Evaluation of the migration of mutagens/carcinogens from PET bottles into mineral water by Tradescantia/micronuclei test, Comet assay on leukocytes and GC/MS.

This study monitored the release of mutagenic/carcinogenic compounds into mineral water (natural and carbonated) from polyethylene terephthalate (PET) bottles, using a plant mutagenicity test which reveals micronuclei formation in Tradescantia pollen cells (Trad/MCN test), a DNA damage assay (Comet assay) on human leukocytes and gas chromatography/mass spectrometry (GC/MS) for the characterisation of migrants. The water samples were collected at a bottling plant and stored in PET bottles for a period ranging from 1 to 12 months. Every month some samples were randomly collected and lyophilised, the residual powders were extracted with organic solvents and then analysed by GC/MS and tested for DNA damage in human leukocytes, or reconstituted with distilled water to obtain concentrates for the exposure of Tradescantia inflorescences. Micronuclei increase in pollen was found only in natural mineral water stored for 2 months. DNA-damaging activity was found in many of the natural and carbonated water samples. Spring water was negative in the plant micronuclei test and the Comet assay, whereas distributed spring water showed DNA-damaging effects, suggesting a possible introduction of genotoxins through the distribution pipelines. GC/MS analysis showed the presence in mineral water of di(2-ethylhexyl)phthalate, a nongenotoxic hepatocarcinogenic plasticizer, after 9 months of storage in PET bottles.

[Genotoxic effects of pesticide-treated vegetable extracts using the Allium cepa chromosome aberration and micronucleus tests]. / Valutazione della genotossicità di frutta e verdura contaminata da pesticidi mediante il test dell'Allium cepa.

The presence of chemical residues in vegetables and fruit is a source of human exposure to toxic and genotoxic chemicals. The mutagenic and carcinogenic action of herbicides, insecticides and fungicides on experimental animals is already known. Several studies have shown that chronic exposure to low levels of pesticides can cause adverse health effects and that many pesticides are mutagenic/carcinogenic. In the present research we monitored concurrently the presence of pesticides and genotoxic compounds extracted from 21 treated vegetables and 8 types of grapes sampled from the markets of a region in Southern Italy. The extracts were analysed for pesticides by gas-chromatography and HPLC, and for genotoxicity with two plant tests in Allium cepa roots: the micronucleus test and the chromosomal aberration test. We found 33 pesticides, some of which are outlawed. Genotoxicity was found in some of the vegetables and grapes tested. Allium cepa tests were sensitive for monitoring genotoxicity in food extracts. The micronucleus test in interphase cells gave much higher mutagenicity than the chromosomal aberration test in anaphase-telophase cells.

The occurrence of cytotoxic Aeromonas hydrophila strains in Italian mineral and thermal waters.

Bacteria of the genus Aeromonas are ubiquitous in aquatic environments, including mineral drinking and thermal waters. Motile species are related to different diseases, mostly gastrointestinal disorders. Criteria for Aeromonas pathogenicity in humans and animals are still unclear and neither is the relationship between production virulence and pathogenicity factors. In the present study, strains of Aeromonas hydrophila, from 61 samples of bottled mineral waters and 23 thermal Italian sources have been isolated and identified by biochemical tests, for toxicity and detection of the aerolysin gene by the Polymerase Chain Reaction (PCR). Six strains were isolated from the mineral waters and were found to be cytotoxic and in possession of the aerolysin gene. For the twelve strains isolated from thermal waters, seven were cytotoxic and eleven contained the aerolysin gene.

Simple method of detecting enteroviruses in contaminated molluscs and sewage by using polymerase chain reaction coupled with a colorimetric microwell detection assay.

The methods normally used for the detection of enteroviruses in environmental samples involve the use of cell cultures, which are expensive and time consuming. The Polymerase Chain Reaction (PCR) is a useful tool for the detection of enteroviruses in several matrixes because primary cell culture is not needed and the increased sensitivity of PCR allows detection of the low numbers of target nucleic acids usually found in environmental samples. A 5-h, user-friendly PCR assay was used to detect enteroviruses in bivalves molluscs (clams) and sewage. Reverse transcription and amplification were performed in a one-step reaction using rTth polymerase. Carryover contamination was prevented with dUTP and uracil N-glycosylase. Detection was performed colorimetrically in a microwell titer plate. This method has greater advantages over conventional methodologies for routinely screening a large number of samples, namely, the rapid acquisition of results and cost effectiveness.

Studies of migration of potentially genotoxic compounds into water stored in pet bottles.

Coloured polyethylene terephthalate (PET) bottles for carbonated beverages were studied for potential migration of genotoxic compounds. A combined approach, using physicochemical methods and a bacterial short-term mutagenicity test (Ames test) was followed. Standard EEC and US FDA tests for total migration of non-volatile migrant compounds into distilled water were performed, together with modified tests, where freeze-drying instead of evaporation of water was used, in order to measure both volatile and non-volatile chemicals. Gas chromatography-mass spectrometry (GC-MS) analysis was performed on these residues. PET bottles filled with naturally carbonated mineral water were also used for long-term total organic carbon (TOC) and mutagenicity migration studies (up to 6 months' storage). Total migration results for PET bottles were within the EEC and US FDA limits. The use of freeze-drying for the elimination of water enabled much higher total migration data (higher than the limits) to be revealed. Some potentially genotoxic compounds (acetaldehyde, dimethyl terephthalate, terephthalic acid) were identified in these migrant compounds by GC-MS analysis. The tests for TOC migration gave a maximum value after 2 wk storage and the mutagenicity tests on non-volatile migrant compounds gave always negative results.

In vitro genotoxicity of dimethyl terephthalate.

Dimethyl terephthalate (DMTP), the para configuration of dimethyl phthalate, is one of the basic monomers used in the synthesis of polyethylene terephthalate (PET) plastics. Human exposure to DMTP may primarily occur during the manufacture of PET fibers and films. The mutagenic potential of dimethyl terephthalate was evaluated using a battery of in vitro short-term tests: the Ames test; DNA single-strand break assays in CO60 cells and in primary rat hepatocytes; UDS in HeLa cells; chromosome aberration and micronucleus assays in human peripheral blood lymphocytes; selective DNA amplification in CO60 and in Syrian hamster embryo cells. The results of this battery of in vitro assays clearly show that DMTP is nongenotoxic. By contrast, other authors have found DMTP to be an in vivo clastogenic compound and suggested that the mechanisms involved in these in vivo effects seem to have nothing in common with genotoxicity and are still unknown.

Leaching of mutagens into mineral water from polyethyleneterephthalate bottles.

Polyethyleneterephthalate (PET) was tested as a source of mutagen contamination from bottles used for beverage packaging. PET bottles were filled with mineral water and stored in daylight and in the dark for different periods of time. The water samples were concentrated and the concentrates (non-volatile compounds) tested for mutagenicity with the Ames test (static tests). Total organic carbon (TOC) leaching was determined concurrently. Leaching of mutagens was also studied using dynamic tests; shaking distilled water in PET bottles. New methods were also used to test the leaching potential of both volatile and non-volatile compounds: directly testing the mutagenicity in unconcentrated water stored in PET bottles and growing Salmonella strains directly in the plastic bottles. The results were positive only for the static test, which identified leaching of mutagens after 1 month of storage in PET bottles. This activity was higher after storage in daylight.

[Bacteriological variations in a medio-mineral water bottled in polyethylene terephthalate containers]. / Variazioni batteriche in un'acqua medio-minerale imbottigliata in contenitori di pet.

Containers made with PET (Polyethylene-terephthalate) are currently more and more employed in packing industry, particularly for the storage of mineral waters. The increasing utilization of such containers is due to the intrinsic properties of this polymer, which was shown particularly suitable for making bottles devoted to the storage of gassed drinks. The resistance of the PET to high pressure, hits by falls and top-to-down loads indeed makes PET bottles unbreakable; their gas-tightness warrants a good gas maintenance during the storage period; the high transparency of the PET allows a good vision of the contents; the light weight of the bottles and the low temperatures required for their production allow a remarkable saving of energy; lastly PET bottles can easily be recycled. Previous microbiological investigations carried out on several mineral waters bottled in glass bottles and non-PET plastic (i.e. PVC) bottles, had shown higher microbial counts in the water samples stored in plastic bottles. In the present work we have studied the growth rates of the bacterial flora in a sample of non gassed medio-mineral water stored in PET bottles, with respect to a control of the same kind of water, stored in glass bottles. Before using, both PET and glass bottles were washed with 5% Desogen, and sterilized by 100 vol. hydrogen peroxide. After the appropriate sterility checks, the bottles were filled directly from the spring with a non gassed medio-mineral water, and then subdivided into four groups, each consisting of the same number of bottles. A the time of bottling, a bacterial count on such water samples at 20 degrees C and 37 degrees C was performed, in order to establish the "zero" value. One of the two groups of PET bottles, and one of the two groups of glass bottles were stored in the darkness, while the other two groups were stored in the light. Afterwards, one bottle from each group was drawn once a week over one year, in order to measure the bacterial concentration in the water. 20 degrees C and 37 degrees C bacterial counts were done after plating in standard agar. The findings of our study show that the bacterial count in both PET- and glass-stored water increases first, but decreases afterwards, though in a non-uniform rate. Further, light exposure weakly don't affects significantly the bacterial growth, even though, in the average, the bacterial count is lower in the bottles exposed to the light.(ABSTRACT TRUNCATED AT 400 WORDS)

An epidemiological study of atopy in children.

A sample of 930 children, between 9 years and 15 years old from six Italian towns, were studied. Their family history of allergy, personal allergy and medical history, current allergy symptoms, exposure to environmental allergens, feeding as infants, and their parents' occupations were recorded. They were skin-prick tested (SPT) with a range of common allergens. A subject was defined as atopic if at least one SPT caused a weal greater than 3 mm diameter. The association between the recorded information and atopy was investigated by logistic multiple regression. Atopy was positively associated with: high density housing; medium or high exposure to environmental allergens; a history of rhinitis, asthma or atopic eczema; male gender; and a history of breastfeeding. It was independent of infectious diseases, vaccinations and operations, social class and family history. Thus, there was no evidence of a genetic factor in atopy, other than sex.