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- Formation of emerging disinfection byproducts in water and evaluation of potential genotoxic effects: the case of chlorinated polycyclic aromatic hydrocarbonsPublication . Rebola, Marlene; Pinto, Miguel; Louro, Henriqueta; Antunes, A.M.M.; José, S.S.; Rocha, M.R.; Silva, Maria João; Cardoso, Ana SofiaDisinfection byproducts (DBPs) are formed when disinfectants used in water treatment plants (WTPs) react with natural (or anthropogenic) organic matter present in the source water. Many studies have addressed health risks posed by a life-time exposure to DBPs through chlorinated drinking water or through dermal or inhalation exposure routes. Experimental studies have revealed genotoxic and carcinogenic effects of some DBPs and epidemiological studies evidenced potential associations between chlorinated drinking water and bladder or colorectal cancer. In addition, a possible link between chlorinated drinking water and reproductive/developmental effects has been hypothesized. Many DBPs have been identified in treated water, which justifies the growing concern about the potential health effects of emerging unregulated DBPs, some of which appear to be more genotoxic, in some assays, than the regulated DBPs. Polycyclic aromatic hydrocarbons (PAHs) are among the most persistent contaminants detected in environmental samples such as river sediments and tap water. Water chlorination can lead to the formation of chlorinated derivatives of PAHs (Cl-PAHs) and the few available toxicological studies have shown that Cl-PAHs possess greater mutagenicity than the corresponding parent PAHs. The present study had two main objectives: 1) identification of the major chlorinated derivatives of benzo[a]pyrene (BaP) and fluoranthene (Fluo) formed as chlorination by-products and 2) evaluation of their potential hazard to humans, through the characterization of their potential genotoxic effects in a human cell line. To synthesize chlorinated standards of PAHs, a newly two phase (water/n-hexane) method was developed for BaP and Fluo. 6-Cl-BaP was obtained as the major chlorination product of BaP, and 3-Cl-Fluo and 1,3-Cl2-Fluo of Fluo. The formation of these BaP and Fluo chlorinated derivatives was also observed under WTPs chlorination conditions after at 0.5 until 24 h of exposure. The effects of equimolar concentrations of 6-Cl-BaP vs. BaP and of 3-Cl-Fluo/1,3-Cl2-Fluo vs. Fluo on cell viability and DNA integrity were assessed by the neutral red uptake (NR) and the comet assay, respectively. Exposure of HepG2 cells to a dose-range of 6-Cl-BaP and BaP showed that both compounds are cytotoxic above 50 µM and that, at the equimolar doses of 100 and 125 µM, 6-Cl-BaP is able to induce a significantly higher level of DNA damage than BaP. On the other hand, no changes of cell viability were observed after exposure to several concentrations of Fluo and its derivatives. Likewise, none of the compounds was able to significantly induce DNA damage. In conclusion, the present data confirmed that chlorinated derivatives of BaP and Fluo are formed during WTPs chlorination procedures and allowed the identification of their major chlorinated derivatives that should be further analysed in drinking water. On the other hand, the results from the comet assay evidenced a higher DNA damaging effect of Cl-BaP comparatively to its parent compound, suggestive of a more potent genotoxic effect. In spite of the negative results found for Fluo and its chorinated products, further genotoxicity studies are still needed to allow a definite conclusion. Although health risks of DBPs are small compared to health risks of waterborne diseases, the identification of hazardous Cl-PAHs in water emphasizes the need of development of new and safer water disinfection methods.
- Portuguese EQAS vision versus experience on pre-analytical phase (2007-2011)Publication . Faria, Ana; Correia, Helena; Gomes, M. AdelinaPortuguese EQAS vision versus experience on pre-analytical phase (2007-2011) Ana Faria, Helena Correia, M. Adelina Gomes Instituto Nacional de Saúde Dr Ricardo Jorge, Lisboa, Portugal (Gabinete de Avaliação Externa da Qualidade Laboratorial) Introduction: According to recent bibliography it's in the pre-analytical phase, that a predominance of errors is observed (32 to 75%) that are not noticed or are overlooked in assessment procedures or result interpretation. It became, therefore, important to monitor its use, as well as the absolute necessity to compare and standardize procedures. In 2007 the Evaluation of Pre-Analytic Phase was implemented, now part of the 77 existing Clinical Programs in the National External Quality Assessment of the National Institute of Health, Portugal. Objective: The main objective of this program is to evaluate the performance of participants in order to improve the performance of clinical laboratories nationwide in the pre-analytical phase. This study evaluated the results obtained in nine surveys by laboratories participating in the PNAEQ. Methods: Between 2007 and 2011 laboratories enrolled in theEvaluation in Pre-Analytic Phase program, 2 surveys per year. We distributed questionnaires, samples, case studies, medical request simulation, sample handling simulation, considerations for compliance of national legislation and guidelines of the General Directorate of Health, contents of collection manual or collection instructions, emergency situations, different conditions for special collection, collections in sampling stations, cases of drugs interference, questions about sample storage and transport , biosafety considerations, equipment and sample material, criteria for sample rejection, control and reagent preparation, record of traceability of the pre-analytical process over the nine surveys. We requested, when applicable, the classification of the recorded errors, with PNAEQ’s comments and clarification. The results were statistically analyzed, frequency charts performed, according with clients answers. In each survey a joint report was prepared with the overall results and pertinent comments. Results: In 5 years of this program 105 laboratories were enrolled with an average participation of 44%. Fifty seven percent enrolled only once, and only 14% maintained their registration with participation in four or more years. The highest percentage of answers received was in surveys that included shipment of samples with simulated clinical history (44 to 65%) or case studies (63%). In these surveys we considered some incomplete comments, but all correct. In patient registration and error detection in simulated medical request, participants submitted their comments on the most appropriate and complete. For questions asked on control records and reagent preparation, questionnaires with accounting errors and biosafety situations, we have not had significant response. We consider we are still in the sensibilization process of laboratories, trying to highlight the importance of participation in this program. In the report cards information is sent on critical points inherent in this phase. Conclusion: Although there has been a good first year in participation, there was a decrease in the number of laboratories enrolled over the years (in 2011less than 1/3 compared to 2007) but with a tendency to an increase active participation in surveys that included sample analysis and case study. Although with an average participation in the first survey of only 27%, the percentage of reported errors is consistent with those described in the literature. On this date there was no indication of automation of the pre analytical phase in laboratories. Laboratories that had an assiduous involvement in 4/ 5 years have the quality management system implemented, a facilitating tool for a response to the program, demonstrating a practice of procedures with records for detection and quantification of errors in different phases of the analytical process. Not reporting errors, does not mean their absence. It was in the information content of the Collection Manual and in simulated analysis with samples sent that we verified the existence of good practice procedures. The practice of recording in order to trace the entire pre analytical procedure will allow its monitoring. We believe that the formative part of this program may warn, encourage and sensitize the participants to errors that may occur at the beginning of the analytical process. It was always proposed that the laboratory staff members jointly reviewed the report card, so that they can draw conclusions and make the self-evaluation. As main objective, we have the evaluation of the impact of errors in future trials.
- Impact of Potentially Contaminated Sediments From The Sado Estuary in Human Health: Cytotoxic And Genotoxic Assays in a Human Cell LinePublication . Pinto, Miguel; Louro, Henriqueta; Costa, Pedro; Caeiro, Sandra; Lavinha, João; Silva, Maria JoãoAs many estuaries in industrialized countries, the river Sado Estuary (W Portugal) is affected by various sources of pollution, such as heavy-industry, urbanism, mining, agriculture and maritime traffic. Mostly classified as a natural reserve, it also remains a privileged site for fishing activities performed by the local population, who not only consume but distribute their fishery. Previous studies revealed sizable amounts of contaminants in the estuary sediments, namely metals, pesticides and polycyclic aromatic hydrocarbons. These compounds can be accumulated in the edible parts of estuarine species with commercial value or local agricultural products and enter the human food chain, posing a public health concern. The present study is part of a broader project whose objective is to evaluate the environmental risk, including ecologic and human health risk, associated with the estuarine benthic environment, complemented with the analysis of a target population from a small village located near the estuary shore. This study aims to assess the cytotoxic and genotoxic potential of sediments from the Sado Estuary through the neutral red uptake assay and the alkaline comet assay (coupled with DNA repair endonucleases) in a human cell line respectively, using multiple samples collected in various points of the Sado Estuary. Sediments were collected from two geochemically distinct and potentially contaminated sites of the Sado Estuary: site F and site C. Total organic and inorganic contaminants were extracted with a mixture of methanol:dicholomethane (1:2) and recovered in DMSO. HepG2 cells were exposed for 48h to concentrations of each extract ranging from 0.1 to 20ul/ml of culture medium. A dose-related decrease in cell viability was observed for extract F from 1ul/ml up to 20ul/ml, indicating sediment contaminant-driven cytotoxicity, whereas no cytotoxicity induction was observed for extract C. Genotoxicity was only found for extract F, collected near a heavy-industrialized site. Also, increased genotoxicity was observed in cells treated with the DNA repair endonuclease FPG, for extract F, suggesting oxidative DNA damage. No significant genotoxity was observed for extract C. Given the difference in cytotoxic and genotoxic effects between both samples, a larger number of samples is necessary to reflect the overall contamination status of the Sado Estuary. Therefore, other samples are currently being analysed in order to obtain a more complete evaluation of the cytotoxic and genotoxic potential of the sediment contaminants from the river Sado Estuary, and a sample from a non-contaminated site will be added as a reference. The results are expected to reflect the Sado Estuary contamination by different anthropogenic pressures.
- Ambiente TérmicoPublication . Pires, A.Acção de formação que visa habilitar os participantes: a caracterizar o ambiente térmico como parâmetro físico presente no local de trabalho e como potencial risco para a saúde; a distinguir casos de conforto térmico e de stresse térmico, com recurso a metodologias de avaliação e interpretação de resultados.