Chemosphere 119 (2015) 1000–1006
Contents lists available at ScienceDirect
Chemosphere journal homepage: www.elsevier.com/locate/chemosphere
Emission of bisphenol analogues including bisphenol A and bisphenol F from wastewater treatment plants in Korea Sunggyu Lee a, Chunyang Liao b, Geum-Ju Song c, Kongtae Ra d, Kurunthachalam Kannan b,f,⇑, Hyo-Bang Moon a,⇑ a
Department of Marine Sciences and Convergent Technology, College of Science and Technology, Hanyang University, Ansan 426-791, Republic of Korea 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, NY 12201-0509, USA c Institute of Environmental and Energy Technology, POSTECH, Pohang 790-784, Republic of Korea d Marine Environment and Conservation Research Division, Korea Institute of Ocean Science and Technology (KIOST), Ansan 426-744, Republic of Korea f Biochemistry Department, Faculty of Science and Experimental Biochemistry Unit, King Fahd Medical Research Center, King Abdulaziz University, PO Box 80216, Jeddah 21589, Saudi Arabia b
h i g h l i g h t s
g r a p h i c a l a b s t r a c t
Occurrence of several BP analogues in
sludge was confirmed. High levels of bisphenol A were found
in industrial WWTP sludge. Emission fluxes of bisphenols via
WWTPs in Korea were estimated. This is the first nationwide survey to
determine BP analogues in sewage sludge of Korea.
a r t i c l e
i n f o
Article history: Received 24 April 2014 Received in revised form 29 August 2014 Accepted 1 September 2014
Handling Editor: Caroline Gaus Keywords: Bisphenol A Bisphenol F Emission Effluent Sludge WWTP
a b s t r a c t Due to the regulation on bisphenol A (BPA) in several industrialized countries, the demand for other bisphenol analogues (BPs) as substitutes for BPA is growing. Eight BPs were determined in sludge from 40 representative wastewater treatment plants (WWTPs) in Korea. Total concentrations of BPs (RBP) in sludge ranged from 70% for industrial. Table S2 shows detailed information on WWTP characteristics such as influent sources, WTC, IE, annual sludge production rate (ASP) and hydraulic retention time (HRT). All samples were poured into pre-cleaned polypropylene (PP) bottles, transported to the laboratory and stored in a freezer at 20 °C until extraction. 2.2. Sample preparation Sludge samples were extracted and analyzed by following the method described elsewhere with some modifications (Liao et al., 2012a,b). In brief, 0.1 g of freeze-dried sample was weighed and transferred into 15-mL PP tube. After spiking with 20 ng of 13 C12-BPA (Cambridge Isotope Laboratories, Andover, MA, USA), the sludge was extracted with a mixture of 5 mL methanol and water (5:3, v/v) by shaking for 60 min. The mixture was centrifuged at 5000g for 5 min (Eppendorf Centrifuge 5804, Hamburg, Germany), and the supernatant was transferred into a glass tube. The extraction step was repeated twice, and the extracts were combined and concentrated to 4 mL under a gentle nitrogen stream. The sample solution was diluted to 10 mL with 0.2% formic acid (pH 2.5), and the extracts were purified with a Rapid Trace SPE workstation (Caliper Life Sciences Inc., Hopkinton, MA, USA). The extract was loaded onto an Oasis MCX cartridge (60 mg/3 cc; Waters, Milford, MA), preconditioned with 5 mL of methanol and 5 mL of water. The cartridge was rinsed with 15 mL of 25% methanol in water and 5 mL of water. The target compounds were eluted with 5 mL of methanol, and the eluate was concentrated to 1 mL prior to liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis. 2.3. Instrumental analysis Concentrations of BPs in sample extracts were determined by an Applied Biosystems API 2000 electrospray triple quadrupole mass spectrometer (ESI-MS/MS; Applied Biosystems, Foster City, CA, USA), coupled with an Agilent 1100 Series HPLC (Agilent Technologies Inc., Santa Clara, CA, USA) equipped with a binary pump and an autosampler. An analytical column (Betasil C18, 100 2.1 mm column; Thermo Electron Corporation, Waltham, MA, USA), connected to a Javelin guard column (Betasil C18, 20 2.1 mm), was used for LC separation. The injection volume was 10 lL. The mobile phase was methanol and water at a flow rate of 0.3 mL min1 with a gradient as follows: 0–2 min, 15% methanol; 2–5 min, 15–50% methanol; 5–8 min, 50% methanol; 8–20 min, 50– 90% methanol; 20–21 min, 90–99% methanol; 21–25 min, 99% methanol; and 25–30 min, 15% methanol. The negative ion multiple reaction monitoring (MRM) mode was used. The MS/MS parameters were optimized by infusion of individual compounds into the mass spectrometer through a flow injection system. Nitrogen was used as
1002
S. Lee et al. / Chemosphere 119 (2015) 1000–1006
both the curtain and collision gas. The quantification was based on an external calibration method and corrected for the recoveries of the internal standard, 13C12-BPA. Total organic carbon (TOC) in sludge was measured using an Elemental Analyzer (Flash 2000 series, Thermo Scientific Co., MA, USA) after removal of inorganic carbon with 1 N HCl. The moisture content in sludge was measured from their weights before and after freeze-drying. The moisture content ranged from 54% to 82% with a mean value of 69% (Table S2).
2.4. Quality assurance and quality control (QA/QC) Procedural blanks were analyzed with every batch of samples to monitor for background contamination. Trace levels of BPA and BPF (approximately 0.27 and 0.02 ng mL1, respectively) were detected in procedural blanks, and the background levels were subtracted from each sample concentrations. To validate the analytical method used in our study, spiked blanks and matrix-spiked samples were analyzed routinely. Recoveries of BPs (40 ng for each compound and 20 ng for 13C12-BPA) in spiked blanks (n = 4) ranged from 65.6 ± 1.53% (mean ± SD) for BPAF to 146 ± 1.36% for BPF. Recoveries of BPs (40 ng for each individual compound and 20 ng for 13 C12-BPA) in matrix-spiked samples (n = 4) ranged from 55.9 ± 12.7% for BPP to 157 ± 9.64% for BPS. Further details regarding QA/QC are provided in the Supporting Information (Table S3). The limit of quantitation (LOQs) were calculated from the lowest acceptable calibration standard and a nominal sample weight of 0.1 g. The calculated LOQs were 0.50 ng g1 for BPA, BPAF, BPAP and BPS; 1.00 ng g1 for BPB and BPP; and 2.00 ng g1 for BPF and BPZ. A midpoint calibration standard was injected after every 20 samples as a check for drift in instrumental sensitivity. Pure solvent (methanol) was injected periodically as a check for carryover of BPs between the samples analyzed. Instrumental calibration was verified daily by the injection of 10 calibration standards at concentrations ranging from 0.02 to 100 ng mL1 and the linearity of the calibration curve (r) was >0.99 for each of the target compounds. The concentrations of BPs in sludge are reported on a dry weight (dw) basis.
2.5. Calculation of emission fluxes of BPs via sludge and effluent discharges The emission flux of BPs through sludge was calculated by multiplying the concentrations (wet weight basis) of BPs in sludge with the annual sludge production rate (tons year1) for each of the studied WWTP (Table S1). The emission flux of BPs through effluent discharges was calculated from the measured concentrations (wet weight basis) of BPs in sludge, the adsorption coefficient, and removal rates (Table S3) during the WWTP processes. The emission fluxes of BPs through sludge (Es) and effluent (Ee) were calculated as follows:
Es ¼
C BPs ASP 365
Ee ¼
ð100 %S %BÞ Es %S
where Es is the emission flux of BPs through sludge (g d1), CBPs is the concentrations of BPs in sludge (ng g1 wet weight) and ASP is the annual sludge production rate (tons year1) for each of the studied WWTP. Ee is the emission flux of BPs through effluent (g d1), %S is the adsorption rate of BPs onto the sludge, and %B is the biodegradation rate of BPs during wastewater treatment. Percapita environmental emissions (mg capita1 d) of BPs through each WWTP were calculated based on the population-served by the plants and the total environmental emission fluxes (sum of Es and Ee). 2.6. Statistical analysis Concentrations below the LOQ were assigned a zero value for the calculation of the mean and median, but were assigned as a value of one-half of the respective LOQ for statistical analyses. One-way ANOVA with Turkey test was performed to assess differences in the concentrations of BPs among three different WWTP types. Spearman rank correlation analysis was performed to investigate the relationships between the concentrations of BPs in sludge and WWTP characteristics such as WTC, IE, ASP and HRT as well as
Table 1 Concentrations of bisphenol analogues (ng g1 dry weight) in sludge from wastewater treatment plants in Korea. BPA
a b c
BPAF
BPAP
BPB
BPF
BPP
BPS
BPZ
RBPs
Domestic WWTPs (n = 16) Mean 292 Median 275 Range 46.4–986 DRb 100
– – – 0
1.98 NAa nd-16.0 12.5
– – – 0
544 554 86.7–1780 100
1.06 NA nd-17.0 6.25
7.06 3.60 nd-34.9 75.0
– – – 0
850 690 140–2100 100
Mixed WWTPs (n = 9) Mean 1050 Median 426 Range 20.6–5593 DR 100
0.70 NA nd-3.59 22.2
– – – 0
– – – 0
699 692 561–947 100
2.53 NA nd-22.8 11.1
63.4 47.3 nd-268 77.8
– – – 0
1820 1340 630–6340 100
Industrial WWTPs (n = 15) Mean 3100 Median 145 Range ndc–25 600 DR 93.3
0.31 NA nd-3.51 13.3
– – – 0
– – – 0
24.8 NA nd-167 33.3
2.71 NA nd-26.2 13.3
74.3 2.47 nd-523 60.0
64.8 NA nd-969 13.3
3270 740 nd-25 700 93.3
Total (n = 40) Mean Median Range DR
0.28 NA nd-3.59 10.0
0.79 NA nd-16.0 5.00
– – – 0
384 249 nd-1780 75.0
2.01 NA nd-26.2 10.0
44.9 3.80 nd-523 70.0
24.3 NA nd-969 5.00
1970 769 nd-25 700 97.5
1520 275 nd-25 600 97.5
NA = Not available. DR = Detection rate (%). nd = Not detected.
1003
S. Lee et al. / Chemosphere 119 (2015) 1000–1006
TOC contents. Statistical analyses were performed using SPSS 18.0 K for Windows. Statistical significance was set at p < 0.05.
3. Results and discussion 3.1. Occurrence and concentrations of BPs in sludge Concentrations of BPs in sludge collected from WWTPs in Korea are summarized in Table 1. The concentrations of total BPs (RBP; the sum of 8 bisphenol analogues) ranged from
Contents lists available at ScienceDirect
Chemosphere journal homepage: www.elsevier.com/locate/chemosphere
Emission of bisphenol analogues including bisphenol A and bisphenol F from wastewater treatment plants in Korea Sunggyu Lee a, Chunyang Liao b, Geum-Ju Song c, Kongtae Ra d, Kurunthachalam Kannan b,f,⇑, Hyo-Bang Moon a,⇑ a
Department of Marine Sciences and Convergent Technology, College of Science and Technology, Hanyang University, Ansan 426-791, Republic of Korea 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, NY 12201-0509, USA c Institute of Environmental and Energy Technology, POSTECH, Pohang 790-784, Republic of Korea d Marine Environment and Conservation Research Division, Korea Institute of Ocean Science and Technology (KIOST), Ansan 426-744, Republic of Korea f Biochemistry Department, Faculty of Science and Experimental Biochemistry Unit, King Fahd Medical Research Center, King Abdulaziz University, PO Box 80216, Jeddah 21589, Saudi Arabia b
h i g h l i g h t s
g r a p h i c a l a b s t r a c t
Occurrence of several BP analogues in
sludge was confirmed. High levels of bisphenol A were found
in industrial WWTP sludge. Emission fluxes of bisphenols via
WWTPs in Korea were estimated. This is the first nationwide survey to
determine BP analogues in sewage sludge of Korea.
a r t i c l e
i n f o
Article history: Received 24 April 2014 Received in revised form 29 August 2014 Accepted 1 September 2014
Handling Editor: Caroline Gaus Keywords: Bisphenol A Bisphenol F Emission Effluent Sludge WWTP
a b s t r a c t Due to the regulation on bisphenol A (BPA) in several industrialized countries, the demand for other bisphenol analogues (BPs) as substitutes for BPA is growing. Eight BPs were determined in sludge from 40 representative wastewater treatment plants (WWTPs) in Korea. Total concentrations of BPs (RBP) in sludge ranged from 70% for industrial. Table S2 shows detailed information on WWTP characteristics such as influent sources, WTC, IE, annual sludge production rate (ASP) and hydraulic retention time (HRT). All samples were poured into pre-cleaned polypropylene (PP) bottles, transported to the laboratory and stored in a freezer at 20 °C until extraction. 2.2. Sample preparation Sludge samples were extracted and analyzed by following the method described elsewhere with some modifications (Liao et al., 2012a,b). In brief, 0.1 g of freeze-dried sample was weighed and transferred into 15-mL PP tube. After spiking with 20 ng of 13 C12-BPA (Cambridge Isotope Laboratories, Andover, MA, USA), the sludge was extracted with a mixture of 5 mL methanol and water (5:3, v/v) by shaking for 60 min. The mixture was centrifuged at 5000g for 5 min (Eppendorf Centrifuge 5804, Hamburg, Germany), and the supernatant was transferred into a glass tube. The extraction step was repeated twice, and the extracts were combined and concentrated to 4 mL under a gentle nitrogen stream. The sample solution was diluted to 10 mL with 0.2% formic acid (pH 2.5), and the extracts were purified with a Rapid Trace SPE workstation (Caliper Life Sciences Inc., Hopkinton, MA, USA). The extract was loaded onto an Oasis MCX cartridge (60 mg/3 cc; Waters, Milford, MA), preconditioned with 5 mL of methanol and 5 mL of water. The cartridge was rinsed with 15 mL of 25% methanol in water and 5 mL of water. The target compounds were eluted with 5 mL of methanol, and the eluate was concentrated to 1 mL prior to liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis. 2.3. Instrumental analysis Concentrations of BPs in sample extracts were determined by an Applied Biosystems API 2000 electrospray triple quadrupole mass spectrometer (ESI-MS/MS; Applied Biosystems, Foster City, CA, USA), coupled with an Agilent 1100 Series HPLC (Agilent Technologies Inc., Santa Clara, CA, USA) equipped with a binary pump and an autosampler. An analytical column (Betasil C18, 100 2.1 mm column; Thermo Electron Corporation, Waltham, MA, USA), connected to a Javelin guard column (Betasil C18, 20 2.1 mm), was used for LC separation. The injection volume was 10 lL. The mobile phase was methanol and water at a flow rate of 0.3 mL min1 with a gradient as follows: 0–2 min, 15% methanol; 2–5 min, 15–50% methanol; 5–8 min, 50% methanol; 8–20 min, 50– 90% methanol; 20–21 min, 90–99% methanol; 21–25 min, 99% methanol; and 25–30 min, 15% methanol. The negative ion multiple reaction monitoring (MRM) mode was used. The MS/MS parameters were optimized by infusion of individual compounds into the mass spectrometer through a flow injection system. Nitrogen was used as
1002
S. Lee et al. / Chemosphere 119 (2015) 1000–1006
both the curtain and collision gas. The quantification was based on an external calibration method and corrected for the recoveries of the internal standard, 13C12-BPA. Total organic carbon (TOC) in sludge was measured using an Elemental Analyzer (Flash 2000 series, Thermo Scientific Co., MA, USA) after removal of inorganic carbon with 1 N HCl. The moisture content in sludge was measured from their weights before and after freeze-drying. The moisture content ranged from 54% to 82% with a mean value of 69% (Table S2).
2.4. Quality assurance and quality control (QA/QC) Procedural blanks were analyzed with every batch of samples to monitor for background contamination. Trace levels of BPA and BPF (approximately 0.27 and 0.02 ng mL1, respectively) were detected in procedural blanks, and the background levels were subtracted from each sample concentrations. To validate the analytical method used in our study, spiked blanks and matrix-spiked samples were analyzed routinely. Recoveries of BPs (40 ng for each compound and 20 ng for 13C12-BPA) in spiked blanks (n = 4) ranged from 65.6 ± 1.53% (mean ± SD) for BPAF to 146 ± 1.36% for BPF. Recoveries of BPs (40 ng for each individual compound and 20 ng for 13 C12-BPA) in matrix-spiked samples (n = 4) ranged from 55.9 ± 12.7% for BPP to 157 ± 9.64% for BPS. Further details regarding QA/QC are provided in the Supporting Information (Table S3). The limit of quantitation (LOQs) were calculated from the lowest acceptable calibration standard and a nominal sample weight of 0.1 g. The calculated LOQs were 0.50 ng g1 for BPA, BPAF, BPAP and BPS; 1.00 ng g1 for BPB and BPP; and 2.00 ng g1 for BPF and BPZ. A midpoint calibration standard was injected after every 20 samples as a check for drift in instrumental sensitivity. Pure solvent (methanol) was injected periodically as a check for carryover of BPs between the samples analyzed. Instrumental calibration was verified daily by the injection of 10 calibration standards at concentrations ranging from 0.02 to 100 ng mL1 and the linearity of the calibration curve (r) was >0.99 for each of the target compounds. The concentrations of BPs in sludge are reported on a dry weight (dw) basis.
2.5. Calculation of emission fluxes of BPs via sludge and effluent discharges The emission flux of BPs through sludge was calculated by multiplying the concentrations (wet weight basis) of BPs in sludge with the annual sludge production rate (tons year1) for each of the studied WWTP (Table S1). The emission flux of BPs through effluent discharges was calculated from the measured concentrations (wet weight basis) of BPs in sludge, the adsorption coefficient, and removal rates (Table S3) during the WWTP processes. The emission fluxes of BPs through sludge (Es) and effluent (Ee) were calculated as follows:
Es ¼
C BPs ASP 365
Ee ¼
ð100 %S %BÞ Es %S
where Es is the emission flux of BPs through sludge (g d1), CBPs is the concentrations of BPs in sludge (ng g1 wet weight) and ASP is the annual sludge production rate (tons year1) for each of the studied WWTP. Ee is the emission flux of BPs through effluent (g d1), %S is the adsorption rate of BPs onto the sludge, and %B is the biodegradation rate of BPs during wastewater treatment. Percapita environmental emissions (mg capita1 d) of BPs through each WWTP were calculated based on the population-served by the plants and the total environmental emission fluxes (sum of Es and Ee). 2.6. Statistical analysis Concentrations below the LOQ were assigned a zero value for the calculation of the mean and median, but were assigned as a value of one-half of the respective LOQ for statistical analyses. One-way ANOVA with Turkey test was performed to assess differences in the concentrations of BPs among three different WWTP types. Spearman rank correlation analysis was performed to investigate the relationships between the concentrations of BPs in sludge and WWTP characteristics such as WTC, IE, ASP and HRT as well as
Table 1 Concentrations of bisphenol analogues (ng g1 dry weight) in sludge from wastewater treatment plants in Korea. BPA
a b c
BPAF
BPAP
BPB
BPF
BPP
BPS
BPZ
RBPs
Domestic WWTPs (n = 16) Mean 292 Median 275 Range 46.4–986 DRb 100
– – – 0
1.98 NAa nd-16.0 12.5
– – – 0
544 554 86.7–1780 100
1.06 NA nd-17.0 6.25
7.06 3.60 nd-34.9 75.0
– – – 0
850 690 140–2100 100
Mixed WWTPs (n = 9) Mean 1050 Median 426 Range 20.6–5593 DR 100
0.70 NA nd-3.59 22.2
– – – 0
– – – 0
699 692 561–947 100
2.53 NA nd-22.8 11.1
63.4 47.3 nd-268 77.8
– – – 0
1820 1340 630–6340 100
Industrial WWTPs (n = 15) Mean 3100 Median 145 Range ndc–25 600 DR 93.3
0.31 NA nd-3.51 13.3
– – – 0
– – – 0
24.8 NA nd-167 33.3
2.71 NA nd-26.2 13.3
74.3 2.47 nd-523 60.0
64.8 NA nd-969 13.3
3270 740 nd-25 700 93.3
Total (n = 40) Mean Median Range DR
0.28 NA nd-3.59 10.0
0.79 NA nd-16.0 5.00
– – – 0
384 249 nd-1780 75.0
2.01 NA nd-26.2 10.0
44.9 3.80 nd-523 70.0
24.3 NA nd-969 5.00
1970 769 nd-25 700 97.5
1520 275 nd-25 600 97.5
NA = Not available. DR = Detection rate (%). nd = Not detected.
1003
S. Lee et al. / Chemosphere 119 (2015) 1000–1006
TOC contents. Statistical analyses were performed using SPSS 18.0 K for Windows. Statistical significance was set at p < 0.05.
3. Results and discussion 3.1. Occurrence and concentrations of BPs in sludge Concentrations of BPs in sludge collected from WWTPs in Korea are summarized in Table 1. The concentrations of total BPs (RBP; the sum of 8 bisphenol analogues) ranged from
