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Archives of Environmental Contamination and Toxicology (v.53, #1)
The Impact of Process Variables on the Removal of PBDEs and NPEOs During Simulated Activated Sludge Treatment by Katherine Langford; Mark Scrimshaw; John Lester (pp. 1-7).
This work illustrates that the removal of some endocrine-disrupting compounds from sewage treatment works effluent is dependent on parameters such as sludge age, influent concentrations, concentrations of co-pollutants and hydraulic retention time as well as physicochemical properties of the compound. Greater nonylphenol polyethoxylates (NPEO) and polybrominated diphenyl ether (PBDE) removal was observed at a higher sludge age, and it appeared that the enzymes required for NPEO degradation were already present. NPEO degradation was reduced in the presence of the more hydrophobic PBDE compounds as sorption of PBDEs occurred, rapidly reducing available sorption sites for NPEOs. The more hydrophobic NP and PBDEs demonstrated little degradation in comparison to longer-chain NPEO compounds. From this research, it is apparent that the principal environmental risk of PBDE contamination after wastewater treatment is via sludge-disposal routes. Treatment of wastewater containing NPEO surfactants poses environmental risks via two routes: some nonylphenolic compounds may pass through into receiving waters and degradation products such as nonylphenol and short-chain ethoxylate compounds will enter the environment via sludge disposal.
Role of Temperature and Hydrochloric Acid on the Formation of Chlorinated Hydrocarbons and Polycyclic Aromatic Hydrocarbons During Combustion of Paraffin Powder, Polymers, and Newspaper by Takumi Takasuga; Norihito Umetsu; Tetsuya Makino; Katsuya Tsubota; Kenneth S. Sajwan; Kurunthachalam Senthil Kumar (pp. 8-21).
Formation of chlorinated hydrocarbons and polycyclic aromatic hydrocarbons (PAHs) were determined using a laboratory-scale incinerator when combusting materials at different temperatures, different concentrations of hydrochloric acid (HCl), and when combusting various types of polymers/newspaper. Polychlorobenzenes (PCBz), polychlorophenols (PCPhs), polychlorinated dibenzo-p-dioxins/furans (PCDD/Fs) and their toxic equivalency (TEQ) and PAHs were highlighted and reported. Our results imply maximum formation of chlorinated hydrocarbons at 400°C in the following order; PCBz≥PCPhs>>PCDFs>PCDDs>TEQ on a parts-per-billion level. Similarly, a maximum concentration of chlorinated hydrocarbons was noticed with an HCl concentration at 1000 ppm with the presence of paraffin powder in the following order; PAHs>PCBz≥PCPhs>>PCDFs>PCDDs>TEQ an a parts-per-billion level. PAHs were not measured at different temperatures. Elevated PAHs were noticed with different HCl concentrations and paraffin powder combustion (range: 27–32 μg/g). While, different polymers and newspaper combusted, nylon and acrylonitrile butadiene styrene (ABS) produced the maximum hydrogen cyanide (HCN) concentration, concentrations of PCDD/FS, dioxin-like polychlorinated biphenyls (DL-PCBs), and TEQ were in a decreasing order: polyvinylchloride (PVC)
Effects of Chlorpyrifos, Carbendazim, and Linuron on the Ecology of a Small Indoor Aquatic Microcosm by M. A. Daam; P. J. Van den Brink (pp. 22-35).
To validate the use of small indoor microcosms for the risk assessment of pesticides, the fate and effects of chlorpyrifos, carbendazim, and linuron were studied in 8.5–liter indoor freshwater microcosms. Functional and structural responses to selected concentrations were evaluated and compared with responses observed in larger-scale model ecosystem studies. Overall, the microcosms adequately displayed the chain of effects resulting from the application, although they did not always predict the exact fate and responses that were observed in larger semifield studies. Because closed systems were used that did not contain sediment and macrophytes, pesticides were relatively persistent in the present study. Consequently, calculated toxicity values were generally more comparable with those reported in studies with long- than with short-term exposure. Carbendazim had a higher overall no-observed-effect concentration (NOEC) compared with experiments performed in larger systems because macroinvertebrate taxa, the most sensitive species group to this fungicide, were not abundant or diverse. Future refinements to the test system could include the addition of a sediment compartment and sensitive macroinvertebrate taxa. However, the simple design offers the potential to perform experiments under more controlled conditions than larger and, consequently, more complex model ecosystems, while maintaining relatively high ecologic realism compared with standard laboratory tests. Further implications for risk-assessment studies are discussed in an ecotoxicologic and methodologic context.
Using a Sediment Quality Triad Approach to Evaluate Benthic Toxicity in the Lower Hackensack River, New Jersey by Mary T. Sorensen; Jason M. Conder; Phyllis C. Fuchsman; Linda B. Martello; Richard J. Wenning (pp. 36-49).
A Sediment Quality Triad (SQT) study consisting of chemical characterization in sediment, sediment toxicity and bioaccumulation testing, and benthic community assessments was performed in the Lower Hackensack River, New Jersey. Chemistry data in sediment and porewater were evaluated based on the equilibrium partitioning approach and other published information to investigate the potential for chemical effects on benthic organisms and communities. Relationships were supported by laboratory toxicity and bioaccumulation experiments to characterize chemical effects and bioavailability. Benthic community results were evaluated using a regional, multimetric benthic index of biotic integrity and four heterogeneity indices. Evidence of slight benthic community impairment was observed in five of nine sediment sample stations. Severe lethal toxicity to amphipods (Leptocheirus plumulosus) occurred in four of these five stations. Although elevated total chromium concentrations in sediment (as high as 1900 mg/kg) were the rationale for conducting the investigation, toxicity was strongly associated with concentrations of polycyclic aromatic hydrocarbons (PAHs) rather than total chromium. PAH toxic units (ΣPAH TU) in sediment and ΣPAH concentrations in laboratory organisms from the bioaccumulation experiment showed a clear dose–response relationship with toxicity, with 0% survival observed in sediments in which ΣPAH TU > 1–2 and ΣPAH concentrations in Macoma nasuta were >2 μmol/g, lipid weight. Metals detected in sediment and porewater, with the possible exception of copper, did not correlate with either toxicity or levels in tissue, likely because acid-volatile sulfide levels exceeded concentrations of simultaneous extracted metals at all sample locations. The study reinforces the value of using multiple lines of evidence approaches such as the SQT and the importance of augmenting chemical and biological analyses with modeling and/or other approaches to evaluate chemical bioavailability and toxicity of sediments.
Acute and Chronic Copper Toxicity to a Saltwater Cladoceran Moina monogolica Daday by Zaosheng Wang; Hainan Kong; Deyi Wu (pp. 50-56).
The objective of this study was to evaluate the sensitivity of a saltwater cladoceran Moina monogolica Daday to copper toxicity. Acute and chronic toxicity tests were conducted according to the American Society for Testing and Materials (ASTM) guidelines. The 24- and 48-h LC50s and their 95% confidence intervals (CIs) were calculated as 154.21 (135.50–182.05) μg Cu/L and 106.07 (99.77– 112.36) μg Cu/L , respectively, based on measured copper concentrations. When compared with different phyla or classes of estuarine organisms, M. monogolica had moderate sensitivity and was suitable to be used as an indicator organism in the Changjiang estuary. Eight end points of longevity, number of broods, brood size, total reproduction, time to first brood, intrinsic rate of natural increase (r m ), net reproductive rate (R 0), and mean generation time (T) used to evaluate chronic effects were affected at chronic copper concentrations. Comparisons of chronic effects showed that toxicity mechanisms of dissolved copper are different from those of dietary exposure. The EC20s, EC50s, and their 95% CIs of end points were calculated using linear regression equations. The geometric mean generated the maximum-acceptable-toxicant concentration of 6.74 μg Cu/L between the no-observed-effect concentration of 5.06 and the lowest-observed-effect concentration of 8.99 μg Cu/L for brood size, total reproduction, r m , and R 0, and 12.1 μg Cu/L between 8.99 and 16.29 μg Cu/L for longevity and number of broods as the end points, respectively. The EC20 of R 0 appeared to be the most sensitive at 3.82 μg Cu/L among the end points. Results showed that aquatic organisms were possibly negatively affected at sites of higher copper concentrations in the Changjiang estuary. Further research is needed to evaluate the direct or indirect effects of copper more comprehensively to protect aquatic organisms and their habitats in the Changjiang estuary.
Bioaccumulation of Metals in Sediment Elutriates and Their Effects on Growth, Condition Index, and Metallothionein Contents in Oyster Larvae by A. Geffard; O. Geffard; J.C. Amiard; E. His; C. Amiard-Triquet (pp. 57-65).
The bioavailability of Cd, Cu, Zn, and Pb from two metal-contaminated sediments (Bidassoa and Dunkerque) was studied using Crassostrea gigas larvae exposed to sediment elutriates. The metal contents within the sediments, the larvae and larval growth, the condition index, and the induction of metallothionein in the larvae were measured. The larval growth and condition index were only affected after exposure to the highest elutriates concentration (5 to 25%) from the most contaminated sediment (Dunkerque). Bioaccumulation of all metals was observed in larvae exposed to Dunkerque elutriatre; only Cu bioaccumulation was observed in the Bidassoa elutriate. The results from larvae exposed to both sediment elutriates show a strong correlation between bioaccumulated metal considered individually or in combination and the metallothionein level in larvae presenting no detrimental effect. On the other hand, in the case of larvae exposed to the highest Dunkerque elutriate concentration and showing the highest metal body burden, we observed a drop in the metallothionein level. These results indicate that metallothionein is a more sensitive indicator of heavy metal pollution than physiological endpoints taken into account in bioassays and could be proposed as an early biomarker of metal exposure in larvae. However, care must be taken with “fault control” due to the toxicological effect on larvae metabolism in the case of substantial contaminant exposure.
Keywords: Sediment elutriates; Crassostrea gigas larvae; Larval growth; Metal bioaccumulation; Metallothionein
Biomarkers Study for Sediment Quality Assessment in Spanish Ports Using the Crab Carcinus maenas and the Clam Ruditapes philippinarum by M.L. Martín-Díaz; J. Blasco; D. Sales; T.A. DelValls (pp. 66-76).
Intermolt crab Carcinus maenas and clam Ruditapes philippinarum were used to determine the toxicity of sediments collected in four Spanish ports (Cádiz, Huelva, Pasajes, and Bilbao) under laboratory conditions during 28 days. Sediment samples were analyzed to determine chemical concentration of metals (As, Cd, Cr, Cu, Fe, Hg, Mn, Ni, Pb, and Zn), polyaromatic hydrocarbons, polychlorinated biphenyls, grain size distribution, and organic matter content. Different biomarkers of exposure of early biological stress were determined after 28-day exposure in crabs and clams, in the hepatopancreas and in the digestive gland, respectively: metallothionein, ethoxyresorufin O-deethylase (EROD), glutathione peroxidase (GPX), glutathione S-transferase (GST), and glutathione reductase activities (GR). The battery of biomarkers tested resulted in showing and linking the bioavailability of various contaminants and sediment characteristics to the toxicity of the different sediments. Significant induction of MTs was observed when organisms were exposed to metal-contaminated sediments (port of Huelva), and induction of EROD and GPX activities after exposure to sediments containing organic compounds (port of Bilbao and Pasajes). Higher induction was shown in biomarkers tested in crabs; nevertheless, only interspecies significant differences were observed in the induction of GR and GST activities. The present work confirms the necessity of using species with different ecological lifestyles for sediment toxicity assessment and validates the use of this set of biomarkers as a potential tool in sediment toxicity assessment.
Fluctuating Asymmetry in Insecticide-Resistant and Insecticide-Susceptible Strains of the Maize Weevil, Sitophilus zeamais (Coleoptera: Curculionidae) by B. Ribeiro; R. N. C. Guedes; A. S. Corrêa; C. T. Santos (pp. 77-83).
Insecticide resistance is an evolutionary response of specific insect populations subjected to exposure and consequent selection by an insecticide. As such, this phenomenon is important as a biomonitoring strategy and also has economical importance in the case of insect-pests by compromising their control. Fluctuating asymmetry (FA) of bilateral traits is a measure of developmental instability also suggested as a monitoring tool for environmental pollution with potential consequences for fitness. Responses to selective agents might have pleiotropic effects influencing development and phenotype, which has yet to be examined for agricultural insecticides and insect-pests. Higher levels of FA are expected in the insecticide-resistant strains, which are usually at a selective disadvantage, relative to susceptible strains, in the absence of the insecticide. Two insecticide-resistant strains and an insecticide-susceptible strain of the maize pest insect Sitophilus zeamais (Coleoptera: Curculionidae) were subjected to FA measurements in 12 traits of wing veins and tibias of 100 individuals of both sexes of each strain. The insecticide-resistant strains showed lower FA than the susceptible strain, in contrast with the initial expectation. An extended period of insecticide selection probably led to the evolution of fitness-modifier genes improving the performance of the resistant genotypes, reducing their FA levels, and leading to their eventual fixation in the population. In addition, one insecticide-resistant strain and the insecticide-susceptible strain showed significant differences in FA between sexes, with more symmetrical males suggesting possible sexual selection by the females. The observed results have potential consequences for insecticide-resistance evolution and dispersal.
Keywords: Insecticide resistance; Developmental instability; Pyrethroid insecticides; Fitness; Adaptative cost
Evaluation of Potentially Nonlethal Sampling Methods for Monitoring Mercury Concentrations in Smallmouth Bass (Micropterus dolomieu) by C. J. Schmitt; W. G. Brumbaugh (pp. 84-95).
We evaluated three potentially nonlethal alternatives to fillet sampling for the determination of mercury (Hg) concentrations in smallmouth bass (Micropterus dolomieu). Fish (n = 62, 226–464 mm total length) from six sites in southern Missouri were captured by electrofishing. Blood samples (1 mL) from each fish were obtained by caudal veinipuncture with a heparinized needle and syringe. Biopsy needle (10 mm × 14 gauge; three cuts per fish; 10–20 mg total dry weight) and biopsy punch (7 mm × 5 mm in diameter, one plug per fish, 30–50 mg dry weight) samples were obtained from the area beneath the dorsal fin. Fillet samples were obtained from the opposite side of the fish. All samples were freeze-dried and analyzed for total Hg by combustion amalgamation atomic absorption spectrophotometry. Mean relative standard deviations (RSDs) of triplicate samples were similar for all four methods (2.2–2.4%), but the range of RSDs was greater for blood (0.4–5.5%) than for the muscle methods (1.8–4.0%). Total Hg concentrations in muscle were 0.0200–0.8809 μg/g wet weight; concentrations in plug, needle, and fillet samples from each fish were nearly identical. Blood Hg concentrations were 0.0006–0.0812 μg/mL and were highly correlated with muscle concentrations; linear regressions between log-transformed blood and fillet Hg concentrations were linear and statistically significant (p < 0.01), and explained 91–93% of the total variation. Correlations between fillet Hg concentrations and fish size and age were weak; together they explained ≤37% of the total variation, and the relations differed among sites. Overall, any of the alternative methods could provide satisfactory estimates of fillet Hg in smallmouth bass; however, both blood and plug sampling with disposable instruments were easier to perform than needle sampling. The biopsy needle was the most difficult to use, especially on smaller fish, and its relative expense necessitates reuse and, consequently, thorough cleaning between fish to prevent cross-contamination.
Immune Status, Carotenoid Coloration, and Wing Feather Growth in Relation to Organochlorine Pollutants in Great Black-Backed Gulls by Jan Ove Bustnes; Kai Ove Kristiansen; Morten Helberg (pp. 96-102).
Previous ecotoxicological studies have documented relationships between residues of various organochlorines (OCs) and immune status, carotenoid colors, and wing feather growth in different bird species. In this study, the density of white blood cells (WBC), carotenoid colors, and length of the same feathers on each wing were measured in breeding great black-backed gulls (Larus marinus) on the coast of northern Norway, and related to the blood residues of five OCs, including HCB (hexachlorobenzene), β-HCH (β-hexachlorocyclohexane), p,p′-DDE (p, p′-dichlorodiphenyldichloroethylene), oxychlordane, and polychlorinated biphenyl (PCB), in addition to ΣOC. Neither, WBC density nor carotenoid colors were significantly related to blood residues of any of the OCs, suggesting that OC levels may have been too low to significantly affect these outcome parameters. However, in the colony where the OC concentrations were highest, there was a weak but significantly positive relationship between the probability of having different length of feathers on each wing and levels of PCB and ΣOC, in males. Thus varying length of the wing primaries may reflect adverse impacts of OCs in great black-backed gulls. However, in gulls with moderate levels of OCs, it is probably not a sensitive indicator of progressing ecological impacts of OCs, since such adverse ecological relationships were found in the breeding colonies where there were no relationships between differences in wing feather lengths and OCs.
Selenium and Metal Concentrations in Waterbird Eggs and Chicks at Agassiz National Wildlife Refuge, Minnesota by Thomas W. Custer; Christine M. Custer; Bruce A. Eichhorst; David Warburton (pp. 103-109).
Exceptionally high cadmium (Cd) and chromium (Cr) concentrations were reported in eggs, feathers, or livers of selected waterbird species nesting at Agassiz National Wildlife Refuge (Agassiz) in 1994. Ten- to 15-day-old Franklin’s gull (Larus pipixcan), black-crowned night-heron (Nycticorax nycticorax), and eared grebe (Podiceps nigricollis) chicks were collected in 1998, 1999, and 2001 at Agassiz and analyzed for selenium (Se) and metals including Cd and Cr. Freshly laid eggs were collected in 2001 from Franklin’s gull, black-crowned night-heron, eared grebe, and pied-billed grebe (Podilymbus podiceps) nests at Agassiz. Based on a multivariate analysis, the pattern of Se and metal concentrations differed among species for eggs, chick feathers, and chick livers. Low Cd and Cr concentrations were measured in eggs, chick livers, and chick feathers of all four species. Mercury concentrations in black-crowned night-heron and eared grebe eggs collected from Agassiz in 2001 were lower than concentrations reported in 1994. Se and metal concentrations, including Cd and Cr, in waterbird eggs and chicks collected at Agassiz in 1998, 1999, and 2001 were not at toxic levels.
Keywords: Metals; Cadmium; Chromium; Mercury; Selenium; Bird eggs; Feathers; Bird livers; Waterbirds
Multiple Cytokine and Acute-Phase Protein Gene Transcription in West Greenland Sledge Dogs (Canis familiaris) Dietary Exposed to Organic Environmental Pollutants by C. Sonne; S. Fonfara; R. Dietz; M. Kirkegaard; R. J. Letcher; S. Shahmiri; S. Andersen; P. Møller (pp. 110-118).
Exposure levels of persistent organic pollutants, such as polychlorinated biphenyls and DDTs, are high in Arctic apex predators and Inuit peoples and are suspected to have negative impacts on their immune systems. We conducted a controlled generational study on liver tissue and ethylenediaminetetraacetic acid blood cytokine and acute-phase protein (APP) mRNA expressions using reverse transcriptionase–polymerase chain reaction in West Greenland sledge dogs (Canis familiaris); contaminated minke whale (Balaenoptera acutorostrata) blubber serves as dietary pollutant source. Two of seven blood cytokine expressions (IL-6 and IL-12) and three of five APP expressions (haptoglobin [HP], heat shock protein, and fatty acid–binding protein [FABP]) were lowest in the exposed group, whereas the remaining five blood cytokine expressions (IL-2, IL-10, IFN-γ, TNF-α, and TGF-β) and two APP expressions (MT1 and MT2) were highest in the exposed group. In liver tissue, three cytokine expressions (IL-10, IFN-γ, and TNF-α) and two APP expressions (MT1 and MT2) were highest in the exposed group, and the remaining cytokine and APP expressions were lowest in the exposed group. Of these, the liver tissue expression of HP and FABP was significantly lowest in the exposed group (both p < 0.05). As a consequence of our findings, we suggest that a daily intake of 50 to 200 g polluted whale blubber is associated with a genotoxic decrease in HP and FABP gene expression in the liver of sledge dog and possibly of other top mammalian marine predators and consumers in the Arctic, indicating a restricted acute-phase reaction and insufficient immune response. Finally, HP and FABP liver expression appear to be new and sensitive biomarkers of organohalogen compound exposure.
Accumulation of Organotin Compounds in Tissues and Organs of Stranded Whales Along the Coasts of Thailand by H. Harino; M. Ohji; G. Wattayakorn; K. Adulyanukosol; T. Arai; N. Miyazaki (pp. 119-125).
Concentrations of butyltin (BT) and phenyltin (PT) compounds were measured in organs and tissues of five species of whales (Bride’s whale [Balaenoptera edeni], false killer whale [Pseudorca crassidens], pygmy sperm whale [Kogia breviceps], short-finned pilot whale [Globicephala macrorhynchus], and sperm whale [Physeter macrocephalus]) found stranded on the coasts of Thailand. The mean concentrations of BTs in various whales were in the range of 0.157 to 1.03 mg kg–1 wet weight, which were higher levels than the reported concentrations in whales from other countries. PT concentrations were also detected in the range of 0.022 to 1.14 mg kg–1 wet weight. The concentrations of BTs and PTs in whales were higher than those in mussels from the coastal area of Thailand. Concentrations of tributyltin (TBT) and triphenyltin (TPT) compounds in whale organs and tissues were also compared, and it was found that TBT concentrations were generally higher in liver and lower in lung. TPT concentrations were higher in liver and blubber and lower in lung. Ratios of TBT degradation products in whale liver, namely monobutyltin (MBT) and dibutyltin (DBT), were higher than the ratios of TBT. TPTs in liver were found to be dominant among PTs. The patterns of BTs and PTs in false killer whale liver were different from those in the other whales by cluster analysis. Their concentrations in false killer whales were the highest among all whales in this study. False killer whales feed on squid and large pelagic fish containing higher concentrations of organotin (OT) compounds, so the differences in patterns and concentrations of OTs in liver between false killer whales and the other whales may be caused by difference in diet.
Time- and Dose-Dependent Effects of Roundup on Human Embryonic and Placental Cells by N. Benachour; H. Sipahutar; S. Moslemi; C. Gasnier; C. Travert; G. E. Séralini (pp. 126-133).
Roundup® is the major herbicide used worldwide, in particular on genetically modified plants that have been designed to tolerate it. We have tested the toxicity and endocrine disruption potential of Roundup (Bioforce®) on human embryonic 293 and placental-derived JEG3 cells, but also on normal human placenta and equine testis. The cell lines have proven to be suitable to estimate hormonal activity and toxicity of pollutants. The median lethal dose (LD50) of Roundup with embryonic cells is 0.3% within 1 h in serum-free medium, and it decreases to reach 0.06% (containing among other compounds 1.27 mM glyphosate) after 72 h in the presence of serum. In these conditions, the embryonic cells appear to be 2–4 times more sensitive than the placental ones. In all instances, Roundup (generally used in agriculture at 1–2%, i.e., with 21–42 mM glyphosate) is more efficient than its active ingredient, glyphosate, suggesting a synergistic effect provoked by the adjuvants present in Roundup. We demonstrated that serum-free cultures, even on a short-term basis (1 h), reveal the xenobiotic impacts that are visible 1–2 days later in serum. We also document at lower non-overtly toxic doses, from 0.01% (with 210 μM glyphosate) in 24 h, that Roundup is an aromatase disruptor. The direct inhibition is temperature-dependent and is confirmed in different tissues and species (cell lines from placenta or embryonic kidney, equine testicular, or human fresh placental extracts). Furthermore, glyphosate acts directly as a partial inactivator on microsomal aromatase, independently of its acidity, and in a dose-dependent manner. The cytotoxic, and potentially endocrine-disrupting effects of Roundup are thus amplified with time. Taken together, these data suggest that Roundup exposure may affect human reproduction and fetal development in case of contamination. Chemical mixtures in formulations appear to be underestimated regarding their toxic or hormonal impact.
Automated Solid Phase Extraction and Quantitative Measurement of 2,3-Dibromo-1-Propanol in Urine Using Gas Chromatography-Mass Spectrometry by G. K. Hemakanthi De Alwis; Larry L. Needham; Dana B. Barr (pp. 134-139).
2,3-Dibromo-1-propanol (DBP) was used as an active flame retardant in the 1970s. It was also used as an intermediate in the preparation of insecticide formulations, pharmaceuticals and the flame retardants tris(2,3-dibromopropyl) phosphate (Tris-BP) and tetrabromobisphenol A bis (2,3-dibromopropyl ether). DBP is also produced in vivo as a metabolic product of Tris-BP in humans. In 1977, sleepwear containing DBP and Tri-BP was banned because of evidence of carcinogenicity animal studies. Although the production of DBP was reduced after 1977, studies show that DBP is still detected in indoor air and dust; hence, the U.S. population may be exposed potentially to DBP. Only a few methods have been reported in the literature for assessing exposure to DBP or Tris-BP by measuring DBP in urine. These methods are based on a labor-intensive and time-consuming liquid-liquid extraction for the isolation of DBP from the urine matrix. To measure urinary DBP in humans, a fast, accurate, and sensitive method was developed with a limit of detection of 0.1 ng/mL and extraction recovery of 96%. This method involves enzymatic cleavage of the DBP-glucuronide or sulfate conjugate, automated solid phase extraction, and analysis by gas chromatography-mass spectrometry using 1,4-dibromo-2-butanol as the internal standard.