Analytical challenges for the analysis of biocides in aqueous and solid environmental matrices Arne Wick, Kathrin Broeder, Michael Schluesener, Thomas Ternes Federal Institute of Hydrology (BfG), Koblenz, Germany Environmental monitoring of biocides in Europe from prioritisation to measurements Workshop, Berlin, 5 6.11.212
Biocides main groups Disinfectants and general biocidal products e.g. Human hygiene biocidal products Triclosan Preservatives e.g. film preservatives Terbutryn Pest control, e.g. insecticides Permethrine Other biocidal products, e.g. antifouling Cybutryne (Irgarol)
Periphyton Biocides Ecotoxicity Mohr et al. (28), Aquatic Toxicol., 9(2), 19-12 Myriophyllum verticillatum EC 5 = 34 ng/l EC 1 = 1 ng/l Copepods Mohr et al. (29), Environ. Sci. Technol., 43, 6838-6843 Example: Irgarol EC 5 = 21 ng/l proposed AA EQS: 2.5 ng/l Radix balthica EC 5 = 9 ng/l Mohr et al. (28), Aquatic Toxicol., 9(2), 19-12 Oehlmann and Watermann (25), UBA Dessau EC 1 = 32 ng/l
Emission routes of biocides Agricultural pesticides Non-agricultural herbicides and insecticides Material protection: coatings, roof sealings, etc. Biocides in PCPs, cleaning and disinfection agents Sludge WWTP surface water/sediment groundwater/soil adapted from Gerecke et al. (22), 48, 37 315
Analytical requirements Broad compound spectrum with different physico chemical properties positively charged: climbazole, imazalil negatively charged: mecoprop, 2,4 D polar: carbendazim (log K OW = 1.5) non polar: triclocarban (log K OW = 5.1) Complex matrices (surface water, wastewater, sludge, sediment) Sensitivity and reproducibility (e.g. LOQ Irgarol < 2.5 ng/l) High throughput Methods based on LC Tandem MS detection Problem: Matrix effects often severe ion suppression using ESI ESI source
Reduction and compensation of matrix effects Reduction: Sample preparation: Enrichment volume for SPE SPE cartridge SPE cartridge wash step Clean up LC MS/MS: Injection Volume Chromatographic conditions Flow rate to MS (split) Ionization source (APCI vs. ESI) Compensation: Use of labeled surrogate standards (isotope dilution technique) Use of standard addition
Analytical method water samples Water sample (.1 1L) ph adjustment (6.5 +/-.2), Addition of surrogate standard Solid Phase Extraction (SPE) SPE tube Vacuum: ~ 4mbar Filtration (<1µm) SPE Box with HLB 2mg cartridges Drying with N 2 Elution 4 x 2 ml MeOH/Acetone (6/4, v/v) Reconstitution to 1 ml Evaporation to.1 ml HPLC MS/MS
Analytical method solid samples Surrogate Freeze-dried sediment ~ 1g standard spike Extract in GW SPE clean-up, HLB PLE cell PLE extract Pressurized Liquid Extraction (PLE) Measurement with LC-MS/MS ASE Parameter: 8 C/1bar Different extraction solvents
Method performance ~ 45 biocides and pesticides, + benzothiazoles and polar UV filters Isothiazolones(e.g. Benzisothiazolone, BIT, Octylisothiazolone, OIT) Imidazole fungicides (e.g. climbazole, imazalil) Triazole fungicides (e.g. propiconazole, tebuconazole) Carbamate fungicides (e.g. carbendazim, IPBC) Phenyl urea herbicides (e.g. diuron, isoproturon) Triazines (e.g. terbutryn, irgarol) Bacteriocides(e.g. triclosan, triclocarban) Insecticides (e.g. DEET, imidacloprid) Matrix: groundwater, surface water, wastewater, partly for sludge, sediments and soils LOQ surface water :.5 5 ng/l, Accuracy: 8 12%, Precision < 25% Determined for every analytical run and matrix!!
Determination of matrix effects Post extraction spike analyte spike SPE extract LC (ESI)MS/MS LC (APCI)MS/MS Matrix effect (ME) = Absolute recovery of a post extraction spike
BZP-3 BT MTBT BZP-2 Mecoprop Propiconazole Imazalil Thiabendazole Terbutryn Irgarol Dimethomorph Fenpropimorph BIT OIT Triclosan Triclocarban Chlorophene 2 15 1 5 Matrix effects (raw wastewater) 24% ESI Matrix effects [%] APCI Matrix effects [%] 315% Diuron Isoproturon Matrix effects [%]
BZP-3 BT MTBT BZP-2 Chlorophene Triclocarban Triclosan Imazalil Thiabendazole Terbutryn Irgarol Dimethomorph Fenpropimorph BIT OIT Mecoprop Propiconazole 2 15 1 5 ESI APCI Matrix effects [%] Relative recovery [%] Matrix effects [%] Relative recovery [%] 315% 2 15 1 5 Diuron Isoproturon Matrix effects [%] Relative recovery [%] Relative Recovery 24%
Example: Thiabendazole intensity [cps] 5.e6 2.e6 1.2e6 5.e5 ESI thiabendazole, MRM 22./175. 6. 7. 8. 9. ME: ~ 24% standard influent spiked with 2 ng ml -1 1.e6 5.e5. 1.2e6 5.e5 APCI thiabendazole, MRM 22./175. 6. 7. 8. 9. ME: ~ 12% 6. 7. 8. 9. 6. 7. 8. 9. 288 1 influent non-spike 6. 7. 8. 9. time [min] S/N = 35, ~ 4 ng/l 368 1 6. 7. 8. 9. time [min] S/N = 9, ~ 4 ng/l LOQ ~ 1 ng/l LOQ ~ 5 ng/l
Direct injection No enrichment prior to measurement with an API 55 Tandem MS No losses during SPE, High Throughput Less matrix effects! Matrix effect [%] 12 1 8 6 4 2 Terbutryn Climbazol Terbutryn Climbazol Direct injection SPE extracts groundwater surface water WWTP effluent WWTP influent Possible drawbacks: Higher LOQs, higher blank values
Extraction of solid samples Many biocides are predominantly sorbed: Quarternary ammonium compounds (QACs), triclosan, triclocarban, chlorophene, many conazoles (e.g. climbazole, ketoconazole, miconazole), etc. Analysis in particulate matter, sediments, biota, soils and sludge is required Challenge: Extractability depends on the physico chemical properties of the compounds (e.g. charge, K OW, etc. ) and the properties of the solid matrix (cation exchange capacity, ph, TOC, etc.) no certified reference materials Spiking of analytes to non polluted groundwater Preparation of reference material Addition to sediment sediment water slurry Evaporation at 3 C
Extraction of solid samples Recovery [%] of climbazole using self prepared reference materials MeOH/H 2 O MeOH MeOH/Acetone soil 79 6 32 sediment 78 51 8 sludge 5 71 2 Recovery using freeze dried soil spiked with climbazole directly prior to extraction with MeOH/acetone: ~ 1 %
Concentration variability Concentration [ng/l] 5 4 3 Average Median Terbutryn (n = 11) 2 1 1 8 loc 1 loc 2 loc 3 loc 4 loc 5 loc 6 Average Median AA EQS = 65 ng/l 6 4 2 loc 1 loc 2 loc 3 loc 4 loc 5 loc 6 Irgarol (n = 11) M1 in the same conc. range AA EQS = 2.5 ng/l
Conclusions Many biocides can be determined simultaneously in various environmental matrices down to the low ng/l and ng/g level. Ionization by APCI can be a valuable alternative, especially if labeled surrogate standards are missing * A compensation of matrix effects is the prerequisite for analytical accuracy and reproducibility. * The relative recoveries and the LOQs should be determined for each sample series and each sample matrix. * Extraction methods for solid matrices should be validated for each individual matrix (e.g. soils, sediments, sludge, biota). Self prepared reference materials are an option for testing extractability. * A sufficient time resolution for sampling is crucial for assessing annual average and maximum concentrations of biocides
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