Submitted by Sandra Pereira (srpereira@ufrrj.br) on 2016-08-01T17:07:57Z No. of bitstreams: 1 2011 - Juliana Mariano Torres.pdf: 1156662 bytes, checksum: 6ae713d0002e13c804fc2eba73bd8f5b (MD5) / Made available in DSpace on 2016-08-01T17:07:57Z (GMT). No. of bitstreams: 1 2011 - Juliana Mariano Torres.pdf: 1156662 bytes, checksum: 6ae713d0002e13c804fc2eba73bd8f5b (MD5) Previous issue date: 2011-07-22 / Conselho Nacional de Desenvolvimento Cient?fico e Tecnol?gico, CNPq. / The increase in life expectancy is a worldwide occurrence that shows the relative success of public health politics, and Brazil is among the countries where people are living longer and in better physical health. But the increase in life expectancy has a negative effect: the appearance of degenerative diseases typical of old age, including several forms of dementia, like Alzheimer?s Disease (AD) that is certainly the most important. It is an irreversible and progressive disease characterized by neuronal deterioration that results in loss of cognitive functions such as memory, communication skills, judgment and reasoning. Donepezil, rivastigmine and galantamine are medicines used for the treatment of AD and act reversibly inhibiting the acetylcholinesterase (AChE). Evidences suggest that the enzyme butyrylcholinesterase (BChE), closely related to AChE, plays a significant role in AD because it is involved in neural functions such as cholinergic co-regulation and non-cholinergic neurotransmission. The aim of this research is to provide new substances with anticholinesterase action by using ?-carboline derivatives from the natural product harmane and carry out a kinetic study to determine the inhibition profile of the enzymes AChE e BChE, which could help in the discovery of new compounds which could be useful in the treatment of AD. Firstly, a screening was carried out with seven ?-carboline derivatives. In a second stage, a kinetic investigation, employing Ellman?s method, was run with these compounds and all of them presented high anticholinesterase action for both AChE and BChE. All seven derivatives presented a non-competitive reversible inhibition. / Torres, Juliana Mariano. ESTUDO CIN?TICO DA ATIVIDADE ANTICOLINESTER?SICA DE DERIVADOS -CARBOL?NICOS DO PRODUTO NATURAL HARMANA. 2011. Disserta??o (mestrado em Qu?mica Org?nica). Instituto de Ci?ncias Exatas, Departamento de Qu?mica, Universidade Federal Rural do Rio de Janeiro, Serop?dica, RJ, 2011. O aumento da expectativa de vida ? um fen?meno mundial que mostra o relativo sucesso de pol?ticas de sa?de p?blica, e o Brasil se inclui entre os pa?ses em que as pessoas est?o vivendo por mais tempo e em condi??es melhores de sa?de. No entanto, o aumento da expectativa de vida tem como efeito negativo o aparecimento de doen?as degenerativas, t?picas de idades mais avan?adas, incluindo-se as v?rias formas de dem?ncia e entre estas, a mais importante ?, sem d?vida, a Doen?a de Alzheimer (DA), patologia irrevers?vel e progressiva caracterizada pela deteriora??o neuronal que resulta em perda de fun??es cognitivas, tais como mem?ria, capacidade de comunica??o, julgamento e racioc?nio. Para o tratamento da DA s?o utilizados f?rmacos como o donepezil, galantamina e rivastigmina, os quais agem inibindo revers?velmente a acetilcolinesterase (AChE). Evid?ncias sugerem que a enzima butirilcolinesterase (BChE), intimamente relacionada com a AChE, tem um papel significante na DA, uma vez que est? envolvida em fun??es neurais tais como a corregula??o da neurotransmiss?o colin?rgica e n?o-colin?rgica. Esta pesquisa pretendeu estudar novas subst?ncias com a??o anticolinester?sica utilizando derivados -carbol?nicos do produto natural harmana, bem como fazer um estudo cin?tico a fim de descobrir qual o perfil de inibi??o das enzimas AChE e BChE, a fim de buscar novos compostos que poderiam ser ?teis no tratamento dos sintomas da DA. Desta forma, foi efetuada preliminarmente uma triagem com 7 derivados -carbol?nicos e posteriormente foi realizada uma investiga??o cin?tica com estes compostos, uma vez que, todos apresentaram alta a??o anticolinester?sica tanto para AChE quanto para BChE. A cin?tica enzim?tica foi estudada segundo o m?todo de Ellman. Contudo, observou-se que todos os 7 derivados apresentaram uma inibi??o revers?vel n?o competitiva.

La present Tesi Doctoral es presenta com una agrupació de quatre publicacions que resumeixen el treball realitzat al Departament de Genètica de la Facultat de Biologia de la Universitat de Barcelona. El principal objectiu és la caracterització funcional de les regions estructurals de la isoforma muscular de l’enzim deubiquitinat USP25, inicialment definides amb eines bioinformàtiques. A més a més, es pretén fer un estudi de noves interaccions moleculars tipus proteïna-proteïna per la cerca de nous substrats o reguladors enzimàtics. La cèl•lula eucariota posseeix, entre altres, un sistema senyalitzador intracel•lular basat en una família de pèptids, el representant dels quals és la ubiquitina. Aquest sistema presenta diferents categories funcionals, entre elles, els enzims deubiquitinants, un centenar a l’espècie humana. Aquests enzims hidrolitzen l’enllaç que uneix la ubiquitina als seus precursors o substrats, mantenint així l’homeostasi d’aquest pèptid dins la cèl•lula. Una alteració de la seva funció pot portar diferents conseqüències depenent de la via metabòlica que es vegi afectada, doncs suposa una desregulació estequiomètrica dels substrats ubiquitinants respecte els no ubiquitinats. L’enzim deubiquitinant USP25 es va descriure en el grup d’investigació d’aquesta tesi durant la cerca de nous gens relacionats amb la síndrome de Down. Un cop caracteritzat funcionalment com una proteasa específica d’ubiquitina, els estudis d’expressió van mostrar l’existència de tres isoformes proteiques, una d’elles, USP25m, restringida al teixit muscular i cardíac. Tenint en compte que en el fenotip dels pacients amb síndrome de Down, entre altres trets, hi ha deficiència cardiovascular i atonia muscular, els esforços del grup es van centrar en la descripció i anàlisi d’aquesta isoforma. Els primers estudis van mostrar la seva situació citosòlica, l’expressió correlativa amb la diferenciació de cèl•lules musculars i la relació específica amb diverses proteines del sarcòmer. En el treball realitzat per la present Tesi Doctoral, s’han caracteritzat funcionalment diferents regions reguladores descrites a nivell bioinformàtic, així com també s’ha analitzat la seva implicació fisiológica a la funció d’USP25m. A més a més, mitjançant un estudi de cerca de nous interactors proteics, s’ha trobat una nova molècula que pertany a la mateixa via senyalitzadora i que es relaciona de manera específica amb USP25m, la lligasa d’ubiquititna MKRN1 (makorin 1) Mitjançant l’ús de diferents construccions amb delecions i mutacions puntuals de la proteïna que afecten a les regions d’interès, s’ha arribat a diferents conclusions, entre elles, que USP25m és monoubiquitinat i té la capacitat d’autodeubiquitinar-se. La monoubiquitinació en regula la seva activitat enzimàtica i es proposa un mecanisme de regulació basat en la conjugació alternativa de SUMO (una altra molècula de la família de la ubiquitina) i ubiquitina, en el mateix residu aminoacídic, la lisina 99 (Lys99). Els dominis d’unió a ubiquitina regulen el reconeixement de substrat i afavoreixen la monoubiquitinació. USP25m oligomeritza dins la cèl•lula i es troba present en diferents formacions proteiques d’elevat pes molecular. S’ha comprovat la relació específica amb la nova lligasa MKRN1 i es suggereixen diferents escenaris moleculars on poden trobar-se inclosos els dos pèptids / The main aim of the present PhD work, titled “Structural domains and new protein interactions of the deubiquitinating enzyme USP25”, is the functional characterisation of the structural domains of the muscle isoform of USP25, USP25m, as well as the analysis of new protein‐protein interactions. The ubiquitin‐proteasome pathway is widely known as the preferential system to get ride of old or non‐functional proteins. Recently, it has become more apparent that this is not the only function. Ubiquitin (Ub) and all ubiquitin–like (UbLs) molecules acted as regulatory tags involved in different cellular events as subcellular localization, enzyme activation, DNA repair, etc. The intricate Ub‐signalling networks require a tight regulation of both conjugation and deconjugationprocesses, which are controlled by ubiquitin ligases and deubiquitinating enzymes (DUBs), respectively. USP25 is a DUB described while looking for novel genes involved in Down syndrome phenotype. First studies showed that it encoded three alternative protein isoforms, one of them, muscle specific. This muscle isoform, USP25m, is a cytosolic protein, upregulated during myogenesis that interacts in a specific manner with different sarcomeric proteins. Using an “in silico” approach, we were able to identify different structural domains, among them three ubiquitin binding domains (UBDs), and we aimed to characterise its role on USP25m function. By generating a collection of deletion and punctual mutants of the regions of interest, we conclude that USP25m is monoubiquitinated and that the UBDs modulate this modification. The preferential site for monoubiquitination is lysine 99 (K99), a residue that has been reported to undergo sumoylation (SUMO conjugation, being SUMO an UbL). According to our results, mutation of the K99 residue diminishes the deubiquitinating function, proposing a mechanistic model for USP25m regulation based on alternative conjugation of Ub and SUMO on the same residue, K99. Futhermore, while seeking new protein interactions of USP25m we identified Makorin Ring finger protein 1 (MKRN1), which belongs to an ubiquitin ligase family, as a putative interactor. We were capable of characterise its interaction and propose different cellular scenarios were they could interact. / The main aim of the present PhD work, titled “Structural domains and new protein interactions of the deubiquitinating enzyme USP25”, is the functional characterisation of the structural domains of the muscle isoform of USP25, USP25m, as well as the analysis of new protein‐protein interactions. The ubiquitin‐proteasome pathway is widely known as the preferential system to get ride of old or non‐functional proteins. Recently, it has become more apparent that this is not the only function. Ubiquitin (Ub) and all ubiquitin–like (UbLs) molecules acted as regulatory tags involved in different cellular events as subcellular localization, enzyme activation, DNA repair, etc. The intricate Ub‐signalling networks require a tight regulation of both conjugation and deconjugationprocesses, which are controlled by ubiquitin ligases and deubiquitinating enzymes (DUBs), respectively. USP25 is a DUB described while looking for novel genes involved in Down syndrome phenotype. First studies showed that it encoded three alternative protein isoforms, one of them, muscle specific. This muscle isoform, USP25m, is a cytosolic protein, upregulated during myogenesis that interacts in a specific manner with different sarcomeric proteins. Using an “in silico” approach, we were able to identify different structural domains, among them three ubiquitin binding domains (UBDs), and we aimed to characterise its role on USP25m function. By generating a collection of deletion and punctual mutants of the regions of interest, we conclude that USP25m is monoubiquitinated and that the UBDs modulate this modification. The preferential site for monoubiquitination is lysine 99 (K99), a residue that has been reported to undergo sumoylation (SUMO conjugation, being SUMO an UbL). According to our results, mutation of the K99 residue diminishes the deubiquitinating function, proposing a mechanistic model for USP25m regulation based on alternative conjugation of Ub and SUMO on the same residue, K99. Futhermore, while seeking new protein interactions of USP25m we identified Makorin Ring finger protein 1 (MKRN1), which belongs to an ubiquitin ligase family, as a putative interactor. We were capable of characterise its interaction and propose different cellular scenarios were they could interact.

Submitted by Automa??o e Estat?stica (sst@bczm.ufrn.br) on 2016-01-13T13:01:42Z No. of bitstreams: 1 PatriciaBatistaBarraMedeirosBarbosa_TESE.pdf: 3182319 bytes, checksum: 19636ea3271cce1b1b4e72c5e6dfa49c (MD5) / Approved for entry into archive by Elisangela Moura (lilaalves@gmail.com) on 2016-01-25T12:17:39Z (GMT) No. of bitstreams: 1 PatriciaBatistaBarraMedeirosBarbosa_TESE.pdf: 3182319 bytes, checksum: 19636ea3271cce1b1b4e72c5e6dfa49c (MD5) / Made available in DSpace on 2016-01-25T12:17:39Z (GMT). No. of bitstreams: 1 PatriciaBatistaBarraMedeirosBarbosa_TESE.pdf: 3182319 bytes, checksum: 19636ea3271cce1b1b4e72c5e6dfa49c (MD5) Previous issue date: 2014-06-09 / A dengue ? a mais importante arbovirose da atualidade, sendo transmitida pela picada do mosquito Aedes aegypti. A aus?ncia de uma vacina profil?tica torna o controle da dengue pautado principalmente no controle do inseto vetor. Todavia os crescentes relatos de resist?ncia e os danos ambientais causados pelos inseticidas qu?micos t?m tornado urgente ? busca por alternativas seguras. Foram preparados 21 extratos brutos (EB) de sementes de plantas da Caatinga. Como extrator foi utilizado o fosfato de s?dio 50mM pH 8.0. Os extratos foram utilizados em bioensaios Aedes aegypti, ensaios de toxicidade e submetidos a uma caracteriza??o parcial para identificar a presen?a de mol?culas bioativas. As an?lises inseticidas foram realizadas por probit e as diferen?as entre os grupos foram constatada por Student?s t-test e ANOVA. Todos os extratos apresentaram atividade larvicida para L1 e L4 e, com exce??o do extrato de G. americana, levaram ao ?bito 100% das larvas (48h). Os EB de M. urundeuva, P. viridiflora, E. velutina, A. cearenses e E. contortisiliquum apresentaram os melhores resultados larvicidas. Do mesmo modo, todos os EB foram repelentes para a postura das f?meas gr?vidas. Os EB de D. grandiflora, E. contortisiliquum, A. cearenses, C. ferrea e C. retusa foram capazes de atrair as f?meas para a posturas quando em baixas concentra??es. Na concentra??o atrativa o EB de E. contortisiliquum foi capaz de levar a ?bito 52% das larvas L1 em 48 h, enquanto que o EB de A. cearenses resultou na morte de 100% dessas larvas em 24h. Os extratos de A. cearenses, P. viridiflora, E. velutina, M. urundeuva e S. brasiliensis tiveram atividade pupicida, enquanto que os extratos de P. viridiflora, E. velutina, E. contortisiliquum, A. cearenses, A. colubrina, D. grandiflora, B. cheilantha, S. spectabilis, C. pyramidalis, M. regnelli e G. americana tiveram a??o adulticidas. Todos os extratos apresentaram toxicidade para C. dubia e os EB de E. velutina e E. contortisiliquum n?o interferiram na viabilidade dos fibroblastos. Considerando todos os ensaios biol?gicos os extratos de A. cearenses, P. viridiflora, E. contortisiliquum, S. brasiliensis, E. velutina e M. urundeuva foram considerados os mais promissores. Em todos os extratos foram identificadas pelo menos duas prote?nas com potencial. O EB de E. contortisiliquum foi selecionado para ser fracionando, j? que n?o apresentou atividade pupicida, indicando que seu mecanismo de a??o larvicida e adulticida resultou da ingest?o de compostos ativos pelos insetos. Al?m disso, esse extrato apresentou o maior teor de prote?na (22 mg/mL) e a maior atividade inibit?ria para as tripsinas das larvas de A. aegypti. Como observado para o EB, as fra??es proteicas de E. contortisiliquum, tamb?m apresentaram atividade larvicida, mas n?o foram pupicida, com destaque para F2 que apresentou maio atividade larvicida e menor toxicidade ambiental do que o EB de origem. A redu??o na atividade proteol?tica das larvas alimentadas com o EB e as fra??es de E. contortisiliquum sugeriu que inibidores de tripsina seriam os respons?vel pela atividade larvicida. Embora o aumento na purifica??o de um inibidor de tripsina de E. contortisiliquum (ITEc) tenha resultado na perda da atividade larvicida, foi constatado que sua aus?ncia reduziu a efic?cia das fra??es, indicando que o ITEc contribui, mas n?o ? suficiente para a atividade larvicida do EB de E. contortisiliquum. Tamb?m n?o foi verificada atividade larvicida e adulticida na fra??o rica em vicilina, e nem evid?ncias da contribui??o dessa mol?cula para a atividade inseticida desse extrato. Os resultados mostram o potencial dos extratos salinos de sementes de planta da Caatinga e, em especial do EB e da F2 de E. contortisiliquum, no controle da popula??o de A. aegypti e indicam que a efic?cia desses extratos deve resultar da a??o conjunta de diferentes compostos ativos, inclusive de natureza proteica, que atuando em diferentes mecanismos s?o capazes de comprometer a viabilidade dos insetos e retardar o surgimento de resist?ncias. / Dengue fever, currently the most important arbovirus, is transmitted by the bite of the Aedes aegypti mosquito. Given the absence of a prophylactic vaccine, the disease can only be controlled by combating the vector insect. However, increasing reports of resistance and environmental damage caused by insecticides have led to the urgent search for new safer alternatives. Twenty - um plant s eed extracts from the Caatinga were prepared , tested and characterized . Sodium phosphate ( 50 mM pH 8.0) was used as extractor. All extracts showed larvicidal and ovipositional deterrence activity . Extracts of D. grandiflora, E. contortisiliquum, A. cearenses , C. ferrea and C. retusa were able to attract females for posture when in low co ncentration . In the attractive concentrations, the CE of E. contortisiliquum and A. cearenses were able to kill 52% and 100% of the larvae respectively . The extracts of A. cearenses , P. viridiflora, E. velutina, M. urundeuva and S. brasiliensis were also pupicides, while extracts of P. viridiflora, E. velutina, E. contortisiliquum , A. cearenses, A. colubrina, D. grandiflora , B. cheilantha , S. spectabilis, C. pyramidalis, M. regnelli e G. americana displayed adulticidal activity. All extracts were toxic to C. dubia zooplankton . The EB of E. velutina and E. contortisiliquum did not affect the viability of fibroblasts . In all extracts were identified at least two potential insecticidal proteins such as enzyme inhibitors, lectins and chitin - binding proteins and components of secondary metabolism . Considering all bioassays , the extracts from A. cearenses, P. viridiflora, E. contortisiliquum , S. brasiliensis, E. velutina and M. urundeuva were considered the most promising . The E. contortisiliquum extracts was the only one who did not show pupicida activity, indicating that its mechanism of action larvicide and adulticidal is related only to the ingesti on of toxic compounds by insect , so it was selected to be fragmenting. As observed for the CE , th e protein fractions of E. contortisiliquum also showed larvicidal activity, highlighting that F2 showed higher larvicidal activity and lower en vironmental toxicity than the CE source. The reduction in the proteolytic activity of larvae fed with crude extra ct and fractions of E. contortisiliquum suggest ed that the trypsin inhibitors ( ITEc) would be resp onsible for larvicidal activity . However the increase in the purification of this inhibitor resulted in loss of larvicidal activity , but the absence of trypsin inhibitor reduced the effectiveness of the fractions , indicating that the ITEC contributes to the larvicidal activity of this extract. Not been observed larvicidal activity and adulticide in rich fraction vicilin, nor evidence of the contribution o f this molecule for the larvicidal activity of the extract. The results show the potential of seeds from plant extracts of Caatinga as a source of active molecules against insects A. aegypti at different stages of its development cycle, since they are comp osed of different active compounds, including protein nature, which act on different mechanisms should result in the death of insec

Submitted by Automa??o e Estat?stica (sst@bczm.ufrn.br) on 2017-10-18T20:27:51Z No. of bitstreams: 1 LucianiPaolaRochaCruz_TESE.pdf: 7850620 bytes, checksum: 76ac66a2a99a5841c825866204febb92 (MD5) / Approved for entry into archive by Arlan Eloi Leite Silva (eloihistoriador@yahoo.com.br) on 2017-10-23T22:34:51Z (GMT) No. of bitstreams: 1 LucianiPaolaRochaCruz_TESE.pdf: 7850620 bytes, checksum: 76ac66a2a99a5841c825866204febb92 (MD5) / Made available in DSpace on 2017-10-23T22:34:51Z (GMT). No. of bitstreams: 1 LucianiPaolaRochaCruz_TESE.pdf: 7850620 bytes, checksum: 76ac66a2a99a5841c825866204febb92 (MD5) Previous issue date: 2017-08-21 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior (CAPES) / Nanocristais de celulose s?o nanoestruturas derivados da celulose, que ? um recurso renov?vel e abundante na natureza. Por apresentarem uma combina??o de propriedades como alta resist?ncia mec?nica e m?dulo de elasticidade, superf?cie reativa e biodegradabilidade, esses materiais t?m recebido grande interesse para aplica??es que incluem desde refor?o em materiais polim?ricos, embalagens alimentares, a aplica??es na ?rea farmac?utica. A produ??o de celulose nanom?trica a partir de fibras de algod?o tem sido relatada em v?rios trabalhos publicados na literatura. O objetivo desta pesquisa foi estudar a obten??o de nanocelulose a partir do res?duo de tecido de fibra de algod?o tingido com corante ?ndigo, devido possuir alto conte?do de celulose, pelas vias de hidr?lise ?cida e enzim?tica. Nanocelulose foi obtida com e sem a realiza??o de pr?-tratamento para remo??o do corante e os efeitos do pr?-tratamento nas caracter?sticas da nanocelulose foram avaliados. Na hidr?lise ?cida, foram avaliadas duas condi??es de tratamento para isolamento de nanocristais de celulose: concentra??es de ?cido sulf?rico de 55% m/m a 60 C ou 65% m/m a 45 C, com tempos de 30 e 60 min. Na hidr?lise enzim?tica foram estudadas as influ?ncias do tipo de complexo enzim?tico (Trichoderma reesei ATCC 26921 ou Aspergillus fumigatus), o tempo (0 a 48h) e a carga enzim?tica (7,5 ou 12 FPU). As suspens?es obtidas ap?s hidr?lise foram caracterizadas pelas t?cnicas de potencial zeta, microscopia de for?a at?mica, microscopia eletr?nica de transmiss?o, espectrofotometria de infravermelho, difra??o de raios X, an?lise termogravim?trica, a??cares redutores totais e cromatografia l?quida de alto desempenho. Os resultados comprovaram a obten??o de nanocelulose a partir do tecido de algod?o tingido com corante ?ndigo, tanto no processo via hidr?lise ?cida, como no de via enzim?tica. As imagens de microscopia indicaram nanocristais de celulose com formato alongado (agulhas) a partir da hidr?lise ?cida. No caso da hidr?lise enzim?tica, as imagens mostraram a presen?a de nanocelulose com formato esf?rico. A hidr?lise ?cida realizada na condi??o de 65% a 45 C e tempo de 60 min resultou em nanocristais com menor comprimento e di?metro, tanto para o tecido pr?-tratado como para o sem pr?-tratamento. Com rela??o a hidr?lise enzim?tica, a realiza??o de pr?-tratamento n?o alterou significativamente as caracter?sticas das estruturas micro e nanocristalina. O tamanho m?dio das nanoceluloses obtidas foram na faixa de 80 a 230nm. Os resultados do processo de hidr?lise enzim?tica indicam que as melhores convers?es de celulose em glicose ocorreram utilizando o complexo enzim?tico produzido por Trichoderma reesei ATCC 26921 com carga de enzima de 12 FPU e tempo de hidr?lise de 48h. Este trabalho demonstrou que nanocristais de celulose podem ser obtidos a partir do tecido de algod?o tingido com corante ?ndigo, sem a necessidade de pr?-tratamento para remo??o do corante, e as caracter?sticas dos nanomateriais obtidos dependem do processo de hidr?lise utilizado. / Cellulose nanocrystals are nanomaterials derived from cellulose, which is a renewable and abundant resource in nature. Due to combination of properties such as high mechanical strength and modulus of elasticity, reactive surface and biodegradability, these materials have received great attention for applications ranging from reinforcement in polymeric materials, food packaging, to applications in the pharmaceutical area. The production of nanometric cellulose from cotton fibers has been reported in several works published in the literature. The objective of this research was to study the production of nanocellulose from indigo-dyed cotton fibers from waste fabric, via acid and enzymatic hydrolysis routes. Nanocellulose was obtained with and without pre-treatment for dye removal and the effects of the pre-treatment on the characteristics of the nanocellulose were evaluated. For the acid hydrolysis, two treatment conditions for the isolation of cellulose nanocrystals were evaluated: sulfuric acid concentrations of 55% m / m at 60 C and 65% m/m at 45 C, for 30 and 60 min. For the enzymatic hydrolysis, the influence of enzyme complex type (Trichoderma reesei ATCC 26921 and Aspergillus fumigatus), time (0 to 48h) and enzymatic load (7.5 and 12 FPU) were studied. The suspensions obtained after hydrolysis were characterized by the techniques of zeta potential, atomic force microscopy, transmission electron microscopy, infrared spectrophotometry, X-ray diffraction, thermogravimetric analysis, total reducing sugars and high performance liquid chromatography. The results demonstrated that nanocellulose was obtained from indigo dyed cotton fibers, in both processing routes evaluated: via acid and enzymatic hydrolysis. Microscopy images indicated needle shaped celulose from the acid hydrolysis. For the enzymatic hydrolysis, the images showed the presence of nanocellulose with spherical shape. The acid hydrolysis carried out at 65% at 45 C for 60 min resulted in nanocrystals of smaller length and diameter, both for the pretreated fabric and the fabric without pretreatment. For the enzymatic hydrolysis, the pretreatment did not affect significantly the characteristics of the micro and nanocrystalline structures. The average size of the nanocellulose obtained was in the range of 80 to 230 nm. The results of the enzymatic hydrolysis suggest that the best cellulose to glucose conversions occurred using the enzymatic complex Trichoderma reesei ATCC 26921 with enzymatic load of 12 FPU and hydrolysis time of 48h. In summary, this study demonstrated that cellulose nanocrytals can be obtained from indigo-dyed cotton fibers from waste fabric, without the need for pretreatment for dye removal, and the characteristics of the nanomaterials obtained depend on the hydrolysis process used.

Submitted by Ricardo Cedraz Duque Moliterno (ricardo.moliterno@uefs.br) on 2015-10-01T23:29:45Z No. of bitstreams: 1 Aspergillus terreus isolamento identifica??o e avalia??o da capacidade catalitica na redu??o de cetonas pr? quirais.pdf: 1820055 bytes, checksum: c5c60d2c4b1c577c84c8b1baaa95660d (MD5) / Made available in DSpace on 2015-10-01T23:29:45Z (GMT). No. of bitstreams: 1 Aspergillus terreus isolamento identifica??o e avalia??o da capacidade catalitica na redu??o de cetonas pr? quirais.pdf: 1820055 bytes, checksum: c5c60d2c4b1c577c84c8b1baaa95660d (MD5) Previous issue date: 2014-09-30 / The object of this inquiry is the isolation, the selection and the potential evaluation of a microorganism with biocatalytic activity in carrying out selective-stereo biorreduction of pro-chial ketones. In the research procedure, Fungi was isolated from soil samples contaminated with lead. Only one of the isolates fungi, identified as Aspergillus terreus, presented good catalytic activity in the reduction of acetophenone. The biomass was grown submerged in broth culture, inoculated with 1,1 x 103 (spores/mL of malt extract) and maintained at 30?C under orbital agitation. Morphological differences and catalytic activity were evaluated in function of the orbit of agitation during the growth process. The catalytic activity in terms of conversion of acetophenone and enantiomeric excess (S), ee_S, in 1-feniletanol, was determinate in conjunction with the growth curve of the fungus. The results were used to construct a graphic showing the variation of quality of the catalyst, according to age of biomass. Apart from acetophenone were tested o-Xacetophenone, p-Xacetophenone and m-Xacetophenone (X = methyl, methoxy, nitro, fluorine, bromine). For reaction with 24 hours growing cells (30?C; 150rpm/r = 50mm), the conversions were between 27% and 97% with ee_S between 41% and 83%. For reactions with cells in suspension in buffered medium (pH = 4.5 ? 6,5) was decrease in activity/selectivity at pH above 5.5. The kinetics of conversion was evaluated for biomasses with different ages and for various concentrations of acetophenone. An amount between 100mg and 200mg of biomass (dry mass) with 168 hours of culture can convert 100mg of acetophenone to 1(S)-feniletanou ee_S (91%) with 98% yield in 72 hours of reaction (93% at 48 hours). The development of this process described above, constitutes the body of this thesis. / O objetivo desta pesquisa ? o isolamento, a sele??o e a avalia??o de um microorganismo com atividade biocatalisadora na biorredu??o estereoseletiva de cetonas pro-quirais. No procedimento de investiga??o, fungos foram isolados de amostras de solo contaminadas com chumbo. Apenas um dos fungos isolados, identificado geneticamente como Aspergillus terreus, apresentou boa atividade catal?tica na redu??o de acetofenona. A biomassa foi cultivada submersa no caldo de cultura, inoculado com 1,1x103 (esporos/mL de extrato de malte) e mantido a 30? sob agita??o orbital. Diferen?as morfol?gicas e de atividade catal?tica foram avaliadas em fun??o da ?rbita de agita??o durante o crescimento. A atividade catal?tica, em termos de convers?o da acetofenona e do excesso enantiom?rico (S), ee_S, no 1-feniletanol, foi determinada em conjunto com a curva de crescimento do fungo. Os resultados foram usados para construir um gr?fico mostrando a varia??o da qualidade do catalisador em fun??o da idade da biomassa. Al?m de acetofenona, foram testadas o-Xacetofenona, p-Xacetofenona e m-Xacetofenona (X=metil, metoxi, nitro, fl?or, bromo). Para 24 horas de rea??o com c?lulas em crescimento (30?C; 150rpm/r = 50mm), as convers?es ficaram entre 94 e 100%, com ee_S ? 98% para acetofenona e para acetofenonas orta ou meta substitu?das. Com substituintes na posi??o para, a convers?o ficou entre 27% e 97%, com ee_S entre 41% e 83%. Para rea??es com c?lulas em suspens?o em meio tamponado (pH = 4,5 ? 6,5), houve queda na atividade/seletividade em pH acima de 5,5. A cin?tica de convers?o foi avaliada para biomassas com idades distintas e para v?rias concentra??es de acetofenona. Uma quantidade entre 100mg e 200mg de biomassa (massa seca) com 168 horas de cultura, pode converter 100mg de acetofenona a 1(S)-feniletanol (ee_S 91%), com 98% de rendimento em 72 horas de rea??o (93% em 48 horas). O desenvolvimento desse processo, acima descrito, constitui o corpo desta Tese.

Made available in DSpace on 2014-12-17T14:07:23Z (GMT). No. of bitstreams: 1 MauricioRB.pdf: 2052566 bytes, checksum: ecabe62a65c1b9db9e09054f3321c5b1 (MD5) Previous issue date: 2007-03-12 / Sugar esters are substances which possess surfactant, antifungical and bactericidal actions and can be obtained through two renewable sources of raw materials: sugars and vegetable oils. Their excellent biodegradability, allied to lhe fact that they are non toxic, insipid, inodorous, biocompatible, no-ionic, digestible and because they can resist to adverse conditions of temperature, pH and salinity, explain lhe crescent use of these substances in several sections of lhe industry. The objective of this thesis was to synthesize and characterize surfactants and polymers containing sugar branched in their structures, through enzymatic transesterification of vinyl esters and sugars, using alkaline protease from Bacillus subtilis as catalyst, in organic medium (DMF).Three types of sugars were used: L-arabinose, D-glucose and sucrose and two types of vinyl esters: vinyl laurate and vinyl adipate. Aiming to reach high conversions from substrates to products for a possible future large scale industrial production, a serie of variables was optimized, through Design of Experiments (DOE), using Response Surface Methodology (RSM).The investigated variables were: (1) enzyme concentration; (2) molar reason of substrates; (3) water/solvent rale; (4) temperature and (5) time. We obtained six distinct sugar esters: 5-0-lauroyl L-arabinose, 6-0-lauroyl D-glucose, 1'-O-lauroyl sucrose, 5-0-vinyladipoyl L-arabinose, 6-0-vinyladipoyl D-glucose and 1 '-O-vinyladipoyl sucrose, being lhe last three polymerizable. The progress of lhe reaction was monitored by HPLC analysis, through lhe decrease of sugar concentration in comparison to lhe blank. Qualitative analysis by TLC confirmed lhe formation of lhe products. In lhe purification step, two methodologies were adopted: (1) chromatographic column and (2) extraction with hot acetone. The acylation position and lhe chemical structure were determined by 13C-RMN. The polymerization of lhe three vinyl sugar esters was possible, through chemical catalysis, using H2O2 and K2S2O8 as initiators, at 60?C, for 24 hours. IR spectra of lhe monomers and respective polymers were compared revealing lhe disappearance of lhe vinyl group in lhe polymer spectra. The molar weights of lhe polymers were determined by GPC and presented lhe following results: poly (5-0-vinyladipoyl L-arabinose): Mw = 7.2 X 104; PD = 2.48; poly (6-0-vinyladipoyl D-glucose): Mw = 2.7 X 103; PD = 1.75 and poly (1'-O-vinyladipoyl sucrose): Mw = 4.2 X 104; PD = 6.57. The six sugar esters were submitted to superficial tension tests for determination of the critical micelle concentrations (CMC), which varied from 122 to 167 ppm. Finally, a study of applicability of these sugar esters, as lubricants for completion fluids of petroleum wells was' accomplished through comparative analysis of lhe efficiency of these sugar esters, in relation to three commercial lubricants. The products synthesized in this thesis presented equivalent or superior action to lhe tested commercial products / ?steres de a??car s?o compostos que possuem a??o surfactante, antif?ngica e bactericida e podem ser obtidos a partir de duas fontes renov?veis de mat?ria-prima: a??cares e ?leos vegetais. Sua capacidade de se biodegradar, aliada ao fato de serem at?xicos, ins?pidos, inodoros, biocompat?veis, n?o-i?nicos, digest?veis e resistirem a condi??es severas de temperatura, pH e salinidade, explicam o crescente emprego destas subst?ncias em diversos setores da ind?stria. O objetivo desta tese foi sintetizar e caracterizar surfactantes e pol?meros, contendo a??cares ramificados em suas estruturas, atrav?s de transesterifica??o enzim?tica de ?steres vin?licos com a??cares, empregando-se protease alcalina de Bac?llus subtilis como catalisador, em meio org?nico (DMF). Foram empregados tr?s tipos de a??cares: L-arabinose, D-glicose e sacarose e dois tipos de ?steres vin?licos: laurato de vinila e adipato de vinila. Para a obten??o de altas convers?es de substratos em produtos, visando uma futura produ??o em larga escala, uma s?rie de vari?veis foram otimizadas, atrav?s de an?lise estat?stica experimental (DOE), por metodologia de resposta de superf?cie (RSM). As vari?veis investigadas foram: (1) a concentra??o de enzima; (2) a raz?o molar entre substratos; (3) a raz?o ?gua/solvente org?nico; (4) a temperatura e (5) o tempo. Foram obtidos seis ?steres de a??car: 5-0-lauroil L-arabinose, 6-0lauroil D-glicose, 1 '-O-Iauroil sacarose, 5-0-viniladipoil L-arabinose, 6-0viniladipoil D-glicose e 1 '-O-viniladipoil sacarose, sendo os tr?s ?ltimos polimeriz?veis. O progresso da rea??o foi monitorado por an?lise em HPLC, atrav?s do decr?scimo da concentra??o de a??car em rela??o ao branco. An?lises qualitativas, por TLC, confirmaram a forma??o dos produtos. Foram obtidas convers?es superiores a 98% na s?ntese do laurato de sacarose. Na purifica??o, foram adotadas duas metodologias: (1) coluna cromatogr?fica e (2) extra??o com acetona a quente. A posi??o de acila??o e a estrutura qu?mica foram determinadas por 13C-RMN. A polimeriza??o dos tr?s ?steres de a??car foi poss?vel, atrav?s de cat?lise qu?mica, empregando-se H2O2 e K2S2O8 como iniciadores, a 60?C, por 24 horas. Espectros de IR dos pol?meros foram comparados com os seus mon?meros, revelando o desaparecimento do grupo vinil. As massas molares dos pol?meros foram determinadas por GPC. Os pol?meros de a??car obtidos apresentaram as seguintes massas molares: poli (5-0-viniladipoil L-arabinose): Mw = 7,2 X 104; PD = 2,48; poli (6-0-viniladipoil D-glicose): Mw = 2,7 x 103; PD = 1,75 e poli (1 '-O-viniladipoil sacarose): Mw = 4,2 X 104; PD = 6,57. Os seis ?steres de a??car foram submetidos a ensaios de tens?o superficial para a determina??o das concentra??es micelares cr?ticas (CMC), que variaram de 122 a 167 ppm. Por fim, um estudo de aplicabilidade dos ?steres n?o polimeriz?veis, como lubrificantes para fluidos de completa??o de po?os de petr?leo foi realizado, atrav?s de an?lise comparativa da efici?ncia destes, em rela??o a tr?s lubrificantes comerciais. Os produtos sintetizados nesta tese apresentaram desempenho equivalente ou superior aos produtos comerciais testados

Disponibiliza??o do trabalho em conte?do parcial, conforme Termo de Autoriza??o. / Submitted by Jos? Henrique Henrique (jose.neves@ufvjm.edu.br) on 2017-03-27T19:34:05Z No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) fernanda_souza_paula_parcial.pdf: 515579 bytes, checksum: f72e2289acf5068c063e2bd00f0fefc3 (MD5) / Approved for entry into archive by Rodrigo Martins Cruz (rodrigo.cruz@ufvjm.edu.br) on 2017-04-24T16:35:34Z (GMT) No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) fernanda_souza_paula_parcial.pdf: 515579 bytes, checksum: f72e2289acf5068c063e2bd00f0fefc3 (MD5) / Made available in DSpace on 2017-04-24T16:35:34Z (GMT). No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) fernanda_souza_paula_parcial.pdf: 515579 bytes, checksum: f72e2289acf5068c063e2bd00f0fefc3 (MD5) Previous issue date: 2017 / O trabalho investiga a utiliza??o de plataformas eletroqu?micas contendo filmes polim?ricos derivados do ?cido 4-aminosalicico (4-AMS) para imobiliza??o da enzima Urato oxidase (UOx) visando aplica??o na quantifica??o de ?cido ?rico (AU) em amostras de urina. Investigou-se a eletrodeposi??o do 4-AMS pelas t?cnicas de voltametria c?clica (VC) e cronoamperometria (CA) sobre eletrodos de grafite (EG). Por VC foram realizados 100 ciclos de potencial na faixa de -0,25 a 1,25 V ? 50 mV/s em solu??o 2,50 mM do mon?mero preparado em H2SO4 0,50 M. Utilizando a CA, a eletropolimeriza??o foi realizada a potencial constante de +0,928 V durante 5600s no mesmo meio reacional utilizado na VC. O poli(4-AMS) obtido por VC e CA mostrou dois pares redox, os quais est?o relacionados a eletroatividade do filme polim?rico, na regi?o de potencial de +0,50/+0,40 V. Contudo, maiores valores de Ipa e Ipc foram obtidos para os eletrodos modificados por VC, sugerindo que estes filmes s?o mais eletroativos. A deposi??o do azul da Pr?ssia (AP), mediador da rea??o de per?xido, foi investigada sobre os EG, com posterior modifica??o com poli(4-AMS). Notou-se que a presen?a do AP n?o altera o perfil voltam?trico da eletropolimeriza??o do 4-AMS. Contudo, quando comparada com a eletropolimeriza??o somente nos EG, obteve-se filmes mais resistivos e com menor eletroatividade. Analisando as propriedades eletroqu?micas e morfol?gicas, por VC conseguiuse filmes mais uniformes, com maior quantidade de material depositado e maior eletroatividade. A eletropolimeriza??o foi realizada tamb?m sobre eletrodos impressos de grafite contendo azul da Pr?ssia (EI/AP), onde posteriormente imobilizou-se 5 U da UOx, e o biossensor foi acoplado a uma c?lula de fluxo num sistema de an?lise por inje??o em fluxo (FIA) de linha ?nica. A vaz?o e o volume da al?a de amostragem foram otimizados em 2,10 mL/min e 200 ?L, repectivamente. O valor de pH da solu??o do analito foi otimizado em 8,27. Medidas de reprodutibilidade mostraram desvio padr?o de 2,15% (n=10). O biossensor respondeu linearmente para AU na faixa de 1,0 x 10-5 a 2,0 x 10-4 M, com limite de detec??o de 3,0 ?M. Amostras de urina foram dilu?das (1:10) e injetadas diretamente no biossensor. A reposta foi reprodut?vel mostrando baixo desvio padr?o para as medidas, e valores encontrados dentro da faixa esperada para o analito em amostras de urina. Testes de adi??o e recupera??o mostraram valores de 97,35% (?2,43). O biossensor mostrou-se bastante promissor para a proposta do trabalho, apresentando resultados muito satisfat?rios para as an?lises e par?metros investigados. / Disserta??o (Mestrado) ? Programa de P?s-Gradua??o em Qu?mica, Universidade Federal dos Vales do Jequitinhonha e Mucuri, 2017. / This work investigates the use of electrochemical platforms containing polymeric films derived from 4-aminosalicylic acid (4-ASA) modified with the enzyme Urate oxidase (UOx) for quantification of uric acid (UA) in urine samples. The electrodeposition of 4-ASA was investigated through Cyclic Voltammetry (CV) and Chronoamperometry (CA) on graphite electrodes (GE). 100 cycles were performed in the range of -0.25 to 1.25 V at 50 mV/s in 2.50 mM monomer solution prepared in 0.50 M H2SO4. Using CA, the deposition was performed at a potential of +0.928 V for 5600 s in the same CV reaction medium. The poly(4-ASA) showed two redox pairs related to the electroactivity of the polymeric film in the potential range of + 0.50 /+ 0.40 V. However, higher values of Ipa and Ipc were obtained for the electrodes modified through CV, suggesting that these films are more electroactive. The deposition of Prussian blue (PB), mediator of the peroxide reaction, was investigated on the GE with subsequent modification with poly (4-ASA). It was observed that the presence of PB does not alter the voltammetric profile of 4-ASA electropolymerization. However, when compared with the electropolymerization in bare GE, more resistive films were obtained with lower electroactivity. Analyzing the electrochemical and morphological properties through CV, more uniform films were obtained, with more material deposited and greater electroactivity. The electropolymerization of poly(4-ASA) was also conducted on screen printed electrodes containing Prussian Blue (SPE/PB), with subsequent immobilization of 5U of Uox. This biosensor was coupled to a flow cell in a Flow Injection Analysis (FIA) system of single line. The flow rate and the sampling loop volume were optimized at 2.10 mL/min and 200 ?L, respectively. The pH value of the analyte solution was optimized at 8.27. Reproducibility measures showed a standard deviation of 2.15% (n = 10). The biosensor responded linearly to UA in the range of 1.0 x 10-5 to 2.0 x 10-4 M, with a detection limit of 3.0 ?M. Urine samples were diluted (1:10) and directly injected over the biosensor. The response was reproducible with low standard deviation and values found within the range expected for the analyte in urine samples. Addition and recovery tests showed values of 97.35% (?2.43). The biosensor is very promising for the work proposal, presenting very satisfactory results for the analyzes and investigated parameters.

Made available in DSpace on 2014-12-17T15:01:12Z (GMT). No. of bitstreams: 1 LeandroFD.pdf: 1531484 bytes, checksum: 8c328385e5fe3dc7294b86f55a82c108 (MD5) Previous issue date: 2007-06-15 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior / The cashew, a fruit from Brazilian Northeast is used to produce juice due to its flavor and vitamin C richness. However, its acceptance is limited due to its astringency. Caju?na is a derivate product appreciated by its characteristic flavor, freshness and lack of astringency, due to tannin removal. Caju?na is a light yellow beverage made from clarified cashew juice and sterilized after bottling. It differs from the integral and concentrated juice by the clarification and thermal treatment steps. Many problems such as haze and excessive browning could appear if these steps are not controlled. The objective of this work was divided into two stages with the aim to supply process information in order to obtain a good quality product with uniform characteristics (sensory and nutritional). Polyphenol-protein interaction was studied at the clarification step, which is an empirical process, to provide values on the amount of clarifying solution (gelatin) that must be added to achieve a complete juice clarification. Clarification essays were performed with juice dilutions of 1:2 and 1:10 and the effect of metabissulfite and tannic acid addition was evaluated. It was not possible to establish a clarification point. Metabissulfite did not influenced the clarification process however tannic acid addition displaced the clarification point, showing the difficulty visual monitoring of the process. Thermal treatment of clarified juice was studied at 88, 100, 111 e 121 ?C. To evaluate the non-enzymatic browning, vitamin C, 5-hidroximetilfurfural (5-HMF) and sugar variation were correlated with color parameters (reflectance spectra, color difference and CIELAB). Kinetic models were obtained for reflectance spectra, ascorbic acid and 5-HMF. It was observed that 5-HMF introduction followed a first order kinetic rate at the beginning of the thermal treatment and a zero order kinetic at later process stages. An inverse correlation was observed between absorbance at 420 nm and ascorbic acid degradation, which indicates that ascorbic acid might be the principal factor on caju?na non-enzymatic browning. Constant sugar concentration showed that this parameter did not contribute directly to the nonenzymatic browning. Optimization techniques showed showed that to obtain a high vitamin C and a low 5-HMF content, the process must be done at 120 ?C. With the water-bath thermal treatment, the 90 ?C temperature promoted a lower ascorbic acid degradation at the expense of a higher 5-HMF level / O caju, frut?fera origin?ria do Nordeste brasileiro ? utilizado para produ??o de suco devido ao seu sabor e ser rico em vitamina C. Entretanto, sua aceita??o ? limitada devido a sua adstring?ncia. Um produto bastante apreciado pelo seu sabor caracter?stico, refrescante e n?o adstringente, devido ? remo??o dos taninos, ? a caju?na. A caju?na ? uma bebida elaborada a partir do suco de caju clarificado e esterilizada no interior de garrafas, apresentando uma colora??o amarelo-?mbar. Ela diferencia-se dos sucos de caju integral e concentrado por meio das etapas de clarifica??o e tratamento t?rmico. Uma s?rie de problemas como turva??o e escurecimento excessivo do produto, pode aparecer se estas etapas n?o forem bem controladas. O objetivo deste trabalho foi dividido em duas etapas, mas com o mesmo prop?sito de fornecer subs?dios que possibilitem o controle do processo de maneira a se obter um produto de qualidade, com caracter?sticas mais uniformes (em termos sensoriais e nutricionais). Na clarifica??o, por se tratar de um processo emp?rico, buscou-se o entendimento das intera??es polifenol-prote?na para fornecer valores adequados de solu??o de clarificante (gelatina) necess?ria para promover a clarifica??o completa do suco. Realizaramse ensaios de clarifica??o com suco de caju dilu?do nas propor??es 1:2 e 1:10 e avaliou-se a influ?ncia da adi??o de metabissulfito e ?cido t?nico ao suco. Pelas t?cnicas utilizadas n?o foi poss?vel determinar um ponto de clarifica??o completa do suco de caju. O metabissulfito n?o influenciou o processo de clarifica??o enquanto que adi??o de ?cido t?nico deslocou o ponto de clarifica??o, mostrando a dificuldade da observa??o deste ponto pelo operador. O efeito do tratamento t?rmico no suco clarificado foi estudado nas temperaturas de 88, 100, 111 e 121 ?C. Para avaliar o escurecimento n?o-enzim?tico, a varia??o de vitamina C, 5- hidroximetilfurfural (5-HMF) e a??cares foram correlacionados com par?metros colorim?tricos (espectro de reflect?ncia, diferen?a de cor e CIELAB). Modelos cin?ticos foram obtidos para a mudan?a no espectro de reflect?ncia, ?cido asc?rbico e 5-HMF. A forma??o de 5-HMF seguiu dois mecanismos cin?ticos: taxa cin?tica de primeira ordem no come?o do tratamento t?rmico e taxa cin?tica de ordem zero em um per?odo mais avan?ado do processo. Observou-se correla??o inversa da absorb?ncia a 420 nm e a perda de ?cido asc?rbico, o que indica que o ?cido asc?rbico pode ser o fator principal que causa o escurecimento da caju?na. A concentra??o constante dos a??cares mostrou que eles n?o influenciaram diretamente o escurecimento n?o-enzim?tico. T?cnicas de otimiza??o mostraram que para se obter uma caju?na com concentra??o elevada de vitamina C e baixo teor de 5-HMF, o tratamento t?rmico deve ser realizado a 120 ?C. No caso da utiliza??o de banho-maria para o tratamento t?rmico, a temperatura de 90 ?C promove uma menor degrada??o de ?cido asc?rbico ? custa de um ?ndice de 5-HMF mais elevado

Made available in DSpace on 2014-12-17T15:01:24Z (GMT). No. of bitstreams: 1 JulianaCSA_DISSERT.pdf: 2885300 bytes, checksum: 18768d46af6895ea630731a4c28b4d2d (MD5) Previous issue date: 2010-12-17 / Conselho Nacional de Desenvolvimento Cient?fico e Tecnol?gico / The food industry is interested in natural products. Anthocyanins are phenolic antioxidants of great importance with health-relevant applications. Several studies have linked the intake of fruits and vegetables with reduced risk of chronic degenerative diseases because of its antioxidant properties. This study aimed to compare different strategies for obtaining natural pigments from red jambo (Syzygium malaccence) and analyze its functional potential. Two different strategies were studied: (1) solid-liquid extraction (SLE) in reactor with controlled parameters, (2) powder obtention. The investigation of the functional potential was conducted taking into account the total phenolic content (TPC), the antioxidant activity (AA), the total anthocyanins concentration (TA) and α-amylase and α-glucosidase inhibition. The best extracts obtained by SLE showed TPC of 174.15 mg GAE/100g, AA of 3.56 μmol Trolox eq/g and TA of 133.59 mg cyd-3-glu/100 g. The best results for the second strategy were TPC of 1024.22 mg GAE/100 g, AA of 29.03 μmol Trolox eq/g and TA of 1193.41 mg cyd-3- glu/100 g. It was observed moderate amylase inhibition (26.30%) and high glucosidase inhibitory activity (97.47%). Skin extracts showed, in general, superior results when compared to whole red jambo, with superior values for dehydrated products. Based on our result, red jambo can be considered as a rich source of phenolic antioxidants, as well on amylase and glucosidase inhibitors / A ind?stria aliment?cia demonstra forte interesse em estudos de extra??o envolvendo produtos naturais. A antocianina ? um fen?lico antioxidante de grande import?ncia e atua??o no organismo dos seres vivos. V?rios estudos relacionam ? ingest?o de frutas e vegetais com a diminui??o do risco e desenvolvimento de doen?as cr?nicodegenerativas em fun??o de suas propriedades antioxidantes. Este trabalho teve como objetivo comparar diferentes estrat?gias de obten??o do corante da casca e do jambo inteiro sem caro?o e analisar seu potencial funcional. Duas diferentes estrat?gias foram estudadas: (1) extra??o s?lido-l?quido em reator enjaquetado com controle de par?metros; (2) obten??o do p?. A investiga??o do potencial funcional foi realizada por meio de an?lises quanto ao teor de compostos fen?licos totais (CFT), a atividade antioxidante (AA), a concentra??o de antocianinas totais (AT) e a inibi??o das enzimas α-amilase e α-glicosidase. Os extratos com os melhores resultados para a estrat?gia 1 foram para CFT de 174,15 mg GAE/100 g, para a AA de 3,56 μmol Trolox eq/g e para AT de 133,59 mg ci-3-gli/100 g. Os melhores valores para a estrat?gia 2 foram para CFT de 1024,22 mg GAE/100 g, para AA de 29,03 μmol Trolox eq/g e para AT de 1193,41 mg ci-3-gli/100 g. A a??o inibit?ria das enzimas α-amilase (26,30%) e α-glicosidase (97,47%) mostraram-se potentes. Os extratos da casca apresentaram, de maneira geral, resultados superiores quando comparados aos valores dos extratos do jambo inteiro e as maiores quantifica??es foram obtidas dos produtos desidratados. As amostras analisadas exibiram fontes satisfat?rias de fen?licos antoci?nicos, com potente capacidade antioxidante e atividade inibit?ria das enzimas α-amilase e α-glicosidase

Made available in DSpace on 2014-12-17T15:42:15Z (GMT). No. of bitstreams: 1 FabioPF_TESE.pdf: 3376603 bytes, checksum: 15dfaa7fe12ca918fd7e1b98c4378dd9 (MD5) Previous issue date: 2011-09-16 / Conselho Nacional de Desenvolvimento Cient?fico e Tecnol?gico / Enzymatic synthesis of peptides using proteases has attracted a great deal of attention in recent years. One key challenge in peptide synthesis is to find supports for protease immobilization capable of working in aqueous medium at high performance, producing watersoluble oligopeptides. At present, few reports have been described using this strategy. Therefore, the aim of this thesis was to immobilize proteases applying different methods (Immobilization by covalent bound, entrapment onto polymeric gels of PVA and immobilization on glycidil metacrylate magnetic nanoparticles) in order to produce water-soluble oligopeptides derived from lysine. Three different proteases were used: trypsin, α-chymotrypsin and bromelain. According to immobilization strategies associated to the type of protease employed, trypsin-resin systems showed the best performance in terms of hydrolytic activity and oligopeptides synthesis. Hydrolytic activities of the free and immobilized enzymes were determined spectrophotometrically based on the absorbance change at 660 nm at 25 ?C (Casein method). Calculations of oligolysine yield and average degree of polymerization (DPavg) were monitored by 1H-NMR analysis. Trypsin was covalently immobilized onto four different resins (Amberzyme, Eupergit C, Eupergit CM and Grace 192). Maximum yield of bound protein was 92 mg/g, 82 mg/g and 60 mg/g support for each resin respectively. The effectiveness of these systems (Trypsin-resins) was evaluated by hydrolysis of casein and synthesis of water-soluble oligolysine. Most systems were capable of catalyzing oligopeptide synthesis in aqueous medium, albeit at different efficiencies, namely: 40, 37 and 35% for Amberzyme, Eupergit C and Eupergit CM, respectively, in comparison with free enzyme. These systems produced oligomers in only 1 hour with DPavg higher than free enzyme. Among these systems, the Eupergit C-Trypsin system showed greater efficiency than others in terms of hydrolytic activity and thermal stability. However, this did not occur for oligolysine synthesis. Trypsin-Amberzyme proved to be more successful in oligopeptide synthesis, and exhibited excellent reusability, since it retained 90% of its initial hydrolytic and synthetic activity after 7 reuses. Trypsin hydrophobic interactions with Amberzyme support are responsible for protecting against strong enzyme conformational changes in the medium. In addition, the high concentration of oxirane groups on the surface promoted multi-covalent linking and, consequently, prevented the immobilized enzyme from leaching. The aforementioned results suggest that immobilized Trypsin on the supports evaluated can be efficiently used for oligopeptides synthesis in aqueous media / S?ntese enzim?tica de pept?deos usando proteases tem atra?do uma enorme aten??o nos ?ltimos anos. Um desafio chave na s?ntese de pept?deos ? encontrar suportes para imobiliza??o de proteases capazes de apresentar um alto desempenho em meio aquoso, produzindo oligopept?deos sol?veis em ?gua, j? que at? o presente momento, pouco tem sido descrito usando essa estrat?gia. Dessa forma, o objetivo dessa tese foi imobilizar proteases usando diferentes m?todos (imobiliza??o por liga??o covalente, aprisionamento em g?is polim?ricos de PVA e imobiliza??o em nanopart?culas magn?ticas de Glicidil) para a produ??o de oligopept?deos derivados da lisina. Tr?s proteases foram utilizadas: tripsina, α-quimotripsina e bromela?na. De acordo com as estrat?gias de imobiliza??o associadas ao tipo de protease empregada, foi provado que os sistemas tripsina-resinas mostraram os melhores desempenhos em termos de atividade hidrol?tica e s?ntese de oligopept?deos. A atividade hidrol?tica das enzimas livres e imobilizadas foi determinada por espectrofotometria com base na mudan?a de absorb?ncia em 660 nm ? temperatura de 25 ?C (Casein method). O rendimento de oligolisina e o c?lculo do grau de polimeriza??o m?dio foram monitorados por RMN H. A protease tripsina foi covalentemente imobilizada em quatro diferentes resinas (Amberzyme, Eupergit C, Eupergit CM and Grace 192). O m?ximo rendimento de prote?na imobilizada foi 92, 82, 60, e 71 mg/g de suporte para cada resina, respectivamente. A efici?ncia desses sistemas (Tripsina-resinas) foi avaliada pela hidr?lise do substrato case?na e a s?ntese de oligolisina em meio aquoso. A maioria dos sistemas foram capazes de catalisar a s?ntese de oligopept?deos, entretanto com diferentes efici?ncias, tais como: 40, 37 e 35% para os suportes Amberzyme, Eupergit C e Eupergit CM, respectivamente, em compara??o com a enzima livre. Esses sistemas produziram olig?meros em somente 1 hora com grau de polimeriza??o m?dio mais alto que a enzima livre. Dentre esses sistemas, Eupergit CTripsina mostrou ser mais eficiente que os outros sistemas em termos de atividade hidrol?tica e estabilidade t?rmica, ao passo que n?o exibiu a mesma efici?ncia como era esperado para a s?ntese de oligolisina. Tripsina-amberzyme provou ser mais eficiente para a s?ntese de oligopept?deos, al?m de exibir um excelente reuso, mantendo 90% de sua atividade hidrol?tica e sint?tica ap?s sete reusos. As intera??es hidrof?bicas da tripsina com o suporte Amberzyme s?o respons?veis por proteger a enzima contra as fortes mudan?as conformacionais no meio reacional. Al?m disso, a alta concentra??o de grupos oxiranos na superf?cie da resina promoveu liga??es covalentes multipontuais e, consequentemente, preveniu a enzima imobilizada do processo de desor??o (Leaching process). Os resultados acima mencionados sugerem que a tripsina imobilizada nesses suportes pode ser eficientemente usada para a s?ntese de oligopept?deos em meio aquoso