<p dir="ltr">This thesis explores consumers' knowledge, perceptions, and handling practices of tree nuts through needs assessments to collect data for the development of tree nut food safety education materials.</p>

La détection d’antigènes à large échelle s’avère être un enjeu majeur dans le secteur de la production alimentaire. Le défi persistant encore à ce jour est la détection spécifique et quantitative des protéines allergènes dans les produits commercialisés, comestibles, cosmétiques ou autres. La technique la plus répandue en industrie pour la détection de matériel biologique spécifique est le test enzyme-linked immunosorbent Assay (ELISA). ELISA permet un dosage de diverses protéines au sein d’un échantillon, mais ne détecte qu’un seul antigène à la fois, ce qui est couteux et peu efficace. La hausse des réactions allergiques notées chez la population de divers pays demande à ce qu’une technique plus efficace et moins couteuse soit développée. Dans le cadre de ce mémoire, une méthode alternative au test ELISA pour la détection d’allergènes de noix sera présentée. Cette nouvelle méthode appelée RIISA (Raman imaging immunosorbent assay), fait intervenir les propriétés de diffusion Raman de nanostructures de carbone, tel que les nanotubes et les nanocornes de carbone, pour les adapter comme sonde optique. L’objectif de ce travail est d’adapter cette méthode pour la détection triplex, permettant la reconnaissance RIISA entre trois couples d’antigène-anticorps, soit d’amande, de cachou et de noisette. Les sondes sont fonctionnalisées d’anticorps permettant la reconnaissance avec la protéine et comportent aussi un colorant encapsulé dans la cavité interne, ce qui permet la différenciation des couples. Le système de détection est couplé à l’imagerie Raman pour permettre une sensibilité plus aigüe du système. Un phénomène de réaction croisée entre les anticorps et les antigènes empêche toutefois de réaliser une détection en triplex. La preuve de concept sera donc démontrée en utilisant les trois monoplexes, et ce, avec divers colorants mettant en évidence la polyvalence du système de détection présenté. / Large scale antigen detection is a major issue in the food production sector. The challenge that still persists nowadays is the specific and quantitative detection of allergenic protein products on the market, whether edible, cosmetic or otherwise. The most widely used technique in the industry for the specific biological detection of allergens is the Enzyme-linked immunosorbent assay (ELISA). This assay detects various proteins in a simple sample. The ELISA test can detect only one allergen at a time and its quantitative detection is limited. In addition, the increase in allergic reactions observed in the population of various countries require a more efficient and less expensive technique to be developed. As part of this master’s thesis, an alternative method to the ELISA assay to detect nut allergens will be presented. This assay, called Raman imaging immunosorbent assay (RIISA), uses carbon nanostructure based nanoprobes consisting of carbon nanotubes and nanohorns. The objective is to develop a triplex detection system using RIISA, that selectively recognizes between three antigen-antibody pairs, namely almond, cashew, and hazelnut. The dye encapsulated nanoprobes are functionalized with antibodies recognizing the protein and for simultaneously differentiating the couples. The presented detection system is coupled with Raman imaging and allows a more acute sensitivity of the system. However, cross reactions between the antibodies and the antigens prevent the triplex detection. The proof of concept will be demonstrated using the three monoplex couples with various dyes that highlight the versatility of the developed system.

Thesis (MEd)--University of Stellenbosch, 2004. / ENGLISH ABSTRACT: The name of the School 'Tot Nut van het Algemeen' appears often in the literature on early Cape education. It is described as an institution of excellence that boasts many famous pupils such as President Jan Brand, Ds JH Neethling and 'Onze Jan' Hendrik Hofmeyr. In this study I explore how the School managed to adapt to political, social and philosophical changes to survive for 70 years. I do this through telling the narrative of its existence and functioning, and investigate the vexing question as to why it was forced to close in 1870. The research document consists of 9 chapters. The introductory chapter provides the orientation for the study. It is followed by a chapter dealing with the factors that led to the establishment of the School, taking into account events both overseas and at the Cape. Chapter 3 focuses on the British occupation of the Cape, with special emphasis on the Anglicisation of schools and the reaction of the colonists to this change of circumstance. Chapter 4 describes the School's activities from 1832, covering among other aspects, its reopening, curriculum and funding. The School's link with the South African College is also explored. In Chapter 5, I discuss the education policies that shaped the School, as well as the ideals of liberalism and democracy in as far as the School practiced them. Chapter 6 deals with the closing of the School, and I offer various reasons for this. In chapter 7, I present cameos of some of the influential teachers, while the School's legacy to Cape society is examined in Chapter 8. The study concludes with a reflection that draws these facts into an integrated view and highlights pertinent insights into the 'Tot Nut' as a worthy institution in the light of the findings revealed in this research. / AFRIKAANSE OPSOMMING: Die naam van die skool 'Tot Nut van het Algemeen' verskyn dikwels in die literatuur oor vroeë onderwys aan die Kaap. Dit word as 'n puik instansie beskryf, met menige bekende oudleerlinge soos President Jan Brand, Ds JH Neethling en 'Onze Jan' Hendrik Hofmeyr. In hierdie studie ondersoek ek hoe die Skool by verskeie politiese, sosiale en filosofiese veranderinge aangepas het om sodoende 70 jaar te kon oorleef. Dit doen ek deur die Skool se bestaan en funksionering te beskryf, en ek spreek die frustrerende kwessie aan waarom dit in 1870 gedwing is om te sluit. Die navorsingsverslag bestaan uit 9 hoofstukke. Die inleiding behels die oriëntasie ten opsigte van die studie. Dit word gevolg deur 'n hoofstuk wat handeloor die faktore wat tot die ontstaan van die Skool gelei het, waar daar na gebeure oorsee sowel as aan die Kaap, gekyk word. Hoofstuk 3 fokus op die Britse besetting, veralop die Anglisasie van die skole en die , koloniste se reaksie daarop. Hoofstuk 4 beskryf die Skool se aktiwiteite vanaf 1832, onder andere sy heropening, die kurrikulum en bevondsing. Die Skool se verwantskap met die Suid- Afrikaanse Kollege word ook bespreek. In Hoofstuk 5 ondersoek ek die opvoedingsbeleid wat die Skool beïnvloed het, asook die ideale van liberalisme en demokrasie in so ver die Skool dit beoefen het. Hoofstuk 6 handeloor die sluiting van die Skool en ek bied verskeie redes daarvoor aan. In Hoofstuk 7 bestaan uit sketse van die vernaamste onderwyspersoneel, terwyl die Skool se bydrae tot die Kaapse samelewing in Hoofstuk 8 voorgelê word. Die studie word afgerond met 'n terugblik wat hierdie feite in 'n integreerde perspektief oor die 'Tot Nut' bymekaar bring en, in die lig van die bevindings wat in hierdie navorsing blootgelê is, kan dit as 'n waardige instansie beskou word.

The Amazon rainforest harbors the planet's highest biodiversity among terrestrial ecosystems; however, the biology and ecology of most of its species are unknown. Niche partitioning is considered a key factor allowing species co-existence, especially for morphological similar species such as spiny rats of the genus Proechimys. I examined the extent to which habitat differentiation, species body mass, and diet could explain the community composition of small mammals at a site in the southeastern Amazon. Moreover, I radio-tagged sympatric species of Proechimys spp. and Mesomys stimulax (an arboreal spiny rat) to obtain detailed autoecology information, including habitat use and use of space. I found support for niche partitioning among species and associated small mammal species with distinct successional phases of gap dynamics. I also observed among Proechimys spp. a typical polygynous organization: females appeared to be territorial against females of any species of their genus; but male territories overlapped with those of several females. Mesomys stimulax showed evidence of monogamy and possible sociality, although sample sizes were small. To identify the importance of small mammals as seed disperser of Brazil nut seeds, I conduct experiments using a combination of fluorescent powder, seed exclosures, and track plates in forests with and without Brazil nut groves, and in forests with and without Brazil nut harvesting. Among small mammals, only Proechimys spp. removed, dispersed, and preyed upon Brazil nut seeds. Proechimys spp. sometimes scatterhoarded these seeds, and hence have the potential to play a significant role in recruiting new Brazil nut trees. I also trapped small mammals and measured forest structures on the same sites used for the seed dispersal experiment, to determine the ecological effects of Brazil nut harvest on small mammal communities; however, my results showed little evidence of changes associated with the seed exploitation. My results highlight the importance of habitat heterogeneity in structuring small mammal communities, and indicate that forest management practices that alter habitats, such as partial logging, also can be expected to alter small mammal composition and diversity. Proechimys spp. have the potential to play an important role in the ecological restoration of intensive exploited Brazil nut groves.

Anopheles gambiae sensu lato mosquitoes are among the dominant malaria vectors in sub-Saharan Africa. However, not much is known about the oviposition behaviour of these species necessary for the development of malaria vector control strategies. With the aim of investigating cues associated with selected oviposition sites, artificial oviposition sites- ponds (soil mixed with water) were set-up in an open field at Mbita, Western Kenya in 2012 and 2013. Ponds were allowed to be colonized by wild An. gambiae s.l.. The numbers of Anopheles early instar larvae were counted and used as a proxy for oviposition preference. Water samples were then analysed for physicochemical, bacterial and chemical profiles. The bacterial profiles were analysed using denaturing gradient gel electrophoresis (DGGE) and the chemical profiles with gas chromatography-mass spectrometry (GC-MS). The detection of possible oviposition cues from oviposition substrates requires sensitive analytical methods. Volatiles detection was improved seven times. The detection of bacteria deoxyribonucleic acid (DNA) bands with DGGE was also improved to a minimum DNA concentration of 50 ng/µl. Results showed that ponds were colonized differently. Fresh ponds were preferred over slightly older ponds. Bacterial analysis revealed a low number of bacteria colony forming units (CFU) in preferred ponds. Some volatiles, including: 6,10-dimethyl-5,9-undecadien-2-one (geranylacetone) and 4-ethylbenzaldehyde, were associated with the oviposition preferred pond. In addition, low pH and high turbidity were associated with the ponds selected for oviposition. Finally, fungi isolated from the rhizomes of nut grass yielded a promising array of volatiles of which one is known to attract oviposition site seeking malaria mosquitoes. This finding opens the door for a cost effective and environmental friendly method of using fungi in an “attract and kill” strategy targeting malaria vectors. / Myggor i Anopheles gambiae sensu lato komplexet tillhör de myggor som är bäst på att sprida malaria parasiter i afrika söder om Sahara. Kunskapen om de här myggornas äggläggningsbeteende är begränsad. Den här kunskapen behövs för att kunna utveckla nya och förbättra tillgängliga malaria vektor kontroll metoder. Nya metoder som kan komplettera de som används idag (insecticides treated nets (ITNs) och indoor residual spraying (IRS)) behövs eftersom de metoderna har problem med resistensutveckling. Två studier utfördes på icipe fältstation i Mbita västra Kenya under 2012 och 2013 med målet att identifiera faktorer som påverkar myggornas äggläggningsbeteende. Baljor fyllda med en blandning av jord och vatten (äggläggningssubstrat) användes för att tillverka artificiella äggläggningsplatser som liknar de vattenpölar som de här myggarterna gärna lägger ägg i. Baljorna koloniserades av vilda myggor och antalet mygglarver som detekterades i baljorna jämfördes och användes som en proxy för äggläggningspreferens. Fysikaliska och kemiska parametrar mättes på jordvattenblandningarna i baljorna och prover togs för att analysera bakteriepopulationer med hjälp av denaturing gradient gel electrophoresis (DGGE) och flyktiga ämnen med hjälp av gas-kromatografi kopplat till mass-spektrometri (GC-MS). För att kunna detektera de låga halter av flyktiga ämnen och bakterier som fanns i de här proverna krävdes det känsliga metoder. Antalet flyktiga ämnen som kunde detekteras ökades sju gånger genom att tillsätta NaCl till vattenproverna innan doften insamlades och termisk desorption användes istället för lösningsmedels desorption. För att förbättra detektionsgränsen för bakterier amplifierades bakterie-DNA i två PCR reaktioner som sedan mixades och koncentrerades. Resultaten från fältstudierna med baljorna visade att de koloniserades olika av Anopheles myggorna. Baljor med nyblandat substrat innehöll dubbelt så många mygglarver som baljor med jord-vattensubstrat som åldrats under en längre tid. Lägre mängd bakterier, lägre pH och högre grumlighet var gemensamt för de baljor som myggorna föredrog. De flyktiga ämnen som detekterades i de olika baljor varierade mellan olika försök och inget ämne fanns med i alla upprepningar av ett försök. Trots det detekterades några ämnen oftare i de baljor som myggorna föredrog att lägga ägg jämfört med de med en mindre mängd mygglarver. De inkluderar geranylacetone och 4-ethylbenzaldehyde. Svampar isolerades från rotstockar av gräs som fanns i den jord som användes för att göra äggläggningssubstraten i fältstudierna. De flyktiga ämnen som avgavs från svampkulturerna analyserades. Bland annat så identifierades ett ämne som fungerar som en äggläggningsattrahent för An. gambiae s.l. myggor.Resultaten från den här avhandlingen kommer att kunna användas för att utveckla miljövänliga ”attract and kill” metoder för att kontrollera malaria myggor. / <p>QC 20160211</p>

Partindo das evidencias encontradas na literatura sobre o desenvolvimento do uso de ferramentas de percussão para quebra de frutos encapsulados, propomos um projeto focalizando não apenas quando as alterações motoras aparecem, mas também na relação dessa trajetoria de mudança com duas possíveis vias de influencia do nicho ecológico sobre a trajetoria de mudança comportamental que da origem ao uso de ferramentas de percussão para quebra de frutos encapsulados em macacos-prego (Sapajus libidinosus). Em primatas, os aspectos físicos e sociais (isto é, artefatos: martelos, bigornas e restos de cocos, e o comportamento dos coespecíficos) canalizam as oportunidades individuais de aprendizagem, permitindo maiores níveis de manipulação de objetos apropriados em contextos apropriados para o surgimento do uso de ferramentas e facilitando a exploração de várias fontes de forrageamento. Por tanto, procuramos determinar o papel de duas variáveis específicas de influência de nicho construídas: reuso dos artefatos e scrounging, no processo de desenvolvimento do repertório manipulativo em um grupo silvestre de macacos-prego (Sapajus libidinosus). Especificamente: 1) Como é a trajetória de mudança na atração e reutilização de artefatos relacionados ao craqueamento de noz (martelos, bigornas e cascas de nozes) e como isso se relaciona com o surgimento de ferramentas de percussão; E, 2) Como relaciona-se o scrounging, no contexto da quebra de frutos, com a emergencia de habilidades manipulativas adequadas para a quebra de frutos encapsulados. Propomos uma pesquisa de campo longitudinal de uma população de macacosprego livres no Brasil, através da observação naturalista. O resultados indicam que não só a idade está relacionada tanto com as mudanças na atração e reuso dos artefatos e com o scrounging, mas também que ambas variaveis estão relacionadas e podem estar em função das mudanças no repertorio manipulative exibido pelos individuos, de tal forma que não só os individuos mais velhos exibem maiores indices de reuso e menores indices de scrounging, como também os indices de variabilidade comportamental e proficiencia na quebra estão intimamente relacionados com os indices de reuso dos artefatos e scrounging / Based on the evidence found in the literature on the development of the use of percussion tools for encapsulated fruit breakage, we propose a project focusing not only \"when\" motor alterations appear, but also on the relation of this trajectory of change with two possible ways of influencing the ecological niche on the trajectory of behavioral change that gives origin to the use of percussion tools to crack nuts in capuchin-monkeys (Sapajus libidinosus). In primates, physical and social aspects (ie, artifacts: hammers, anvils and remains of coconuts, and co-specific behavior) channel individual learning opportunities, allowing greater levels of manipulation of appropriate objects in appropriate contexts for the emergence of use of tools and facilitating the exploration of various foraging sources. Therefore, we sought to determine the role of two niche-specific variables constructed: artifact reuse and scrounging, in the process of developing the manipulative repertoire in a wild group of monkey-prey (Sapajus libidinosus). Specifically: 1) How is the trajectory of change in the attraction and reuse of artifacts related to nut cracking (hammers, anvils and walnut shells) and how does this relate to the emergence of percussion behavior. And, 2) How scrounging is related, in the context of the fruit break, with the emergence of adequate manipulative skills for the breaking of encapsulated fruits. We propose a longitudinal field survey of a population of free monkey-natives in Brazil, through naturalistic observation. The results indicate that not only age is related both to changes in attraction and reuse of artifacts and to scrounging, but also that both variables are related and may be due to changes in the manipulative repertoire exhibited by individuals, such that not only do older individuals exhibit higher reuse rates and lower scrounging rates, but also the indices of behavioral variability and proficiency in the break are closely related to the artifact reuse indices and scrounging

Nanopartículas lipídicas têm sido desenvolvidas para aplicação tópica de fármacos e ativos cosméticos. Neste trabalho, foi proposta a primeira aplicação de um lipídeo natural não-refinado biodegradável e biocompatível - manteiga de cupuaçu (Theobroma grandiflorum) - para a preparação de nanopartículas lipídicas, as quais foram denominadas teosferas. As teosferas foram preparadas por emulsificação-evaporação do solvente (EES) e por homogeneização à alta pressão (HAP), apresentando tamanho nanométrico e distribuição granulométrica estreita quando preparadas por ambos os métodos. O trabalho teve continuidade com a preparação de teosferas pelo método de EES utilizando manteiga de cupuaçu ou sua mistura com óleo de castanha do Brasil (Bertholletia excelsa) - também derivado da biodiversidade Amazônica - visando a incorporação de antioxidantes. Idebenona (IDB) foi selecionada por sua conhecida ação antioxidante e pela sua utilização em formulações cosméticas antienvelhecimento. IDB foi incorporada nas teosferas com eficiência de encapsulação superior a 99%, sendo que os estudos de liberação in vitro mostraram que a liberação de IDB a partir das teosferas foi mais lenta em comparação à IDB livre. Estes experimentos foram capazes ainda de demonstrar as características elásticas das teosferas. Além disso, foi evidenciada in vitro a atividade antioxidante superior das teosferas contendo IDB em relação ao ativo livre. Visando possibilitar a aplicação tópica de teosferas contendo IDB, em um trabalho subseqüente, suspensões de teosferas preparadas por HAP foram incorporadas em géis hidrofílicos. As formulações apresentaram características pseudoplásticas e demonstraram efeito oclusivo in vitro, o qual foi dependente da composição dos colóides. Finalmente, os estudos de permeação in vitro utilizando pele humana demonstraram que teosferas e nanocápsulas de núcleo lipídico, utilizadas neste estudo de modo comparativo, modificaram a permeção da IDB, permitindo a acumulação do ativo nas camadas superficiais da pele. / Lipid nanoparticles have been developed for administration of active substances to the skin, both for pharmaceutical and cosmetic uses. In the present work, we proposed the first use of a none-refined natural biodegradable and biocompatible lipid – Cupuaçu seed butter (Theobroma grandiflorum) – for the preparation of lipid nanoparticles, which were called theospheres. Theospheres were prepared by emulsification-solvent evaporation (ESE) and by high pressure homogenization technique (HPH), presenting size in nanometrical range and narrow particle size distribution for both methods. Taking these results into account, the next step of this work was the preparation of theospheres by ESE method using Cupuaçu seed butter with or without Brazil nut (Bertholletia excelsa) seed oil - another ingredient derived from an Amazonian fruit - intending the encapsulation of an antioxidant. Idebenone (IDB) has been selected due to its known antioxidant action and because it has been used in antiaging cosmetic formulations. IDB was incorporated in the theospheres presenting encapsulation efficiency higher than 99%. The in vitro release evaluation demonstrated that the release of IDB from theospheres was lower than that of free drug. Besides, the in vitro release study highlighted the elastic characteristics of theospheres. Additionally, IDB-loaded theospheres showed higher antioxidant activity compared to free IDB. Viewing the cutaneous administration, theosphere suspensions prepared by HPH technique were incorporated into hydrogels. The rheograms of the semi-solid formulations exhibited a non-Newtonian behavior presenting pseudoplastic characteristics. In vitro occlusion study highlighted the dependence of the occlusive effect on the lipidic composition of the theospheres. Finally, in vitro human skin permeation studies showed that theospheres and lipid-core nanocapsules, used in this study in a comparative way, changed the permeation of IDB, increasing the accumulative amount of IDB in the upper skin layer.

The current research was done to evaluate the nutritive values and the ruminal degradation of dry matter (DM) and crude protein (CP) from three non-conventional oil cakes, viz: amarula (Sclerocarya birrea) (AOC), macadamia (Integrifolia) (MOC) and baobab (Adansonia digitate L.) (BOC). The oil cakes were collected from biodiesel producers in Limpopo Province, transported to the ARC-Animal Production campus, where proximate and ruminal nutrient degradation analysis were conducted. Triplicates samples from each oil cake were analyzed for the nutritive values, mineral and amino acids contents. Three rumen cannulated mid-lactating (days in milk; DIM: 180±5) Holstein cows weighing 667±43 kg body weight were allocated to determine the in situ ruminal dry matter (DM) and crude protein (CP) degradation. The cows were offered a totally mixed ration (TMR) (60 concentrate: 40 forage ratio) that was compounded according to their daily nutrient requirements, and were milking was done twice per day at 12 hrs intervals. The three oil cake samples were ground using a 2-mm screen after which sub-samples (6.5 g) were put in 10 x 20 cm; 50 μm pore size polyester bags to achieve 15 mg/cm² (ratio of the sample size to surface area). The bags were then fistulated in each cow’s rumen in triplicate for a period of 2, 4, 8, 16, 24, or 48 hrs. After being incubated, the bags were removed from the rumen and washed with cold (4°C) water in 20-L buckets. Following immersing in cold water, the bags were machine washed until clean water was obtained. The bags were then dried at 60 °C in an oven for 48 hrs. The dried bags were individually weighed, and the content of each bag were removed and stored into glass vial until analysis. The remaining two duplicate sets of each sample were rinsed using cold water in order to determine solubility at 0 hrs. The AOC had higher (P<0.05) ether extract (EE) and CP content than both BOC and MOC. Macadamia oilcake (MOC) and BOC had higher (P<0.05) fractions of fibre (NDF, ADF and ADL) compared to the AOC. The AOC had greater (P<0.05) content of essential amino acids than in the BOC and MOC. Additionally, AOC had a high (P<0.05) phosphorus, but low calcium and potassium concentration. While AOC had high effective degradability of DM, it also had high water soluble as well as DM and CP rapidly degradable fractions. Effective degradation of CP was higher in AOC and BOC than in MOC. However, BOC had a high insoluble but degradable fraction of CP. Further work to determine the toxicology of these non-conventional oil cakes and animal feeding experiments is needed / Agriculture and  Animal Health / MSc. Agriculture

The Amazon rainforest harbors the planet's highest biodiversity among terrestrial ecosystems; however, the biology and ecology of most of its species are unknown. Niche partitioning is considered a key factor allowing species co-existence, especially for morphological similar species such as spiny rats of the genus Proechimys. I examined the extent to which habitat differentiation, species body mass, and diet could explain the community composition of small mammals at a site in the southeastern Amazon. Moreover, I radio-tagged sympatric species of Proechimys spp. and Mesomys stimulax (an arboreal spiny rat) to obtain detailed autoecology information, including habitat use and use of space. I found support for niche partitioning among species and associated small mammal species with distinct successional phases of gap dynamics. I also observed among Proechimys spp. a typical polygynous organization: females appeared to be territorial against females of any species of their genus; but male territories overlapped with those of several females. Mesomys stimulax showed evidence of monogamy and possible sociality, although sample sizes were small. To identify the importance of small mammals as seed disperser of Brazil nut seeds, I conduct experiments using a combination of fluorescent powder, seed exclosures, and track plates in forests with and without Brazil nut groves, and in forests with and without Brazil nut harvesting. Among small mammals, only Proechimys spp. removed, dispersed, and preyed upon Brazil nut seeds. Proechimys spp. sometimes scatterhoarded these seeds, and hence have the potential to play a significant role in recruiting new Brazil nut trees. I also trapped small mammals and measured forest structures on the same sites used for the seed dispersal experiment, to determine the ecological effects of Brazil nut harvest on small mammal communities; however, my results showed little evidence of changes associated with the seed exploitation. My results highlight the importance of habitat heterogeneity in structuring small mammal communities, and indicate that forest management practices that alter habitats, such as partial logging, also can be expected to alter small mammal composition and diversity. Proechimys spp. have the potential to play an important role in the ecological restoration of intensive exploited Brazil nut groves.

Knowledge of the dissociation constants of the ionizable protons of weak acids in aqueous media is of fundamental importance in many areas of chemistry and biochemistry. The pKa value, or equilibrium dissociation constant, of a molecule determines the relative concentration of its protonated and deprotonated forms at a specified pH and is therefore an important descriptor of its chemical reactivity. Considerable efforts have been devoted to the determination of pKa values by different experimental techniques. Although in most cases the determination of pKa values from experimental is straightforward, there are situations where interpretation is difficult and the results ambiguous. It is, therefore, not surprising that the capability to provide accurate estimates of the pKa value has been a central goal in theoretical chemistry and there has been a large effort in developing methodologies for predicting pKa values for a variety of chemical systems by differing quantum chemical techniques. A prediction accuracy within 0.5 pKa units of experiment is the desirable level of accuracy. This is a non-trivial exercise, for an error of 1 kcal/mol in estimates of the free energy value would result in an error of 0.74 pKa units. In this thesis ab initio Car-Parrinello molecular dynamics (CPMD) has been used for investigating the Brϕnsted acid-base chemistry of weak acids in aqueous solution. A key issue in any dissociation event is how the solvating water molecules arrange themselves spatially and dynamically around the neutral and dissociated acid molecule. Ab initio methods have the advantage that all solvent water molecules can, in principle, be con- sidered explicitly. One of the factors that has inhibited the widespread use of ab initio MD methods to study the dissociation reaction is that dissociation of weak acids are rare events that require extremely long simulation times before one is observed. The metady- namics formalism provides a solution to this conundrum by preventing the system from revisiting regions of configuration space where it has been in the past. The formalism allows the system to escape the free-energy minima by biasing the dynamics with a history dependent potential (or force) that acts on select degrees of freedom, referred to as collective variables. The bias potentials, modeled by repulsive inverted Gaussians that are dropped during propagation, drive the system out of any free-energy minima and allow it to explore the configurational space by a relatively quick and efficient sampling. The the- sis deals with a detailed investigation of the Brϕnsted acid-base chemistry of weak acids in aqueous solutions by the CPMD-metadynamics procedure. In Chapter 1, current approaches for the theoretical estimation of pKa values are summarized while in Chapter 2 the simulation methodology and the metadynamics sampling techniques used in this study are described. The potential of the CPMD-metadynamics procedure to provide estimates of the acid dissociation constant (pKa) is explored in Chapter 3, using acetic acid as a test sys- tem. Using the bond-distance dependent coordination number of protons bound to the dissociating carboxylic groups as the collective variable, the free-energy profile for the dissociation reaction of acetic acid in water was computed. Convergence of the free-energy profiles and barriers for the simulations parameters is demonstrated. The free-energy profiles exhibit two distinct minima corresponding to the dissociated and neutral states of the acid and the deterrence in their values provides the estimate for pKa. The estimated value of pKa for acetic acid from the simulations, 4.80, is in good agreement with the experiment at value of 4.76. It is shown that the good agreement with experiment is a consequence of the cancellation of errors, as the pKa values are computed as the divergence in the free energy values at the minima corresponding to the neutral and dissociated state. The chapter further explores the critical factors required for obtaining accurate estimates of the pKa values by the CPMD-metadynamics procedure. It is shown that having water molecules sufficient to complete three hydration shells as well as maintaining water density in the simulation cell as close to unity is important. In Chapter 4, the CPMD-metadynamics procedure described in Chapter-3 has been used to investigate the dissociation of a series of weak organic acids in aqueous solutions. The acids studied were chosen to highlight some of the major factors that influence the dissociation constant. These include the influence of the inductive effect, the stabilization of the dissociated anion by H-bonding as well as the presence of multiple ionizable groups. The acids investigated were aliphatic carboxylic acids, chlorine-substituted carboxylic acids, cis- and trans-butenedioic, the isomers of hydroxybenzoic acid and ophthalmic acids and its isomers. It was found that in each of these examples the CPMD-metadynamics procedure correctly estimates the pKa values, indicating that the formulism is capable of capturing these influences and equally importantly indicating that the cancellation of errors is indeed universal. Further, it is shown that the procedure can provide accurate estimates of the successive pKa values of polypro tic acids as well as the subtle difference in their values for different isomers of the acid molecule. Changes in protonation-deprotonation of amino acid residues in proteins play a key role in many biological processes and pathways. It is shown that CPMD simulations in conjunction with metadynamics calculations of the free energy profile of the protonation- deprotonation reaction can provide estimates of the multiple pKa values of the 20 canonical α-amino acids in aqueous solutions in good agreement with experiment (Chapter 5). The distance-dependent coordination number of the protons bound to the hydroxyl oxygen of the carboxylic and the amine groups is used as the collective variable to explore the free energy profiles of the Brϕnsted acid-base chemistry of amino acids in aqueous solutions. Water molecules, sufficient to complete three hydration shells surrounding the acid molecule were included explicitly in the computation procedure. The method works equally well for amino acids with neutral, acidic and basic side chains and provides estimates of the multiple pKa values with a mean relative error with respect to experimental results, of 0.2 pKa units. The tripeptide Glutathione (GSH) is one of the most abundant peptides and the major repository for non-protein sulfur in both animal and plant cells. It plays a critical role in intracellular oxidative stress management by the reversible formation of glutathione disulfide with the thioldisulfide pair acting as a redox buffer. The state of charge of the ionizable groups of GSH can influences the redox couple and hence the pKa value of the cysteine residue of GSH is critical to its functioning. In Chapter 6, it has been reported that ab initio Car-Parrinello Molecular Dynamics simulations of glutathione solvated by 200 water molecules, all of which are considered in the simulation. It is shown that the free-energy landscape for the protonation - deprotonation reaction of the cysteine residue of GSH computed using metadynamics sampling provides accurate estimates of the pKa and correctly predicts the shift in the dissociation constant values as compared to the isolated cysteine amino acid. The dissociation constants of weak acids are commonly determined from pH-titration curves. For simple acids the determination of the pKa from the titration curves using the Henderson-Hasselbalch equation is relatively straightforward. There are situations, however, especially in polyprotic acids with closely spaced dissociation constants, where titration curves do not exhibit clear inflexion and equivalence stages and consequently the estimation of multiple pKa values from a single titration curve is no longer straightfor- ward resulting in uncertainties in the determined pKa values. In Chapter 7, the multiple dissociation constant of the hexapeptide glutathione disulfide (GSSG) with six ionizable groups and six associated dissociation constants has been investigated. The six pKa values of GSSG were estimated using the CPMD-metadynamics procedure from the free-energy profiles for each dissociation reaction computed using the appropriate collective variable. The six pKa values of GSSG were estimated and the theoretical pH-titration curve was then compared with the experimentally measured pH-titration curve and found to be in excellent agreement. The object of the exercise was to establish whether interpretation of pH-titration curves of complex molecules with multiple ionizable groups could be facilitated using results of ab initio molecular dynamics simulations.