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No / Ethylmethanesulphonate (EMS), benzidine (BZD) and benzo[a]pyrene (B[a]P) were assayed for ability to induce mutation at the hprt locus of wild-type L5178Y mouse lymphoma cells. EMS was assayed in the absence of metabolic activation, B[a]P in the presence of metabolic activation (S-9 mix) and BZD both in the absence and presence of S-9. Treatment with EMS minus S-9 and B[a]P plus S-9, especially when the S-9 content of the incubation was 2% (v/v), produced strong dose-related increases in mutant frequency. BZD failed to induce mutation at the hprt locus, either in the absence or presence of S-9.

In this study, a well biofilm-forming bacterial strain was isolated from oil contaminated water and sediment samples collected in petroleum storage Duc Giang, Hanoi. It was identified as Paracoccus sp. DG25 and registered in the GenBank database with the accession numbers KJ608354. Several biophysical and bio-chemical conditions for the biofilm formation of the strain were estimated such as pH, temperature, carbon sources and nitrogen sources. As the results the biofilm forming capacity was highest at pH 7, 37 oC, on maltose and supplemented with KNO3. Using these optimal conditions, the formed biofilm degraded 76.07 % of pyrene after 7 day-incubation, with the initial concentration of 300 ppm by high-performance liquid chromatography (HPLC) analysis. To our knowledge, there is rare publication on pyrene degradation by biofilm-forming bacteria. Therefore, the obtained results show that biofilm formed the strain Paracoccus sp. DG25 may considerably increase the degrading efficiency of pyrene and may lead to a new approach to treat polycyclic aromatic hydrocarbons containing in petroleum oil contaminated water in Vietnam. / Trong nghiên cứu này, từ các mẫu đất và nước nhiễm dầu lấy tại kho xăng Đức Giang, Hà Nội, chúng tôi đã phân lập được chủng vi khuẩn có khả năng tạo màng sinh học tốt. Chủng vi khuẩn này đã được phân loại và định tên là Paracoccus sp. DG25 với số đăng ký trên ngân hàng Gen là KJ608354. Chúng tôi cũng đã nghiên cứu một số điều kiện hóa lý ảnh hưởng tới khả năng hình thành màng sinh học như pH, nhiệt độ, nguồn Carbon và nguồn Nitơ. Kết quả cho thấy, chủng DG25 có khả năng tạo màng tốt nhất ở các điều kiện pH 7, 37 oC, nguồn Carbon là maltose và nguồn Nitơ là KNO3. Sử dụng các điều kiện tối ưu này để tạo màng và đánh giá khả năng phân hủy pyrene của màng tạo thành. Bằng phương pháp sắc ký lỏng cao áp, chúng tôi đã đánh giá được hàm lượng pyrene bị phân hủy sau 7 ngày nuôi tĩnh bởi màng sinh học của chủng DG25 lên tới 76,07 % với nồng độ ban đầu là 300 ppm. Cho tới nay, chưa có nhiều công bố về hiệu quả phân hủy pyrene của các chủng vi khuẩn tạo màng sinh học. Do vậy, kết quả đạt được này mở ra khả năng sử dụng màng tạo thành bởi chủng DG25 để nâng cao hiệu quả phân hủy pyren và có thể mở ra phương pháp mới nhằm xử lý các hợp chất hydrocarbon thơm có trong nước ô nhiễm dầu ở Việt Nam.

Reconnu par le Centre international de recherche sur le cancer (CIRC) comme cancérigène chez l’être humain, le benzo[a]pyrène (BaP) est un des hydrocarbures aromatiques polycycliques (HAP) les plus étudiés. Souvent rencontrés dans de nombreux milieux de travail, les HAP sont un groupe de polluants omniprésents dans l’environnement, formé par des processus de combustion incomplets. Bien que le BaP présente un risque élevé pour la santé des travailleurs, il n’existe aucun biomarqueur spécifique au composé permettant le suivi et la surveillance d’exposition au BaP dans un lieu de travail. Le 3-hydroxybenzo[a]pyrène (3-OHBaP) est le métabolite du BaP le plus développé comme biomarqueur. Ce métabolite est principalement excrété dans les fèces, ainsi qu’une quantité infime dans l’urine, ce qui le rend difficile à mesurer. De plus, le 3-OHBaP montre une certaine rétention rénale, un facteur qui rend plus compliqué son utilisation en tant que biomarqueur car il oblige de prendre plusieurs critères en compte dans l’analyse de sa cinétique temporelle. Par ailleurs, le 1-hydroxypyrène (1-OHP) est un métabolite urinaire du pyrène souvent utilisé comme biomarqueur d’exposition aux HAP. Neanmoins, il s’agit du métabolite d’un HAP non-cancérigène et par conséquent sa capacité de démontrer le risque de cancer associé à une exposition donnée est faible. Ce mémoire visait à détecter et à évaluer l'exposition au BaP en suivant ses métabolites urinaires: le 4,5-dihydrodiol-benzo[a]pyrène et le 7,8-dihydrodiol-benzo[a]pyrène (le 4,5-diolBaP et le 7,8-diolBaP; les diolBaP). L’évaluation du 4,5-diolBaP et du 7,8-diolBaP s’est déroulée dans deux études: La première étude, surnommée « l’expérience du shampooing », portait sur un volontaire qui s’est exposé aux HAP dans un environnement contrôlé en utilisant un shampooing à base de goudron de houille. Cette étude a été conçue afin d’évaluer l'évolution temporelle du 7,8-diolBaP chez l'homme et de vérifier son potentiel en tant que biomarqueur d'exposition par comparaison avec le 1-OHP dans le même cadre expérimental. Elle a été réalisée avec deux expériences. La première portait sur une seule exposition et la seconde sur une exposition multiple. La deuxième étude, surnommée « l’étude des travailleurs », reposait sur une analyse comparative du 4,5-diolBaP, du 7,8-diolBaP, du 1-OHP et du 3-OHBaP dans un milieu de travail. Cette étude avait pour objectif d’évaluer les compétences des diolBaP dans un contexte réel à côté des biomarqueurs établis d'exposition au BaP et aux HAP. Cinq travailleurs d'une usine de production d'anodes en carbone ont accepté de participer à cette étude. Dans le cadre de ces deux études, les échantillons d'urine étaient analysés par la chromatographie en phase liquide à ultra-haute performance (UHPLC) couplée à la fluorescence. « L’expérience du shampooing » : L’expérience de l’exposition unique et celle de l’exposition multiple ont révélé une élimination de façon mono-exponentielle du 7,8-diolBaP, identique à celle du 1-OHP, avec des concentrations dans le même ordre de grandeur. Nous avons également confirmé un taux d’élimination plus rapide pour le 1-OHP en regardant ses pics. Le 7,8-diolBaP augmente en valeur maximale après chaque exposition, et cette découverte a mis en évidence une accumulation tout au long de la semaine, alors que pour le 1-OHP, le deuxième pic est plus grand, mais le troisième est plus petit, montrant ainsi moins d'accumulation pendant la même période temporelle. « L’étude des travailleurs »: Selon les résultats, la méthode analytique utilisée était incapable de discerner correctement le 4,5-diolBaP des autres contaminants urinaires éluant pendant le même temps de rétention. Le 7,8-diolBaP, quant à lui, élue à des concentrations urinaires d'un ordre de grandeur similaire au 1-OHP tel que vu chez tous les travailleurs évalués. Chez certains travailleurs, la concentration urinaire du 7,8-diolBaP était toujours plus élevée avant le début d'un quart de travail avec l'élimination qui avait lieu pendant le quart de travail pour fournir une valeur de concentration inférieure à la fin du quart de travail. Cependant, la concentration du 1-OHP a eu une hausse immédiate avec l'exposition, culminant à la fin de chaque quart de travail. Pour les autres travailleurs, les concentrations du 7,8-diolBaP et du 1-OHP étaient systématiquement plus élevées à la fin du quart de travail. Il est probable que ces variations indiquent les différentes voies d'exposition. Le présent mémoire a montré le potentiel du 7,8-diolBaP en tant que biomarqueur d'exposition spécifique au BaP et par conséquent, il fournit un point de départ pour explorer la quantification du lien entre l'exposition au BaP et ses effets néfastes sur la santé de l'être humain. L’utilisation de la spectrométrie de masse est nécessaire à confirmer l’identité des diolBaP avant d'aller de l'avant. / Benzo[a]pyrene (BaP) is one of the more commonly studied polycyclic aromatic hydrocarbons (PAHs), a group of omnipresent pollutants in the environment formed through incomplete combustion processes. It is listed as a confirmed carcinogen to human beings by the International Agency for Research on Cancer (IARC) and is highly present in many workplaces. Although presenting a significant health risk to workers, there are currently no convenient compound-specific biomarkers that enable the tracking and monitoring of occupational exposure to BaP. 3-Hydroxybenzo[a]pyrene (3-OHBaP) is the most developed, as a biomarker, amongst the metabolites of BaP. It’s mostly excreted with the faeces, presenting in trace amounts in urine, which makes it difficult to measure; it is also demonstrates renal retention, which adds a layer of complexity in its use as a biomarker because there are many factors to take into consideration when looking at its kinetic time course. 1-Hydroxypyrene (1-OHP), in the other hand, is a urinary metabolite of pyrene that serves as a good representative of PAH presence. It is, however, the metabolite of a non-carcinogenic PAH, and is not fully capable of representing the cancer risk posed in a given scenario. This thesis sought to detect and assess BaP exposure through tracking its urinary metabolites: 4,5-dihydrodiol-benzo[a]pyrene and 7,8-dihydrodiol-benzo[a]pyrene (4,5-diolBaP and 7,8-diolBaP; diolBaPs). 4,5-DiolBaP and 7,8-diolBaP were evaluated through two studies: The first study, the “shampoo experiment”, featured a volunteer who self-exposed to PAHs in a controlled setting by using a coal-tar-based shampoo. The study consisted of two experiments. The first focused on a single exposure and the second on multiple exposures. This study was set to evaluate the time course of 7,8-diolBaP in humans and verifying its potential as a biomarker of exposure through a comparison with 1-OHP in the same experimental framework. The second study consisted of a comparative analysis of 4,5-diolBaP, 7,8-diolBaP, 1-OHP and 3-OHBaP in an occupational setting, evaluating the competency of the diolBaPs in a real-world setting alongside established biomarkers of BaP and PAH exposure. Five workers at a carbon anode production plant volunteered to participate in this study. For both of these studies, the urine samples were analysed by ultra-high-pressure liquid chromatography (UHPLC) coupled with fluorescence. “Shampoo Experiment”: The single and multiple exposures revealed a monoexponential elimination on the part of 7,8-diolBaP, identical to 1-OHP, with similar magnitudes of concentration. 1-OHP was also confirmed to undergo a more rapid elimination from the system, where after each exposure for 7,8-diolBaP, the ensuing peak value is higher. This finding demonstrated evidence of accumulation of 7,8-diolBaP throughout the week, whereas with 1-OHP, the second peak is larger, then the third one is smaller, thus showing less accumulation over the same time frame. “Worker Study”: The results showed that the analytical method used was unable to properly discern 4,5-diolBaP from other urinary contaminants eluting during the same retention time. 7,8-DiolBaP, on the other hand, eluted at urinary concentrations that were a similar order of magnitude to 1-OHP, as can be seen in all of the workers evaluated. For some workers, the urinary concentration of 7,8-diolBaP was consistently at its peak prior to the start of a shift and elimination took place during the shift, to provide a lower concentration value at the end of the shift. With 1-OHP, the rise was immediate with exposure, peaking at the end of every shift. For other workers, both 7,8-diolBaP and 1-OHP are consistently higher at the end of the shift. These variations are likely to indicate different routes of exposure. This thesis showed the potential use of 7,8-diolBaP as a compound-specific biomarker of exposure for BaP and thus provides a starting point in exploring the quantification of BaP exposure and negative health effects in humans. Confirmation of the compound’s identity is needed through the use of mass spectrometry.

L'obésité est une maladie définie par une accumulation de masse grasse dans le tissu adipeux ayant des conséquences néfastes pour la santé. Les causes de l’obésité sont multiples. Dans un travail récent, il y a été démontré le rôle de la pollution environnementale dans la prise de poids. Dans ce travail, les hypothèses selon lesquelles les récepteurs adrénergiques situés à la surface des cellules adipeuses seraient le siège de l’action des polluants aromatiques polycycliques ont été vérifiées par le dosage de plusieurs agonistes et antagonistes spécifiques et non spécifiques en présence ou non du benzo[a]pyrène sur des récepteurs humains et de cellules d’hamster chinois (CHO). Les quantités d’AMPc obtenues montrent que les HAP ne se déposent pas sur les récepteurs β1, β2, β3 adrénergiques.Cette accumulation se fait au niveau des phospholipides de la membrane cytoplasmique des cellules. Ce qui cause une rigidité des membranes.Cette observation tend à renforcer l'hypothèse selon laquelle le benzo[a]pyrène induirait une inhibition de la lipolyse par l'accumulation au niveau de la bicouche de phospholipides et des changements de conformation de la bicouche de phospholipides dans les environs des récepteurs à sept domaines transmembranaires qui sont β-adrénergiques.La liaison de la bicouche phospholipidique avec les HAP utilisés est une réaction exothermi-que avec un faible dégagement de chaleur / Obesity is a disease defined by an accumulation of fat in adipose tissue with adverse consequences for health. The causes of obesity are many.In recent work, there was demonstrated the role of environmental pollution in weight gain.In this work, the assumptions that the adrenergic receptors on the surface of fat cells would home to the accumulation of polycyclic aromatic pollutants have been verified by measurement of several agonists and antagonists specific and non-specific in the presence or absence of benzo[a]pyrene receptors on human cells and Chinese hamster (CHO). The amounts of cAMP obtained showed that PAHs are not deposited on β-receptors, β1, β2, β3 adrenergic receptors.This accumulation occurs at the cytoplasmic membrane phospholipids of the cells. What cau-ses stiffness of the membranes. This observation tends to reinforce the hypothesis that benzo [a]pyrene induce an inhibition of lipolysis by the accumulation in the phospholipid bilayer and conformational changes of the bilayer phospholipids in the vicinity of receptors seven transmembrane domains which are β-adrenergic receptors

Setenta e três fungos representantes de grupos taxonômicos distintos, isolados de cnidários, coletados no litoral norte do estado de São Paulo, foram submetidos ao teste de descoloração do corante RBBR e ao teste de MIC. Treze fungos selecionados no teste de descoloração foram submetidos ao teste de degradação de pireno e benzo[a]pireno. Sete fungos mostraram resultados satisfatórios de degradação, entretanto, o fungo Aspergillus sulphureus (8.2A) apresentou os melhores resultados: degradou 99,7% de pireno e 73,6% de benzo[a]pireno. Com relação ao teste de MIC, 21 isolados inibiram o crescimento das bactérias: B. subtillis; P. aeruginosa, S.aureus, E. coli EPEC e V. parahaemolyticus. Penicillium sp. (12.12) apresentou o melhor resultado (0,0078 mg/mL). Os 23 isolados que apresentaram potencial biotecnológico foram caracterizados taxonomicamente. Os resultados do presente trabalho sugerem que fungos filamentosos associados a cnidários possuem potencial para aplicação biotecnológica, em particular para biorremediação de ambientes marinhos contaminados com HPAs. / Seventy three fungi representatives of distinct taxonomic groups, isolated from cnidarians collected from north coast of São Paulo State, were submitted to RBBR dye decolorization tests and to MIC. Thirteen fungal selected from the decolorization test were submitted to pyrene and benzo[a]pyrene degradation assays. Seven fungi showed satisfactory PAH degradation results. Among them Aspergillus sulphureus (8.2A) presented the best results degrading pyrene (99.7%) and benzo[a]pyrene (73.6%). Regarding the MIC experiments, 21 isolates were able to inhibit the growth of B. subtilis, P. aeruginosa, S. aureus, E. coli EPEC and V. parahaemolyticus. Penicillium sp. (12.12) presented the best result (0,0078 mg/mL). The 23 isolates that showed potential biotechnologic were taxonomically characterized. Results derived from the present work suggest that cnidarians-derived filamentous fungi have a great potential for biotechnological application, in particular on bioremediation of marine environments contaminated by PAHs.

Le Benzo(a)pyrène (BaP), appartenant à la famille des hydrocarbures aromatiques polycycliques (HAP) est cancérigène pour l’homme. Nous avons développé une approche protéomique quantitative nanoLC-MS/MS label-free pour identifier des biomarqueurs liés à l’exposition au BaP dans le sécrétome des cellules hépatiques humaines exposées au BaP vs. des cellules non exposées et exposées au Benzo(e)pyrène (BeP). Le BeP, agent non classifié comme cancérigène pour l’homme, est choisi comme contrôle négatif afin de distinguer les protéines spécifiques du BaP de celles des HAP. 847 protéines ont été identifiées et quantifiées, et 55 ont été fortement surexprimées avec un ratio supérieur à 5 : la plupart de ces protéinessurexprimées sont précoces et liées au cancer. Une validation ultérieure de l'expression de ces protéines dans le plasma de la population exposée au BaP aidera dans le développement de biomarqueurs qui permettront d'améliorer la détection précoce, le pronostic et prévention. / Benzo(a)pyrene (BaP) belongs to a class of polycyclic aromatic hydrocarbon and is reported as a potent human carcinogen. We performed a nanoLC-MS/MS-based label-free quantitative proteomics approach to identify potential biomarkers of exposure in the secretome of BaPtreated vs. non-treated and Benzo(e)pyrene (BeP)-treated human hepatoma cell line HepG2.BeP-treated cells were chosen as a negative control to distinguish the BaP-specific from the HAP-specific regulated proteins: BeP is not classifiable as to its carcinogenicity to humans. 847 proteins have been identified and quantified and 55 proteins were seen as being highly upregulated with a fold change of at least 5. Most of these up-regulated proteins were focused incancer-related activities. Further validation of expression of these proteins in the plasma of BaP-exposed population will assist in the development of biomarkers that will greatly improve early detection, prognosis, prediction of treatment response and prevention.

Setenta e três fungos representantes de grupos taxonômicos distintos, isolados de cnidários, coletados no litoral norte do estado de São Paulo, foram submetidos ao teste de descoloração do corante RBBR e ao teste de MIC. Treze fungos selecionados no teste de descoloração foram submetidos ao teste de degradação de pireno e benzo[a]pireno. Sete fungos mostraram resultados satisfatórios de degradação, entretanto, o fungo Aspergillus sulphureus (8.2A) apresentou os melhores resultados: degradou 99,7% de pireno e 73,6% de benzo[a]pireno. Com relação ao teste de MIC, 21 isolados inibiram o crescimento das bactérias: B. subtillis; P. aeruginosa, S.aureus, E. coli EPEC e V. parahaemolyticus. Penicillium sp. (12.12) apresentou o melhor resultado (0,0078 mg/mL). Os 23 isolados que apresentaram potencial biotecnológico foram caracterizados taxonomicamente. Os resultados do presente trabalho sugerem que fungos filamentosos associados a cnidários possuem potencial para aplicação biotecnológica, em particular para biorremediação de ambientes marinhos contaminados com HPAs. / Seventy three fungi representatives of distinct taxonomic groups, isolated from cnidarians collected from north coast of São Paulo State, were submitted to RBBR dye decolorization tests and to MIC. Thirteen fungal selected from the decolorization test were submitted to pyrene and benzo[a]pyrene degradation assays. Seven fungi showed satisfactory PAH degradation results. Among them Aspergillus sulphureus (8.2A) presented the best results degrading pyrene (99.7%) and benzo[a]pyrene (73.6%). Regarding the MIC experiments, 21 isolates were able to inhibit the growth of B. subtilis, P. aeruginosa, S. aureus, E. coli EPEC and V. parahaemolyticus. Penicillium sp. (12.12) presented the best result (0,0078 mg/mL). The 23 isolates that showed potential biotechnologic were taxonomically characterized. Results derived from the present work suggest that cnidarians-derived filamentous fungi have a great potential for biotechnological application, in particular on bioremediation of marine environments contaminated by PAHs.

Interest into the effects of weak static magnetic fields on chemical reactions involving spin correlated radical pairs has increased over the last few decades, particularly as scientists have become more curious about the mechanisms by which animals can sense and respond to small variations in the Earth's weak (50 µT) magnetic field. The magnetosensitivity of radical pairs, as dictated by the radical pair mechanism, lies at the heart of the most heavily supported hypothesis of this magnetoreception phenomenon. This thesis is concerned with the spectroscopic investigations of isotropic and anisotropic magnetic field effects in fluorescent systems. First of all, an introduction to spin chemistry and magnetoreception is presented. In chapter 3, the effects of weak radiofrequency oscillating fields when applied in combination with weak static fields are explored in isotropic solutions. The validity of the high-field model, typically used to describe spin dynamics in magnetic resonance, is tested and the effects of orientation and field strength on magnetic field effects are discussed in detail. In Chapter 4, a range of exciplex systems are studied by fluorescence methods and their energetics are explored. The factors which determine the formation of an exciplex, i.e. the complex equilibrium between the exciplex and the spin-correlated radical pair,are considered and used to assess the existence and magnitude of MFEs. Radical pair systems investigated, using MARY spectroscopy, with respect to their potential to act as model chemical compasses are introduced in chapter 5. Solid-state media are used to align the exciplex systems to detect any magnetic field direction dependence. Finally, in chapter 6, AMELIA, an experiment which can directly measure the anisotropic magnetic field response of a system, is presented and applied successfully to systems to detect directly the anisotropic field response of a photoexcited anthracene crystal.

Long interspersed nuclear element-1 (L1) is a genetic element that mobilizes throughout the mammalian genome via retrotransposition and damages host DNA via mutational insertions, chromosomal rearrangements, and reprogramming of gene expression. The cellular mechanisms responsible for aberrant L1 expression during cancer pathogenesis are unclear. Previously, we have shown that L1 reactivation in several human cell lines is dependent upon the activation of aryl hydrocarbon receptor (AhR), a ligand-activated transcription factor member of the PAS superfamily of proteins. We also showed that ectopic expression of L1 reprograms the HepG2 genome leading to epithelial-to-mesenchymal transition (EMT). Here we present evidence that reactivation of L1 and modulation of EMT in HepG2 cells by the AhR ligand benzo(a)pyrene (BaP) is effected through the canonical TGF-β1 signaling pathway. BaP increased TGF-β1 mRNA, SMAD2 phosphorylation and decreased expression of E-Cadherin. The functional relevance of these interactions and the involvement of TGFBR1/ALK5 and SMAD2/3 were confirmed by siRNA interference. Furthermore, expression of L1-encoded ORF1p was positively correlated with the activation of TGF-β1 signaling in human hepatocarcinoma samples at various stages of malignant progression. These results indicate that ligand-mediated AhR activation regulates L1 via canonical TGF-β1 signaling and raise important questions about the molecular etiology of human hepatocarcinomas.