Submitted by Automa??o e Estat?stica (sst@bczm.ufrn.br) on 2017-02-13T19:44:49Z No. of bitstreams: 1 GrazieneLopesDeSouza_DISSERT.pdf: 1409638 bytes, checksum: b2258a2f626dcdfadfc00445de366ca7 (MD5) / Approved for entry into archive by Arlan Eloi Leite Silva (eloihistoriador@yahoo.com.br) on 2017-02-14T21:07:25Z (GMT) No. of bitstreams: 1 GrazieneLopesDeSouza_DISSERT.pdf: 1409638 bytes, checksum: b2258a2f626dcdfadfc00445de366ca7 (MD5) / Made available in DSpace on 2017-02-14T21:07:26Z (GMT). No. of bitstreams: 1 GrazieneLopesDeSouza_DISSERT.pdf: 1409638 bytes, checksum: b2258a2f626dcdfadfc00445de366ca7 (MD5) Previous issue date: 2015-06-26 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior (CAPES) / Anacardium occidentale Linn ? uma ?rvore nativa do Brasil, popularmente conhecida como cajueiro. Estudos mostram que a fra??o Acetato de Etila (AcOEt) obtida a partir do extrato acetona das cascas do caule de A. occidentale apresenta atividade anti-inflamat?ria, sugerindo-se os taninos como respons?veis por essa a??o. Sendo uma das 71 esp?cies da Rela??o Nacional de Plantas Medicinais de Interesse ao Sistema ?nico de Sa?de (RENISUS), ? importante que sejam desenvolvidas metodologias anal?ticas para o estudo dos seus constituintes. Nesse contexto, a Cromatografia L?quida de Alta Efici?ncia (CLAE) constitui uma metodologia de import?ncia para a an?lise de amostras de origem vegetal. Este estudo teve como objetivo desenvolver e validar uma metodologia anal?tica por CLAE para a an?lise da fra??o AcOEt do extrato acetona das cascas do caule de A. occidentale. As condi??es cromatogr?ficas foram estabelecidas utilizando como fase m?vel ?cido ac?tico a 0,3% e acetonitrila. Como ponto de partida para o desenvolvimento do m?todo cromatogr?fico, foi utilizado um gradiente explorat?rio, que permite analisar todas as faixas de polaridade. O gradiente de elui??o foi ent?o ajustado at? a obten??o de um cromatograma com adequada separa??o e par?metros cromatogr?ficos satisfat?rios. Para a identifica??o dos compostos, foi realizada a an?lise de padr?es e, pela observa??o dos tempos de reten??o e espectros de absor??o das subst?ncias, foi poss?vel eleger as que estariam presentes na amostra. Foi poss?vel sugerir a presen?a de catequina, epicatequina, epigalocatequina e ?cido g?lico na fra??o AcOEt de A. occidentale. Para a valida??o do m?todo, foram realizados os testes de linearidade, precis?o, exatid?o, robustez, limites de detec??o e quantifica??o. O m?todo foi avaliado como sendo linear, preciso e exato para os compostos estudados. Dentre as condi??es de an?lise alteradas para o estudo da robustez, o m?todo n?o apresentou robustez para a an?lise do ?cido g?lico, quando a temperatura da coluna foi de 35?C, mas as demais condi??es alteradas n?o promoveram mudan?as estatisticamente significativas para este analito. Conclui-se que o m?todo cromatogr?fico ? adequado para a an?lise da fra??o AcOEt do extrato acetona de A. occidentale. / Anacardium occidentale Linn is a tree native to Brazil, popularly known as cashew. Studies show that the Ethyl Acetate fraction (EtOAc) obtained from the acetone:water extract of the A. occidentale stem bark has anti-inflammatory activity, suggesting tannins as responsible for this action. As one of 71 species of the National List of Medicinal Plants of Interest to the Unified Health System (RENISUS), it is important that analytical methodologies for the study of its constituents are developed. In this context, the High Performance Liquid Chromatography (HPLC) is a method of importance to the analysis of samples of plant origin. This study aimed to develop and validate an analytical methodology by HPLC for analysis of EtOAc fraction of acetone:water extract from the stem bark of A. occidentale. The chromatographic conditions were established using acetic acid 0.3% (v/v in water) and acetonitrile as the mobile phase. As a starting point for the development of chromatographic procedures, exploratory gradient was used, which allows to analyze all polarity groups. The gradient elution was then adjusted to obtain a chromatogram with adequate separation and satisfactory chromatographic parameters. For identification of the compounds, the pattern analysis was carried out, and by observing the retention times and absorption spectra of the substances was possible to choose those which would be present in the sample. It could suggest the presence of catechin, epicatechin, epigallocatechin and gallic acid in the EtOAc fraction of A. occidentale. To validate the method, linearity tests were performed, precision, accuracy, robustness, limits of detection and quantification. According to Resolution 899/2003 (ANVISA), the method was assessed to be linear, precise and accurate for the studied compounds. Among the changed analysis conditions for studying the robustness of the method, when the column temperature was 35?C analysis did not present robustness for gallic acid, but when other conditions were changed it did not promote statistically significant changes to this analyte. It follows that the chromatographic method is suitable for analysis of gallic acid, catechin and epicatechin in EtOAc fraction of the acetone:water extract of the A. occidentale.

Made available in DSpace on 2014-12-17T14:16:30Z (GMT). No. of bitstreams: 1 MarceloVPA_DISSERT.pdf: 3152615 bytes, checksum: 2a3513c0df769468fec98423bf2dba29 (MD5) Previous issue date: 2013-03-21 / Tuberculosis is a serious disease, but curable in practically 100% of new cases, since complied the principles of modern chemotherapy. Isoniazid (ISN), Rifampicin (RIF), Pyrazinamide (PYR) and Chloride Ethambutol (ETA) are considered first line drugs in the treatment of tuberculosis, by combining the highest level of efficiency with acceptable degree of toxicity. Concerning USP 33 - NF28 (2010) the chromatography analysis to 3 of 4 drugs (ISN, PYR and RIF) last in average 15 minutes and 10 minutes more to obtain the 4th drug (ETA) using a column and mobile phase mixture different, becoming its industrial application unfavorable. Thus, many studies have being carried out to minimize this problem. An alternative would use the UFLC, which is based with the same principles of HPLC, however it uses stationary phases with particles smaller than 2 ?m. Therefore, this study goals to develop and validate new analytical methods to determine simultaneously the drugs by HPLC/DAD and UFLC/DAD. For this, a analytical screening was carried out, which verified that is necessary a gradient of mobile phase system A (acetate buffer:methanol 94:6 v/v) and B (acetate buffer:acetonitrile 55:45 v/v). Furthermore, to the development and optimization of the method in HPLC and UFLC, with achievement of the values of system suitability into the criteria limits required for both techniques, the validations have began. Standard solutions and tablets test solutions were prepared and injected into HPLC and UFLC, containing 0.008 mg/mL ISN, 0.043 mg/mL PYR, 0.030 mg.mL-1 ETA and 0.016 mg/mL RIF. The validation of analytical methods for HPLC and UFLC was carried out with the determination of specificity/selectivity, analytical curve, linearity, precision, limits of detection and quantification, accuracy and robustness. The methods were adequate for determination of 4 drugs separately without interfered with the others. Precise, due to the fact of the methods demonstrated since with the days variation, besides the repeatability, the values were into the level required by the regular agency. Linear (R> 0,99), once the methods were capable to demonstrate results directly proportional to the concentration of the analyte sample, within of specified range. Accurate, once the methods were capable to present values of variation coefficient and recovery percentage into the required limits (98 to 102%). The methods showed LOD and LOQ very low showing the high sensitivity of the methods for the four drugs. The robustness of the methods were evaluate, facing the temperature and flow changes, where they showed robustness just with the preview conditions established of temperature and flow, abrupt changes may influence with the results of methods / A tuberculose ? uma doen?a grave, por?m cur?vel em praticamente 100% dos casos novos, desde que obedecidos os princ?pios da moderna quimioterapia. S?o considerados f?rmacos de 1? linha no tratamento ? tuberculose: isoniazida, pirazinamida, etambutol e rifampicina. De acordo com USP 33 - NF28 (2010) as an?lises cromatogr?ficas para 3 dos 4 f?rmacos (isoniazida, pirazinamida e rifampicina) duram em m?dia 15 minutos e mais 10 minutos para a obten??o do 4? f?rmaco (etambutol) utilizando outra coluna, com outra mistura de fase m?vel, tornando esta aplica??o na pr?tica industrial desfavor?vel. Uma das alternativas ? utilizar o CLUE, o qual baseia-se nos mesmos princ?pios da CLAE, por?m utiliza fases estacion?rias com part?culas menores que 2 ?m. Dessa forma pretende-se com o presente estudo desenvolver e validar novos m?todos anal?ticos para determina??o simult?nea de tuberculost?ticos por CLAE/DAD e CLUE/DAD. Para isto, foi realizado um screening anal?tico, o qual verificou que ? necess?rio um gradiente de sistema de fase m?vel A (tamp?o acetato:metanol 94:6 v/v) e B (tamp?o acetato:acetonitrila 55:45 v/v). Posteriormente, ao desenvolvimento e otimiza??o do m?todo em CLAE e CLUE com a obten??o dos valores de adequabilidade do sistema dentro dos limites de aceita??es vigente para ambos as t?cnicas, as valida??es deram-se in?cio. Solu??es padr?es e solu??es testes dos comprimidos foram preparadas e injetadas no CLAE e CLUE, contendo isoniazida, pirazinamida, etambutol e rifampicina nas concentra??es de 0,008, 0,043, 0,030 e 0,016 mg.mL-1, respectivamente. A valida??o dos m?todos anal?ticos foram realizadas para: especificidade / seletividade, intervalos da curva anal?tica, linearidade, limite de detec??o, limite de quantifica??o, exatid?o, precis?o (repetibilidade, precis?o intermedi?ria) e robustez. Os m?todos foram adequados para determina??o dos 4 f?rmacos separadamente sem interfer?ncia nos demais. Precisos, devido ao fato de que os m?todos demonstraram que mesmo com varia??o de dias, al?m da repetibilidade, os valores ficaram dentro da faixa preconizada na legisla??o vigente. Lineares (R > 0,99), ou seja, os m?todos foram capazes de demonstrar que os resultados obtidos eram diretamente proporcionais ? concentra??o do analito na amostra, dentro de um intervalo especificado. Exatos, uma vez que os m?todos foram capazes de apresentar valores de coeficiente de varia??o e porcentagem de recupera??o dentro dos limites exigidos (98 a 102%). Os m?todos mostraram LD e LQ com n?veis baixos demonstrando que os m?todos possuem elevada sensibilidade aos quarto f?rmacos. A robustez foi avaliada frente ?s altera??es de temperatura e fluxo, onde os m?todos demonstraram-se robustos apenas nas condi??es previamente estabelecidas de temperatura e fluxo, altera??es bruscas podem acarretar influ?ncia nos resultados dos m?todos