Phthalate esters (phthalates) such as dibutylphthalate (DBP) are commonly used as plasticisers and pesticides in a variety of products such as children‟s plastic toys, food packaging, cosmetics, medical equipment (including surgical equipment), and acaricides. Because of their widespread use phthalates are ubiquitous environmental contaminants that humans are commonly exposed to. Phthalates are known endocrine-disrupting chemicals (EDCs) that are well known to cause male reproductive defects such as cryptorchidism (failed descent of the testes) and hypospadias (malformations in the urethra) in a range of different species if they are exposed in utero. They do this by reducing testosterone production in Leydig cells, which are the primary site of testosterone biosynthesis in the male. Because phthalates are dose-additive they are considered to share the same mechanism of toxicity. However, the details of phthalates mechanism of toxicity are not fully understood. The aim of this research was to investigate the effects of DBP on the steroidogenesis pathway using the cultured rat Leydig cell cancer line R2C as a Leydig cell model. R2C cells were exposed to a range of DBP concentrations (10 μg/mL, 5 μg/mL, 1 μg/mL, and 0.1 μg/mL) and their steroid hormone production was analysed using reverse phase HPLC. R2C cells did not synthesise testosterone at detectable levels. However, DBP exposure stimulated cortisol biosynthesis at all concentrations but caused no change in progesterone biosynthesis. This cortisol stimulation in Leydig cells has not been observed before. Because cortisol and testosterone compete for precursors an increase in cortisol synthesis could starve testosterone synthesis of precursors. On top of this it has been shown that glucocorticoids including cortisol have an adverse effect on Leydig cell development reducing steroid production and even causing apoptosis. This could explain how DBP and other phthalates can cause male developmental defects such as cryptorchidism and hypospadias.

Orientadores: Helena Teixeira Godoy, Marisa Padula / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos / Made available in DSpace on 2018-08-23T04:09:43Z (GMT). No. of bitstreams: 1 Oliveira_WellingtondaSilva_M.pdf: 956439 bytes, checksum: 3f0a6d0735f71f0d97e159bbf8fce38f (MD5) Previous issue date: 2013 / Resumo: No Brasil, mais de 50% da população utiliza mamadeiras durante a amamentação de crianças. Após a proibição do uso do bisfenol A em mamadeiras de policarbonato, outros polímeros como o polipropileno e o Tritan® passaram a ser utilizados com essa finalidade. Embora estes materiais sejam liberados para uso como material de contato com alimentos há uma lacuna no que diz respeito à migração de compostos oriundos do plástico utilizado na fabricação de mamadeiras. Estudos voltados para a identificação de substâncias não intencionalmente adicionadas e migrantes, em mamadeiras, são necessários para avaliar a segurança no uso destes materiais. Diante disso, o objetivo deste estudo foi avaliar o potencial de migração dos materiais utilizados na confecção de mamadeiras. Foram avaliadas 32 mamadeiras, de 3 marcas diferentes, feitas de Tritan (A) e polipropileno (B, CT e CP). Entre as marcas, três eram transparentes e com desenhos (A, B, CT), e uma era completamente pigmentada (CP). Inicialmente, foi feito a caracterização das mamadeiras e confirmação do material por espectroscopia no infravermelho por transformada de Fourier (FT-IR). Na caracterização foram avaliadas a espessura mínima, o diâmetro, o volume, a massa e altura das mamadeiras. Em seguida, foram realizados ensaios de migração de acordo com a RDC 51/2010 da Agência Nacional de Vigilância Sanitária (ANVSA), que recomenda a utilização de etanol 50% como simulante de leite. A simulação foi realizada mantendo a mamadeira em contato com o simulante a 70°C por 2h. Neste ensaio foi feita a identificação dos migrantes com base na biblioteca de espectros de massas do National Institute of Standards and Technology (NIST). Um método para determinação de di(2-etilhexil) ftalato (DEHP) e dibutil ftalato foi validado para os parâmetros limite de detecção (LD) e quantificação (LQ), linearidade, repetitividade e precisão intemediária. O perfil de antioxiandantes foi determinado utilizando um cromatógrafo líquido com detector de arranjo de diodos. Com base na caracterização, foi observado que as mamadeiras das marcas B, CT e CP apresentaram maior uniformidade nos parâmetros avaliados que as mamadeiras da marca A, que apresentaram dimensões com coeficientes de variação maiores que 20%, indicando falta de padrão em mamadeiras de um mesmo lote. Na seleção e identificação de migrantes foi observada a migração de mais de 20 compostos, dentre eles o DEHP e o DBP. O método validado apresentou limites de detecção e quantificação de 16 µg/L e 52 µg/L para o DBP e 10 µg/L e 30 µg/L para o DEHP. O método mostrou-se linear nas faixas de 52-660 µg/L para o DBP e 52-1100 µg/L para o DEHP, com boa repetitividade e precisão intemediária com CV abaixo de 20%. Todas as mamadeiras apresentavam concentrações de DEHP abaixo do LQ. Em relação ao DBP, somente as mamadeiras da marca CP apresentaram migrações acima do limite de migração específa (0,3 mg/kg) recomendado pela ANVISA. Em todas as mamadeiras foram encontrados os antioxidantes Irganox 1010 e Irgafos 168. Estes compostos não têm restrição de uso, podendo ser usados em materiais de contato com alimentos. Das três marcas de mamadeiras analisadas, somente as marcas B e CT apresentaram resultados satisfatórios para utilização em contato com o leite. As mamadeiras da marca A deformaram após o ensaio de simulação. As mamadeiras da marca CP apresentaram níveis de migração acima do limite de migração especifica (0,3 mg/kg) preconizado pela ANVISA / Abstract: In Brazil, more than 50% of the population uses baby bottles to feed children. After banning the use of bisphenol A in polycarbonate baby bottles, other polymers such as polypropylene and Tritan® have been used for this purpose. Although these materials were approved for using as food contact material, there is a vacuity with regard to the migration of plastic derived compounds used in the manufacture of baby bottles. Studies aimed at the identification of non-intentionally added substances and migrants, in baby bottles, are needed to assess the safety of these materials. Thus, the aim of this study was to evaluate the migration potential of materials used in the baby bottles manufacture . Thirty-two baby bottles of three different brands made of Tritan (A) and polypropylene (B, CT, and CP) were evaluated. Among the brands, three were transparent and drawings (A, B, CT), and one was completely pigmented (CP). Initially, were made a characterization the material baby bottles and then its materials were confirmed by infrared spectroscopy Fourier transform (FT-IR). The characterization was based on minimum thickness, diameter, volume, mass and height of the baby bottles. Then, migration assays were performed according to the RDC 51/2010 of the National Agency for Sanitary Vigilance (ANVISA), which recommends the use of 50% ethanol as a milk simulant. The simulation was done by putting the baby bottles in contact with the simulant at 70 ° C for 2 h. In this assay the identication of the migrant was done based on the mass spectra library of the National Institute of Standards and Technology (NIST). A method for the determination of di (2-ethylhexyl) phthalate (DEHP) and dibutyl phthalate (DBP) was validated for the parameters detection (LOD) and quantification (LOQ) limit, linearity, repeatability and accuracy-intermediate. The antioxiandants profile was determined using a liquid chromatograph with a diode array detector. The materials used in the baby bottles manufacture, as declared on the labels were confirmed. Based on characterization, it was observed that the baby bottles of brands B, CT, and CP showed greater uniformity in the parameters evaluated than brand A, which had dimensions with variation coefficients (CV) greater than 20%, indicating the lack of standard in the same lot of baby bottles. The selection and identification of migrants showed the migration of more than 20 compounds, including DEHP and DBP. The method validated in this study showed a detection and quantification limits of 16 µg/L and 52 µg/L for DBP and 10 µg/L and 30 µg/L for DEHP. The method was linear in the range of 52-660 µg/L for DBP and 52-1100 µg/L for DEHP, with good repeatability and accuracy-intermediate with CV below 20%. All baby bottles had concentrations of DEHP below the LOQ. Regarding DBP, only the baby bottles of brand CP showed migration above specific migration limit (0.3 mg / kg) recommended by ANVISA. In all bottles were found the antioxidants Irganox 1010 and Irgafos 168. These compounds have no use restriction and can be used in food contact materials. Taking into account the brands of baby bottles analyzed, only the brands B and CT showed satisfactory results for use in contact with milk. The baby bottles of brand A deformed after the simulation test. The baby bottles of the brand CP showed migration levels above the specific migration limit (0.3 mg/kg) recommended by ANVISA / Mestrado / Ciência de Alimentos / Mestre em Ciência de Alimentos

Orientador: Samuel Monroe Cohen / Resumo: O aumento da incidência de distúrbios testiculares e a possível influência de substâncias químicas ambientais, como o dibutilftalato (DBP) e a acrilamida (AA), exigem a identificação de modos de ação. A maioria dos estudos de toxicologia reprodutiva utiliza amostras de RNA provenientes de todo o testículo para avaliar a expressão genica; entretanto, análises de tipos celulares isolados poderiam gerar resultados mais específicos. Entre as células germinativas testiculares, as espermatogônias são importantes pois representam o início da espermatogênese. Este estudo objetivou, 1) estabelecer técnica de isolamento de espermatogônias; 2) aplicar esta técnica para verificar possíveis alterações na expressão gênica (Pou5f1, Kitlg, Mki-67, Bak1 e Spry4) em testículos de ratos pré-púberes (DPN24) e púberes (DPN45) após exposição in utero e pós-natal ao DBP ou à AA. A técnica foi eficiente para o isolamento das espermatogônias. A exposição ao DBP levou à redução do peso corporal da ninhada ao nascer, da distância anogenital dos filhotes machos no DPN4 e ao aumento da frequência de retenção de mamilos no DPN14. Os pesos relativos dos testículos expostos ao DBP estavam reduzidos apenas no DPN24. Animais expostos ao DBP mostraram níveis reduzidos de expressão de Pou5f1 e Mki67 no DPN24, e de Pou5f1 e Spry4 no DPN45. A exposição a AA reduziu a expressão de Pou5f1, Mki67 e Spry4, embora não significativamente. Nossos resultados sugerem que DBP atue reduzindo a proliferação celular e prejudi... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: The increased incidence of testicular disorders in young men and the possible influence of environmental chemicals, such as dibutyl phthalate (DBP) and acrylamide (AA), requires experimental models for identifying modes of action. Most published reproductive toxicologic studies use RNA samples from the total testis to evaluate testicular gene expression; however, analyses of isolated cell types could provide a more specific tool. Among testicular germ cells, spermatogonia are critical since they represent the onset of spermatogenesis. This study aimed, 1) to establish a technique for spermatogonia isolation; 2) to apply this isolation technique to verify possible gene expression alterations (Pou5f1, Kitlg, Mki-67, Bak1 and Spry4) in prepubertal post-natal day, (PND24) and pubertal (PND45) testes after in utero and postnatal exposure to DBP or AA. The technique was efficient for isolation of a majority of spermatogonia. In utero DBP exposure led to reduced litter body weight at birth, reduced anogenital distance of male pups on PND4, and increased frequency of male nipple retention on PND14 compared to controls. DBP-exposed relative testes weights were reduced only at PND24 compared to control but they did not differ at PND45. DBP-exposed animals showed reduced expression levels of Pou5f1 and Mki67 on PND24, and reduced expression of Pou5f1 and Spry4 on PND45. AA exposure reduced expression of Pou5f1, Mki67 and Spry4 at PND45 although not significantly. Our results suggest tha... (Complete abstract click electronic access below) / Doutor

There is general concern whether human exposure to selected Canadian bottled waters and the chemicals which may leach from them, could potentially cause deleterious effects. This research was designed to determine whether exposures to bottled water and plastic leachates caused toxicity to the freshwater Cnidarian Hydra viridissima (green hydra). Three chemicals used in the production of polycarbonate and polyethylene plastics, bisphenol A (BPA) and two phthalate esters: dibutyl phthalate (DBP) and bis (2- ethylhexyl) phthalate (DEHP) along with one type of commercial bottled water were investigated. One brand of bottled water was analyzed over four months (stored in light and dark conditions) along with lab water similarly stored in glass, polycarbonate and polyethylene bottles. Following 2, 4, 8 and 16 weeks in each of the two treatments, hydra bioassays were conducted. Chronic toxicity tests were also conducted on the two phthalates and BPA. The chronic toxicity tests showed that BPA caused effects on hydra morphology and population at low doses and DBP and DEHP both showed signs of hormesis.

Orientador: Rejane Maira Góes / Texto em português e inglês / Dissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-24T19:26:40Z (GMT). No. of bitstreams: 1 Negrin_AnaCarolina_M.pdf: 3573186 bytes, checksum: 84ff9397d827717b776f6497a21bc510 (MD5) Previous issue date: 2014 / Resumo: Desreguladores endócrinos (DE), como o di-n-butil ftalato (DBP), podem alterar o panorama dos hormônios esteroides ou sua ação, comprometendo o desenvolvimento testicular e a capacidade reprodutiva na vida adulta. Estudos toxicológicos mostram que ratos adultos, quando expostos a altas doses de ftalatos durante a fase de diferenciação sexual, exibem diversas anomalias reprodutivas, como agenesia do epidídimo e redução na produção diária de espermatozoides. Dados mostram que exposições a 2mg/kg/dia de DBP durante a gestação e a lactação são suficientes para prejudicar o desenvolvimento das células germinativas. Além disso, há relatos de que DE contrinuem para o aumento da adipogênese por causar alterações na sinalização celular de adipócitos. É conhecido que a obesidade masculina pode afetar a qualidade do sêmen e aumenta as taxas de infertilidade. Considerando que os ftalatos podem se acumular no tecido adiposo, ficamos interessados em avaliar os efeitos da exposição prolongada a baixas doses de DBP a as possíveis interferências do excesso de lipídeos na dieta para a função testicular e os parâmetros espermáticos de gerbilos adultos. Gerbilos fêmeas adultas, alimentadas com dieta balanceada (4% de lipídeos) ou hiperlipídica (20% de lipídeos) por oito semanas, foram acasaladas com machos normais. A prole masculina foi dividida em grupos controle (C), di-n-butil ftalato (Ph), dieta hiperlipídica (HF) e hieta hiperlipídica mais di-n-butil ftalato (HFPh). DBP (5 mg/kg/dia) foi administrado na água de beber às mães durante a gravidez e lactação e aos filhotes do desmame até a idade adulta (14 semanas). A resposta do testículo foi avaliada por meio de análises microscópicas e esterológicas, da sensibilidade de suas principais populações celulares a andrógenos e estrógenos, e da produção espermática. Também foram examinados os efeitos sobre a reserva espermática, o tempo de trânsito dos espermatozoides pelo epidídimo e a motilidade espermática. Isoladamente, baixas doses de DBP resultaram em obesidade e dislipidemia nos animais adultos. Nenhuma alteração foi observada na estrutura testicular, mas a síntese de estrógeno pelo testículo foi reduzida, resultando em tendência a diminuição na eficiência espermática. A redução no estrógeno intratesticular, após exposição ao DBP, foi acompanhada de um aumento de ~70% na expressão de ER? nos testículos, que pode ser uma resposta adaptativa aos baixos níveis deste hormônio. A ingestão crônica de dieta hiperlipídica não induziu os gerbilos adultos à obesidade, mas causou uma leve queda na eficiência espermática. Esta redução não está relacionada a alterações na estrutura testicular ou na sua capacidade esteroidogênica, mas podem estar ligadas ao comprometimento da sinalização testicular, já que foi verificada uma redução no conteúdo de AR. A exposição combinada à dieta hiperlipídica e a baixas doses de DBP atuaram sinérgica e negativamente na síntese de testosterona intratesticular, prejudicando a eficiência espermática e aumentando o tempo de trânsito dos espermatozoides pelo epidídimo. A motilidade espermática não sofreu alteração frente as exposições isoladas ou combinadas. Este estudo demonstrou que o ambiente nutricional pode interferir na resposta dos testículos frente aos ftalatos e proporciona novas informações para o entendimento das consequências da exposição aos DE para a redução na reverva espermática e fertlidade humana / Abstract: Endocrine disrupting chemicals (EDC), as di-n-butyl phthalate (DBP), can alter the scenary of steroid hormones or their action, impairing the testicular development and reproductive capacity in adulthood. Toxicological studies show that adult rats, when exposed to high doses of phthalates during sexual differentiation, exhibit several reproductive anomalies, such as agenesis of the epididymis and reduction in daily sperm production. Data show that exposures to 2mg/kg/day of DBP during gestation and lactation periods are enough to impair the development of the male germ cells. Furthermore, there are reports that EDC contribute to the increase of the adipogenesis by altering cell signaling of adipocytes and lipid metabolism. It is known that the male obesity can affect semen quality and increases infertility rates. Whereas that phthalates can accumulate in adipose tissue, we were interested in evaluating the effects of prolonged exposure to low doses of DBP and possible interferences of excess dietary fat for testicular function and spermatic parameters of adult gerbils. Adult female gerbils, fed high-fat (20% fat) or balanced diet (4% fat) for eight weeks, were mated with normal males. The male offspring was divided into control (C), di-n-butyl phthalate (Ph), high-fat diet (HF) and high-fat diet plus di-n-butyl phthalate (HFPh) groups. DBP (5 mg/kg/day) was administered in drinking water to pregnant and breastfeeding mothers and to offspring from weaning up to adulthood (14-week-old). Testis response was evaluated by means of microscopic and stereological analyses, sensitivity of its major cell populations to androgens and estrogens, steroidogenic capacity and spermatic efficiency. We also examined the effects on sperm reserves, sperm transit time through the epididymis and sperm motility. Alone, low doses of DBP resulted in obesity and dislipidemy at adulthood. No histological change was observed in testicular structure of these animals, but there was reduction in intratesticular estrogen synthtesis, resulting in a tendency to decrease in sperm production. The decrease in intratesticular estrogen after lifetime DBP exposure was accompanied by a ~70% increment in ER? content in the testis, that might be an adaptative response to low estrogen levels. Chronic intake of high-fat diet did not induce obesity in aldult gerbils, but led to a slight decrease in spermatic efficiency. This decrease was not associated with changes in testicular structure or steroidogenic capacity, because serum or intratesticular testosterone and estrogen concentrations were not altered, but it can be linked to an unbalanced in androgen receptor signaling, since it was observed a decrease in AR content. The combined exposure to high-fat diet and to low doses of DBP acted synergical and negatively on intratesticular testosterone synthesis, impairing the spermatic efficiency and increasing the sperm transit time. The sperm motility was not changed in isolated or combined exposures. This study demonstrated that fat nutritional environment may adversely affect the response of testes to phthalates, and provide new informations for understanding the consequences of exposures to ECD in decreasing human sperm counts and fertility / Mestrado / Biologia Celular / Mestra em Biologia Celular e Estrutural

Orientador: Rejane Maira Góes / Dissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-23T14:41:50Z (GMT). No. of bitstreams: 1 Jesus_MarianaMarcielode_M.pdf: 3839823 bytes, checksum: 5cdcc794169929d299d222355a85e2d1 (MD5) Previous issue date: 2013 / Resumo: O resumo poderá ser visualizado no texto completo da tese digital / Abstract: The abstract is available with the full electronic document / Mestrado / Biologia Celular / Mestra em Biologia Celular e Estrutural

<p>Uvod: Poslednjih godina u fokusu istraživača je efekat sintetskih jednjenja na endokrini sistem. Estri ftalne kiseline se koriste u procesu plastifikacije, kao industrijski rastvarači, lubrikanti, aditivi u tekstilnoj industriji, u pesticidima, kozmetičkim proizvodima. Raste broj dokaza da je tiroidna žlezda podložna dejstvu endokrinih disruptora. Tiroidni hormoni imaju važnu ulogu u regulaciji rasta, tkivne diferencijacije, energetskog metabolizma, reprodukcije i formiranja centralnog nervnog sistema. Brojna istraživanja ukazala su da ftalati deluju kao EDs. Ciljevi istraživanja: 1. Procena izloženosti populacije mono-etilheksil-ftalatu (MEHP) i mono-etil-ftalatu (MEP). 2. Evaluacija razlika u nivou pokazatelja tirodine funkcije između ftalat pozitivnih i ftalat negativnih ispitanika i između gojaznih i normalno uhranjenih ftalat pozitivnih ispitanika. 3.Utvrđivanje razlika u serumskom nivou leptina gojaznih ispitanika sa i bez pozitivnih ftalatnih metabolita i procena povezanosti leptina sa MEP i MEHP i pokazateljima tiroidne funkcije. Izbor ispitanika i metod rada: Istraživanje je sprovedeno kao studija preseka, obuhvatilo je 201 ispitanika. Ispitanici su podeljeni u grupu MEP/MEHP pozitivnih i negativnih i na podgupe normalno uhranjenih i gojaznih. Od antropometrijskih mera određena je telesna visina, telesna masa, obim struka i indeks telesne mase. Laboratorijske analize: jutarnji uzorak urina za određivanje MEP i MEHP; na&scaron;te uzet uzorka venske krvi za FT4, FT3, TSH i leptin. Statististička analiza sprovedena je na softverskom paketu SPSS. Rezultati: Polovina stanovni&scaron;tva je izložena ftalatima. MEP dovodi do povi&scaron;enja FT4 samo u subpopulaciji gojaznih. Nije utvrđen statistički značajan uticaj MEP na FT3. Kod gojaznih MEP pozitivnih osoba ženskog pola povi&scaron;en je TSH. MEHP uzrokuje sniženje FT4 kod normalno uhranjenih ispitanika, a kod normalno uhranjenih mu&scaron;karaca snižava FT3. Nije utvrđen uticaj MEHP na tirotropin. U gojaznih nije ustanovljen uticaj DEHP i DEP na leptinsku sekreciju.Uočena je tendencija negativne korelacije leptina i FT4 kod gojaznih, dok uticaja na FT3 i TSH nema. Zaključak: Na&scaron;a populacija je u velikoj meri izložena ftalatima. Potvrđeno je da MEP i MEHP imaju uticaj na pojedine indikatore tiroidne funkcije. Ftalati u na&scaron;em istraživanju ne uzrokuju poremećaj leptinske skrecije, a leptin ima blag uticaj jedino na FT4.</p> / <p>Introduction: Effects of synthesized chemicals on endocrine system has been in the focus in the last years. Phthalates are used in plasticization, as industrial solvents, lubricants, textile industry additives, in pesticides and cosmetic products. Evidence for thyroid disruption is growing. Thyroid hormones (TH) have an important role in regulation of growth, tissue differentiation, energy metabolism, reproduction and central nervous system formation. Studies show phthalates can cause endocrine disruption. Aims: 1. Estimation of burden of mono-ethyl phthalate (MEP) and di-2-ethylheksyl phthalate(MEHP) in the population. 2. Evaluation of differences in TH and TSH in MEP/MEHP positive and negative participants, as in obese and lean MEP/MEHP positive participants. 3. Evaluation of differences in leptin in obese MEP/MEHP positive and negative subjects and evaluation of the connection between leptin, MEP, MEHP and thyroid indicators. Patients and methods: This was a cross-sectional study that comprised 201 subjects divided into MEP/MEHP positive and negative group, further subdivided in obese and lean. Anthropometric parameters done: body height, body weight, waist and body mass index. Laboratory tests done: morning urine sample analysis for MEP/MEHP and venous sample analysis for free thyroxine (FT4), free tri-iodothyronine (FT3), thyroid stimulating hormone (TSH) and leptin. Statistical analysis was done in SPSS. Results: Half of subjects were exposed to phthalates. MEP induced an increase in FT4 in obese participants and had no influence on FT3. TSH was increased in obese MEP positive female subjects. MEHP induced a decrease in FT4 in lean participants and a decrease of FT3 in lean males. There was no correlation between MEHP and TSH. Influence of MEP/MEHP on leptin secretion. A tendency for negative correlation between leptin and FT4 was seen. There was no influence of leptin on FT3 and TSH. Conclusion: Our population is greatly exposed to phthalates. MEP and MEHP influence certain thyroid indicators i.e. cause thyroid disruption. Phthalates do not influence leptin secretion in our study. There is a mild effect of leptin on FT4.</p>