Urinary Phthalate Metabolites in Common Bottlenose Dolphins (Tursiops truncatus) From Sarasota Bay, FL, USA

Abstract

Phthalates are chemical additives to common consumer goods including cleaning products, cosmetics, personal care products, and plastic. Because they are not chemically bound to these products and are widely used, the potential for environmental contamination is significant. Phthalates and their metabolites have been associated with endocrine disruption and reproductive impairment, among other adverse health effects, in laboratory animals and human epidemiologic studies. Common bottlenose dolphins (Tursiops truncatus) are vulnerable to environmental pollutants due to their apex position in the food chain, long life spans, and habitat overlap with developed coastal areas. The objective of this study was to quantify phthalate metabolite concentrations in urine collected from bottlenose dolphins in Sarasota Bay, Florida, during May 2016 (n = 7) and May 2017 (n = 10). Screening of nine phthalate monoester metabolites in bottlenose dolphin urine was performed by liquid chromatography tandem mass spectrometry using methods adapted from those used for analyzing human samples. At least one phthalate metabolite was detected in 71% of the dolphins sampled across both years, with the highest concentrations detected for monoethyl phthalate (MEP; GM = 5.4 ng/ml; 95%CI: 1.3–22.0 ng/ml) and mono‐(2‐ethylhexyl) phthalate (MEHP; GM = 1.9 ng/ml; 95%CI: 1.1–3.2 ng/ml). These data demonstrate exposure to two of the most commonly used phthalates in commercial manufacturing, diethyl phthalate (DEP) and di‐2‐ethylhexyl phthalate (DEHP). This study establishes methods for urinary detection of phthalate metabolites in marine mammals and provides baseline data to address a significant and growing, yet poorly understood, health threat to marine wildlife.

Sentinels of synthetics – a comparison of phthalate exposure between common bottlenose dolphins (Tursiops truncatus) and human reference populations

“…This study relied upon bottlenose dolphin urinary metabolite concentrations reported in Hart et al [ 30 ] (sample years 2016–2017), as well as results from analyses conducted on samples collected 2010–2015 and 2018–2019. Urine samples were opportunistically collected aseptically via catheterization from bottlenose dolphins during routine health assessments conducted under permit from the National Oceanic and Atmospheric Administration’s (NOAA) National Marine Fisheries Service (NMFS) between 2010 and 2019, as described in Wells [ 42 ] and Hart et al [ 30 ]. Archived urine samples for the years 2010–2015 were retrieved from the Sarasota Dolphin Research Program’s specimen bank, where urine samples were stored frozen at -80°C.…”

“…Archived urine samples for the years 2010–2015 were retrieved from the Sarasota Dolphin Research Program’s specimen bank, where urine samples were stored frozen at -80°C. The protocol for sample collection and storage for years 2016–2019 are described in Hart et al [ 30 ]. All urine samples were screened for eight phthalate metabolites including: monomethyl phthalate (MMP), monoethyl phthalate (MEP), monoisobutyl phthalate (MiBP), monobutyl phthalate (MBP), monobenzyl phthalate (MBzP), mono-(2-ethylhexyl) phthalate (MEHP), mono-(2-ethyl-5-oxohexyl) phthalate (MEOHP), and mono-(2-ethyl-5-hydroxyhexyl) phthalate (MEHHP).…”

“…All urine samples were screened for eight phthalate metabolites including: monomethyl phthalate (MMP), monoethyl phthalate (MEP), monoisobutyl phthalate (MiBP), monobutyl phthalate (MBP), monobenzyl phthalate (MBzP), mono-(2-ethylhexyl) phthalate (MEHP), mono-(2-ethyl-5-oxohexyl) phthalate (MEOHP), and mono-(2-ethyl-5-hydroxyhexyl) phthalate (MEHHP). Methods for analyzing phthalate metabolite concentrations in bottlenose dolphin urine were based on protocols established in Hart et al [ 30 ] and conducted at the NOAA National Centers for Coastal Ocean Science (NCCOS) lab in Charleston, SC, USA. Briefly, urine samples were spiked with 13 C-labeled internal standards prior to an enzymatic de-glucuronidation step.…”

“…Compounds were separated using a C18 analytical column (Waters XBridge 50 mm x 2.1 mm; 2.5 μm particle size) with a gradient mobile phase of HPLC water with 0.1% acetic acid and acetonitrile with 0.1% acetic acid. Analytes of interest were identified using multiple reaction monitoring (MRM; S2 Table ) and quantified against standard calibration curves [ 17 , 30 ], and the limit of detection (LOD) was determined for each metabolite based on the lowest point of the calibration curve that could be detected on the instrument, divided by the volume of urine extracted. Complete details of the instrumental methodology is detailed in Hart et al [ 30 ].…”

“…Analytes of interest were identified using multiple reaction monitoring (MRM; S2 Table ) and quantified against standard calibration curves [ 17 , 30 ], and the limit of detection (LOD) was determined for each metabolite based on the lowest point of the calibration curve that could be detected on the instrument, divided by the volume of urine extracted. Complete details of the instrumental methodology is detailed in Hart et al [ 30 ]. As reported in Hart et al [ 30 ] quality assurance and control methods included laboratory spikes, laboratory blanks, field blanks, a standard reference material (SRM 3672), and matrix spikes.…”