Phthalates and bisphenols migration in Mexican food cans and plastic food containers.

The presence of endocrine disruptors bisphenol-A, bisphenol-A-dimethacrylate, bisphenol-A-diglycidyl-ether, phthalic-acid, dibutyl-phthalate, diethyl-phthalate and dioctyl-phthalate was determined in vegetable cans, baby bottles and microwaveable containers from the Mexican market. Gas-Chromatography-Mass-Spectrometry was used for the identification and High-Performance-Liquid-Chromatography with UV/Visible light and fluorescence detectors was used for the quantification. Endocrine disruptors were found in all samples. PA and DOP were the substances most commonly found, and maximum concentrations were 9.549 and 0.664 µg/kg, respectively from a jalapeno peppers can. Bisphenol A, phthalic-acid, bisphenol-A-dimethacrylate, bisphenol-A-diglycidyl-ether, dioctyl-phtalate and dibutyl-phthalate were found in baby bottles and microwaveable containers.

Oestrogenicity of paper and cardboard extracts used as food containers.

Bisphenol-A (BPA), dibutyl phthalate (DBP), and di-2-ethylhexyl phthalate (DEHP), which are common chemical residues in food-packaging materials, were investigated in paper and cardboard containers used for take-away food. The oestrogenicity of aqueous extracts was tested in E-Screen bioassay and analysis carried out by high-performance liquid chromatography (HPLC) and gas chromatography-mass spectrometry (GC/MS). Oestrogenicity was demonstrated in 90% of extracts (geometric mean [GM] = 11.97 pM oestradiol equivalents g(-1)). DEHP, DBP, and BPA (GM = 341.74, 37.59, and 2.38 ng g(-1) of material) were present in 77.50, 67.50, and 47.50% of samples, respectively. In bivariate analyses, no significant association was found between the levels of these chemicals and oestrogenicity in cardboard/paper extracts. A close-to-significant association was found between oestrogenicity and DBP (beta = 1.25; p = 0.06) in paper extracts, which reached statistical significance in multivariate analysis (beta = 1.61; p = 0.03). Paper and cardboard used in food packaging may contribute to the inadvertent exposure of consumers to endocrine-disrupting chemicals.

Environmental and lifestyle factors for organochlorine exposure among women living in Southern Spain.

Southern Spain has the largest area of intensive greenhouse agriculture in Europe, and may constitute a special case of occupational and female exposure, because this type of farming requires considerable pesticide use and employs many women. We measured adipose tissue levels of 14 organochlorine pesticides in 458 women living in this area and analyzed the relationship between pesticide level/presence and sociodemographic characteristics, reproductive history or life-style factors that may influence this exposure. Pesticide presence was determined by gas chromatography with electron-capture detector. All fat samples were positive for 1 residue. DDT or metabolites were found in 98.25%, with mean value of 660 ngg(-1) of lipid. p,p'-DDE level was higher in women who were older, with lower educational level or obese. Almost 70% had measurable levels of endosulfan and/or metabolites, with a mean total value of 38.8 ngg(-1) of lipid. Endosulfan-I exposure was higher in women with shorter residence in rural settings and more frequent in women with 3 children. 52.62% were exposed to 1 of aldrin-dieldrin-endrin group. Endrin was more frequent in women who were younger, with higher educational level or no agricultural working experience; dieldrin was more frequent in women who were older, with lower educational level or more children. Finally, lindane residues were found in 39.30%. Lindane levels were higher in women who breastfed longer or had more children. Research is required on women occupationally exposed to a selected group of organochlorine pesticides, especially those of reproductive age, as a basis for preventive action.

Human exposure to endocrine disrupters: standardisation of a marker of estrogenic exposure in adipose tissue.

In many epidemiological studies based on the direct measurement of exposure to organochlorines, the chemicals of concern are determined directly from adipose tissue samples. Although the measurement of all possible organochlorines, their metabolites, isomers and congeners may be desirable, it is expensive and time-consuming and many chemicals with hormonal activity may not yet have been identified. Testing systems are therefore required to screen for estrogenicity and to identify appropriate biomarkers of human exposure. To address this issue, we developed and standardised a method to assess the total estrogenic xenobiotic burden in human adipose tissue. The method extracts and separates the more lipophilic xenoestrogens from ovarian estrogens, with a subsequent bioassay determination of the cumulative effect of the xenoestrogens. It was applied to 400 women, using 200 mg of adipose tissue: 65% of samples showed measurable estrogenicity in the fraction where most non-polar xenoestrogens eluted, and 76% of fractions where ovarian estrogens eluted were positive for estrogenicity. Residues of 16 organochlorine pesticides were determined. No correlation was found between pesticide content and estrogenicity of the samples. The high percentage of positive samples suggests that the method is sensitive enough to be used as a biomarker of human exposure to estrogenic xenobiotics and can be applied in epidemiological studies.

Analysis of organochlorine pesticides in human milk: preliminary results.

In the face of evidence of human milk contamination by organochlorine pesticides, an analysis was performed on samples of milk obtained from healthy lactating women in the provinces of Granada and Almeria in Southern Spain. The samples were obtained by the Neonate Section of the Department of Pediatrics of Granada University Hospital (Neonatology Division) and by the Neonatal Service of Poniente Hospital in El Ejido, Almería. A liquid-liquid extraction procedure was performed. The cleaning of the sample before gas chromatography-mass spectrometry (GC-MS) used silica Sep-Pak. Among other pesticides, aldrin, dieldrin, DDT and its metabolites, lindane, methoxychlor and endosulfan were identified. The presence of these products was confirmed by mass spectrometry. The identification and quantification of these organochlorine molecules is important because they have estrogenic effects.

Determination of bisphenol A and related aromatic compounds released from bis-GMA-based composites and sealants by high performance liquid chromatography.

Most of the composites and sealants used in dentistry are based on bisphenol A diglycidylether methacrylate (Bis-GMA). Reports revealed that in situ polymerization is not complete and that free monomers can be detected by different analytic methods. Concerns about the estrogenicity of bisphenol A (BPA) and other aromatic components leached from commercial products have been expressed. We studied biphenolic components eluted from seven composites and one sealant before and after in vitro polymerization using HPLC and gas chromatography/mass spectrometry and we investigated how pH modifications affect the leaching of these components. We found BPA (maximal amount 1.8 microg/mg dental material), its dimethacrylate derivative (Bis-DMA, 1.15 microg/mg), bisphenol A diglycidylether (6. 1 microg/mg), Bis-GMA (2.0 microg/mg), and ethoxylate and propoxylate of bisphenol A in media in which samples of different commercial products were maintained under controlled pH and temperature conditions. Our results confirm the leaching of estrogenic monomers into the environment by Bis-GMA-based composites and sealants in concentrations at which biologic effects have been demonstrated in in vivo experimental models. The main issue with implications for patient care and dentist responsibility is to further determine the clinical relevance of this estrogenic exposure.

Development of a marker of estrogenic exposure in human serum.

The deleterious, disruptive effects of estrogen mimics on the endocrine system were discovered after the compounds were released into the environment. Their chemical structure does not obviously resemble that of steroid hormones; hence, their estrogenic effects were totally unexpected. In addition to occupational exposures, environmental estrogens may have played a role in decreasing the quantity and quality of human semen during the last 50 years and in increasing the incidences of testicular cancer and cryptorchidism in men and breast cancer in women and men in industrialized countries. Testing the environmental estrogen hypothesis will require developing appropriate biomarkers of exposure and measuring these biomarkers at developmental points where exposure is critical. We report the ongoing development of a method to extract and separate xenoestrogens from ovarian estrogens with human serum as a source, followed by determination of xenoestrogen concentration by a bioassay. We also critically assess bioassays currently available to measure the cumulative effect of xenoestrogens, e.g., (a) the E-SCREEN assay, which measures the proliferative effect of estrogens on their target cells, and (b) the induction by estrogens of specific gene products, such as progesterone receptor and pS2.

The E-screen assay: a comparison of different MCF7 cell stocks.

MCF7 human breast cancer cells have been studied extensively as a model for hormonal effects on breast cancer cell growth and specific protein synthesis. Because the proliferative effect of natural estrogen is considered the hallmark of estrogen action, it was proposed that this property be used to determine whether a substance is an estrogen. The E-screen assay, developed for this purpose, is based on the ability of MCF7 cells to proliferate in the presence of estrogens. The aim of our study was to characterize the response of four MCF7 cell stocks (BUS, ATCC, BB, and BB104) and determine which of them performed best in the E-screen test. The four stocks assayed were distinguishable by their biological behavior. In the absence of estrogen, MCF7 BUS cells stopped proliferating and accumulated in the G0/G1 phase of the cell cycle; estrogen receptors increased, progesterone receptors decreased, and small amounts of pS2 protein were secreted. Of all the MCF7 stocks tested, MCF7 BUS cells showed the highest proliferative response to estradiol-17 beta: cell yields increased up to sixfold over those of nontreated cells in a 144-hr period. The differences between estrogen-supplemented and nonsupplemented MCF7 BUS cells were due mostly to G0/G1 proliferative arrest mediated by charcoal dextran-stripped serum. MCF7 BUS cell stocks and others showing a similar proliferative pattern should be chosen for use in the E-screen test, or whenever a proliferative effect of estrogen is to be demonstrated.

Xenoestrogens released from lacquer coatings in food cans.

We present data showing that some foods preserved in lacquer-coated cans and the liquid in them may acquire estrogenic activity. Hormonal activity was measured using the E-screen bioassay. The biological activity of vegetables packed in cans was a result of plastic monomers used in manufacturing the containers. The plastic monomer bisphenol-A, identified by mass spectrometry, was found as a contaminant not only in the liquid of the preserved vegetables but also in water autoclaved in the cans. The amount of bisphenol-A in the extracts accounted for all the hormonal activity measured. Although the presence of other xenoestrogens cannot be ruled out, it is apparent that all estrogenic activity in these cans was due to bisphenol-A leached from the lacquer coating. The use of plastic in food-packaging materials may require closer scrutiny to determine whether epoxy resins and polycarbonates contribute to human exposure to xenoestrogens.