Occurrence, distribution and fate of pharmaceuticals as chemical markers of contamination from urban sources in the vulnerable area of the Ebro Delta (Spain) Mira Čelić 1, Meritxell Gros 1, Marinella Farre 2, Damia Barceló 1,2, Mira Petrović 1,3 ICRA-Catalan Institute for Water Research, Water Quality Area 1, Girona, Spain IDAEA-CSIC, Department of Environmental Chemistry 2, Barcelona, Spain Catalan Institution for Research and Advanced Studies 3, Barcelona, Spain
OUTLINE o Introduction o Sources and routes of entry of pharmaceuticals o INTEGRA-COAST project o Objectives of the study o Study area and sampling sites o Analysis o Target pharmaceuticals in water samples o Extraction of water samples o Target pharmaceuticals in sediment samples o Extraction of sediment samples o Quality parameters obtained during analysis o Results o Occurrence of PhACs in water samples o Removal during wastewater treatment processes o Occurrence of PhACs in sediment o Conclusion
Sources and routes of entry of pharmaceuticals INTRODUCTION PhACs are the contaminants of anthropogenic origin with the biggest input into the environment. After their consumption they are discharged into wastewaters. PhACs are partially removed during wastewater treatment processes, being discharged into receiving water bodies. Changes in river flow have one of the greatest effects on water quality (point source pollution is controlled by river flow). Up to now there have been a lot of studies on their occurrence in waste and surface waters but information about their presence in sea and coastal areas is still sparse. sea water
INTEGRA-COAST PROJECT INTEGRA-COAST study microplastics, nanomaterials, emerging organic contaminants (pharmaceuticals and personal care products, new pesticides, perfluoroalkyl substances and siloxanes) and marine biotoxins in a vulnerable coastal area. The main goal of INTEGRA-COAST is to perform an integrated study of the fate, behaviour, and the river transport of emerging pollutants, nanomaterials and microplastics in estuaries, wetlands and coastal waters and to identify specific organic contaminants used as chemical markers of wastewater pollution.
OBJECTIVES INTRODUCTION To assess the impact of WWTP discharges in coastal areas and protected ecosystems, with special focus on the contamination by pharmaceuticals. To study the fate and transport of a large number of multiple-class pharmaceuticals in riverine and coastal ecosystems (e.g. occurrence in water and sediments). To evaluate possible seasonal fluctuations in the occurrence and behavior of these contaminants in these ecosystems. Identify relevant pharmaceuticals as markers of urban pollution in coastal areas with the objective to include them in risk assessment schemes and future monitoring programs.
Study area the Ebro Delta INTRODUCTION wetland area of 320 km 2 the third largest delta in the Mediterranean Sea with high biological productivity highly relevant for conservation has a typical Mediterranean climate (with rainfall concentrated in autumn and spring (200 300 mm) and intense summer drought (<50 mm)) c.a. 13% of its total surface is composed of natural lagoons, bays and marshes agricultural is one of the main activities such as rice and orchards. Three sampling campaigns: October-November 2015 (autumn) February-April 2016 (winter) June-July 2016 (spring-summer) Total number of 213 samples: 87 waters and 71 sediments
Sampling: 29 hot-spots sites WWTP2 WWTP1 wastewater IN/OUT emissary canals Ebro river lagoons sea water (water/sediment) Wastewater treatment plants (WWTP) Edar Amposta (WWTP1) Served population: 19 805 14 262 Population equivalent (PE): 27.500 28.921 Flow (m 3 /dan): 5.500 6.310 Type of the treatment: Biological with activated sludge Sant Carles de la Ràpita (WWTP2) Biological with elimination of N with tertiary treatment
Target pharmaceuticals in water samples ANALYSIS Analgesics/antiinflammatories Ketoprofen Naproxen Ibuprofen Indomethacine Acetaminophen Salicylic acid Diclofenac Phenazone Propylphenazone Piroxicam Tenoxicam Meloxicam Oxycodone Codeine Psychiatric drugs Carbamazepine 2-Hydroxycarbamazepine 10,11-epoxycarbamzepine Acridone Sertraline Citalopram Venlafaxine Olanzapine Trazadone Fluxotine Norfluxotine Paroxetine Diazepam Lorazepam Alprazolam Antibiotics Erithromycin Azithromycin Clarithromycin Tetracycline Ofloxacin Ciprofloxacin Sulfamethoxazole Trimethoprim Metronidazole Metronidazole-OH Dimetridiazole Ronidazole Cefalexin To treat asthma Salbutamol Histimine H2 receptor antagonists Loratadine Desloratadine Ranitidine Famotidine Cimetidine Cholesterol lowering agents Atrovastatine Pravastatine Mevastatine Antihelmintics Albendazole Thiabendazole Levamisole Β-Blocking agents Atenolol Sotalol Metoprolol Propranolol Carzalol Nadalol Antihypertensives Amlodipine Losartan Ibersartan Valsartan Diuretic Hydroclorothiazide Furosemide Torasemide Lipid regulators and cholesterol lowering drugs Bezafibrate Gemfibrozil Pravastatin Fluvastatin Atrovastatin X-ray contrast agents Iopromide Anticoagulant Warfarin Prostatic hyperplasia Tamsulosin Antiplatelet agent Clopidogrel Sedation and muscle relaxation Xylizine Tranquilizer Azaperone Azaperol Synthetic glucocoticoid Dexamethasone Calcium channel blockers Diltiazem Verapamil Norverapamil Antidiabetic Glibenclamide Total=81 PhACs
Extraction of water samples ANALYSIS SAMPLE PRE-TREATMENT FILTRATION 0.7 µm/ 0.45 µm Addition of 0.1% Na 2 EDTA Elution with 6 ml MeOH Addition of 34 isotopically labelled standards EXTRACTION & PRE-CONCENTRATION 500 ml sea water 100 ml river water and canals 50 ml effluent 25 ml influent Surrogate standards SPE OASIS HLB Evaporation to dryness 1mL MeOH/H 2 O (10:90, v/v) HPLC-MS/MS LC colum Purospher RP-18 (125x2mm) (5 µm) Quantification of target compounds: MRM mode: two transitions [M+H]+ > 2 Products in PI [M-H]- > 2 Products in NI HPLC-ESI-MS/MS (QqLIT) Gros et al., Journal of Chromatography A, 1248 (2012) 104-121 Analytical Protocol >> liquid chromatography coupled to tandem mass spectrometry
Target pharmaceuticals in sediment samples ANALYSIS Analgesics/antiinflammatories Acetaminophen Diclophenac Ibuprofen Indomethacine Ketoprofen Naproxen Mefenamic acid Lipid regulators and cholesterol lowering drugs Bezafibrate Fenofibrate Gemfibrozil Mevastatin Pravastatin Atrovastatin Macrolide antibiotics Erytromicin Roxithromycin Clarithromicin Josamycin Tylosin A Β-blockers Atenolol Sotalol Metoprolol Timolol Nadalol Pindolol Histimine H2 receptor antagonists Ranitidine Famotidine Cimetidine Psychiatric drugs Diazepam Lorazepam Carbamazepine Sulfonamid antibiotics Sulfamethazine Diuretic Hydrohlorothiazide Furosemide Β-agonists Clenbuterol Salbutamol Antihypertensive Nifuroxazide Enalapril Antidiabetic Glibenclamide Other antibiotics Trimethoprim Chloramphenicol Metronidazole Phenazone type of drugs Phenazone Barbiturates Butalbital Total=43 PhACs
Extraction of solid samples ANALYSIS pressurized liquid extraction Sample pre-treatment Sample extraction freeze-drying (-40 ºC, 0.044 bar) store at -20 ºC 1g of sediment sample; T=100ºC methanol/water, 1/2 v/v 3 static cycles (5 min) total flush volume 100% of cell 60s of nitrogen purge PLE - ASE 300 Dionex PLE extract (~22 ml) is diluted in 500 ml of HPLC water + 15mL Na 2 EDTA SPE extraction and clean-up SPE OASIS HLB HPLC-MS/MS (QqLIT) Elution with 8mL MeOH Reconsituted with 1mL MeOH/H 2 O (25:75, v/v) Addition of 34 isotopically labelled standards liquid chromatography coupled to tandem mass spectrometry Jelić et al., Talanta 80 (2009) 363-371
Quality parameters obtained during analysis, recoveries (%) and method detection limits (MDL) ANALYSIS Recovery, % (n=3), ± RSD Sea Lagoons Canal Effluent Influent Analgesics/antinflammatories 59-114% 50-120% 56-146% 57-112% 59-110% Lipid regulators and cholesterol lowering statin drugs 58-101% 60-96% 42-117% 70-99% 88-103% Psychiatric drugs 63-114% 51-124% 41-147% 61-113% 55-115% Histamine H1 and H2 receptor antagonist 62-124% 52-80% 40-95% 87-119% 50-112% β-blocking agents 71-126% 60-90% 57-120% 56-88% 57-109% Duretic 85-101% 68-113% 53-91% 55-98% 63-76% Antidiabetic 87% 104% 97% 103% 112% Antihypertensives 73-102% 60-90% 41-101% 51-85% 51-85% Antiplstelet agent 91% 95% 86% 90% 60% Prostatic hyperplasia 110% 50-97% 100% 112% 56% To treat asthma 72% 82% 73% 117% 108% Anticoagulant 82% 123% 90% 76% 62% X-ray contrast agent 102% 52-85% 100% 94% 70% Anti helmintics 55-120% 50-110% 50-115% 74-98% 60-112% Synthetic glucocorticoid 80% 60-118% 82% 74% 65% Sedation and muscle relaxation 85% 123% 88% 83% 87% Tranquilizer 90-92% 52-85% 93-100% 92-102% 76-92% Antibiotics 63-121% 57-128% 53-116% 66-116% 61-120% Calcium channel blocker 86-120% 60-127% 64-96% 83-115% 69-74% MDL (ng/l) 0.01-7.2 0.01-9 0.03-15.2 0.2-26 0.2-50 Sediment Recovery (%) Analgesics/antiinflam matories 60-118% Phenayone type drugs 87-110% Lipid regulators and cholesterol lowering statin drugs 77-115% Psychiatric drugs 80-105% Histamine H1 and H2 receptor antagonist 87-104% Macrolide antibiotics 68-95% Sulfonamid antibiotics 59-100% Other antibioptics 75-93% β-blockers 90-114% β-agonist 84-111% Diuretic 71-84% Antidiabetic 89-116% MDL (ng/g) 0.01-3.20
Elimination efficiency, % RESULTS Less then < 10 % removal >80% (good removal) citalopram tamsulosin diazepam losartan valsartan propanolol albendazole furosemide codeine hydrochlorothyazide oxycodone thiabendazole clarithromycin desloratadine diltiazem trimethoprim sulfamethoxazole atorvastatin nadolol gemfibrozil atenolol ranitidine salicylic acid naproxen ibuprofen bezafibrate 1 st Campaing Low to moderate removal rates were obtained for most compounds, except for analgesics and anti-inflammatories (>80%). 0 10 20 30 40 50 60 70 80 90 100 % removal WWTP_1 st campaign
Elimination efficiency, % RESULTS >80% (good removal) tamsulosin diazepam losartan carbamazepine valsartan propanolol furosemide codeine clopidrogel hydrochlorothyazide azaperone oxycodone thiabendazole clarithromycin desloratadine diltiazem trimethoprim sulfamethoxazole ciprofloxacine atorvastatin nadolol gemfibrozil atenolol ranitidine salicylic acid bezafibrate naproxen 2 nd Campaing The pharmaceuticals belonging to the same therapeutic groups show the similar removal for both sampling campaing. 0 10 20 30 40 50 60 70 80 90 100 % removal _2 nd campaign
Concentration (ng/l) Concentration (ng/l) Concentration of pharmaceuticals in WWTP effluents and in receiving natural water bodies 18000,00 16000,00 14000,00 12000,00 WWTP1 Calcium channel blocker Antibiotics Tranquilizer 10000,00 8000,00 6000,00 4000,00 2000,00 0,00 WWTP1 effluent 1st Campaing Downstream WWTP1 WWTP1 effluent 2nd Campaing Downstream WWTP1 Synthetic glucocorticoid Anti helmintics X-ray contrast agent Anticoagulant To treat asthma Prostatic hyperplasia Wastewater effluents are main source of pharmaceuticals in river waters Important - Dilution factor 14000,00 12000,00 WWTP2 the natural water bodies receiving effluents discharge has the same chemical profile as wastewater effluents, detected in µg/l range. 10000,00 8000,00 6000,00 4000,00 2000,00 0,00 WWTP2 effluent Emissary Sant Carles WWTP2 effluent Emissary Sant Carles 1st Campaing 2nd Campaing
Concentration (ng/l) Canals 10000,00 1000,00 100,00 10,00 Concentration range 1 464.2 ng/l dried Canals are used to assist in the growing of agricultural crops, thus water quality requirements are essential. Water from this canals could be significant route of crop contamination. 1,00 1 2 1 2 1 2 1 2 1 2 1 2 Canal A Canal B Canal C Canal D1 Canal D2 Canal D3 Analgesics/ant-inflammatories Psychiatric drugs β-blocking agents Antidiabetic Antiplstelet agent To treat asthma Synthetic glucocorticoid Antibiotics Lipid regulators and cholesterol lowering statin drugs Histamine H1 and H2 receptor antagonist Duretic Antihypertensives Prostatic hyperplasia Anti helmintics Tranquilizer Calcium channel blocker
Concentration (ng/l) Concetration (ng/l) Sea water and lagoons 10000,0 1000,0 100,0 10,0 1,0 Sea water 1 2 1 2 1 2 1 2 1 2 1 2 Bay Alfalcs A (beach) Bay Alfalcs B (shore) Bay Alfalcs C (inside) Concentration range 1-20 ng/l Bay Fangar A (beach) Bay Fangar B Bay Fangar C (shore) (inside) Antibiotics Tranquilizer Antihypertensives Duretic β-blocking agents Psychiatric drugs Analgesics/antinflammatories Iopromide tiabendazole Atenolo Sotalol Hydrochlorothyazid Ibersartan varsartan ibuprofen acetaminophen salicylic acid phenazone 1000,0 100,0 10,0 1,0 Lagoons Salycilic acid the most abundant compound with max conc. 252.8 ng/l 0,1 1 2 1 2 1 2 1 2 1 2 1 2 Tancada A (shore) Tancada A (inside) Buda A (shore) Buda B (inside) Encanizada A (shore) Encanizada A (inside)
Frequency of detection in water samples diltiazem levamisol clopidrogel sea water lagoons canals glibenclamide furosemide cimetidine alprazolam atorvastatin bezafibrate diclofenac naproxen clarithromycin sotalol 10,11-epoxyCBZ prophyphenazone ketoprofen azaperone venlafaxine iopromide thiabendazole valsartan acetaminophen ibuprofen salicylic acid phenazone valsartan irbersartan thiabendazole iopromide atenolol clarithromycin Analgesics/antiinflammatories Antihypertensives Antihelmintics X-ray contrast agents β-blocking agents Antibiotics actaminophen 0 20 40 60 80 100
Concentration (ng/g) Analgesics/antiinflammatories β-blockers Concentration (ng/g) Analgesics/antiinflammatories β-blockers Sediment samples with influence of WWTPs discharges 25,000 Lipid regulators and cholesterol lowering statin drugs The total concentration levels were up to 20 ng/g 20,000 WWTP2 Sulfonamide antibiotics 20,000 Antibiotics 18,000 Lipid regulators Antidiabetic 16,000 Antidiabetic WWTP1 14,000 Β-agonist 15,000 12,000 10,000 10,000 8,000 6,000 5,000 4,000 2,000 0,000 Upstream of WWTP1 Downstream WWTP1 0,000 Emissary SCr WWTP2 Canal C (WWTP2) Concentration range 0.09-7.08 ng/g Concentration range 0.01-5.98 ng/g
Concentration(ng/g) Concentration (ng/g) Concentration (ng/g) Sediment samples 16,00 14,00 12,00 10,00 8,00 6,00 4,00 2,00 0,00 Canals 0.01-4.54 ng/g ketoprofen Canal A Canal B Canal C Canal D1 Canal D4 8,00 7,00 6,00 5,00 4,00 3,00 2,00 1,00 0,00 Sea Bay Alfalcs Bay Alfalcs A (beach) B (shore) 0.11-2.81 ng/g Bay Alfalcs C (open sea) ranitidine Bay Fangar A (beach) Bay Fangar B (shore) Bay Fangar C (open sea) 9,00 8,00 7,00 6,00 5,00 4,00 3,00 2,00 1,00 Lagoons 0.06-3.42 ng/g ketoprofen Analgesics/antiinflammatories Phenayone type drugs Lipid regulators and cholesterol lowering statin Psychiatric drugs Histamine H1 and H2 receptor antagonist Macrolide antibiotics Sulfonamid antibiotics Other antibioptics β-blockers β-agonist Diuretic Antidiabetic 0,00 Tancada A (shore) Tancada A (inside) Buda A (shore) Buda B (inside) Encanizada A (shore) Encanizada A (inside) Canal Vell A (shore)
Frequency of detection in sediment samples metoprolol clarithromycin sulfamethaxzine cimetidine sea lagoon canal carbamazepine gemfibrozil naproxen sotalol atrovastatin actaminophen diclofenac bezafibrate ibuprofen Indomethacine furosemide metronidazole pravastatin phenazone ibuprofen ketoprofen chloramphenicol erythromycin ranitidine erythromycin chloramphenicol Histamine H2 receptor antagonists ranitidine bezafibrate diclofenac 0 20 40 60 80 100
CONCLUSION o Levels of PhACs detected in waste, river, canals, lagoons and sea water indicate that they are widespread pollutants along the Ebro Delta. o Wastewater treatment plants proven to be an important source of pollution for water bodies, as well as for sediment. o The compounds reaching the sea water coming from WWTP discharges were from the several therapeutical groups: analgesics/anti-inflammatories (such as acetaminophen, ibuprofen, salicylic acid, phenazone), β-blocking agents (atenolol, sotalol, ibersartan, valsartan) and diuretics drugs (iopromide, thiabendazole). o The compounds most widely detected in sediments were: diclofenac, bezafibrate, ranitidine, ibuprofen, erythromycin, chloramphenicol.
ACKNOWLEDGMENTS This work is supported by: The Spanish Ministry of Economy and Competitiveness through the coordinated project TRANSFORM COAST (CGL- 2014-56530-C4-4-R) Generalitat de Catalunya (Consolidated Research Groups 2014 SGR 291 ICRA and 2014 SGR 418 Water and Soil Quality Unit). M.Celic acknowledges for grant for PhD received from the Spanish Ministry of Economy and Competitiveness (BES-2015-072297).
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