Article pubs.acs.org/est
Mass Loading and Removal of Select Illicit Drugs in Two Wastewater Treatment Plants in New York State and Estimation of Illicit Drug Usage in Communities through Wastewater Analysis Bikram Subedi† and Kurunthachalam Kannan*,†,‡ †
Wadsworth Center, New York State Department of Health, and Department of Environmental Health Sciences, School of Public Health, State University of New York at Albany, Empire State Plaza, P.O. Box 509, Albany New York 12201-0509, United States ‡ Experimental Biochemistry Unit, King Fahd Medical Research Center, King Abdulaziz University, P.O. Box 80216, Jeddah 21589, Saudi Arabia S Supporting Information *
ABSTRACT: Sewage epidemiology is a rapidly expanding field that can provide information on illicit drug usage in communities, based on the measured concentrations in samples from wastewater treatment plants (WWTPs). In this study, select illicit drugs (six drugs and eight metabolites) were determined on a daily basis for a week in wastewater, suspended particulate matter (SPM), and sludge from two WWTPs in the Albany area in New York State. The WWTP that served a larger population (∼100 000, with a flow rate of 83 300 m3/d) showed 3.2 (methadone) to 51 (3,4-methylenedioxyamphetamine; MDA) times higher mass flows of illicit drugs than did the WWTP that served a smaller population (∼15 000, with a flow rate of 6850 m3/d). The consumption rate of target illicit drugs in the communities served by the two WWTPs was estimated to range from 1.67 to 3510 mg/d/1000 people. Between the dissolved and particulate phases, the fraction of methadone, 2-ethylidene-1,5dimethyl-3,3-diphenylpyrrolidine (EDDP), amphetamine, and MDA sorbed to SPM ranged from 34.3% to 41.1% of the total mass in the waste stream. The removal efficiencies of illicit drugs from the two WWTPs ranged from 4% (norcocaine) to 99% (cocaine); however, methamphetamine, methadone, and EDDP showed a negative removal in WWTPs. The environmental emission of illicit drugs from WWTP discharges was calculated to range from 0.38 (MDEA) to 67.5 (EDDP) mg/d/1000 people. Other markers such as caffeine, paraxanthine, nicotine, and cotinine were found to predict the concentrations of select illicit drugs in raw wastewater (r2 = 0.20−0.79; p ≤ 0.029).
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the occurrence of illicit drugs in wastewater from Spain,7−9 France,10,11 The Netherlands,12 Belgium,13,14 and the UK.1 Despite these findings, a very limited number of studies have determined the occurrence and fate of illicit drugs in WWTPs in the USA.6,15,16 Further, studies that describe the fate of illicit drugs in WWTPs made measurements only in the aqueous fraction, owing to the high water solubility of some drugs (Table S1), and the fraction of drugs sorbed to suspended particulate matter (SPM) was given less consideration.6,17 The estimation of the loadings of illicit drugs in WWTPs based only on the aqueous fraction can underestimate the actual load,18,19 particularly for those drugs that have high Koc values [for example, methadone and 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP) have log Koc values of 4.86 and 5.67, respectively, Table S1]. A few studies in Europe have reported the occurrence of illicit drugs in SPM and sewage sludge.1,20−22
INTRODUCTION The occurrence of illicit drugs in the environment has been recognized as an important ecosystem and public health issue, mainly due to these drugs’ high psychoactive properties and unidentified effects on nontarget organisms.1 The United Nations Office on Drugs and Crime (UNODC) estimated that there are between 165 and 315 million illicit-drug users globally and reported 210 000 illicit-drug related deaths in 2010.2 An estimated 14.9% of the U.S. population ≥12 years of age used illicit drugs in 2011.3 Cocaine is one of the major illicit drugs, second to cannabis, with an estimated 13.3 to 19.7 million global users in 2010.4 An estimated 94 tons of cocaine (estimated global manufacture of 776−1051 tons) were seized in 2011 in the USA.2 Following ingestion, drugs get into wastewater in different forms, as metabolites and/or unchanged, through human excretion. Wastewater treatment plants (WWTPs) are the primary sources of illicit drugs and their metabolic and transformation products to the environment.5 Select illicit drugs were found in WWTP effluent and influent,6 surface water,7,8 and drinking water7 at sub-ng/L to μg/L levels. Several studies have reported © 2014 American Chemical Society
Received: Revised: Accepted: Published: 6661
April 7, 2014 May 22, 2014 May 27, 2014 May 27, 2014 dx.doi.org/10.1021/es501709a | Environ. Sci. Technol. 2014, 48, 6661−6670
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chemical-spike or lag periods in WWTPs.15 The 24-h composite samples can underestimate the loadings of unstable drugs in WWTPs. Nevertheless, most studies that determined the fate of illicit drugs in WWTPs used 24-h composite samples.18,26,27 In our study, 24-h composite wastewater samples, including raw wastewater (influent), primary treated wastewater (primary effluent), secondary treated wastewater (effluent), and sludge were collected over a seven-day period, from Friday, July 12, to Thursday, July 18, 2013, consecutively from two WWTPs in the Albany area in New York State. The wastewater influents were collected after the initial treatment that included screening of large-sized debris followed by grit removal. The primary effluents were collected after the primary settling to remove settleable solids, grease, and scum. Primary treatments typically remove ∼25−35% biological oxygen demand (BOD5), ∼40−60% total suspended solids (TSS), and produce primary sludge.28 The wastewater effluents were collected after the secondary biological treatment with activated sludge, followed by final clarification. Secondary biological treatments remove ∼85−95% of the remaining BOD5, as well as TSS, and produce secondary sludge. The two plants are denoted as WWTPA (population served ∼15 000) and WWTPB (population served ∼100 000), with a treatment capacity of 2.5 and 35 MGD, respectively. Both WWTPs used activated sludge treatment, and the average daily wastewater inflow was 1.81 MGD for WWTPA (hydraulic retention: 24 h) and 22 MGD for WWTPB (hydraulic retention: 16 h). The plants received predominantly the domestic waste with ≈5% industrial waste in WWTPA and ≈25% industrial waste in WWTPB. Detailed information on the two WWTPs is provided in Table S2. The strategies of wastewater treatment employed in these WWTPs were described elsewhere.28 The activated sludge (3.9% solid, determined gravimetrically) samples were collected for 7 days in WWTPA; however, activated sludge and dewatered sludge (21.7% solid, determined gravimetrically) samples were collected from WWTPB for four consecutive days within a sampling week. The activated sludge samples from both WWTPs were the combined sludge produced after the primary and secondary treatments. All samples were collected in certified precleaned amber glass jars with Teflon-faced caps, shipped to the laboratory, and stored in a freezer at 4 °C until extraction. Sample Preparation. Wastewater samples were analyzed by following the methods described elsewhere with some modifications.19,29 Briefly, wastewater samples (100 mL) were centrifuged at 5000 rpm for 10 min, and the supernatant was filtered through a glass fiber filter (37 mm, pore size, 1 μm; GE Osmonics Inc., Minnetonka, MN) to separate SPM. The aqueous phase was spiked with a mixture of labeled internal standards of illicit drugs (10 ng) and human excretion markers (50 ng), mixed well, and allowed to equilibrate for ∼30 min at room temperature. The aqueous samples were extracted by passage through Oasis HLB 6 cm3 (200 mg; Waters, Milford, MA) solid phase extraction (SPE) cartridges. Prior to use, the cartridges were conditioned with 5 mL of methanol and 5 mL of milli-Q water, and wastewater samples were loaded at ∼1 mL/min. Cartridges were allowed to dry for ∼30 min under vacuum and then eluted with 6 mL of methanol followed by 3 mL of a mixture of acetone, methanol, and ethyl acetate (2:2:1 v/v/v). Cartridges also were eluted with 3 mL of methanol containing 5% ammonia. The eluents were combined and concentrated to ∼100 μL under a gentle stream of nitrogen
However, no previous studies have reported the occurrence of illicit drugs in SPM and sewage sludge from the USA. The usage of illicit drugs in a community has been estimated typically from survey questionnaires, crime statistics, drug seizures, and self-reported information.18,23 However, the estimation of usage based on survey questionnaires can have a high degree of uncertainty, which often results in an underestimation. The concept of “sewage epidemiology” utilizes the measured concentrations of illicit drugs in centralized WWTPs to back-calculate drug usage in local communities.1,20,24 Analysis of illicit drugs in WWTPs, a complement to the classic socio-epidemiological approach, provides authorities with nearreal time statistics to identify usage trends and rates, emerging hot spots of drug abuse, and effectiveness of countermeasures.20,25 The usefulness and limitations of the sewage epidemiology approaches have been reviewed in detail elsewhere.25 In this study, select illicit drugs (six drugs and eight metabolites) were analyzed in 24 h composite wastewater influent, primary effluent, final effluent, and sludge collected daily for 7 days in two WWTPs in the Albany area in New York State. Wastewater samples were filtered to determine the sorption of drugs to SPM; and the fraction of illicit drugs sorbed to the SPM was determined for the first time in wastewater samples in the USA. The data were utilized to estimate the mass loading of illicit drugs to WWTPs. Per-capita consumption of illicit drugs was calculated based on the mass loadings, excretion profiles of parent drug and its metabolites, and the population served by the WWTPs. The removal efficiencies of illicit drugs were determined for the first time based on the measured concentrations in influent, SPM, and effluent. The concentrations of illicit drugs determined in sewage sludge and wastewater effluent were used to calculate the total environmental emission from WWTPs. Select chemical markers of human excretion, such as caffeine and nicotine, were determined in wastewaters, and their use in the prediction of loadings of illicit drugs in WWTPs was examined.
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MATERIALS AND METHODS Reagents and Chemicals. Standard stock solutions (100 or 1000 μg/mL) of individual illicit drugs, including cocaine (CCN), benzoylecgonine (BEG), norcocaine (NCCN), cocaethylene/benzoylecgonine ethyl ester (CCE), morphine (MPH), morphine-3-β-D-glucuronide (M3G), morphine-6-β-D-glucuronide (M6G), methadone (MTD), 2-ethylidene-1,5-dimethyl-3,3diphenylpyrrolidine (EDDP, a metabolite of MTD), amphetamine (APT), methamphetamine (MAPT), 3,4-methylenedioxyamphetamine (MDA), 3,4-methylenedioxymethamphetamine (MDMA), 3,4-methylenedioxyethylamphetamine (MDEA), paraxanthine (PXT), and their corresponding deuterated internal standards, including CCN-D3, BEG-D8, NCCN-D3, CCE-D8, MPH-D6, M3G-D3, M6G-D3, MTD-D9, EDDP-D3, APT-D8, MAPT-D8, MDA-D5, MDMA-D5, and MDEA-D5 were purchased from Cerilliant (Round Rock, TX. Nicotine (NCT), NCT-D3, cotinine (CTN), CTN-D3, caffeine (CFI), 13C3−CFI, and PXT−D3 were purchased from Sigma-Aldrich (St. Louis, MO). The purity of all standards was ≥98.2%. Formic acid (98.2%) was from Sigma-Aldrich. Ultrapure water was prepared using a milli-Q ultrapure system (Barnstead International, Dubuque, IA). All standard stock solutions were stored at −20 °C. Sample Collection. Grab samples of wastewater can overor underestimate the daily loadings of illicit drugs due to the 6662
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at 35 °C using a TurboVap Evaporator (Zymark, Inc., Hopkinton, MA). The final volume of the extract was adjusted to 1 mL with methanol in an amber glass vial, and 10 μL of the extract was injected into HPLC-MS/MS. Sludge and SPM were analyzed by following the method described elsewhere with some modifications.30 Briefly, ∼0.1 g of freeze-dried sludge was spiked with a mixture of internal standards (10 ng) and allowed to equilibrate for ∼30 min at room temperature. Spiked sludge samples were vortex-mixed for 1 min and extracted with 6 mL of methanol−water mixture (5:3 v/v) using an ultrasonic bath (Branson Ultrasonics 3510RDTH; Danbury, CT) for 30 min. Extracts were centrifuged at 4500 rpm for 5 min (Eppendorf Centrifuge 5804, Hamburg, Germany), the supernatant was collected in a polypropylene tube, and the extraction was repeated with 6 mL of methanol. The extracts were combined and concentrated to ∼1 mL under a gentle stream of nitrogen. The concentrated extract was diluted with milli-Q water to ∼12 mL and purified by passage through Oasis HLB (6 cm3, 200 mg) cartridges, as described above for wastewater samples. The entire amount of SPM residue obtained by centrifugation and filtration of 100 mL of wastewater (as described above) was transferred to a clean and preweighed polypropylene tube along with a preweighed glassfiber filter, freeze-dried, and analyzed. The final volume of the extract was adjusted to 1 mL with methanol in an amber glass vial, and 10 μL of the extract was injected into HPLC-MS/MS for analysis. Instrumental Analysis. Target chemicals were analyzed using an API 2000 electrospray triple quadrupole mass spectrometer (ESI-MS/MS; Applied Biosystems, Foster City, CA), interfaced with an Agilent 1100 Series HPLC system (Agilent Technologies, Santa Clara, CA). The autosampler was maintained at 4 °C. The analytes were separated using an Ultra Biphenyl column (100 mm × 2.1 mm, 5 μm) (Restek Corporation, Bellefonte, PA). Methanol and water (0.1% formic acid) were used as mobile phases; a description of the mobile phase gradient flow is presented in Tables S3 and S4. Target analytes were determined by multiple reaction monitoring (MRM) in positive ionization mode. Detailed information on the MS/MS transitions is provided in the Supporting Information (Table S5). Analyte peak identification was based on the retention time (±0.05 min) and the ratio of qualitative to quantitative transition-ion responses (±30%). The quantitation of illicit drugs was performed by the isotope dilution method. Sevento nine-point calibration curves (concentrations ranging from 0.1 to 1000 ng/L for wastewater and 0.1 to 1000 ng/g for solid samples) were prepared and used for the quantification of illicit drugs. The calibrations curves were prepared by plotting a concentration-dependent response factor for each target analyte (peak area of analyte divided by peak area of the corresponding labeled internal standard) versus the response-dependent concentration factor (concentrations of analyte divided by concentration of internal standard). The regression coefficients (r2) from equal weighting quadratic regression were ≥0.99 for all target analytes. Quality Assurance and Quality Control. Quality assurance and quality control (QA/QC) protocols have been described elsewhere.29−31 Briefly, continuing calibration verification standards injected before and after every batch of sample analysis showed target analyte recoveries of 100 ± 30%. A method blank was analyzed with every batch of samples. The concentrations of target illicit drugs in method blanks were below the corresponding limit of quantification (LOQ).
The concentrations in solid matrices are presented on a dryweight basis, unless stated otherwise. One sample was selected randomly for matrix spike (MS) and matrix spike duplicate (MSD) analyses, with each batch of samples analyzed. Target analytes and their corresponding internal standards were spiked (10−50 ng), and passed through the entire analytical procedure. The background concentrations of analytes in samples were subtracted to calculate the recoveries. The average relative recoveries (n = 4) of drugs in wastewater samples ranged from 64 ± 33% (APT) to 121 ± 28% (CCE); however, relatively higher recoveries of CCN (162 ± 19%), which may have resulted from its elevated concentrations in samples, were found (i.e., our spike level was ∼50% of the actual concentrations in samples). Similarly, the average relative recoveries (n = 4) of illicit drugs in sludge and SPM ranged from 53 ± 33% (NCCN) to 139 ± 1.0% (M3G). Method Validation. The analytical method was validated by the spiking of the target analytes and calculation of recoveries after passage through the entire analytical procedure. Approximately, 100 mL of wastewater samples (n = 3) was spiked with 10 ng of target analytes and extracted through HLB cartridges as described above. Extracts were concentrated to ∼100 μL, spiked with 10 ng of labeled internal standards, and adjusted to a final volume of 1 mL with methanol. The average (n = 3) ± relative standard deviation of recoveries of target analytes was 101 ± 27% (Figure S1). The LOQs and limits of detection (LODs) were determined as the minimum concentration of analytes in sample extracts that provided a signal-tonoise ratio ≥10 and ≥3, respectively. LOQs and LODs of target drugs ranged 1−10 and 0.1−5.0 ng/L, respectively, in wastewater samples and 1−10 and 0.1−1.0 ng/g, respectively, in solid samples (Table 1). The relative standard deviation from seven consecutive analyses of one of the MS samples (see above) was ≤22% for all target analytes. Sewage Epidemiology Back-Calculations. The measured concentrations of illicit drugs in wastewater influent, SPM, stability of illicit drugs in wastewater, population served by the WWTPs, human excretion rate, and the daily inflow rate of wastewater at the two WWTPs were used to estimate drug usage at the community level. The fraction of analytes sorbed to the SPM and the loadings of illicit drugs to WWTPs were calculated using the following equations (eqs 1 and 2), as reported by Baker and Kasprzyk-Hordern1 and Baker et al.32 PSPM =
((CSPM × MSPM)/Vw ) × 100 ((CSPM × MSPM)/Vw ) + C W
(1)
⎞ ⎛ 100 mass load = Cw × F × ⎜ ⎟ ⎝ 100 + stability ⎠ ⎛ ⎞ 100 1 ×⎜ ⎟× ⎝ 100 − PSPM ⎠ 1000
(2)
where PSPM is the percentage of sorption of analyte to SPM, CSPM is the concentration of analytes in SPM (ng/g), MSPM is the mass of SPM analyzed (g), VW is the volume of wastewater (L) used to filter out MSPM, CW is the concentration of analyte in wastewater influent (ng/L), mass load is the amount of individual illicit drug introduced into WWTP (mg/d), F is the daily flow rate of wastewater influent (m3/d) over a 24 h period, and stability is a measure of stability change (%) of analyte after 72 h (Table S1).18 6663
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