumb structure. The formulation PB11 presented similar chewing to traditional pan bread. The aroma and flavor ranged from the light of rice flourish to neutral, with remark for formulas PB02 and PB05, where the levels of modified cassava starch and powdered and acidified egg albumen were maximum and water was minimum, whose flavors were unpleasant. Once determined the significant variables the central composite rotational design 24 including eight star points and four replicates at the central point was used to determine which independent variables - dried and acidified egg albumen, water, enzyme maltogenic amylase and modified cassava starch ¿ had a significant effect on the average height of central slice and crumb firmness on days 01, 03, 07 and 10 of gluten-free loaves based on rice flour. As a result of this work, it was found that the average height of the central slice was positively and significantly impacted (p<0.05) by water in the form of a parabolic shape with a maximum at the level 1.1. Regarding the firmness, it was significantly, linear and positively affected (p <0.10 on day 01 and p <0.05 on the other days) by the modified cassava starch, dried and acidified egg albumen. The quadratic and linear terms of the water on the firmness were significantly positive and negative, respectively (p <0.10 on day 01 and p <0.05 on the other days). On the contrary expected, the enzyme maltogenic amylase showed no effect on firmness. All formulations showed good proofing and visual aspects, lacking the flavor that ranged from slightly rice flour to bitter. Once the fungal protease was not studied in the previous step, and it was noted as significant by the Plackett and Burman planning, the central composite rotational design 22 trials including 4 star points and 4 replicates at the central point was employed to determine which independent variables - (carboxymethyl) cellulose (cmc) and fungal protease (eprt) - had a significant effect on the height of the central slice, the crumb firmness on days 01, 03, 07 and 10, specific volume and proofing time of gluten-free loaves based on rice flour. As a result of this work, it was found that the average height of the center slice was positively and significantly impacted (p <0.05) by eprt in the form of a parabolic shape by cmc in linear way, with a maximum at the level 2.0 and 0.8, respectively; and also the linear relationship of eprt and cmc. Regarding the firmness, it was significant and positively affected by the cmc linear and quadratic term for days 01, 03 and 07 after produced (p <0.05, p<0.05 and p<0.10), while the firmness of 10 days was affected negatively only by the linear term of the cmc (p <0.10). The specific volume was positively affected (p <0.10) by cmc linearly, while the quadratic term of cmc positively affected the fermentation (p <0.10). All formulations showed good bread development and good chewing characteristics. The taste remained neutral, with the exception of the experiment with the lowest level of cmc which collapsed and presented bitterness. From these results mathematical models for firmness on days 01, 03, 07 and 10, shelf life, the average height of the center slice, bulk and fermentation time were created. The models were tested and validated at optimum firmness in 01 days of shelf life and average height of the center slice, cmc = 0.8 and eprt = 1.41 (encoded variables). The sensorial analysis has shown that the gluten-free pan bread has presented characteristics very close to regular wheat loaf (p<0.01). It was concluded that it is possible to produce sliced bread based on recipe of rice flour with typical features of a traditional loaf, but that still presents opportunities of improvement in the texture and flavor / Doutorado / Tecnologia de Alimentos / Doutor em Tecnologia de Alimentos

This diploma thesis deals with methods of determining and treatment of coeliac disease. In the theoretical part of the work sums up informations about coeliac disease - history, prevalence, etiopathogenesis, immunopathogenesis, possibilities of diagnosis, discusses about gluten-free diet, mentions legislation of the Czech Republic and the European Union and compares expensiveness of gluten-free diet and normal diet. The experimental part of this thesis compares PCR, ELISA and indirect immunofluorescence methods for assesment of determination of genetic predisposition and antibodies in coeliac disease (greatest emphasis was placed on the comparison of antibodies against gliadin and deamidated gliadin antibodies in IgA and IgG).

The only treatment for coeliac disease, a common autoimmune disorder, is life-long adherence to a gluten-free diet. However, the replacement of wheat gluten, a key structural and functional component in bread, poses a major technological challenge for food scientists. The use of non-wheat cereal proteins, as alternatives to gluten, shows much promise in gluten-free bread making. Literature has shown that when zein, the maize prolamin protein, is subjected to wet heat above its glass transition temperature (Tg), the protein becomes viscoelastic, rubbery and dough-like. Gluten-like fibrils are visible, which form complex protein networks similar to those found in wheat dough. The resulting zein dough has viscoelastic characteristics and can be successfully used with hydrocolloids to produce gluten-free bread. This project examined the influence of wet heat treatment and dilute organic acids (lactic acid and acetic acid) on the dough-making quality of non-wheat cereal proteins, such as kafirin and zein. Zein was the only non-wheat cereal protein to show any physical change when it was subjected to wet heat treatments, forming a dough-like substance. Acidification of the zein dough prepared at 40°C with concentrations of 0.7, 1.3 and 5.4% (v/v) organic acid in distilled water solutions, showed that the higher the concentration of acid used, the greater its effect on the dough's rheological properties. Tensile tests using a Keifer rig on zein dough showed that as the concentration of organic acid was increased from 0.7 to 1.3 and to 5.4% (v/v) the dough become softer and increasingly more extensible. The dough also exhibited less resistance to extension and reduced elasticity. CLSM revealed that the zein doughs contained a protein network, made up of fine protein fibrils, which became smoother and more homogenous as the concentration of acid was increased. Although SDS-PAGE revealed that no oligomerization took place with acid addition, iv FTIR showed that zein dough prepared with distilled water at 40°C had elevated levels of β-sheets. When organic acids were added in increasing levels, corresponding increases in the quantities of α-helices in the protein were observed. Alveography showed that zein-based doughs prepared with dilute organic acids retained gases well and that the concentration of dilute organic acids influenced dough distensibility (biaxial extensibility) and stability (the ability of the dough to retain gas). Low concentration of acids (0.7 and 1.3%) increased dough stability to levels similar to that of strong wheat flour, 103 mm H2O, but higher concentrations of acids (5.4%) led to a marked reduction in dough stability. Thus, by increasing zein dough functionality to such an extent, the apparent usefulness of the doughs and their ability to retain gases produced during fermentation is reduced. Simple distensibility tests on zein doughs showed that added organic acids promoted ‘clumping’ of the fine protein fibrils in the dough network into pronounced fibres. This would account for the decreased dough stability when high levels (5.4%) of organic acids were used. Baking trials with zein doughs were not successful as adequate leavening was impossible without an acid-tolerant leavening agent. It is believed that dilute organic acids influence the rheological properties of zein dough by creating a positively charged environment, in which the protein is partially solubilized. The higher the level of organic acid used, the greater the positive net charge and the more pronounced the effect on the protein network structure. Organic acids could also improve fluidity of the zein dough by acting as plasticizers. From this work it can be seen that although a protein network is present in all zein-based doughs, the ability of this network to retain gases is dependant on the level of organic acids present. The functional properties of zein-doughs made with low levels of organic acids (0.7 and 1.3%) shows potential in the production of gluten-free bread for individuals suffering from coeliac disease. / Dissertation (MSc)--University of Pretoria, 2013. / gm2014 / Food Science / unrestricted

Leukocyte extravasation is mediated in part by tissue specific chemotactic cytokines (chemokines) and specific chemokine receptors expressed on the surface of circulating cells. C-C chemokine ligand CCL25 is expressed exclusively in the intestine and thymus and mediates chemotaxis by cells expressing receptor CCR9. This chemokine and receptor pair may be relevant in the pathogenesis of intestinal inflammation, in diseases such as Crohn’s disease (CD) and coeliac disease. In this thesis I investigated CCR9 expression in situ, in tissues affected by intestinal inflammation, and also examined the effects of CCR9 antagonist treatment in patients. In vitro I investigated CCR9 function using human peripheral blood T cells enriched for CCR9 by cell sorting or all-trans retinoic acid treatment. Using tissues collected as part of a clinical trial in CD testing CCR9 antagonist, CCX282-B, I investigated ways of measuring if treatment reduced the number of CCR9 expressing cells in the intestinal mucosa. However, in situ staining for CCR9 by immunohistochemistry was unsuccessful, and in this thesis, I explored reasons why this might be the case. Treatment with CCX282-B did however, show a tendency to reduce T cell density in the intestinal mucosa, although results were highly variable between individuals. In an examination of human CCR9 function in vitro I demonstrate for the first time that CCL25 stimulates CCR9 surface internalization. These data clarify the observation that CCR9 staining by IHC produces poor results in tissues where ligand is abundant, such as the intestine and thymus. I describe a novel technique for measuring calcium flux in two populations simultaneously by flow cytometry, which confirmed that in a heterogeneous population of cells, only CCR9 expressing cells respond to CCL25 by calcium flux. Variability in clinical trials is partly created by the use of concomitant medications, and in CD, corticosteroids are widely used. For the first time I show that glucocorticoids (GC) impair CCR9 mediated chemotaxis, calcium flux and intracellular signalling without changes to CCR9 mRNA and surface protein expression. Reduced CCR9 mediated signalling was accompanied by an enhanced expression and function of co-expressed CXCR4, demonstrating that the effects of GC were receptor-specific and not mediated by non-specific toxicity or inhibition of cell signalling. In a second study CCX282-B was tested in patients with coeliac disease, and in this trial, there was no reported concomitant use of GCs. It was confirmed that dietary gluten stimulates significant T cell recruitment to the intestinal mucosa with a pronounced accumulation of intraepithelial lymphocytes (IEL) and a rise in the frequency of FoxP3 expressing cells. Patients on CCX282-B had lower IEL counts, and an equivalent proportion of FoxP3 expressing T cells, suggesting that CCR9 blockade restricted the recruitment of effector T cell subsets. This thesis confirms that the accumulation of T cells is central to inflammation in the intestine and that modulating chemokine receptor function may affect this. Furthermore, this thesis demonstrates that the function of CCR9 is suppressed by GCs, which are widely used therapeutically and therefore could identify a novel mechanistic basis for their activity in CD.

<p>Coeliac disease is a chronic inflammatory disease treated with a gluten-free diet, excluding barley, rye and wheat. Hence, there is a demand for methods able to detect gluten in foods in order to ensure correct labeling of products. According to the Codex Alimentarius Commission, 20ppm gluten is the maximum amount allowed in food labeled gluten-free.</p><p>PCR can detect DNA from cereals in food. Four real-time PCR-systems,</p><p>using TaqMan®-probes for detection of barley, oat, rye and wheat were optimized and evaluated. Evaluations were carried out using seeds. Primers were targeted to genes coding for prolamines, seed storage proteins. PCR-systems targeted to barley, oat and wheat were shown to be specific for the cereals corresponding to each system. The system targeted to rye showed cross-reactions with durum wheat and spelt wheat. Detection limits were 50pg, corresponding to <10 haploid genome copies for each cereal. All systems were able to detect 250ppm amounts of DNA, most likely even smaller amounts are detectable. All systems showed an amplification efficiency of ≥95%.</p><p>Systems for detection of barley, oat and wheat are ready for further evaluation, using food products as samples. The rye system however, needs to be re-designed before further evaluation can take place.</p>