Fate and Toxicity of Pesticides in the Ebro Delta

SPANISH COUNCIL OF SCIENTIFIC RESEARCH Institute of Environmental Assessment and Water Research (IDÆA)

Fate and toxicity of pesticides in the Ebro Delta

Marianne Köck Schulmeyer - IDAEA-CSIC Marinel.la Farré - IDAEA-CSIC Antoni Ginebreda - IDAEA-CSIC / ACA Asunción Navarro - IDAEA-CSIC Elena Martínez - IDAEA-CSIC Miren López de Alda - IDAEA-CSIC Damià Barceló - IDAEA-CSIC / ICRA

Water toxicity and pesticides loads in water and seafood in the Ebro river delta during rice cultivation

OVERVIEW OVERVIEW

INTRODUCTION Marine bacteria Vibrio fischeri OBJETIVES INTRODUCTION

SITE DESCRIPTION OBJETIVES and SAMPLES SITE DESCRIPTION and SAMPLES METHODOLOGY METHODOLOGY - Pesticides analysis in Zooplancton water - Pesticides analysis in water Daphnia magna - Ecotoxicity bioassays - Ecotoxicity Bioassays - Pesticides analysis in - Pesticides analysis in shellfish shellfish RESULTS RESULTS Oysters and -Pesticides in water - Pesticides in water mussel - Toxicity in water -Toxicity in water - Pesticides in shellfish Alga Selenastrum -Pesticides in shellfish capricornutum CONCLUSIONS CONCLUSIONS Acknowledgements Acknowledgements Water toxicity and pesticides loads in water and seafood in the Ebro river delta during rice cultivation

INTRODUCTION OVERVIEW

INTRODUCTION The Ebro river OBJETIVES SPAIN (NE) EUROPE SITE DESCRIPTION -It is Spain’s most voluminous river (910 km) and SAMPLES -It flows through many cities before METHODOLOGY discharging in a delta on the Mediterranean Sea in the province of Tarragona. - Pesticides analysis in water The Ebro delta - Ecotoxicity bioassays - Pesticides analysis in shellfish -It is one of the largest wetland areas (320 km²) in the western Mediterranean region. RESULTS

-Pesticides in water -It hosts numerous beaches, marshes and EBRO RIVER -Toxicity in water salt pans that provide habitat for over 300 species of birds. -Pesticides in shellfish -A network of canals and irrigation ditches are CONCLUSIONS helping to maintain the ecologic and economic resources of the Ebro Delta. Acknowledgements EBRO RIVER DELTA

Water toxicity and pesticides loads in water and seafood in the Ebro river delta during rice cultivation

INTRODUCTION OVERVIEW

INTRODUCTION

OBJETIVES -It is in intensive agricultural use for rice, fruit and vegetables. SITE DESCRIPTION and SAMPLES -The rice growing areas are situated in the rice crops Delta del Ebro Natural Park where there is METHODOLOGY also shellfish farming. - Pesticides analysis in water -Pesticides typical of rice crops frequently - Ecotoxicity bioassays used here include propanil, bentazone, MCPA and molinate. The insecticides fenitrothion - Pesticides analysis in shellfish and malathion are employed occasionally if necessary. RESULTS draining channel -Recently, the seafood farmers have -Pesticides in water complained about a loss of production in the -Toxicity in water periods of rice cultivation which has raised -Pesticides in shellfish also public concern about the toxicity of the water in this area

CONCLUSIONS

Acknowledgements mussel farming Water toxicity and pesticides loads in water and seafood in the Ebro river delta during rice cultivation

OBJETIVES OVERVIEW

INTRODUCTIONOBJETIVES

OBJETIVES Main objective

SITE DESCRIPTION To investigate the role of pesticides in the mortality of and SAMPLES shellfish in the Ebro river during the main growing season

METHODOLOGY of rice - Pesticides analysis in water - Ecotoxicity bioassays Sub-objectivess - Pesticides analysis in shellfish 9 To assess residual concentrations of pesticides in water RESULTS and biota.

-Pesticides in water 9To assess toxicity of water samples. -Toxicity in water -Pesticides in shellfish 9To investigate potential correlations between pesticides and ecotoxicity data with shellfish mortality episodes. CONCLUSIONS 9To identify possibly responsible compounds.

Acknowledgements

Water toxicity and pesticides loads in water and seafood in the Ebro river delta during rice cultivation

SITE DESCRIPTION and SAMPLES OVERVIEW

INTRODUCTION A total number of 108 samples have been studied, of which 54 are from SOUTHERN hemidelta (three sample points) and the other 54 from NORTHERN hemidelta (three OBJETIVES sample points).

HEOD: OLLES draining HEIMD: ILLA DE MAR SITE DESCRIPTION HEM: FANGAR bay SITE DESCRIPTION channel draining channel and SAMPLES • Sampling: METHODOLOGY 14 April-12 June 2008 - Pesticides analysis in water SPAIN • Measurements and - Ecotoxicity bioassays matrices: - Pesticides analysis in shellfish -7 pesticides in oyster & RESULTS mussel (11 samples) -Pesticides in water -23 pesticides in water -Toxicity in water (108 samples) -Pesticides in shellfish HDCD: CAMPREDÓ draining channel EBRO RIVER DELTA -3 toxicity tests in water CONCLUSIONS (108 samples) HDM: ALFACS bay HDAD: ALA draining Acknowledgements channel Water toxicity and pesticides loads in water and seafood in the Ebro river delta during rice cultivation

SITE DESCRIPTION and SAMPLES OVERVIEW Physical-chemical parameters (Fangar bay) INTRODUCTION

OBJETIVES Temperatura de l'aigua o Water temperatureºC ( C)

SITE DESCRIPTION 35.00 Mitjana and SAMPLES 30.00 ¾Temperature (°C) 25.00 20.00

METHODOLOGY 15.00 ¾Conductivity (µS/cm) - Pesticides analysis in 10.00 water 5.00 0.00 - Ecotoxicity bioassays ¾Dissolved oxygen (mg/L) - Pesticides analysis in shellfish ¾pH 08/05/2008 10/05/2008 13/05/2008 15/05/2008 17/05/2008 20/05/2008 22/05/2008 25/05/2008 27/05/2008 29/05/2008 01/06/2008 03/06/2008 05/06/2008 08/06/2008 10/06/2008 13/06/2008 15/06/2008 17/06/2008 21/06/2008 26/06/2008 01/07/2008 06/07/2008 10/07/2008 15/07/2008 20/07/2008 25/07/2008 29/07/2008 03/08/2008 08/08/2008 13/08/2008 18/08/2008 22/08/2008

ConductivityConductivitat (µS/cm) RESULTS ¾Turbidity (NTU) µS/cm -Pesticides in water 60.00 Mitjana -Toxicity in water ¾Redox potential (mV) 50.00 40.00 -Pesticides in shellfish 30.00 Conductivity decrease input of shallow water 20.00 CONCLUSIONS 10.00

0.00 Acknowledgements 08/05/2008 10/05/2008 13/05/2008 15/05/2008 17/05/2008 20/05/2008 22/05/2008 25/05/2008 27/05/2008 29/05/2008 01/06/2008 03/06/2008 05/06/2008 08/06/2008 10/06/2008 13/06/2008 15/06/2008 17/06/2008 21/06/2008 26/06/2008 01/07/2008 06/07/2008 10/07/2008 15/07/2008 20/07/2008 25/07/2008 29/07/2008 03/08/2008 08/08/2008 13/08/2008 18/08/2008 22/08/2008

Water toxicity and pesticides loads in water and seafood in the Ebro river delta during rice cultivation

METHODOLOGY Pesticides analysis in water OVERVIEW

INTRODUCTION TARGET ANALYTES OBJETIVES

Solubilidad Grupo Compuesto log Kow MW Formula molecular SITE DESCRIPTION agua (mg/L) Triazinas Atrazina 2.50 215.7 33 C H ClN and SAMPLES 8 14 5 Simazina 2.10 201.7 6.2 C7H12ClN5

Cianazina 2.22 240.7 170 C9H13ClN6 Desetil-atrazina 1.51 187.6 3200 C H ClN METHODOLOGY 6 10 5 Terbutilazina 3.21 229.7 8.5 C9H16ClN5 Deisopropilatrazina 1.15 173.6 670 C H ClN - Pesticides analysis analysis in 5 8 5 Trifluralina 5.34 335.29 0.184 C13H16F3N3O4 inwater water Órganofosforados Diazinón 3.30 304.3 60 C12H21N2O3PS 3 - Ecotoxicity bioassays Dimetoate 0.70 229.3 23.3x10 C5H12NO3PS2 Fenitrotión 3.43 277.2 14 C9H12NO5PS

- Pesticides analysis in Malatión 2.75 330.4 145 C10H19O6PS2 shellfish Chloroacetanilidas Alaclor 3.09 269.8 170.31 C14H20ClNO2 Metolaclor 2.90 283.8 488 C15H22ClNO2

Fenil Ureas Diurón 2.68 233.1 42 C9H10Cl2N2O RESULTS Isoproturón 2.87 206.2 65 C12H18N2O Linurón 3.20 249.10 75 C9H10Cl2N2O2

-Pesticides in water Clortolurón 2.41 212.67 70 C10H13ClN2O Tiocarbamato -Toxicity in water Molinato 2.88 187.30 990 C9H17NOS Ácidos Mecoprop 3.13 214.65 620 C10H11ClO3

-Pesticides in shellfish 2,4 D 2.81 221.04 677 C8H6Cl2O3 Bentazona 2.34 240.28 500 C10H12N2O3S

MCPA 3.25 200.62 630 C9H9ClO3 CONCLUSIONS Anilida Propanil 3.30 218.10 130 C9H9Cl2NO

Acknowledgements Water toxicity and pesticides loads in water and seafood in the Ebro river delta during rice cultivation

METHODOLOGY Pesticides analysis in water OVERVIEW

INTRODUCTION EBRO RIVER DELTA OBJETIVES

SITE DESCRIPTION and SAMPLES

METHODOLOGY Liquid Chromatography Data - Pesticides analysis analysis in Water samples (LC-ESI(+)-MS/MS) inwater water

500ppt - Ecotoxicity bioassays Delta320 1: MRM of 6 Channels ES+ 8.77 174 > 104 100 4.30e4 % 1 Delta320 1: MRM of 6 Channels ES+ - Pesticides analysis in 10.67 188.1 > 146 100 1.08e5 % shellfish 0 Delta320 1: MRM of 6 Channels ES+ 10.99 230.1 > 199 100 4.21e4 % 1 Delta320 2: MRM of 12 Channels ES+ 12.75 202.1 > 132 100 8.58e4 RESULTS % 1 Delta320 2: MRM of 12 Channels ES+ 12.77 241.1 > 214 100 7.09e4 -Pesticides in water % 1 Delta320 2: MRM of 12 Channels ES+ 13.81 213.1 > 72.2 100 1.07e5 % -Toxicity in water 1 Delta320 2: MRM of 12 Channels ES+ 14.25 207.2 > 72.2 100 2.28e5 % -Pesticides in shellfish 0 Delta320 2: MRM of 12 Channels ES+ 14.34 216.1 > 174 100 1.67e5 % 0 Time Filtration 2.50 5.00 7.50 10.00 12.50 15.00 17.50 20.00 22.50 25.00 27.50 CONCLUSIONS On-line Solid Fase Extraction (0.4 um nylon filters) Results (sample volume = 5+5mL) Acknowledgements (sorbent: *PLRP-s for ESI+) (sorbent: Hysphere Resin GP for ESI-)

Water toxicity and pesticides loads in water and seafood in the Ebro river delta during rice cultivation

METHODOLOGY Pesticides analysis in water OVERVIEW

Compuesto Rt (min) SRM Cone(V) Coll(eV) Relación* INTRODUCTION LC-MS/MS Modo de ionización negativo 7.9 SRM 1: 239 → 132 35 25 1.91 Bentazona SRM 2: 239 → 197 35 20 conditions SRM 1: 199 → 141 20 15 3.62 OBJETIVES MCPA 9.9 SRM 2: 201 → 143 15 15 SRM 1: 219 → 161 15 10 7.77 2,4-D 10.0 SRM 2: 219 → 125 15 30 SRM 1: 213 → 141 15 15 3.10 SITE DESCRIPTION Mecoprop 10.3 SRM 2: 215 → 143 15 15 SRM 1: 216 → 160 30 15 1.71 and SAMPLES Propanil 16.9 SRM 2: 218 → 162 30 15 SRM 1: 262 → 152 25 30 5.50 Fenitrotión 21.1 -Column: SRM 2: 262 → 122 25 20 METHODOLOGY Purospher STAR Modo de ionización positivo SRM 1: 174 → 104 25 20 Deisopropil atrazina 9.0 1.24 - Pesticides analysis in RP-18e (125x2 SRM 2: 174 → 132 25 15 - Pesticides analysis SRM 1: 188 → 146 25 15 Desetil atrazina 10.9 3.06 inwater water mm, 5µm, Merck) SRM 2: 188 → 104 25 25 SRM 1: 230 → 199 15 10 Dimetoato 11.2 1.83 - Ecotoxicity bioassays SRM 2: 230 → 125 15 20 SRM 1: 202 → 132 30 20 Simazina 12.9 1.03 - Mobile phase: SRM 2: 202 → 124 30 20 - Pesticides analysis in SRM 1: 241 → 214 30 15 Gradient Cianazina 12.9 12.16 shellfish SRM 2: 241 → 174 30 20 SRM 1: 213 → 72 25 15 ACN/H2O (0.2 Clortolurón 14.0 11.03 SRM 2: 213 → 140 25 25 SRM 1: 207 → 72 25 15 mL/min) Isoproturón 14.4 95.6 RESULTS SRM 2: 207 → 165 25 25 SRM 1: 216 → 174 30 15 Atrazina 14.5 3.52 -Pesticides in water SRM 2: 216 → 132 30 20 SRM 1: 233 → 72 15 15 Diurón 14.8 1.96 SRM 2: 235 → 72 15 30 -Toxicity in water SRM 1: 230.7 → 174 20 15 Terbutilazina 17.1 4.26 SRM 2: 230.7 → 132 20 25 SRM 1: 249 → 160 25 20 -Pesticides in shellfish Linurón 17.6 1.08 SRM 2: 249 → 182 25 15 SRM 1: 188 → 126 25 10 Molinato 19.1 2.68 SRM 2: 188 → 83 25 20 SRM 1: 284.8 → 252 25 15 CONCLUSIONS Metolaclor 19.9 2.39 SRM 2: 284.8 → 176 25 25 SRM 1: 353 → 227 25 15 Malatión 19.9 1.62 SRM 2: 353 → 195 25 15 SRM 1: 270 → 238 15 10 Acknowledgements Alaclor 20.1 4.03 SRM 2: 270 → 162 15 25 SRM 1: 305 → 169 25 20 Diazinon 23.6 1.51 SRM 2: 305 → 153 25 20 *Relación SRM1/SRM2 Water toxicity and pesticides loads in water and seafood in the Ebro river delta during rice cultivation

METHODOLOGY Pesticides analysis in water OVERVIEW

INTRODUCTION METHOD OBJETIVES VALIDATION

SITE DESCRIPTION LOD (ng/L) LOQ (ng/L) and SAMPLES %R (n=5) HPLC Agua superficial HPLC Agua superficial Atrazina 80 0.585 0.630 1.561 1.679 METHODOLOGY Simazina 73 0.483 0.306 1.287 0.816 - Pesticides analysis in Cianazina 89 0.654 0.371 1.744 0.988 - Pesticides analysis Desetil-atrazina 44 0.300 0.688 0.801 1.835 inwater water Terbutilazina 91 1.143 1.688 3.049 4.501 - Ecotoxicity bioassays Deisopropilatrazina 86 0.794 1.368 2.118 3.648 Diazinon 102 0.060 0.069 0.160 0.185 - Pesticides analysis in Dimetoate 75 0.294 0.999 0.784 2.664 shellfish Fenitrotión 36 5.786 7.583 15.428 20.223 Malatión 64 0.738 1.641 1.967 4.377 Alaclor 97 0.910 6.909 2.428 18.423 Metolaclor 100 0.284 1.088 0.757 2.901 RESULTS Diurón 91 0.611 1.057 1.628 2.819 -Pesticides in water Isoproturón 93 0.079 0.152 0.210 0.407 Linurón 90 1.076 1.472 2.868 3.924 -Toxicity in water Clortolurón 91 0.320 0.345 0.854 0.922 Molinato 96 0.776 3.716 2.070 9.908 -Pesticides in shellfish Mecoprop 101 0.330 0.394 0.879 1.051 2,4 D 80 0.483 0.368 1.288 0.981 Bentazona 79 0.052 0.145 0.138 0.387 CONCLUSIONS MCPA 92 0.297 0.320 0.793 0.854 Propanil 81 0.218 0.688 0.582 1.834

Acknowledgements

Water toxicity and pesticides loads in water and seafood in the Ebro river delta during rice cultivation

METHODOLOGY Pesticides analysis in water OVERVIEW

INTRODUCTION NI mode

500ppt OBJETIVES Delta40 1: MRM of 9 Channels ES- 9.67 239.1 > 132 100 6.64e5

% SITE DESCRIPTION Bentazona and SAMPLES 0 Delta40 1: MRM of 9 Channels ES- 10.77 199 > 141 100 1.36e5 METHODOLOGY % MCPA - Pesticides analysis in - Pesticides analysis 0 water Delta40 1: MRM of 9 Channels ES- in water 10.85 219 > 161 100 9.53e4 - Ecotoxicity bioassays % 2,4-D - Pesticides analysis in 0 shellfish Delta40 1: MRM of 9 Channels ES- 11.06 213 > 141 100 1.40e5 RESULTS % Mecoprop -Pesticides in water 0 Delta40 2: MRM of 4 Channels ES- 16.84 216 > 160 100 -Toxicity in water 1.34e5 -Pesticides in shellfish % Propanil 0 Delta40 2: MRM of 4 Channels ES- 20.93 262.3 > 152.2 100 CONCLUSIONS 2.65e3

% Fenitrotión

Acknowledgements 14 Time 2.50 5.00 7.50 10.00 12.50 15.00 17.50 20.00 22.50 25.00 27.50 30.00

Water toxicity and pesticides loads in water and seafood in the Ebro river delta during rice cultivation

METHODOLOGY Pesticides analysis in water OVERVIEW

PI mode INTRODUCTION

500ppt Delta320 1: MRM of 6 Channels ES+ 8.77 174 > 104 OBJETIVES 100 Deisopropil atrazina 4.30e4 % 1 Delta320 1: MRM of 6 Channels ES+ 10.67 188.1 > 146 100 SITE DESCRIPTION Desetil atrazina 1.08e5 % and SAMPLES 0 Delta320 1: MRM of 6 Channels ES+ 10.99 230.1 > 199 100 4.21e4 % Dimetoato 1 METHODOLOGY Delta320 2: MRM of 12 Channels ES+ 12.75 202.1 > 132 100 500ppt 8.58e4 Simazina Delta08 2: MRM of 12 Channels ES+ % 15.00 233 > 72 100 - Pesticides analysis in 8.89e4 - Pesticides analysis 1 Diurón Delta320 2: MRM of 12 Channels ES+ % water 12.77 241.1 > 214 in water 100 1 Cianazina 7.09e4 Delta08 3: MRM of 12 Channels ES+ % 17.37 230.2 > 174 100 1.17e5 - Ecotoxicity bioassays 1 Terbutilazina % Delta320 2: MRM of 12 Channels ES+ 13.81 213.1 > 72.2 100 0 Clorotolurón 1.07e5 Delta08 3: MRM of 12 Channels ES+ - Pesticides analysis in 17.89 249.1 > 160 % 100 5.35e4 1 % Linurón shellfish Delta320 2: MRM of 12 Channels ES+ 14.25 207.2 > 72.2 1 100 Isoproturón 2.28e5 Delta08 3: MRM of 12 Channels ES+ 19.38 188.1 > 126 % 100 6.95e4 0 % Molinato Delta320 2: MRM of 12 Channels ES+ RESULTS 14.34 216.1 > 174 1 100 1.67e5 Delta08 3: MRM of 12 Channels ES+ Atrazina 20.22 284.2 > 252 % 100 1.85e5 -Pesticides in water 0 Time % Metolaclor 2.50 5.00 7.50 10.00 12.50 15.00 17.50 20.00 22.50 25.00 27.50 0 Delta08 3: MRM of 12 Channels ES+ 20.18 353.5 > 227.3 -Toxicity in water 100 1.16e4 % Malatión 4 -Pesticides in shellfish Delta08 3: MRM of 12 Channels ES+ 20.54 270.2 > 238 100 Alaclor 3.19e4 % 2 Delta08 4: MRM of 2 Channels ES+ CONCLUSIONS 23.91 305.2 > 169 100 Diazinon 8.99e5 % 0 Time 14.00 16.00 18.00 20.00 22.00 24.00 26.00 28.00 Acknowledgements

Water toxicity and pesticides loads in water and seafood in the Ebro river delta during rice cultivation

METHODOLOGY Pesticides analysis in water OVERVIEW

INTRODUCTION Criteria for positive confirmation OBJETIVES Standard aqueous solution: SITE DESCRIPTION and SAMPLES SRM 1: 216 → 174 SRM 2: 216 → 132 METHODOLOGY Concentration: 5 ng/L

- Pesticides analysis analysis in inwater water - Ecotoxicity bioassays Sample: SRM 1: 216 → 174 - Pesticides analysis in shellfish SRM 2: 216 → 132 Concentration: 2.97 ng/L RESULTS -Pesticides in water -Toxicity in water -Pesticides in shellfish

CONCLUSIONS Rtsample = Rtstandard ± 2% Acknowledgements Ratio SRM1/SRM2sample = Ratio SRM1/SRM2standard ± 20-50 % Water toxicity and pesticides loads in water and seafood in the Ebro river delta during rice cultivation

METODOLOGY Ecotoxicity bioassays OVERVIEW

INTRODUCTION

OBJETIVES Toxicity assay Toxicity assay Vibrio fischeri Selenastrum SITE DESCRIPTION capricornutum and SAMPLES according to ISO 11348-3 according to METHODOLOGY ISO 8692 - Pesticides analysis in water

- Ecotoxicity bioassays bioassays - Pesticides analysis in shellfish

RESULTS -Pesticides in water Toxicity assay -Toxicity in water Daphnia magna -Pesticides in shellfish according to ISO 6341 CONCLUSIONS

Acknowledgements

Water toxicity and pesticides loads in water and seafood in the Ebro river delta during rice cultivation

METODOLOGY Pesticides analysis in shellfish OVERVIEW

INTRODUCTION TARGET PESTICIDES (7) OBJETIVES

SITE DESCRIPTION Use type Chemical class Compound and SAMPLES Herbicides Anilides Propanil METHODOLOGY - Pesticides analysis in Thiocarbamates Molinate water - Ecotoxicity bioassays Acids Bentazone - Pesticides analysis analysis in inshellfish shellfish Insecticides Pyrethroids Cypermethrin

RESULTS Organophosphorus Malathion -Pesticides in water -Toxicity in water Maloxon (malathion TP) -Pesticides in shellfish Fenitrothion

CONCLUSIONS Fenitrothion oxon

Acknowledgements SS: acetochlor D11, alachlor D13 y ethyl parathion D10 Water toxicity and pesticides loads in water and seafood in the Ebro river delta during rice cultivation

METODOLOGY Pesticides analysis in shellfish OVERVIEW

INTRODUCTION EBRO RIVER DELTA OBJETIVES

SITE DESCRIPTION and SAMPLES

METHODOLOGY

- Pesticides analysis in SPE GC/MS-EI water (Florisil) (HP-5MS, 30m x 0,25mm) - Ecotoxicity bioassays Oysters and mussels (0.25 g) - Pesticides analysis analysis in inshellfish shellfish

RESULTS -Pesticides in water data -Toxicity in water -Pesticides in shellfish

CONCLUSIONS Lyophilization and Pressurized Liquid Acknowledgements trituration Extraction (ASE) Results (MeOH:DCM, 1:1)

Water toxicity and pesticides loads in water and seafood in the Ebro river delta during rice cultivation

METODOLOGY Pesticides analysis in shellfish OVERVIEW

INTRODUCTION METHOD VALIDATION OBJETIVES

SITE DESCRIPTION and SAMPLES

METHODOLOGY - Pesticides analysis in water - Ecotoxicity bioassays

- Pesticides analysis analysis in inshellfish shellfish

RESULTS -Pesticides in water -Toxicity in water -Pesticides in shellfish

CONCLUSIONS

Acknowledgements Water toxicity and pesticides loads in water and seafood in the Ebro river delta during rice cultivation

RESULTS Pesticides in water OVERVIEW

INTRODUCTION Frequency of detection and average and maximum concentration of individual pesticides OBJETIVES

SITE DESCRIPTION and SAMPLES

METHODOLOGY - Pesticides analysis in water - Ecotoxicity bioassays - Pesticides analysis in shellfish

RESULTS --Pesticides Pesticides in in water water -Toxicity in water -Pesticides in shellfish

CONCLUSIONS

Acknowledgements

HEIMD, draining channel in the hemidelta left; HDAD, draining channel in the hemidelta right

Water toxicity and pesticides loads in water and seafood in the Ebro river delta during rice cultivation

RESULTS Pesticides in water OVERVIEW

10000 INTRODUCTION HEIMD Mortality episodes OBJETIVES 8000

SITE DESCRIPTION 6000 and SAMPLES

4000 METHODOLOGY concentration (ng/L) concentration - Pesticides analysis in 2000 water

- Ecotoxicity bioassays 0 10000 - Pesticides analysis in HDAD shellfish 8000

RESULTS 6000 --Pesticides Pesticides in in water water -Toxicity in water 4000 -Pesticides in shellfish concentration (ng/L) concentration

2000 CONCLUSIONS

0 Acknowledgements 2104. 2404. 14/04. 17/04. 28/04. 02/05. 05/05. 08/05. 13/05. 15/05. 19/05. 22/05. 26/05. 29/05. 02/06. 05/06. 09/06. 12/06.

HEIMD, draining channel in the hemidelta left; HDAD, draining channel in the hemidelta right Water toxicity and pesticides loads in water and seafood in the Ebro river delta during rice cultivation

RESULTS Pesticides in water OVERVIEW

malatión 2000,00 INTRODUCTION HEM bentazona 1800,00 molinato MCPA Mortality episodes OBJETIVES 1600,00 fenitrotión terbutilazina 1400,00 alaclor propanil SITE DESCRIPTION 1200,00 diurón simazina and SAMPLES 1000,00 dimetoate 2,4D 800,00 desetilatrazina

600,00 atrazina METHODOLOGY mecoprop 400,00 diazinon - Pesticides analysis in deisopropilatrazina 200,00 linurón water metolaclor 0,00 isoproturón clorotolurón - Ecotoxicity bioassays cianazina 1704HEM 2104HEM 2404HEM 0205HEM 0505HEM 0805HEM 1305HEM 1505HEM 1905HEM 2205HEM 2605HEM 2905HEM 0206HEM 0506HEM 0906HEM 1206HEM

- Pesticides analysis in malatión 1600,00 shellfish HDM bentazona molinato 1400,00 MCPA fenitrotión RESULTS 1200,00 terbutilazina alaclor

1000,00 propanil --Pesticides Pesticides in in water water diurón simazina 800,00 -Toxicity in water dimetoate 2,4D -Pesticides in shellfish 600,00 desetilatrazina atrazina 400,00 mecoprop diazinon deisopropilatrazina CONCLUSIONS 200,00 linurón metolaclor 0,00 isoproturón Acknowledgements clorotolurón 1704HDM 2104HDM 2404HDM 2804HDM 0205HDM 0505HDM 0805HDM 1305HDM 1505HDM 1905HDM 2205HDM 2605HDM 2905HDM 0206HDM 0506HDM 0906HDM 1206HDM cianazina

HEM, hemidelta left (Fangar bay); HDM, hemidelta right (Alfacs bay)

Water toxicity and pesticides loads in water and seafood in the Ebro river delta during rice cultivation

RESULTS Pesticides in water OVERVIEW

INTRODUCTION The five most abundant pesticides in water

OBJETIVES MALATHION 5825.33 ng/L* SITE DESCRIPTION MCPA and SAMPLES 4196.70 ng/L*

METHODOLOGY MOLINATE - Pesticides analysis in 3589.83 ng/L* water BENTAZONE - Ecotoxicity bioassays 3286.38 ng/L* FENITROTHION - Pesticides analysis in shellfish 1196.95 ng/L*

RESULTS --Pesticides Pesticides in in water water herbicide herbicide herbicide -Toxicity in water insecticide insecticide -Pesticides in shellfish

CONCLUSIONS

* Maximum individual concentrations. Acknowledgements Water toxicity and pesticides loads in water and seafood in the Ebro river delta during rice cultivation

RESULTS Pesticides in water OVERVIEW

INTRODUCTION Malathion levels in water OBJETIVES

SITE DESCRIPTION and SAMPLES

METHODOLOGY - Pesticides analysis in water - Ecotoxicity bioassays HEM - Pesticides analysis in 6000.00 HDM shellfish 5000.00 HDCD HEIMD 4000.00 HEOD HDAD RESULTS 3000.00 HDAD HEOD 2000.00 HDCD --Pesticides Pesticides in in water water HEIMD 1000.00 HDM -Toxicity in water HEM 0.00

-Pesticides in shellfish 4 4 4 4 4 5 5 5 5 5 5 5 5 5 6 6 6 6 /0 /0 /0 /0 /0 /0 /0 /0 /0 /0 /0 /0 /0 /0 /0 /0 /0 /0 4 7 1 4 8 2 5 8 3 5 9 2 6 9 2 5 9 2 1 1 2 2 2 0 0 0 1 1 1 2 2 2 0 0 0 1 CONCLUSIONS

Acknowledgements

Water toxicity and pesticides loads in water and seafood in the Ebro river delta during rice cultivation

RESULTS Pesticides in water OVERVIEW

INTRODUCTION Maximum cumulative levels (MCL) and percentage of samples with MCL > 5000 ng/L

OBJETIVES 17% 0% SITE DESCRIPTION Max: 9217 ng/L and SAMPLES Max: 1982 ng/L

METHODOLOGY - Pesticides analysis in 39% water Max: 9798 ng/L - Ecotoxicity bioassays - Pesticides analysis in shellfish

RESULTS 18% 22% - Pesticides in water -Pesticides in water Max: 6200 ng/L Max: 6530 ng/L -Toxicity in water -Pesticides in shellfish 0%

CONCLUSIONS Max: 1580 ng/L

Acknowledgements Water toxicity and pesticides loads in water and seafood in the Ebro river delta during rice cultivation

RESULTS Pesticides in water and their THEORETICAL toxicity OVERVIEW T = Σ Ci × TUi INTRODUCTION theoretical Vibrio fischeri Daphnia Magna Selenastrum

OBJETIVES Pesticide EC50 TU EC50 TU EC50 TU (μg/L) (μg/L) (μg/L) (μg/L) (μg/L) (μg/L) Acids SITE DESCRIPTION mecoprop --- 100 1.000 10300 0.010 and SAMPLES 2,4 D --- 126 0.794 24200 0.004 bentazone --- 64000 0.002 4500 0.022 MCPA --- 190000 0.001 80000 0.001 METHODOLOGY Anilide and Chloroacetamides - Pesticides analysis in alachlor 160000 0.001 10000 0.010 10 10.000 metolachlor 17000 0.006 23500 0.004 55 1.818 water propanil --- 2000 0.050 50 2.000 - Ecotoxicity bioassays Organophosphates diazinon 84000 0.001 0.6 166.667 10000 0.010 - Pesticides analysis in dimethoato 55500 0.002 1800 0.056 35000 0.003 shellfish fenitrothion 1500 0.067 50 2.000 800 0.125 malathion 37000 0.003 0.7 142.857 18000 0.006 Phenylureas RESULTS diuron 43240 0.002 8600 0.012 2.4 41.667 isoproturon 96000 0.001 580 0.172 21 4.762 --Pesticides Pesticides in in water water linuron 5500 0.018 310 0.323 67 1.493 chlortoluron 45000 0.002 6700 0.015 1 100.000 -Toxicity in water Thiocarbamate -Pesticides in shellfish molinate 2500 0.040 1830 0.055 70 1.429 Triazines atrazine 39800 0.003 50 2.000 21 4.762 CONCLUSIONS simazine 10000 0.010 38 2.632 2 50.000 cyanazine 78500 0.001 23 4.348 27 3.704 desethylatrazine 81860 0.001 50 2.000 27 3.704 Acknowledgements terbuthylazine ----- 5000 0.020 16 6.250 deisopropylatrazine 12740 0.008 5000 0.020 25 4.000

EC50: effective concentration that produces a specific measurable effect in 50% of the test organisms

TU: Toxic Unit = 100 / EC50

Water toxicity and pesticides loads in water and seafood in the Ebro river delta during rice cultivation

RESULTS Pesticides in water and their related toxicity OVERVIEW

malathion INTRODUCTION Theoretical toxicity - D. Magna diazinon fenitrothion 900 simazine The most molinate OBJETIVES 800 malathion (98 %) desethylatrazine representative 700 atrazine diazinon (almost 2%) 2,4D organism. SITE DESCRIPTION 600 mecoprop terbuthylazine and SAMPLES 500 propanil bentazone Theoretical 400 diuron alachlor toxicity values 300 METHODOLOGY dimethoato 200 MCPA predicted in water - Pesticides analysis in isoproturon 100 linuron water deisopropylatrazine from pesticides 0 metolachlor - Ecotoxicity bioassays chlortoluron levels 14/04 17/04 21/04 24/04 28/04 02/05 05/05 08/05 13/05 15/05 19/05 22/05 26/05 29/05 02/06 05/06 09/06 12/06 cyanazine - Pesticides analysis in shellfish diuron Theoretical toxicity - V. Fischeri Theoretical toxicity - S. Capricornotum simazine terbuthylazine molinate 25 0.16 alachlor RESULTS fenitrothion molinate (69 %) molinate 0.14 malation diuron (36 %) propanil --Pesticides Pesticides in in water water 20 fenitrothion (21 %) simazine simazine (26 %) chlortoluron 0.12 desethylatrazine malathion (8 %) diuron terbuthylazine (17%) -Toxicity in water atrazine 0.10 alachlor 15 bentazone -Pesticides in shellfish atrazine fenitrothion 0.08 dimethoato deisopropylatrazine 10 isoproturon deisopropylatrazine 0.06 malathion metolachlor metolachlor CONCLUSIONS linuron 0.04 desethylatrazine 5 MCPA diazinon 0.02 diazinon linuron mecoprop Acknowledgements 0 0.00 chlortoluron 2,4D dimethoato

isoproturon 14/04 17/04 21/04 24/04 28/04 02/05 05/05 08/05 13/05 15/05 19/05 22/05 26/05 29/05 02/06 05/06 09/06 12/06

14/04 17/04 21/04 24/04 28/04 02/05 05/05 08/05 13/05 15/05 19/05 22/05 26/05 29/05 02/06 05/06 09/06 12/06 cyanazine cyanazine Water toxicity and pesticides loads in water and seafood in the Ebro river delta during rice cultivation

RESULTS Toxicity in water OVERVIEW 90

80 INTRODUCTION Experimental toxicity - D. Magna Mortality episodes 70 occurred often 1-3 days OBJETIVES 60 after water aqute % I 50 toxicity tests indicated 40

SITE DESCRIPTION 30 toxicity and SAMPLES 20 Ecotoxicological 10 responses of tests METHODOLOGY 0 100 Experimental toxicity - S. capricornutum & - Pesticides analysis in 90 14/04. 14/04. 17/04. 17/04. 28/04. 28/04. 02/05. 02/05. 05/05. 05/05. 08/05. 08/05. 13/05. 13/05. 15/05. 15/05. 19/05. 19/05. 22/05. 22/05. 26/05. 26/05. 29/05. 29/05. 02/06. 02/06. 09/06. 09/06. shellfish mortality t! water 80 en 70 episodes m % I ee - Ecotoxicity bioassays 60 gr A 50 d - Pesticides analysis in oo shellfish 40 G 30

20 RESULTS 10 0 -Pesticides in water 100 Samples with I% > 30%

14/04. 14/04. 17/04. 17/04. 28/04. 28/04. 02/05. 02/05. 05/05. 05/05. 08/05. 08/05. 13/05. 13/05. 15/05. 15/05. 19/05. Experimental19/05. 22/05. 22/05. 26/05. 26/05. toxicity29/05. 29/05. -02/06. V. Fischeri02/06. 09/06. 09/06. 90 are considered TOXIC --Toxicity Toxicity in in water water 80 SAMPLES -Pesticides in shellfish 70 % I 60 50 Toxic samples: for CONCLUSIONS 40 Selenastrum indicates 30 possible residues of 20 herbicides and metals Acknowledgements 10

0 14/04. 14/04. 17/04. 17/04. 28/04. 28/04. 02/05. 02/05. 05/05. 05/05. 08/05. 08/05. 13/05. 13/05. 15/05. 15/05. 19/05. 19/05. 22/05. 22/05. 26/05. 26/05. 29/05. 29/05. 02/06. 02/06. 09/06. 09/06.

Water toxicity and pesticides loads in water and seafood in the Ebro river delta during rice cultivation

RESULTS Toxicity in water OVERVIEW

INTRODUCTION Theoretical toxicity vs Experimental toxicity

900 90

OBJETIVES 800 S TUs Daphnia magna 80 Zooplancton

700 % Inhibición muestras 70 Daphnia magna SITE DESCRIPTION 600 60 and SAMPLES 500 50 TU % I 400 40 METHODOLOGY 300 30 200 20

- Pesticides analysis in 100 10 water 0 0 - Ecotoxicity bioassays 0.20 100

Marine bacteria 0.18 90 - Pesticides analysis in S TUs teoricas shellfish Vibrio fischeri 0.16 % Inhibición muestras 80

0.14 70

RESULTS 0.12 60

0.10 50 -Pesticides in water TU % I 0.08 40

--Toxicity Toxicity in in water water 0.06 30 -Pesticides in shellfish 0.04 20

0.02 10

0.00 0 CONCLUSIONS

% Inhibition measured and toxicity units ent! Acknowledgements reem calculated as a simple additive approach d Ag Goo Water toxicity and pesticides loads in water and seafood in the Ebro river delta during rice cultivation

RESULTS Pesticides in shellfish OVERVIEW Levels (mg/kg) of pesticides in biota INTRODUCTION Samples Molinate Propanil Fenitrothion Malathion Bentazone Cipermetrine Maloxon Fenitrothion Oxon 1 n.d n.d 3.49 n.d n.d n.d 3.62 n.d OBJETIVES 2 n.d n.d n.d n.d n.d n.d n.d n.d 3 n.d n.d n.d n.d n.d n.d n.d n.d 4 n.d n.d n.d n.d n.d n.d n.d n.d SITE DESCRIPTION 5 n.d n.d n.d n.d n.d n.d n.d n.d 6 n.d n.d n.d n.d n.d n.d n.d n.d and SAMPLES 7 n.d n.d n.d 53.12 n.d n.d n.d n.d 8 n.d n.d n.d n.d n.d n.d n.d n.d 9 n.d n.d n.d n.d n.d n.d n.d n.d METHODOLOGY 10 n.d n.d n.d n.d n.d n.d 2.53 n.d 11 n.d n.d n.d n.d n.d n.d 4.59 n.d - Pesticides analysis in water Data Left side bay (HEM) Right side bay (HDM) Sample Results (mg/kg) Sample Results (mg/kg) - Ecotoxicity bioassays 17/04 oyster n.d. mussel 3.46 (fenitrothion), - Pesticides analysis in * mortality 3.62 (maloxon) shellfish 07/05 oyster* n.d. episodes 15/05 oyster n.d. mussel n.d. 26/05 oyster** n.d. RESULTS 27/05 oyster* 53.12 (malathion) 29/05 oyster* n.d. -Pesticides in water 09/06 oyster 2.53 (maloxon) mussel 4.59 (maloxon) -Toxicity in water n.d.: not detected *mortality episodes --Pesticides Pesticides in in shellfish shellfish Pesticide aquatic species (scientific name) LC50 (µg/L) classification **big mortality episode (around 60 %) Maloxon Fish Medaka (Oryzias latipes) 365 Highly toxic Carpa (Cyprinus carpio) 1600 - CONCLUSIONS Malathion Mussel (Mytilus galloprovincialis) 6000 Moderately toxic Oyster (Crassostrea virginica) 2660 Moderately toxic >>5.8 ug/L ELS Carpa (Cyprinus carpio) 5886 Moderately toxic V Acknowledgements W LE Fenitrothion Mussel (Mytilus edulis) 123000 No acute toxicity LO Fish Medaka (Oryzias latipes) 4911 Moderately toxic >>1.2 ug/L Carpa (Cyprinus carpio) 3938 Moderately toxic

Water toxicity and pesticides loads in water and seafood in the Ebro river delta during rice cultivation

RESULTS Pesticides in shellfish OVERVIEW

INTRODUCTION

OBJETIVES

SITE DESCRIPTION and SAMPLES

METHODOLOGY - Pesticides analysis in water - Ecotoxicity bioassays - Pesticides analysis in shellfish

RESULTS -Pesticides in water -Toxicity in water --Pesticides Pesticides in in shellfish shellfish

CONCLUSIONS

Acknowledgements Water toxicity and pesticides loads in water and seafood in the Ebro river delta during rice cultivation

CONCLUSIONS OVERVIEW

INTRODUCTION 9 With respect to the Maximum Allowable Concentration (MAC) OBJETIVES stipulated by the EU for 5 of the 22 pesticides in inland and other surface waters (Directive 2008/105/EC), only alachlor was found to SITE DESCRIPTION exceed the proposed limit of 700 ng/L in two of the samples and SAMPLES investigated. 9 The northern bay is more contaminated than the southern bay METHODOLOGY (larger in size) and in both bays the inner sites are more - Pesticides analysis in contaminated than the outer sites (dilution). water - Ecotoxicity bioassays 9Most of toxicity responses were explained using an - Pesticides analysis in additive approach with concentrations of pesticides shellfish measured by LC-MS/MS and TU of each pesticide (100/EC ). RESULTS 50 -Pesticides in water 9Modulation of the toxicological responses can be attributed -Toxicity in water to synergic effects, matrix effects and possible presence of metals. -Pesticides in shellfish

CONCLUSIONS 9 The most representative organism was, as expected, Daphnia magna, a micro Acknowledgements crustacean.

Water toxicity and pesticides loads in water and seafood in the Ebro river delta during rice cultivation

CONCLUSIONS OVERVIEW

INTRODUCTION 9 The most serious mortality episodes occurred in the left side (north) of the delta, OBJETIVES which was also identified as the most polluted area.

SITE DESCRIPTION and SAMPLES 9Good agreement was found 9Good agreement was also METHODOLOGY between ecotoxicological found between water discharges data using standard from rice crops (as deducted - Pesticides analysis in bioassays and mortality from water conductivity) and water episodes occurred in shellfish mortalities - Ecotoxicity bioassays farms - Pesticides analysis in shellfish

9 Among the compounds analyzed in water discharges from RESULTS river, the most problematic compounds were: -Pesticides in water - in terms of abundance and ubiquity: bentazone, MCPA, -Toxicity in water propanil, malathion and molinate -Pesticides in shellfish - taking into account toxicity: malathion, diazinon, molinate and fenitrothion CONCLUSIONS

Acknowledgements Water toxicity and pesticides loads in water and seafood in the Ebro river delta during rice cultivation

ACKNOWLEDGEMENTS OVERVIEW

INTRODUCTION

OBJETIVES

SITE DESCRIPTION and SAMPLES

METHODOLOGY This work has been supported by the Catalan Water Agency - Pesticides analysis in water and by the Spanish Ministry of Science and Innovation through - Ecotoxicity bioassays the projects CEMAGUA [CGL2007-64551/HID] and SCARCE - Pesticides analysis in shellfish [CSD2009-00065], and reflects the author’s view.

RESULTS -Pesticides in water -Toxicity in water -Pesticides in shellfish Merck is acknowledged for the gift of LC columns, and Biotage CONCLUSIONS for the gift of SPE cartridges. Acknowledgements

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