Advances in the sample preparation techniques of human body fluids in toxicological analysis

S210

Abstracts / Toxicology Letters 205S (2011) S180–S300

triptyline, imipramine, amitriptyline, doxepin, dezypramine, and clomipramine (as internal standard). For separation and detection of analytes the HPLC-DAD system was used with C-18 column (thermostated to 50 ◦ C) and with phosphoric buffer (pH 2.36, with 0.1% diethylamine)/acetonitrile (1:1, v/v) as the mobile phase. The MAE technique was optimized in terms of kind of extraction solvent (four different mixtures) and temperature (three levels). The optimum extraction conditions were found to be: a mixture of n-hexane/isoamyl alcohol (99:1, v/v) as the extraction solvent, temperature of 60 ◦ C and irradiation for 3 min (2 min ramping temperature and then holding a steady temperature for 1 min). On the basis of validation study limit of quantification, precision (RSD) and recovery of the method developed were found in the ranges (depending on an analyte): 0.04–0.15 ␮g/mL, 1.57–4.34% and 94–105%, respectively. Validation parameters indicate that the extraction procedure used is reliable and sensitive the quantitative analysis of the examined drugs in human serum. Therefore, the method developed may be recommended for clinical and forensic laboratories for preparation of serum samples for the determination of TCADs. Meanwhile, the usefulness of the MAE technique in the field of forensic toxicology is still tested in our laboratory. doi:10.1016/j.toxlet.2011.05.721

P2098 Advances in the sample preparation techniques of human body fluids in toxicological analysis 1 , A. Garbacik 1 , K. ´ R. Wietecha-Posłuszny 1,∗ , M. Wozniakiewicz Madej 2 , P. Ko´scielniak 1 1

Faculty of Chemistry, Laboratory for Forensic Chemistry, Jagiellonian University, Krakow, Poland, 2 Faculty of Chemistry, Toxicological and Pharmaceutical Analysis Group, Jagiellonian University, Krakow, Poland The aim of this work is to present the development of novel sample preparation techniques recently introduced in toxicological analysis. Sample preparation is a persistent challenge in the field of forensic analysis of biological materials. Present demands are focused on limitation of the sample volume, high selectivity and efficiency, environmental friendliness, and reduction of time and cost. All these demands are fulfilled by such techniques as microwave-assisted extraction (MAE), cloud-point extraction (CPE) and microextraction by packed sorbent (MEPS). All of them are developed at our laboratory (Laboratory for Forensic Chemistry) and successfully applied to isolation of psychotropic drugs from different biological matrices. Extract are analyzed using the HPLC-DAD system. Application of the MAE technique is exemplified on the extraction of tricyclic antidepressants (TCADs) from human blood using a mixture of n-hexane/isoamyl alcohol (99:1, v/v) as the extraction solvent at 60 ◦ C. The MEPS technique (with bins containing C8-SCX sorbent) was used to isolate TCADs from human serum. The CPE technique was exploited to extraction of desipramine, promazine and chlorpromazine form human plasma. For CPE conditions, the non-ionic surfactant Triton X-114 as the extraction medium was used and the drugs were back extracted from the micelle-rich phase into n-hexane/isoamyl alcohol (99:1, v/v). For all tested drugs and materials, the extraction efficiency were ranged from 45 to 100%. The best results in terms of extraction recovery (80–100%) and repeatability (from 5 to 10%) were obtained for microwave-assisted extraction and microextraction by packed sorbent. doi:10.1016/j.toxlet.2011.05.722

Mixtures P2099 Interpreting biomonitoring data for di-n-butyl and di-(2-ethylhexyl) phthalate metabolites in urine using a physiologically based pharmacokinetic model and reverse dosimetry: Estimation of cumulative in utero exposure J. Campbell 1,∗ , R. Clewell 2 , M. Andersen 3 , H. Clewell 1 1

Center for Human Health Assessment, The Hamner Institutes for Health Sciences, Research Triangle Park, USA, 2 Institute for Chemical Safety Sciences, The Hamner Institutes for Health Sciences, Research Triangle Park, USA, 3 Institute for Chemical Safety Sciences, The Hamner Institutes, Research Triangle Park, USA Purpose: At high doses, several phthalate esters, including di-nbutyl and di-2-ethylhexyl phthalate (DBP and DEHP respectively), disrupt normal male reproductive development in rats. Biomonitoring results confirm phthalate exposure in humans of all ages based on serum, urine, cord blood and breast milk measurements from representative samplings in the U.S. and Europe. We used PBPK modeling to examine the correlation between the biomonitoring data and environmental exposure. Methods: Human PBPK models for DBP and DEHP and their primary metabolites – mono-n-butyl and mono-2ethylhexyl phthalate (MBP and MEHP) – during pregnancy were developed from published rodent PBPK models. To estimate daily intake of DBP and DEHP from data on population based measures of urinary metabolite levels, Monte Carlo analyses were performed to generate an inverted distribution between urine concentration and steady state exposure. Results: The predicted distribution of daily intake for DBP in the U.S. (NHANES III) had a median of 0.65 ␮g/kg/day with a 95 percentile of 5.0 ␮g/kg/day. For DEHP the estimated daily intakes were higher median of 1.86 ␮g/kg/day and a 95 percentile of 20.4 ␮g/kg/day. Applying forward dosimetry with the pregnancy PBPK model, the 95 percentile exposure concentration resulted in an average fetal plasma concentration of 6.77 ␮g/L for MBP and 1.4 ␮g/L for MEHP. These values can be compared to an average concentration of 1680 ␮g/L for MBP and 71.2 ␮g/L for MEHP in rat fetal plasma, simulated from the points of departure of 30 and 5 mg/kg, respectively, for developmental effects. doi:10.1016/j.toxlet.2011.05.723

P2100 Serum liver enzyme levels in Wistar rats 28 days orally exposed to the mixture of BDE209 and cadmium M. Curcic 1,∗ , S. Stankovic 2 , V. Jacevic 3 , S. Jankovic 4 , K. Durgo 5 , V. Milovanovic 6 , S. Vucinic 7 , B. Antonijevic 1 1

Department of Toxicology, Faculty of Pharmacy, Belgrade, Serbia, Department of Medical Biochemistry, Faculty of Pharmacy, Belgrade, Serbia, 3 Poison Control Center, Military Medical Academy, Belgrade, Serbia, 4 Institute of Meat Hygiene and Technology, Belgrade, Serbia, 5 Faculty of Food Technology and Biotechnology, Zagreb, Croatia, 6 Serbian Chemical Agency, Belgrade, Serbia, 7 Clinic of Emergency and Clinical Toxicology, National Poison Control Centre, Belgrade, Serbia 2

During the last 10 years, toxicology of mixtures has undergone a remarkable and productive development. Decabromodiphenyl ether (BDE209) and heavy metal cadmium (Cd) are the most abun-