Development of a new microextraction method and on-line derivatization coupled with GC-MS for analyzing of five metabolites of synthetic pyrethroids in urine samples

摘要

Sample preparation step is usually the most expensive and time consuming part of laboratory work during the assessment of exposure to environmental chemicals. In order to reduce expenses and/or increase throughput of analytical methods much efforts are made for miniaturization and automatization. The main goal of this work was to simplify and automate the sample preparation technique before GC-MS analysis of urinary biomarkers of pyrethroid exposure. A novel microextraction by packed sorbent (MEPS) method coupled with gas chromatography-mass spectrometry (GC-MS) was developed for determination of five urinary metabolites of synthetic pyrethroids: cis-2,2-dimethyl-3-(2-chloro-3,3,3-trifluoro-1-propenyl)-cyclopropanecarboxylic acid, cis/trans-3-(2,2-dichlorovinyl)-2,2-dimethylcyclopropane-1-carboxylic acids, cis-(2,2-dibromovinyl)-2,2-dimethylcyclopropane-1-carboxylic acid and 3-phenoxybenzoic acid. MEPS is a miniaturized solid phase extraction (SPE) utilizing a manually operated semiautomatic syringe equipped with a needle combined with SPE sorbent. Several significant factors affecting MEPS performance like: sample pH, type of extraction packing, type and volume of washing solvent, number of draw-eject cycles, volume and type of elution solvent were optimized. MEPS was performed using C18 solid phase, which was conditioned with methanol (4x50μL) and 2% formic acid in water (3x20uL). Subsequently the enzymatically hydrolyzed urine sample (3x100μL) was loaded and the bed was washed with 30% methanol in water (3x50uL). Then the sorbent was dried under vacuum and finally analytes were simultaneously derivatized and eluted with the mixture of 1% 1,1,1,3,3,3-hexafluoroisopropanol and 2% diisopropylcarbodiimide in n-hexane (2x40uL). Forty microliters of the extract were injected into GC-MS system using large volume injection mode (LVI). Optimized method was then validated and LODs in the range of 0.06 - 0.42 ng mL-1, correlation coefficient above 0.990 and precision below 17% were obtained for all analytes. The proposed method is very fast, simple and environment friendly. In comparison with reference extraction method (liquid-liquid extraction) the volume of sample and organic solvents and time of extraction procedure were significantly decreased. The new approach can be used to routine monitoring studies of evaluation of human exposure to synthetic pyrethroids.