monitoring of the non-specific metabolites of organophosphate pesticide in amniotic fluid of...

S156 Abstracts / Toxicology Letters 189S (2009) S57–S273

which progressed to maximal severity of seizures and fascicula-tions within 60 min. At this time interval, biomarkers of cerebralROS generation (F2-isoprostanes, F2-IsoPs; F4-neuroprostanes, F4-NeuroPs) were elevated to 142% and 225% of control, respectively.In addition, elevation of citrulline (a marker of RNS) by a 4- to6-fold resulted in significant (p < 0.001) depletion of high-energyphosphates (ATP and phosphocreatine). Quantitative morphome-tric analysis of pyramidal neurons of the hippocampal CA1 region1 h post DFP exposure revealed significant decreases in dendriticlengths and spine density to 30% and 58% of control, respec-tively. Pretreatment of rats, 30 min prior to DFP exposure, withmemantine (18 mg/kg, s.c.) in combination with atropine sul-fate (16 mg/kg, s.c.), markedly attenuated DFP-induced increasesin F2-IsoPs, F4-NeuroPs, citrulline and declines in high-energyphosphates and their metabolites. Furthermore, memantine pre-treatment also protected neurons in the CA1 hippocampal areafrom DFP-induced dendritic degeneration. These results suggestthat memantine provides neuroprotection by suppressing cerebraloxidative/nitrosative stress involving multiple mechanisms (sup-ported by NINDS NS057223).

doi:10.1016/j.toxlet.2009.06.761

Y19Cotinine biomarker validation of self reported smoking statusamong Greek adolescents: The HELENA study

Constantine Vardavas 1,∗, Manolis Tzatzarakis 1, Maria Plada 1,Aristeidis Tsatsakis 2, Alina Papadaki 2, Wim Saris 3, LuisMoreno 4, Anthony Kafatos 2

1 University of Crete, Department of Social Medicine, Faculty ofMedicine, Heraklion, Crete, Greece, 2 University of Crete, Departmentof Forensic Sciences & Toxicology, Medical School, Heraklion, Greece,3 University of Maastricht, Nutrition and Toxicology Research inMaastricht (NUTRIM), Department of Human Biology, Maastricht,Netherlands, 4 Universidad de Zaragoza, Escuela Universitaria deCiencias de la Salud, Saragoza, Spain

Question: A major problem in evaluating active smoking status,especially among adolescents, is its accurate verification, as adoles-cents usually underreport their current smoking status. We aimedat validating their self reported smoking status among adolescentsof the HELENA, Heraklion cohort using serum cotinine.

Methods: As part of the European HELENA study, 400 adoles-cents from Crete, Greece were randomly selected and contactedout of which 341 agreed to participate, of which 311 were withinthe valid age range (77.8% response rate) and questionnaire datawas collected through personal interviews. Blood samples werecollected from a random sample of 111 adolescents (of the 142randomly selected, 78.2%) of which 106 were analysed for theircotinine concentrations with GC/MS.

Results: In addition to the adolescents who were self reportedcurrent smokers (at least one cigarette during the last month), thecotinine analysis revealed another four participants with cotininelevels above that of the cut-off (cotinine>15 ng/ml), with serumcotinine levels of 28.86, 38.54, 132.61, and 290.97 ng/ml, respec-tively, despite them having reported that they were non-smokers.Moreover, the analysis indicated that 4% of the non-smokingadolescents were actually smokers, despite their self reported non-smoking status.

Conclusions: The applied questionnaire procedure successfullyidentified 96% of the active smokers, with only a small percentageof smokers found to report otherwise. This information provides

support of the procedures validity in assessing the lifestyle habitsof the adolescents participating in the HELENA study.

doi:10.1016/j.toxlet.2009.06.762

Y20Monitoring of the non-specific metabolites of organophosphatepesticide in amniotic fluid of pregnant women in the region ofCrete

Manolis Tzatzarakis 1,∗, Dimitrios Koutroulakis 2, Stavros Sifakis 2,Matthaios Kavalakis 1, Maria Tutudaki 1, Nikitas Mantas 2,Ourania Koukoura 2, Manolis Kokkinakis 1, Ioannis Mataliotakis 2,Aristidis Tsatsakis 2

1 University of Crete, Laboratory of Toxicology, Medical School,Heraklion, Greece, 2 University Hospital, Obstetrics and Gynecology,Fetal Maternal Unit, Heraklion, Greece

Purpose: The levels of the non-specific metabolites of organophos-phate pesticides (DAPs) in amniotic fluid were determined in thepresent study, in order to assess exposure of pregnant womento organophosphate pesticides. The examined metabolites weredimethylphosphate (DMP), diethylphosphate (DEP), diethylthio-phosphate (DETP), and diethyldithiophosphate (DEDTP). Thesemetabolites are polar water soluble substances, can be detected inseveral biological samples like urine, blood, meconium, hair andare considered as biomarkers of exposure to a great number oforganophosphate pesticides.

Method: During 2007–2008, 200 amniotic samples were col-lected from pregnant women, permanent residents of Crete. Thecollection of amniotic samples was performed at 17–24 weekof pregnancy, during amniocentesis for fetal karyotype investi-gation. An amount of 5 ml of amniotic fluid was collected andstored at −20 ◦C until analysis. DAPs were extracted from amni-otic fluid by liquid–liquid extraction, derivitized and analyzed bygas chromatography–mass spectrometry (GC–MS).

Results: The mean values (±SD) and the range of DMP, DEP,DETP and DEDTP in positive amniotic samples were 14.63 ±38.27 ng/ml (0.22–252.01), 8.65 ± 29.54 ng/ml (0.17–356.85),12.60 ± 119.95 ng/ml (0.12–1449.87), and 0.34 ± 0.67 ng/ml(0.05–5.13), respectively. DEP was the metabolite with thehigher frequency of detection (90.5%) followed by DETP (74.0%),DMP (60.0%), and DEDTP (39.5%).

Conclusion: The high frequency of detection of all DAPs is prob-ably an indication of the widespread use of organophosphatepesticides in the region of Crete. The correlation of the detectedlevels of pesticide metabolites in amniotic fluid with a varietyof parameters of pregnancy outcome including congenital abnor-malities, pregnancy-associated complications (intrauterine growthrestriction, preterm labor, and stillbirth), birth weight, and neona-tal morbidity may be valuable in prenatal screening and pregnancysurveillance. Further studies and more data are required before theaforementioned goal is achieved.

doi:10.1016/j.toxlet.2009.06.763