S118 Abstracts / Toxicology Letters 189S (2009) S57–S273 Liver samples were taken, in which parameters of the glutathione redox system, namely reduced glutathione (GSH) concentration and glutathione-peroxidase (GSHPx) activity were measured. The free radical generating ability of T-2 toxin was measured by a direct reactive oxygen metabolites (dROMs) test, and also a meta-stable end product of the lipid peroxidation processes, malondialde- hyde (MDA) concentration was determined. From the 1st week lower feed consumption and also weight gain was recorded in T-2 toxin treated group, which resulted significantly lower live weight at the end of experiment. T-2 toxin treatment increased the amount/activity of glutathione redox system during the 1st week, which is shown by the significantly elevated GSH concentration and GSHPx activity of liver. At 2nd week the T-2 toxin treatment resulted decrease in GSH concentration and GSHPx activity of liver compared to control. Although the reactive oxygen metabolite con- centration was elevated in T-2 toxin treated group during the first 2 weeks, no significant changes were found in the MDA concen- tration in liver of juvenile carps. Results show that the biological antioxidant system was able to eliminate the harmful peroxidative effect of T-2 toxin in common carps. doi:10.1016/j.toxlet.2009.06.408 B08 Intracellular glutathione level as indicator for cellular stress: Method development Katja Schmalbach 1,∗ , Sabrina Jäger 2 , Leane Lehmann 1 1 University Würzburg, Institute for Pharmacy and Food Chemistry, Würzburg, Germany, 2 University Karlsruhe, Institute of Applied Biosciences, Karlsruhe, Germany Glutathione (GSH) biosynthesis is mediated by various transcrip- tion factors activated by cellular stress binding to their responsive elements in the promoter region of the key enzyme of GSH biosynthesis, -glutamylcysteine synthetase (GCS). Therefore, the intracellular GSH level is a sensitive indicator for cellular stress. Thus, the aim of this work was to develop a fast and practicable method for measuring relative GSH levels in living cells. The method is based on the reaction of the non-fluorescent monochlorobimane (MCB) and GSH forming a fluorescent product. The measurement of intracellular fluorescence intensity (FI) was performed by means of flow cytometry in cultured V79 cells. The modal value of FI was used to quantify the fluorescence signal. The method was optimized with respect to (i) stained cell density (0.7–13 mio cells/ml), (ii) staining time (15–90min), and (iii) cells to be analyzed by flow cytometry (2.5 × 10 4 –2.5 × 10 5 ). The FI was not affected by any of the parame- ters analyzed. Therefore, (i) time-consuming determination of cell number prior to staining could be omitted, (ii) an individual exper- imental design was possible, and (iii) a measuring time less than 60 s per sample could be realized. Thus, a multiwell application is possible. The method was calibrated using absolute GSH level determined by the method of Ellman. With this calibrated flow cytometric method to determine the total intracellular GSH content a fast and practicable indicator test for cellular stress was established. Supported by Deutsche Forschungsgemeinschaft grant Le 13/29-7. doi:10.1016/j.toxlet.2009.06.409 B09 An investigation of the effects of addictive drugs on oxidative stress in humans Ahmet Sayal 1,∗ , Cemal Akay 1 , Bu˘ gra Soykut 1 , Ays ¸ e Eken 1 , Onur Erdem 1 , Ahmet Aydin 1 , Mustafa Kemal Cetin 2 , Nesrin Dilbaz 2 1 Gulhane Military Medical Academy, Pharmaceutical Toxicology, Ankara, Turkey, 2 Numune Research and Education Hospital, AMATEM, Ankara, Turkey Addictive drugs can cause toxicity and permanent damages in brain, nervous system and other systems. The aim of the present study was to determine the toxicological effects of opiates, one of the most widely used and the most dangerous addictive drugs, mea- suring the oxidative stress status. Antioxidant enzyme activities and malondialdehyde (MDA) levels were measured as oxidative stress parameters. Study group was composed of individuals diag- nosed as heroin addictives, while people with no drug addiction were classified as control group. When the study and control group (288.17 ± 68.12 nmol/ml MDA level and 371.18 ± 86.25 U/ml SOD activity) was compared each other, we observed a statistically significant increase in MDA levels (323.42 ± 45.73 nmol/ml) and decrease in SOD activity (321.80 ± 85.07 U/ml) for the addictive group (p < 0.05). We also investigated the effects of smoking and compared to addictive smoker group and control smokers finding a significant increase in MDA levels and a significant decrease in SOD levels due to the effects of opiates. It was stated that, using opiates intravenously or nasally had no effects on the activity of antioxidant enzymes. However, an important increase in lipid per- oxidation was observed for the individuals using opiates over 10 years. It was concluded from the results of the study that the opiates seemed to have toxicological effects on cellular damage biomark- ers such as oxidative stress enzymes and lipid peroxidation. On this account, it can be suggested that antioxidant supplement besides pharmacological and psychiatric approaches would inhibit the tox- icological effects of addictive drugs and increase the life quality of patients during and after the therapy. doi:10.1016/j.toxlet.2009.06.410 B10 Differences in metabolism and GSH conjugation of 2-methyl- 1,4-naphthoquinone in vivo and in vitro suggest oversensitivity for oxidative stress-induced toxicity in vitro Ursula Lutz 1,∗ , Amy V. Pointon 2 , Joel D. Parry 3 , Lars Oliver Klotz 4 , Timothy W. Gant 2 , Werner K. Lutz 1 1 University of Würzburg, Institute of Toxicology, Würzburg, Germany, 2 University of Leicester, Medical Research Council Toxicology Unit, Leicester, United Kingdom, 3 GSK R&D Ltd., Investigative Preclinical Toxicology, Safety Assessment, Ware, United Kingdom, 4 Heinrich-Heine-Universität Düsseldorf, Institut für Physiologische Chemie I, Düsseldorf, Germany Menadione (2-methyl-1,4-naphthoquinone; CAS-RN 58-27-5) is often used as a model compound to investigate quinone toxicity resulting from (i) oxidative stress by ROS associated with redox cycling and (ii) arylation of cellular thiols. Glutathione represents a cellular defence against both mechanisms by providing reducing equivalents for inactivating peroxides and by acting as a conju- gation partner to protect more critical nucleophilic sites. To date redox action and metabolism with menadione have been studied