Elevated Accumulation of Parabens and their Metabolites in Marine

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Elevated Accumulation of Parabens and their Metabolites in Marine Mammals from the United States Coastal Waters Jingchuan Xue, Nozomi Sasaki, Madhavan Elangovan, Guthrie Diamond, and Kurunthachalam Kannan Environ. Sci. Technol., Just Accepted Manuscript • Publication Date (Web): 17 Sep 2015 Downloaded from http://pubs.acs.org on September 22, 2015

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Environmental Science & Technology

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Elevated Accumulation of Parabens and their Metabolites in Marine

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Mammals from the United States Coastal Waters

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Jingchuan Xue1, Nozomi Sasaki2, Madhavan Elangovan1, Guthrie Diamond1, Kurunthachalam Kannan1,3,*

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1

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, Albany, NY 12201, United States 2

Department of Fisheries and Wildlife, School of Agriculture Science, Oregon State University, Corvallis, OR 97331, United States 3

Biochemistry Department, Faculty of Science and Experimental Biochemistry Unit, King Fahd Medical Research Center, King Abdulaziz University, Jeddah, Saudi Arabia

*Corresponding author: K. Kannan Wadsworth Center Empire State Plaza, P.O. Box 509 Albany, NY 12201-0509 Tel: +1-518-474-0015 Fax: +1-518-473-2895 E-mail: [email protected]

Submission to: ES&T

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ACS Paragon Plus Environment


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Abstract: The widespread exposure of humans to parabens present in personal care products

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is well known. Nevertheless, little is known about the accumulation of parabens in marine

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organisms. In this study, six parabens and four common metabolites of parabens were

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measured in 121 tissue samples from eight species of marine mammals collected along the

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coastal waters of Florida, California, Washington, and Alaska. Methyl paraben (MeP) was the

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predominant compound found in the majority of the marine mammal tissues analyzed, and

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the highest concentration found was 865 ng/g (wet weight [wet wt]) in the livers of bottlenose

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dolphins from Sarasota Bay, FL. 4-Hydroxybenzoic acid (4-HB) was the predominant

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paraben metabolite found in all tissue samples. The measured concentrations of 4-HB were

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on the order of hundreds to thousands of ng/g tissue, and these values are some of the highest

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ever reported in the literature. MeP and 4-HB concentrations showed a significant positive

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correlation (p < 0.05), which suggested a common source of exposure to these compounds in

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marine mammals. Trace concentrations of MeP and 4-HB were found in the livers of polar

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bears from the Chuckchi Sea and Beaufort Sea, which suggested widespread distribution of

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MeP and 4-HB in the oceanic environment.

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Key words: parabens; MeP; 4-HB; marine mammals; sea otter; dolphin; liver

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Introduction

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The esters of p-hydroxybenzoic acid (hereafter referred to as parabens) are a class of

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chemicals that include methyl-paraben (MeP), ethyl-paraben (EtP), propyl-paraben (PrP),

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butyl-paraben (BuP), heptyl-paraben (HeP), and benzyl-paraben (BzP). As the most

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commonly employed preservatives, parabens are used in processed foods, cosmetics, and

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pharmaceuticals.1-3 MeP and PrP are the most widely used parabens and are often used in

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combination due to their synergistic antimicrobial action. P-hydroxybenzoic acid

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(4-hydroxybenzoic acid [4-HB]) is the major metabolite of several parabens in both animals

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and

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(3,4-dihydroxybenzoic acid [3,4-DHB]), methyl protocatechuate (OH-MeP), and ethyl

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protocatechuate (OH-EtP).8

humans.1,

4-7

Other

metabolites

of

parabens

include

protocatechuate

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Concern over the safety of parabens was triggered by a study that showed an

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association between the incidence of breast cancer and the use of cosmetics that contain

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parabens.9 Although the debate continues,10,11 several studies have shown that parabens and

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their metabolites possess estrogenic activity. 12,13 Parabens were also reported to exert adverse

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effects on male reproductive system in rats.14

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Several studies have reported the occurrence of parabens and their metabolites in

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human specimens, such as urine and plasma.15-20 Parabens were recently found in human

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adipose tissue, with 4-HB found at concentrations of up to 17,400 ng/g.21 Accumulation of

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parabens and their metabolites in human adipose tissue provided new insights into their

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bioaccumulation potential. Little is known on the occurrence and accumulation of parabens

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and their metabolites in wildlife, especially in marine organisms. Parabens are metabolized to

3



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4-HB by esterases present in the intestine and liver,22 and 4-HB is expected to concentrate in

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the livers of marine mammals. The objective of this study was to investigate the

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accumulation of parabens and their metabolites, for the first time, in the tissues of marine

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mammals collected from the coastal and open ocean waters of the United States.

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Materials and Methods

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Standards and Reagents

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Analytical standards of MeP (~98%), BuP (~98%), and 4-HB (~98%) were purchased from

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Cambridge Isotope Laboratories (Andover, MA, USA). 3,4-DHB (≥97%), OH-MeP (~97%),

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and OH-EtP (~97%) were purchased from Sigma-Aldrich (St. Louis, MO, USA). 13C6-4-HB

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was purchased from Cambridge Isotope Laboratories (Andover, MA, USA). Analytical

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standards of EtP, PrP, BzP, and HeP were purchased from AccuStandard Inc. (New Haven, CT,

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USA). D4-HeP and D4-BzP were obtained from C/D/N Isotopes Inc (Pointe-Claire, Quebec,

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Canada). Mixed solutions of 13C6-MeP and 13C6-BuP were purchased from Sigma-Aldrich (St.

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Louis, MO, USA). The stock solutions of target analytes and internal standards (ISs) were

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prepared at 1 mg/mL in methanol and stored at -20° C until use. The chemical structures of

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the target analytes and ISs are shown in Table S1. Methanol (HPLC grade), acetone (ACS

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grade), and acetonitrile (ACS grade) were purchased from Mallinckrodt Baker (Phillipsburg,

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NJ, USA). Milli-Q water was purified by an ultrapure water system (Barnstead International;

96

Dubuque, IA, USA).

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Sample Collection

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A total of 121 tissue samples from eight species of marine mammals were analyzed in this

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study. The tissue samples were collected as part of our previous investigations on

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bioaccumulation of chemical contaminants in marine mammals, as reported elsewhere.23-25

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The marine mammals analyzed were pygmy sperm whale (Kogia breviceps), clymene

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dolphin (Stenella clymene), rough-toothed dolphin (Steno bredanensis), striped dolphin

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(Stenella coeruleoalba), bottlenose dolphin (Tursiops truncatus), southern sea otter (Enhydra

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lutris nereis), northern sea otter (Enhydra lutris kenyoni), and polar bear (Ursus maritimus),

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from the coastal waters of Florida, California, Washington, and Alaska. Details of sample

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collection have been described elsewhere.23-25 Briefly, tissues of marine mammals had been

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obtained from federal or state agencies or university laboratories. Livers and the blubber of

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cetaceans were collected by Mote Marine Laboratory, Sarasota, FL, from dolphins stranded

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along the Florida coast. Livers, kidneys, and brains of sea otters from California, Washington,

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and Alaska were collected through a stranding network coordinated by the U.S. Fish and

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Wildlife Service, with the cooperation of other state and federal agencies, and were obtained

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from the National Wildlife Health Center, Madison, WI. Livers of polar bears from Alaska

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were obtained from the native subsistence hunters and the U.S. Fish and Wildlife Service in

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Anchorage, AK. Information regarding collection date, age, sex, and the cause of death were

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recorded, when available. All samples were collected from 1994 to 2004. Generally, tissues

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were collected from the carcasses at the time of necropsy and were wrapped in aluminum foil,

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placed in airtight plastic bags, and frozen immediately at -20° C until analysis. Sampling

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locations of different species are shown in Figure 1.

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Sample Preparation

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Parabens are present in many cosmetic products, and contamination can arise during necropsy,

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storage, and analysis. The surface of each tissue sample was removed with clean scissors and

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tweezers, and only the inner portion of the tissue from each sample was used for analysis.

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Extraction and purification procedures were similar to those previously reported for human

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adipose tissue samples, with minor modifications.26 Briefly, 200 to 300 milligrams of tissue

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were accurately weighed and spiked with 50 ng of IS mixture (13C6-MeP,

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d4-BzP, and 13C6-4-HB). After equilibration for 30 min at room temperature, 5 mL of acetone

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were added to the sample. The mixture was homogenized in a mortar and then transferred to a

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15 mL polypropylene (PP) tube by washing with 2 mL of mixture of methanol and

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acetonitrile (1:1, v/v). The combined extracts were shaken in an oscillator shaker for 60 min

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and then centrifuged at 5000 x g for 5 min (Eppendorf Centrifuge 5804, Hamburg, Germany).

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The supernatant was then transferred to a new PP tube, and the mixture was concentrated to

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near dryness under a gentle nitrogen stream. One milliliter of a mixture of methanol and

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acetonitrile (1:1, v/v) was added, and an ultra-low temperature (-20° C) incubation was

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employed to separate lipids from the organic solvent layer. After storage at 20° C for 30 min

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and immediate centrifugation at 5000 x g for 5 min, the supernatant was transferred into a

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vial for analysis.

13

C6-BuP, d4-HeP,

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Instrumental Analysis

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Chromatographic separation was carried out using an Agilent 1100 Series HPLC system

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(Agilent Technologies Inc., Santa Clara, CA, USA). Identification and quantification of target

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analytes was performed with an Applied Biosystems API 2000 electrospray triple quadrupole

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mass spectrometer (ESI-MS/MS; Applied Biosystems, Foster City, CA, USA). A Zorbax

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SB-Aq (150 mm × 2.1 mm, 3.5 µm) column, serially connected to a Javelin guard column

146

(Betasil C18, 20 mm × 2.1 mm, 5 µm), was used for the separation. The injection volume

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was 10 µL, and the mobile phase comprised methanol (A) and Milli-Q water that contained

148

0.4% (v/v) acetic acid (B). The target compounds were separated by gradient elution at a flow

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rate of 300 µL/min starting at 5% (v/v) A, held for 3 min, increased to 85% A within 2 min,

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held for 1 min, further increased to 98% A within 2 min, held for 7 min, and reverted to 5% at

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the 16th min and held for 7 min. The MS/MS system was operated in the negative ion

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multiple-reaction monitoring mode. The compound specific MS/MS parameters are shown in

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Table S1. Nitrogen was used as the curtain and the collision gas at flow rates of 20 and 2 psi,

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respectively. The electrospray ionization voltage was set at -4.5 kV. The source heater was set

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at 450° C. The nebulizer gas (ion source gas 1) was set at 55 psi, and the heater gas (ion

156

source gas 2) was set at 70 psi. The data acquisition was performed at a scan speed of 25 ms

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and a resolving power of 0.70 full width at half maximum. Typical LC-MS/MS

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chromatograms of select analytes in marine mammal tissues are shown in Figure S1.

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Quality Assurance and Quality Control (QA/QC)

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Quantification was performed by the isotope dilution method based on the responses of

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13

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HeP), d4-BzP (for BzP), and

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standard calibration curve, with concentrations ranging from 0.1 to 200 ng/mL (1,000 ng/mL

C6-MeP (for MeP, OH-MeP, EtP, and OH-EtP),

13

C6-BuP (for BuP and PrP), d4-HeP (for

13

C6-4-HB (for 4-HB and 3, 4-DHB). An 11- to 13-point

7



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for 3, 4-DHB and 4-HB), was used for the quantification of each target analyte. A calibration

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curve was constructed by plotting the concentration-response factor for each target analyte

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(analyte peak area divided by corresponding IS peak area) versus the response-dependent

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concentration factor (concentration of analyte divided by concentration of IS). The regression

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coefficients (r) were ≥0.99 for all calibration curves. The lowest concentration on the

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calibration curve with a signal-to-noise ratio (S/N) of >10 was regarded as the instrumental

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limits of quantification (LOQs). Method LOQs (MLOQs) were determined from the

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post-matrix spiked 6-point calibration curves or estimated as 10 times the S/N ratio. As a

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check for instrumentation drift in response factors, a midpoint calibration standard was

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injected after every 10 samples. To prevent carryover of target chemicals from sample to

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sample, a pure solvent (methanol) was injected after every 10 samples.

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Several procedural blanks were analyzed with each batch of samples to evaluate

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contamination arising from laboratory materials and solvents. Efforts were taken to minimize

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background levels of target analytes. All glassware and materials used in the analysis were

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thoroughly rinsed with acetone, methanol, and Milli-Q water and baked at 300° C prior to use.

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Trace levels of MeP (mean: 0.52 ng/ml), PrP (0.87 ng/ml), 3,4-DHB (13.6 ng/ml), and 4-HB

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(3.86 ng/ml) were found in procedural blanks (Table S2). The concentrations of target

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analytes detected in procedural blanks were subtracted from sample values. Other QA/QC

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protocols include pre-extraction matrix spike (MS), pre-extraction matrix spike duplicate

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(MSD), and post-extraction matrix spike (MM), which were analyzed to determine accuracy

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and precision (results presented below). For each matrix analyzed (for both tissues and

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species), three samples were randomly selected, and 50 ng of target analytes and ISs were

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spiked into MS, MSD, and MM to determine recoveries. Precision was evaluated as a relative

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percentage difference (RPD), which was calculated using the following equation (1).

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ܴܲ‫[ܦ‬%] =

190

where ‫ܥ‬ெௌ and ‫ܥ‬ெௌ஽ are the concentrations determined in MS and MSD, respectively.

|஼ಾೄ ି஼ಾೄವ |×ଵ଴଴ ಴ಾೄ శ಴ಾೄವ మ

-----(1)

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Data Analysis

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The MS/MS data were acquired with Analyst software, Version 1.4.1 (Applied Biosystems,

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Foster City, CA, USA). Statistical analyses were performed with statistics software package

195

R v.3.1.0 and Microsoft Excel 2007. For the calculation of geometric mean and arithmetic

196

mean, concentrations below the LOQ were substituted with a value equal to half of the LOQ.

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To examine the relationship between analytes in the same tissue or the same analytes in

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different tissues, Spearman (when data followed a normal distribution after logarithmic

199

transformation) or Pearson (when data did not follow a normal distribution after logarithmic

200

transformation) correlation analysis was used. Only those samples with detectable

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concentrations of both chemicals or in both tissues were used in correlation analysis. To

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investigate the difference between means for the same analyte in different categories, a

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Student’s t-test (for those chemicals that followed a normal distribution after logarithmic

204

transformation) or Mann-Whitney U test (for those chemicals that did not follow a normal

205

distribution

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Quantile-Quantile (Q-Q) plot were used to determine if the data were normally distributed or

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not. The concentrations of target analytes in marine mammal tissues are presented on a wet

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weight (wet wt) basis, unless stated otherwise. Statistical significance was set at p < 0.05.

after

logarithmic

transformation)

was

used.

9


Shapiro-Wilk

test

and


209 210

Method Performance

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The MLOQs of target analytes in various matrices (with respect to both tissues and species)

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ranged from 0.82 to 20.5 ng/g (Table S3), except for 4-HB, which was elevated in a few

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matrices. Relative recoveries of analytes were estimated by dividing the ratio of the analyte

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response to the IS response in MS/MSD samples (average) by the same ratio determined in

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the MM sample. Relative recoveries of analytes were variable between matrices. Mean

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relative recoveries ranged from 80% to 127% for MeP, EtP, PrP, BuP, BzP, and HeP; from 47%

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to 79% for 3,4-DHB; from 109% to 156% for 4-HB; and from 11% to 85% for OH-MeP and

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OH-EtP. Calculation of absolute recovery was based on the comparison between the response

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of the analyte in the MS/MSD sample (average) to the response in the MM sample. Low

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recoveries observed for OH-MeP, OH-EtP, and 3,4-DHB were due to matrix interference, and

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the recoveries of corresponding ISs were similar to those of the parent compounds. Therefore,

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correction for IS recoveries enabled accurate analysis of OH-MeP, OH-EtP, and 3,4-DHB in

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tissues. The RPDs between MS and MSD analyses were below 30% for all target analytes.

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Results and Discussion

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For the purpose of discussion, the samples were grouped into four geographical regions:

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Florida coast, California coast, Washington coast, and Alaskan waters.

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Florida Coast. Livers and the blubber of dolphins from the Gulf of Mexico, Sarasota Bay,

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Anna Maria Sound, Palma Sola Bay, Venice Inlet, and Tampa Bay and livers of whales from

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the Atlantic Ocean and the Anclote River were analyzed from the Florida coast. Of the six

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parent parabens determined, MeP was the most predominant compound (detected in 12 of 20

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samples), found in the livers of small cetaceans collected from the Florida coast, the Gulf of

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Mexico, and the Atlantic Ocean (Table 1). 4-HB was the predominant metabolite (detected in

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all samples) among the four metabolites measured (Table 1). PrP, OH-MeP, and 3,4-DHB

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were found in 5%-25% of the samples analyzed (Table S4).

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Concentration of MeP in the livers of the cetaceans ranged from rough-toothed dolphin ~ striped dolphin > pygmy sperm whale ~ clymene dolphin.

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The mean hepatic concentration of MeP in the bottlenose dolphin was 159 ± 260 ng/g, wet wt

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(n = 10). Greater concentrations of MeP in the bottlenose dolphin than in other cetaceans can

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be explained by this mammal’s near-shore feeding habits. The hepatic concentrations of 4-HB

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were much higher than those of MeP, at a range of 875 to 26,500 ng/g, wet wt. 4-HB

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concentrations were found in the decreasing order of rough-toothed dolphin > striped dolphin

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~ bottlenose dolphin > pygmy sperm whale ~ clymene dolphin. The mean hepatic

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concentration of 4-HB in the rough-toothed dolphin was 12,000 ± 14,800 ng/g, wet wt (n = 2),

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which was higher than that in the bottlenose dolphin (7,230 ± 8,430 ng/g, wet wt). Although

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4-HB is formed through the metabolic transformation of parabens, this compound has been

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reported to occur naturally in certain plants, including seagrasses.27 The concentrations of PrP,

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OH-MeP, and 3,4-DHB in cetacean livers were one to four orders of magnitude lower than

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those of MeP and 4-HB (Table S4).

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4-HB was found in all (100%) dolphin blubber samples analyzed at concentrations

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that ranged from 57.0 to 2,180 ng/g, wet wt (Table 1). 3,4-DHB was detected in the blubber

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of 4 of 17 individuals (23.5%), at concentrations that ranged from 0.05) (Figure S2a).

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The concentrations of MeP and 4-HB in the livers and the blubber (4-HB only) of

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female bottlenose dolphins were greater than those in males, but the difference was not

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statistically significant (p > 0.05) (Figure S3). No significant correlation was found between

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hepatic MeP/4-HB concentrations and age of female bottlenose dolphins (age data were

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available only for female bottlenose dolphins) (Figure S2b, c). These results suggest that

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accumulation features of MeP and 4-HB are different from those of persistent organic

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pollutants, such as polychlorinated biphenyls, for which adult females generally contain

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lesser concentrations than do adult males.28 A significant positive correlation was found

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between MeP and 4-HB concentrations in the livers of cetaceans (r = 0.79, p = 0.002) (Figure

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2a), which may suggest their co-exposures or metabolic transformation from MeP to 4-HB in

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the livers of cetaceans.

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The concentrations of MeP and 4-HB in livers of dolphins collected from the

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near-shore waters of Florida (Sarasota Bay, Anna Maria Sound, Palma Sola Bay, Venice Inlet,

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and Tampa Bay) were over 2-fold greater than those collected from the open waters in the

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Gulf of Mexico (Figure 3). Similarly, the concentrations of 4-HB in the blubber of dolphins

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collected from near-shore areas were over 2-fold greater than those collected from the Gulf of

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Mexico (Figure 3). These results may suggest greater exposures to parabens in near-shore

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coastal waters than in offshore waters. The primary sources of parabens in marine mammals

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are postulated to be wastewater discharges in coastal areas. Although 4-HB is found naturally

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in certain seagrasses, a significant correlation between MeP and 4-HB in dolphins suggests

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the significance of anthropogenic sources of exposure in marine mammals.

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California Coast. Livers, brains, and kidneys of southern sea otters collected from California

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coast were analyzed. MeP was found in the livers of 18 of 25 individuals and in all brain and

286

kidney samples (Table 2). 4-HB was found in all livers, brains, and kidneys (Table 2). It

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should be noted that 3,4-DHB was detected in 8 of 10 kidneys, with a mean concentration of

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76.7 ± 39.8 ng/g, wet wt, which was higher than the concentration found in livers (DR: 20%;

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mean: 20.7 ±3 4.0 ng/g, wet wt) and brains (DR: 30%; mean: 31.0 ± 17.7 ng/g, wet wt)

290

(Table S5–S7). EtP, PrP, OH-MeP, and OH-EtP were found less frequently (DR: 8%–30%) at

291

concentrations below 12 ng/g, wet wt (Table S5–S7).

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Mean hepatic concentrations of MeP and 4-HB in sea otters were 31.0 ± 24.7 (range:

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2-fold greater than those in adults (Figure S7a). A significant

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341

positive correlation between hepatic MeP and 4-HB concentrations was found in northern sea

342

otters (r = 0.82, p = 0.0006, Figure 2e).

343 344

Alaskan Waters. Livers of northern sea otters from Prince William Sound, Kachemak Bay,

345

and Pow Island and polar bears from the Chuckchi and the Beaufort Seas were analyzed

346

(Table 3, S9 and S10). MeP was found in the livers of northern sea otters from Alaska at

347

concentrations that ranged from 10.3 to 686 ng/g, wet wt (mean: 179 ± 270 ng/g), which were

348

approximately 2-fold greater than the concentrations found in sea otters from the Washington

349

coast. MeP concentrations in livers of three sea otters (mean: 595 ± 78.6 ng/g, wet wt), one

350

from Pow Island and two from Prince William Sound, were much higher than those in

351

remaining samples (mean: 23.1 ± 10.9 ng/g, wet wt). The mean hepatic 4-HB concentration

352

in livers of sea otters collected from Alaska was 4,830 ± 4,960 ng/g, wet wt, which was

353

2-fold lower than those found in sea otters from the Washington coast. A significant positive

354

correlation was found between hepatic MeP and 4-HB concentrations in sea otters from

355

Alaska (r = 0.92, p = 0.001, Figure 2f).

356

MeP was detected in 2 of 10 polar bear livers, and 4-HB was found in all polar bear

357

livers. The concentrations of MeP and 4-HB in polar bears were much lower than those found

358

in cetaceans and sea otters from other U.S. coastal locations. The mean hepatic concentration

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of 4-HB in polar bears was 465 ± 234 ng/g, wet wt (Table 3). The presence of MeP and 4-HB

360

in livers of polar bears from the Chuckchi and the Beaufort Seas suggests widespread

361

distribution of 4-HB in remote marine areas. The natural sources of 4-HB is a source of

362

exposure in polar bears and other marine mammals, but further studies are needed to

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ascertain the contribution of anthropogenic sources of these compounds in remote marine

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environments.

365

In summary, this is the first study to report the occurrence of MeP and 4-HB in marine

366

mammals. Occurrence of parabens and their metabolites in marine mammals from remote

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marine locations suggests widespread distribution of these chemicals in the environment. The

368

concentrations of 4-HB found in dolphins and sea otters were some of the highest values ever

369

reported in the literature. Although 4-HB has been reported to occur naturally in plants, a

370

significant correlation between MeP and 4-HB in all of the marine mammal species analyzed

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from all of the geographic locations suggests a significant contribution from anthropogenic

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sources of these chemicals in marine mammals. Further studies are needed to evaluate the

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sources, pathways, and toxic effects of parabens and 4-HB in the marine environment.

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Associated Content

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Supporting Information Available

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Additional information as noted in the text. This material is available free of charge via the

377

Internet at http://pubs.acs.org.

378

References

379 380 381 382 383 384 385 386 387 388 389 390

1.

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473

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Table 1. Concentrations (ng/g, wet weight) of Parabens and Their Metabolites in Livers of Cetaceans Found Stranded Along the Florida Coastal Waters

species

sex

age

date

n

stranded

Bottlenose

Liver water body

MeP

Blubber* 4-HB

4-HB

mean±SD

range

mean±SD

range

mean±SD

range

188±285

< 41.1 - 865

7980±9280

875 - 26500

229±196

62 - 662

74.5±41.8

44.9 - 104

4230±3760

1570 - 6890

84.7±22.8

64.3-105

98.0±67.9

50 - 146

8620±7620

3230 - 14000

167±155

57 - 276

120±112

< 41.1 - 199

11900±14800

1480-22400

1140±1480

91.3-2180

< 41.1

< 41.1

1450±720

916-2270

423±66.5

350-480

31.4±9.45

< 20.5 - 37.7

2520±1270

1450-3920

n.a.**

n.a.

Gulf of Mexico, 2;

dolphin

2 n.a.; F

18

3

M

n.a.

3

Pygmy sperm

11/27/ -

2

F

Gulf of Mexico

12/3/1995

Anna Maria Sound, 1; Tampa Bay, 1

6/15/1995

Gulf of Mexico

8/16/1994-8

Atlantic Ocean, 2;

/23/2000

Anclote River, 1

whale * MeP was found in only one blubber **n.a.: not available

20


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Table 2. Concentrations (ng/g, wet weight) of Parabens and Their Metabolites in Livers, Kidneys, and Brains of Sea Otters Collected from the California Coastal Waters, and in Livers of Sea Otters from the Washington Coastal Waters sex

county

n

collection date

5

8/19/1994-7/2/1999

age

body condition

MeP

postmortem state

mean±SD

4-HB range

mean±SD

range

16800±10200

6930-27700

7330±5790

3050-20200

7930±6230

1090-15700

17200±13400

3700-35300

Liver (California) San Luis Obispo

M

Monterey

Santa Cruz San Luis Obispo

F

Monterey Santa Cruz

7

8/23/1994-11/27/1995

4

9/30/1994-10/9/1995

4

3/20/1995-11/2/1995

4

1

9/6/1994-11/27/1995

2/23/1995

Immature, 2;

Poor, 1; good,

Poor, 1; fair,

Adult, 3

3; emaciated, 1

2; good, 2

Suckling, 1;

Emaciated, 1;

immature, 1;

poor, 1; fair, 2;

Fair, 2;

Subadult, 2;

Good, 2;

good, 5

adult, 3

excellent, 1

Subadult, 3;

Poor, 2; fair, 1;

Fair, 1;

adult, 1

good, 1

good, 1

Immature, 2;

Emaciated, 1;

Fair, 2;

Adult, 2;

poor, 2; good, 1

good, 2

Immature, 2; adult, 2 Adult

Poor, 1;

Poor, 1;

emaciated, 1;

22.4±13.3

23.4±10.6 33.2±24.6

< 10.26-54.8 < 10.26-44.1 < 10.26-36.1 < 10.26-67.5

56.9±46.2

29.1-126

19600±10700

10200-31100

< 10.26

n.a.*

5690

n.a.

121±159

29.9-360

23800±30000

2460-66400

Good, 2

41.9±19.2

28.3-55.4

7980±5690

3950-12000

good, 3

fair, 1

30.9±17.1

Poor

Good

Poor, 2; fair, 1;

Fair, 1;

good, 1

good, 3

Kidney (California) Santa

4

9/30/1994-10/9-1995

2

11/18/1994-8/1-1995

Monterey

1

11/27/1995

Subadult

Excellent

Good

84.5

n.a.

2480

n.a.

San Luis

1

3/20/1995

Adult

Emaciated

Fair

12.2

n.a.

3580

n.a.

Cruz M

Subadult, 1;

San Luis Obispo

Adult, 3 Immature, 1;

Emaciated, 1;

adult, 1

good, 1

21



F

Page 22 of 28

Obispo Monterey

2

11/28/1994-8/8/1995

Immature, 1;

Emaciated, 1;

poor, 1;

adult, 1

poor, 1

good, 1

Subadult, 1;

Poor, 2; fair, 1;

Fair, 1;

Adult, 3

good, 1

good, 3

Immature, 1;

Emaciated, 1;

fair,1;

adult, 1

good, 2

Good, 2

Subadult

Good

Fair

Immature, 1;

Emaciated, 1;

poor, 1;

adult, 1

poor, 1

good, 1

n.a. n.a.

152±112

72-231

33300±18700

20000-46500

27.1±33.6

5.99-77.2

3240±5780

200-11900

18.8±11.7

9.16-31.8

1610±1610

156-3340

Brain (California) Santa

4

9/30/1994-10/9-1995

3

11/18/1994-7/2/1999

Monterey

1

8/23/1994

Monterey

2

11/28/1994-8/8/1995

Cruz M

F

San Luis Obispo

14

458

6.80±0.07

6.75-6.85

1590±63.6

1540-1630

n.a.

92.4±109

< 20.5-358

6890±8940

722-32600

n.a.

95.2±91.5

< 20.5-228

12900±14600

329-31200

Liver (Washington) M F

Pacific coast,

13

7/16/2000-5/5/2004

5

7/3/2000-4/12/2004

WA

Subadult, 3; Adult, 10 Adult, 5

*n.a.: not available

22


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Table 3. Concentrations (ng/g, wet weight) of Parabens and Their Metabolites in Livers of Northern Sea Otters and Polar Bears from the Alaskan Coastal Waters sex

n

collection date

age

MeP

cause of death

4-HB

mean±SD

range

mean±SD

range

197±277

< 20.5-686

5160±5100

1110-17600

sea otter* M

13

n.a.***

1

4/11/1996-2003

n.a.

Pup, 2; Subadult,

Hunted, 6;

1; Adult, 5; old

Stranded, 3;

adult, 2

n.a., 1

n.a.

n.a.

21.8

n.a.

1520

n.a.

n.a.

4.57±1.16

< 4.10-6.94

395±180

229-719

n.a.

7.30±6.40

< 4.10-16.9

569±293

280-862

polar bear** M

6

1/26/1995-6/3/2002

F

4

6/6/1994-12/4/2000

Sub-adult, 3; adult, 3 Cub, 1; sub-adult, 2; adult, 1

*Sea otters were from Prince William Sound, Kachemak Bay, Pow Island; **Polar bears were collected from the Chuckchi Sea and the Beaufort Sea. ***n.a.: not available.

23



Figure 1. Map showing collection locations for marine mammal samples

24


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Page 25 of 28


Figure 2. Correlations between concentrations of MeP and 4-HB in various marine mammal tissues. (a) livers of cetaceans from Florida coastal waters; (b) livers of sea otters from California coastal waters; (c) brains of sea otters from California coastal waters; (d) kidneys of sea otters from California coastal waters; (e) livers of sea otters from Washington coastal waters; (f) livers of sea otters from Alaska waters. Note: only those samples with measurable levels of chemicals are presented.

25



Figure 3. Concentrations (mean±SD) of MeP and 4-HB in livers, and 4-HB in blubber of dolphins from near-shore Florida coastal waters and Gulf of Mexico.

26


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Figure 4. Concentrations (mean±SD) of MeP and 4-HB in livers of female and male sea otters from California (a) and Washington (b) coastal waters.

27



TOC ART

28


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Elevated Accumulation of Parabens and their Metabolites in Marine

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