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Pesticides in air and in precipitation and effects on plant communities Appendix IVPrinciples for analysis of water samples collected in 1998:The water sample is passed through a solid phase extraction column (SPE-column) and the retained compounds are eluted with an organic solvent. The solvent is evaporated in a vacuum centrifuge and the residue is redissolved in a suitable solvent for HPLC. The compounds are separated on a HPLC column and detected in a mass spectrometer after electrospray ionisation (ESI) Chemicals, solutions and standards: Methanol, HPLC grade ( Rathburn ) Stock solutions, standards and internal standards: Stock solutions are made by weighing of 50 mg standard, which is dissolved in 50 ml acetonitril. Stock solutions are stored in refrigerator and are registered in a stock solution record. Standards are made by dilution in acetonitril and final dilution in A-eluent or in a solvent comparable to HPLC-start conditions. Standard concentrations should cover the concentration range for the samples after preconcentration, for instance 10, 50 ,100 and 200 m g/L. 13C- or 2H-labelled internal standards are recommended for adding to the samples before preconcentration to checking recovery. HPLC-Eluents: For analysis of isoproturon the following gradient system is used: A1-eluent: methanol/10 mM Ammonium acetate in milliQ-water 10/990, For phenoxyalkanoic acid herbicides and other acidic compounds: A1-eluent: methanol/ 20 mM acetic acid 10/90, The eluents are filtered through 0.2 m m millipore filter (type Fluropore, FG), Millipore. The LC-MS system was composed by a Hewlett-Packard LC-MSD 1100 system with a binary gradient pump system. The HPLC-column was a Hypersil BDS 250 x 2.1 mm column. The same linear gradient profile were used for both acidic compounds and isoproturon compounds with different eluents, as described above:
Mass spectrometry: Phenoxyalkanoic acid herbicides, bentazon and – metabolites together with DNOC was detected by MS with electrospray inlet (ESI), negative mode and selected ion monitoring (SIM):
Isoproturon and degradation products were detected by MS with electrospray inlet (ESI),positive mode and selected ion monitoring (SIM):
Sample preparation: 50 ml preservation solution is added to the sample flasks before they are placed on the locations. The samples are stored at 4 oC if sample preparation takes places within a few days. Otherwise the samples are frozen at –18 oC until sample preparation will take place. The sample volume is registered and eventually internal standard is added The sample is passed through a glass fibre prefilter and a glass fibre filter and the filters are rinsed with 5 ml methanol. The filtrate or 250 ml water sample ( + 1,25 ml Methanol ) (+ 6,25 ml preservation solution to recovery and blanks) are added to Porapak Rdx solid phase extraction columns, which are preconditioned with 10 ml acetonitril, 10 ml Methanol and 20 ml Milli-Q water. The water sample is applied under vacuum pressure with a flow about 10-20 ml/min. The columns must never run dry during the application. Afterwards the columns are air dried with vacuum suction in further 20 min. The columns are eluted with 5 ml methanol/acetonitril 1/1 in the following way: 1 ml is passed through the column, 2 minutes standing and the rest of the solvent is passing without vacuum into a vial with 50 m L propylene glycol as keeper. The sample volume is reduced to 50 m l in a vacuum centrifuge. The propylene glycol residue is diluted with 500 µl 10% methanol and is ready for HPLC-analysis.
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