Anal Bioanal Chem (2006) 385: 1444–1456 DOI 10.1007/s00216-006-0569-0 ORIGINAL PAPER Syrago-Styliani E. Petropoulou . Anthony Tsarbopoulos . Panayotis A. Siskos Determination of carbofuran, carbaryl and their main metabolites in plasma samples of agricultural populations using gas chromatography–tandem mass spectrometry Received: 2 March 2006 / Revised: 17 May 2006 / Accepted: 18 May 2006 / Published online: 25 July 2006 # Springer-Verlag 2006 Abstract A gas chromatography–tandem mass spectro- metric (GC-MS/MS) method has been developed for the determination of carbofuran (2,3-dihydro-2,2-dimethyl- benzofuran-7-yl methylcarbamate), carbaryl (1-naphthyl- N-methylcarbamate) and their main metabolites in human blood plasma. Optimization of the isolation of the com- pounds from plasma matrix included the precipitation, denaturation and digestion of plasma proteins. Derivatiza- tion was achieved by the use of trifluoroacetic acid anhydride and was optimized for temperature, time and volume of derivatization agent. In the proposed method, a mild precipitation technique was applied using β-mercap- toethanol and ascorbic acid in combination with solid- phase extraction technique using Oasis HLB (Hydrophobic Lipophilic Balance) cartridges for further clean up of samples. Carbamate linkage was not hydrolyzed to its phenol product, but both carbamate phenol and ketones were transformed into trifluoroacetyl derivatives in order to become volatile compounds and were determined using tandem mass spectrometry. The linearity of the method was shown for nine concentrations in the range of 0.50–250 ng mL -1 in fortified plasma aliquots. Limits of detection (LODs) for all compounds ranged from 0.015–0.151 ng mL -1 . Inter-day and intra-day assays (RSD) for all com- pounds, at three concentration levels of 2.5, 25 and 100 ng mL -1 (n=3) in fortified plasma samples were less than 18%. Accuracy (%E r ) was calculated at three concentration levels, 8, 80 and 160 ng mL -1 (n=3), and ranged from -12.0 to 15.0%. Matrix effect was evaluated so mean recoveries were calculated for all compounds and ranged from 81–107%. Specificity for the use of this method to biological monitoring studies was achieved including four main metabolites of CF, 1-naphthol and 2-naphthol from the naphthalene metabolism pathways, and both the parent compound of carbofuran and carbaryl. The proposed method was applied to plasma samples of pesticide users. Keywords N-methylcarbamates . Tandem mass spectrometry . Biological monitoring . Biomarkers of exposure . Human plasma samples Introduction One of the major types of worldwide pollutants is the pesticides. Pesticide use has increased over the last two decades in agriculture, but also in households and in other areas such as golf courses. This broad use of pesticides, among other chemicals, may be necessitated by modern civilization, but it is also of concern to hygienists, epidemiologists, physicians and other health officials due to likely adverse health effects caused by the exposure of humans to these chemicals. Pesticides include a variety of compounds such as insecticides, nematocides, and herbi- cides. Since the middle of last century when pesticide use started, humans have gained many benefits from them, but over the years have forgotten that they were deliberately developed to harm living species [1, 2]. Preventive medicine considers that it is extremely important to determine residues of pesticides and their mixtures in food, air and water to estimate the magnitude of human exposure. According to the latest report, released from the U.S. Department of Health and Human Services of the Centers for Disease Control and Prevention (CDC) in Atlanta, Georgia, concentrations of environmental chemi- S.-S. E. Petropoulou . A. Tsarbopoulos Bioanalytical Laboratory, GAIA Research Center, The Goulandris Natural History Museum, 100 Othonos str., 145 62 Kifissia, Greece S.-S. E. Petropoulou . P. A. Siskos (*) Analytical Chemistry Laboratory, Department of Chemistry, National and Kapodistrian University of Athens, Panepistimioupoli, 15771 Zographos, Athens, Greece e-mail: siskos@chem.uoa.gr Tel.: +30-210-7274311 Fax: +30-210-7274750 A. Tsarbopoulos Laboratory of Instrumental Analysis, Department of Pharmacy, University of Patras, 265 04 Rio, Greece