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Identifier 000347424
Title Δείκτες χρόνιας έκθεσης σε φυτοφάρμακα : μελέτη εναπόθεσης οργανοφωσφορικών, καρβαμιδικών και οργανοχλωριομένων εντομοκτόνων στις τρίχες
Author Τουτουδάκη, Μαρία
Thesis advisor Τσατσάκης, Αριστείδης
Abstract The main aim of the present study was the investigation of the possibility to use hair analysis as a tool for the assessment of chronic and low level exposure to pesticides, employing in vitro and in vivo experiments. The studied substances belong to the families of organophosphates, carbamates and organochlorines. A literature review revealed that no such studies have been conducted in the past. The disposition of organochlorine pollutants and pesticides in hair was also studied. The organochlorine pesticides under investigation are no longer used and the PCBs are byproducts of industrial activities. Exposure of the organisms to the aforementioned substances occurs either through the diet or from the environment. The first and crucial step was the development and validation of the analytical methods for the detection and quantification of the analytes of interest in a difficult matrix like hair. The pesticide diazinon was the model compound for the study of organophosphates incorporation in hair. Sample preparation included methanolic extraction of the pesticide from the hair matrix, at 37 0C overnight, followed by liquid-liquid extraction with ethyl acetate, evaporation of the solvent under nitrogen, resuspension of the residue in 50 μl of methanol and analysis by GC-MS or GC-NPD. GC-MS analysis was performed in full scan and SIM mode. Analysis in SIM mode offered much better selectivity and sensitivity. Two ionisation modes were also utilised; electronic ionisation and negative chemical ionisation with methane. Chemical ionisation mode offered much lower detection limits than the electronic ionisation mode. The disposition of diazinon in hair from the environment was studied using fortified hair samples. Three parameters that would probably affect the detected concentration of diazinon in hair were studied; the concentration of diazinon in the exposure media, the time of exposure and the colour of the hair. Initially hair samples coming from the same person were immersed in aqueous solutions diazinon at different concentrations, for varying time periods. The samples were analysed by EI GC-MS following a decontamination rinse with water, drying, pulverization, methanolic extraction in an ultrasonic bath, liquid-liquid extraction with ethyl acetate, evaporation of the solvent and resuspension in 50 μl of methanol. It was found that the concentration of the pesticide detected in hair was proportional to the concentration of the exposure media and to the time of exposure. The same experiment was repeated using hair of different shades brown, coming from different people. The hair samples were immersed in aqueous solution of the pesticide at the same concentration and for the same time period. Our results indicated that in the case of external disposition of the pesticide in hair, the concentration of the pesticide is not related to the melanin content and hence the colour of the hair. On the other hand, other factors like the lipid content of the hair or the diameter of the hair fiber may play a more important role in the amount of pesticide detected. The next step was the study of incorporation of diazinon in hair in vivo. Rats and rabbits were used as our experimental animals. It is obvious by the difference in the literature reported LD50 values that rabbits are more susceptible to diazinon toxicity than rats. Only a very small fraction of the administered doses was detected in the hair of the animals. The concentration of the pesticide detected in the hair of the rats was much higher than that detected in the hair of the rabbits even though the rabbits received higher doses than the rats. This may be explained by the different metabolic profile of the two animal species. Two methods were used for the detection of methomyl in the hair of experimental animals. The screening of the samples was performed by ELISA while the results were confirmed by HPLC. Even though ELISA gave acceptable results with methomyl there was significant background noise. Analysis with GC-MS was not possible due to the instability of the molecule The exposure of the population to certain organochlorine compounds through the diet was also studied. The compounds studied were two organochlorine pesticides, DDT and lindane, and the following PCB congeners: PCB 28, 52, 99, 101, 118, 138, 149, 153, 156, 170, 180 and 187. A comparative study of the population exposure of three countries, Greece, Romania and Belgium was also conducted. The hair matrix was dissolved by incubation in HCl, and the samples were initially liquid-liquid extracted in a mixture of hexane: dichloromethane (4:1) and subsequently cleaned by SPE. Finally they were analysed by GCECD and GC-MS for confirmation. The Greek samples were found to be the most contaminated ones by organochlorine pesticides while they carried the smallest PCBs burden. Belgian samples were more contaminated by PCBs but carried a smaller pesticide burden while the samples from Romania contained less pesticide than the Greek ones but more than the Belgian ones. Three sensitive and selective analytical methods were developed and validated during the study of incorporation of diazinon in hair. They were based on GC-MS and GC-NPD detection and they were successful in detecting and quantitating diazinon in hair in the pg/mg level. It was confirmed by the in vivo studies in white rats and rabbits that diazinon may be detected in hair once it enters blood circulation, at concentrations proportional to the administered dose. When the detected pesticide concentrations in hair were compared, it became evident that metabolic species differences influence the quantity of non metabolized pesticide circulating in the blood and hence the concentration detected in hair. The in vitro studies indicated that diazinon is deposited in hair not only in the case of exposure through the blood stream, but also in the case of external exposure. The concentration of the exposure media as well as the time of exposure does play a role in the detected pesticide concentration. On other hand the colour of the hair is not such a crucial parameter as it happens when substances are incorporated in the hair shaft from the sweat and sebum that bathe the shaft during the hair growth. The incorporation of methomyl in hair was studied using rabbits as experimental animals. It was confirmed that once it enters the blood stream it is detected in hair. Segmental hair analysis that was performed indicated that methomyl binds to specific sites and does not migrate along the hair shaft. This can be used in human exposure studies to obtain an accurate record of exposure, bearing in mind that human head hair grows approximately 1 cm/month.. The main conclusion reached by the study of the organochlorine compounds was that contamination of the samples with the aforementioned substances was universal and that organochlorine concentration detected in the samples was related to the origin of the examined population. The Greek and occupationally exposed samples carried more DDTs, and less PCBs. The HCHs burden was similar for all the samples. In summary, during the present study sensitive and selective methods for the detection of minute quantities of pesticides in complex matrices were developed and validated, it was confirmed that the metabolic profile of an organism influences the concentration of pesticides detected in hair, and when external contamination of hair was studied, it was deduced that the concentration of pesticide in the exposure media as well as the time of exposure influences the concentration of the detected pesticides in hair. Segmental hair analysis that was performed in the case of methomyl and may be performed in human hair may give an accurate record of past exposure to pesticides. As a future development, hair disposition of certain characteristic metabolites of pesticides should be studied. Also it would be interesting to examine hair disposition of other commonly used pesticides. This is a potentially valuable additional tool for the assessment of chronic exposure to pesticides.
Physical description 115 σ. : πιν. ; 30 εκ.
Language Greek
Subject Pesticides adverse effects
Issue date 2007-07-26
Collection   Faculty/Department--School of Medicine--Department of Medicine--Doctoral theses
  Type of Work--Doctoral theses
Permanent Link https://elocus.lib.uoc.gr//dlib/f/8/6/metadata-dlib-b786d84b007f223fdc848b474811aba0_1247125888.tkl Bookmark and Share
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