Spelling suggestions: "subject:"[een] COMMERCIAL FORMULATION"" "subject:"[enn] COMMERCIAL FORMULATION""
1 |
Embryotoxicité de contaminants métalliques et organiques chez l'escargot Helix aspersa / Embryotocixity of mettallic and organic chemicals in the land snail Helix aspersaBaurand, Pierre-Emmanuel 26 September 2014 (has links)
Les oeufs d’escargot terrestre de l’espèce petit-gris Helix aspersa (syn. Cantareusaspersus) peuvent être utilisés pour évaluer l’écotoxicité de substances chimiques pures ou enmélange. La mesure des effets embryotoxiques classiquement réalisée est le succès d’éclosionaprès 15 à 20 jours d’exposition (Druart et al., 2012). Cependant, les mécanismes impliquésdans la mise en place des effets toxiques à différents niveaux d’organisation biologique chezl’embryon ne sont pas connus. Des oeufs d’escargots ont été exposés à des solutions decontaminants métallique (Cd) ou organiques (pesticides: le Round Up® flash, le Corail® et laBouillie Bordelaise) selon deux modalités différentes (en continu sur la totalité dudéveloppement embryonnaire ou sur une période de 24 heures) afin de 1/ déterminer denouveaux paramètres de mesure au cours du développement embryonnaire pouvant rendrecompte d’un effet toxique, 2/ détecter des effets génotoxiques de divers contaminants(solution métallique de Cd ou de formulations commerciales de pesticides) par la méthodeRandom Amplified Polymorphic DNA (RAPD) et 3/ d’étudier des systèmes de défense métalspécifiques(métallothionéines).Les paramètres morphologiques et physiologiques suivis au cours d’expositionscontinues au Cd ont montré des effets néfastes sur le rythme cardiaque, la durée del’incubation, la taille et le poids à l’éclosion chez les exposés à la plus forte concentrationtestée. Chez ces derniers des signes de fragmentation de l’ADN ont également été détectés enfin d’exposition. Le couplage de la méthode RAPD avec un système d’électrophorèse hauterésolution (SHR) a permis de détecter des effets génotoxiques suite à des expositionscontinues au Cd, au Round Up® et au Corail®. L’étude par PCR quantitative de l’expressiondes gènes des métallothionéines (MTs) a mis en évidence une expression constitutive des MTsainsi qu’un haut niveau d’expression du gène mixte CdCuMT chez les embryons non exposés.Chez les embryons exposés au Cd durant 24 heures, une surexpression du gène spécifiqueCdMT a été mise en évidence alors qu’aucune augmentation significative des taux detranscrits des 2 autres isogènes étudiés (CuMT et CdCuMT) n’a été démontrée.Les résultats de toxicité du Cd basés sur le taux d’éclosion et l’expression des gènes desMTs ont démontré que des facteurs comme le régime d’exposition (24 heures ou en continu)ou le stade de développement (âge des embryons lors de l’exposition) peuvent modulerl’embryotoxicité des substances chimiques.206Les données obtenues durant cette étude intégrative permettent de proposer un largepanel de paramètres de mesure des effets toxiques des substances chimiques chez l’embryond’escargot terrestre H. aspersa au niveau individuel (rythme cardiaque, taille, durée dedéveloppement et succès d’éclosion) et au niveau moléculaire (expression de gènes dessystèmes de défense, détection des signes de génotoxicité et de la fragmentation de l’ADN)pour l’évaluation de la toxicité des substances chimiques. L’approche RAPD-SHR, bien quenécessitant une certaine expertise pour l’analyse des profils d’amplifications obtenus, apparaîtadaptée pour une détection rapide et efficace du potentiel embryogénotoxiques de substancesvariées (métaux, pesticides. / The land snail species Helix aspersa (syn. Cantareus aspersus) eggs can be used to assess theecotoxicity of chemicals. Measurement of embryotoxic effect is classically based on hatching successafter 15-20 days of exposure (Druart et al., 2012). However, the mechanisms involved in toxic effectsin embryos at different levels of biological organization are not known. Eggs of snails were exposedto solutions metallic contaminants (Cd) or organic (pesticides: Round Up® Flash, Corail® andBordeaux Mixture) in two different regimes (continuous over the entire embryonic development orduring a period of 24 hours), in order to 1 / identify of new endpoints of toxic effect measurementsduring embryonic development, 2 / detect of genotoxic effects of metal solution (Cd) or threepesticides commercial formulations by Random Amplified Polymorphic DNA method (RAPD) and 3 /study metal-specific defense systems (metallothionein).Morphological and physiological parameters monitored during Cd continuous exposures showedadverse effects on heart rate, duration of incubation, size and weight of new hatchlings exposed to thehighest concentration tested. In the latter, signs of DNA fragmentation were detected at the end ofexposure. Coupling the RAPD with a high-resolution electrophoresis system (SHR) has enabled todetect genotoxic effects of Cd, Round Up® and Corail® after continuous exposures. Quantitative PCRstudy of metallothioneins (MTs) gene expression has showed constitutive expression of MTs genesand a high level of mRNA for the mixed gene CdCuMT in unexposed embryos. In embryos exposedto Cd for 24 hours, an overexpression of the specific gene CdMT has been demonstrated whereas thetwo other isogenes (CuMT and CdCuMT) didn’t show significant induction of expression rates.The toxicity results based on the hatching rate and MTs genes expression obtained with Cd haveshowed that factors such as the exposure regime (24 hours or continuous) or the stage of development(age of embryos upon exposure) can modulate embryotoxicity of chemicals. This thesis provides awide range of endpoints usable at the individual level (heart rate, height, hatching monitoring) and atthe molecular level (gene expression of defense systems, detection of genotoxicity signs and DNAladdering) for the assessment of the ecotoxicity of chemical substances. The RAPD-SHR, althoughrequiring some expertise to analyze profiles obtained, appears suitable for rapid and efficientdetection of potential embryogenotoxic effects of various substances (metals, pesticides).
|
2 |
[en] DEVELOPMENT AND COMPARATIVE STUDY OF SPECTROFLUORIMETRIC AND VOLTAMMETRIC METHODOLOGIES FOR THE DETERMINATION OF THALIDOMIDE IN ONE COMMERCIAL FORMULATION, URINE AND BLOOD SERUM / [pt] DESENVOLVIMENTO E ESTUDO COMPARATIVO DE METODOLOGIAS ESPECTROFLUORIMÉTRICA E VOLTAMÉTRICA PARA A DETERMINAÇÃO DE TALIDOMIDA EM UM FÁRMACO, URINA E SORO SANGÜÍNEOCARLOS EDUARDO CARDOSO 21 July 2003 (has links)
[pt] No presente trabalho foram desenvolvidas duas metodologias
analíticas para a determinação de talidomida, um composto
de reconhecida importância farmacológica. As metodologias
espectrofluorimétrica e voltamétrica desenvolvidas foram
comparadas em termos de desempenho analítico.As
características fluorescentes e eletroquímicas da
talidomida foram estudadas para que se encontrassem
condições experimentais que fornecessem máximo sinal
fluorescente do analito em solução na temperatura ambiente e
possibilidade de pré-concentração do analito no eletrodo de
mercúrio.Nas condições experimentais otimizadas para a
talidomida, limites de detecção compreendidos entre 10-6 e
10-9 g L-1 foram obtidos para o método
espectrofluorimétrico e voltamétrico, respectivamente.
Faixas lineares dinâmicas entre 2 e 4 ordens de grandeza
foram alcançadas dependendo do método e do interferente
presente na amostra. Esses parâmetros de mérito se mostraram
adequados para o problema proposto. O possível efeito
interferente de substâncias geralmente usadas em
associação com o analito foi estudado, e estratégias para
minimização das interferências foram desenvolvidas.
Enquanto a tetraciclina não interferiu no método
espectrofluorimétrico, o uso combinado de meio ácido e
irradiação UV da amostra foi necessária para permitir a
quantificação do analito em presença de sulfanilamida. Na
voltametria, as interferências da tetraciclina e da
sulfanilamida puderam ser compensadas pelo uso da
quantificação pelo método de adição do analito.
Nas determinações dos fluídos biológicos, o uso da extração
em coluna de sílica C18 mostrou-se bastante eficiente na
separação do analito dos interferentes da matriz para o
método espectrofluorimétrico e para o voltamétrico, no caso
do soro sangüíneo. Para a urina, apenas a clarificação da
amostra com sulfato de amônio foi suficiente no caso da
determinação voltamétrica.Os métodos desenvolvidos foram
testados na dosagem de talidomida presente em uma
formulação comercial e em amostras de urina e soro sangüíneo
enriquecidas com o analito. Para tal, utilizaram-se os
procedimentos de curva de calibração (espectrofluorimetria)
e método da adição do analito (voltametria). Em todos os
casos, as recuperações obtidas estiveram compreendidas na
faixa de 96,5 a 107,6 %, dentro da faixa de recuperação
estabelecida pela Farmacopéia dos Estados Unidos da América. / [en] In the present work two analytical methodologies were
developed aiming the determination of thalidomide, an
important pharmacological compound. The developed
spectrofluorimetric and the voltammetric based analytical
methodologies were compared in terms of analytical
performance. The thalidomide fluorescent and the
electrochemical characteristics were studied in order to
find experimental conditions for maximum fluorescence in
solution and at room temperature and to allow analyte pre-
concentration on the mercury electrode. Using the optimized
experimental conditions, limits of detection between
10-6 to 10-9 g L-1 were achieved respectively for the
spectrofluorimetric method and for the voltammetric method.
Dynamic linear ranges between 2 and 4 orders of magnitude
were obtained depending on the method utilized and the
interferent substances present in the sample. Those
parameters of merit were suitable for this proposed
analytical problem. The potential interference effect from
substances usually used in association with thalidomide,
were studied and strategies for the minimization of such
interferences were developed. While no interference in the
spectrofluorimetric method was observed for tetracycline,
the combined use of acidic medium and UV irradiation of the
samples was necessary to allow the analyte determination in
the presence of sulfanilamide. For the voltammetric method,
interferences from tetracycline and sulfanilamide could be
compensated by quantifying thalidomide using the analyte
addition method. For the determination in biological
fluids, the use a solid-liquid extraction on a C18 column
was found to be very effective for the analyte separation
and elimination of matrix interferences for the
spectrofluorimetric method and for the voltammetric method
developed for blood serum. For urine samples, a clean-up
step using ammonium sulfate was found to be sufficient for
the voltammetric determination of thalidomide.
The developed methodologies were tested by determining the
thalidomide content in a commercial pharmaceutical
formulation and in analyte spiked biological fluids using
calibration curves (spectrofluorimetric) and analyte
addition method (voltammetric). In all cases, the
recoveries were between the 96,5 and 107,6 %, within the
recovery range considered adequate according to the
United States Pharmacopoeia.
|
Page generated in 0.0396 seconds