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The extraction of bitumen from surface mining in the Athabasca Oil Sands Region (AOSR) produces large quantities of oil sands process‐affected water (OSPW) that needs to be stored in settling basins near extraction sites. Chemical constituents of OSPW are known to impair bone health in some organisms, which can lead to increased fracture risk and lower reproductive fitness. Naphthenic acid fraction components (NAFCs) are thought to be among the most toxic class of compounds in OSPW; however, the effect of NAFCs on osteoblast development is largely unknown. In this study, we demonstrate that NAFCs from OSPW inhibit osteoblast differentiation and deposition of extracellular matrix, which is required for bone formation. Extracellular matrix deposition was inhibited in osteoblasts exposed to 12.5–125 mg/L of NAFC for 21 days. We also show that components within NAFCs inhibit the expression of gene markers of osteoblast differentiation and function, namely, alkaline phosphatase (Alp), osteocalcin, and collagen type 1 alpha 1 (Col1a1). These effects were partially mediated by the induction of glucocorticoid receptor (GR) activity; NAFC induces the expression of the GR activity marker genes Sgk1 (12.5 mg/L) and p85a (125 mg/L) and inhibits GR protein (125 mg/L) and Opg RNA (12.5 mg/L) expression. This study provides evidence that NAFC concentrations of 12.5 mg/L and above can directly act on osteoblasts to inhibit bone formation and suggests that NAFCs contain components that can act as GR agonists, which may have further endocrine disrupting effects on exposed wildlife. Surface mining in the Athabasca Oil Sands Region (AOSR) produces large quantities of oil sands process‐affected water (OSPW), constituents of which are known to impair bone health in some organisms. Naphthenic acid fraction components (NAFCs) are thought to be responsible for most of the toxic effect of OSPW, but their effects on bone are unknown. In this study, we demonstrate that NAFCs extracted from the AOSR inhibit osteoblast differentiation and may act via the glucocorticoid receptor. [ABSTRACT FROM AUTHOR]

Our study evaluated whether exposure to naphthenic acid fraction compounds (NAFCs) extracted from oil sands process–affected waters (OSPW) has adverse effects on fish embryos that persist into later life. We exposed fathead minnow (Pimephales promelas) embryos to concentrations of NAFCs found in OSPW (2.5–54 mg/L) for 7 days (1 day postfertilization to hatch), then raised surviving larvae in outdoor mesocosms of uncontaminated lake water for 1 month. Embryos exposed to NAFCs were more likely to exhibit malformations (by up to 8‐fold) and had slower heart rates (by up to 24%) compared to controls. Fish raised in uncontaminated lake water following exposure to NAFCs as embryos, were 2.5‐fold less likely to survive during the larval stage than control fish. These fish also showed up to a 45% decrease in swim activity and a 36% increase in swim burst events during behavioral tests relative to controls. We conclude that exposure to NAFCs during the embryonic stage can have lasting effects on fish survival, physiology, and behavior that persist at least through the larval stage. These findings of delayed mortalities and persistent sublethal effects of embryonic NAFC exposure are relevant to informing the development of regulations on treated OSPW releases from mining operations. Environ Toxicol Chem 2022;41:1319–1332. © 2022 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC. [ABSTRACT FROM AUTHOR]

The aim of the present study was to evaluate the effects of exposure to Chilean pulp mill effluent extracts on developing postfertilized medaka embryos before and after sex definition relative to sex steroids (testosterone and 17beta‐estradiol) and a wood phytoestrogen (beta‐sitosterol). Our study included 2 waterborne semichronic exposure experiments, using a 24‐h post fertilization (hpf) unknown‐sex FLFII (female leucophore free) group and a second 72‐hpf FLFII phenotypic sex‐identified group (male autofluorescence leucophore) strain of medaka embryos. Chronic exposure of both FLFII strain embryo groups showed similar delay in time to hatch and decreased hatchability. Teratogenic responses such as vertebral malformation (fusion, incomplete formation, and lack of vertebral formation process) and pericardial edema were observed in both experiments, with a high percentage related to FLFII fluorescent leucophore–identified males. In addition, high mortality associated with severe malformations was observed in male and female embryos exposed to testosterone. Our research has demonstrated that exposure to Chilean mill effluent extracts caused severe male medaka embryotoxicity (in postfertilized embryos) before and after sex definition and, irrespective of the experimental group and effluent treatment, suggests partial removal following secondary treatment. Furthermore, differences in the severity and type of teratogenic effects with previous experiments (d‐rR medaka strain), are associated with the unique phenotypes of this medaka mutant strain. Environ Toxicol Chem 2021;40:2297–2305. © 2021 SETAC [ABSTRACT FROM AUTHOR]

Rationale: The objective of this study was to identify unique chemical tracers of oil sands process-affected water (OSPW) to enable definitive discrimination of tailings pond seepage from natural bitumen-influenced waters from the Canadian Alberta McMurray formation. Methods: The approach involved comparing unknowns from an unprecedented sample set of OSPW (n = 4) and OSPW-affected groundwaters (n = 15) with natural bitumen-influenced groundwaters (n = 20), using high-performance liquid chromatography/electrospray ionisation high-resolution mass spectrometry (HPLC/ESI-HRMS) operated in both polarities. Results: Four unknown chemical entities were identified as potential tracers of OSPW seepage and subsequently subjected to structural elucidation. One potential tracer, tentatively identified as a thiophene-containing carboxylic acid [C15H23O3S]-, was only detected in OSPW and OSPW-affected samples, thereby showing the greatest diagnostic potential. The remaining three unknowns, postulated to be two thiochroman isomers [C17H25O3S]+ and an ethyl-naphthalene isomer [C16H21]+, were detected in one and two background groundwaters, respectively. Conclusions: We advanced the state of knowledge for tracers of tailings seepage beyond heteroatomic classes, to identifying diagnostic substances, with structures postulated. Synthesis of the four proposed structures is recommended to enable structural confirmations. This research will guide and inform the Oil Sands Monitoring Program in its efforts to assess potential influences of oil sands development on the Athabasca River watershed. [ABSTRACT FROM AUTHOR]

Effects‐directed analysis (EDA) is used to identify the principal toxic components within a complex mixture using iterative steps of chemical fractionation guided by bioassay results. Bioassay selection can be limited in EDA because of the volume requirements for many standardized test methods, and therefore, a reduced‐volume acute toxicity test that also provides whole‐organism responses is beneficial. To address this need, a static, 7‐d, water‐only, reduced‐volume method (50 mL, 10 organisms) was developed for Hyalella azteca that substantially decreases the volume requirements of standard‐volume acute test exposures (200–500 mL of test solution, 15–20 organisms) while maintaining water quality and meeting control survival criteria. Standard‐ and reduced‐volume methods were compared by conducting concurrent toxicity tests with 2 inorganic toxicants (KCl and CdCl2) and 2 organic mixtures of naphthenic acid fraction components (NAFCs) to evaluate test performance. There was no difference between methods when comparing the median lethal concentrations (LC50s) for KCl and both NAFC mixtures (p > 0.05). The LC50s for CdCl2 were statistically different (p = 0.0002); however, this was not considered biologically meaningful because the difference between LC50s was <2‐fold. In conclusion, the reduced‐volume H. azteca test method generated results comparable to standard‐volume test methods and is suitable for use in situations where limited testing material is available, such as when conducting EDA. Environ Toxicol Chem 2020;39:2221–2227. © Her Majesty the Queen in Right of Canada 2020. Reproduced with the permission of the Minister of Environment and Climate Change Canada. [ABSTRACT FROM AUTHOR]

Introduction: Neurogenic orthostatic hypotension (nOH) is associated with neurodegenerative conditions, may cause symptoms of end‐organ hypoperfusion, increases fall risk, and can negatively impact quality of life. Droxidopa is approved for the treatment of symptomatic nOH in adults. As the largest subpopulation of patients with nOH has a diagnosis of Parkinson disease (PD), the efficacy and tolerability of droxidopa in patients with PD and nOH were examined using integrated clinical trial data. Methods: Post hoc analyses included data from the phase 3, randomized, placebo‐controlled clinical trials of droxidopa (two short‐term [1−2 weeks] trials and one medium‐term [8−10 weeks] trial) in the subset of participants with PD and symptomatic nOH. Efficacy was assessed using standing blood pressure (BP) measurements and the Orthostatic Hypotension Questionnaire (OHQ), a patient‐reported evaluation of nOH symptoms (Orthostatic Hypotension Symptom Assessment [OHSA]), and their impact (Orthostatic Hypotension Daily Activity Scale [OHDAS]). Results: The analysis included 307 patients with PD (droxidopa, n = 150; placebo, n = 157). Compared with placebo, droxidopa significantly improved the OHQ composite score (P = 0.014), the OHSA composite score (P = 0.022), and the OHDAS composite score (P = 0.029) from baseline to end of study/week one. We found significant increases in standing mean systolic/diastolic BP for droxidopa versus placebo (P = 0.003/0.002). Adverse event (AE) rates were qualitatively similar between groups; the most frequently reported AEs in the droxidopa groups included headache, dizziness, nausea, and hypertension. Conclusions: These post hoc analyses suggest that droxidopa provides meaningful clinical benefits and is well tolerated in the treatment of symptomatic nOH in patients with PD. [ABSTRACT FROM AUTHOR]

Abstract: A double‐blind, 4‐period crossover study (NCT01327066) was conducted to assess the effect of the novel norepinephrine prodrug droxidopa on the QT interval in in healthy subjects. Subjects were randomized to receive a single dose of droxidopa 600 mg (maximal dose) and 2000 mg (supratherapeutic dose) compared with the positive control, moxifloxacin 400 mg, and placebo, each separated by a 3‐day washout period. Patients were monitored by continuous Holter monitoring, and electrocardiograms (ECGs) were extracted 0.5–23 hours after dosing. Blood samples for pharmacokinetic analysis were collected before dosing and after ECG data collection. The primary end point was the time‐matched placebo‐adjusted change from baseline in the individually corrected QT (QTcI). The time‐averaged QTcI mean placebo‐corrected changes from baseline for droxidopa 600 and 2000 mg were 0.1 milliseconds (90%CI, ‐0.9 to 1.0 milliseconds) and 0.3 milliseconds (90%CI, ‐0.6 to 1.3 milliseconds), respectively, and 9 milliseconds (90%CI, 8.4–10.3 milliseconds) for moxifloxacin. This study found no effect of either dose of droxidopa on cardiac repolarization using QTcI. Analysis of the pharmacokinetic/pharmacodynamic relationship and cardiac repolarization showed no association with droxidopa exposure. There were no clinically relevant effects of droxidopa on heart rate, atrioventricular conduction, or cardiac depolarization identified. No morphologic ECG changes were observed. [ABSTRACT FROM AUTHOR]

The authors examined the potential of pulp mill effluent from pulp-producing countries (Canada, Brazil, New Zealand) to affect fish reproduction. Specifically, the estrogenic effects in juvenile rainbow trout ( Oncorhynchus mykiss) pulse-exposed to 11 different mill effluent extracts (intraperitoneal injections of solid-phase extraction-dichloromethane nonpolar fraction). The results indicated that effluent extracts were estrogenic in juvenile trout irrespective of the gender, as reflected by increasing level of plasma vitellogenin (VTG; Brazil > New Zealand > Canada). Despite the high variability observed among mills, differences in VTG levels were related to the type of mill process (kraft > elementary chlorine-free kraft > thermomechanical pulping). Moreover, effluent treatments did not appear to significantly decrease VTG induction. A consistent estrogenic effect was observed in those mills that process a combination of feedstocks (softwood and hardwood), with the highest increase in VTG related to eucalyptus feedstock. The results demonstrate significant estrogenic effects of pulp mill effluents on chronically exposed juvenile trout, suggesting that in vivo metabolic activation of precursors is necessary to cause the observed increases in VTG levels. This molecular estrogenic response provides a useful starting point for predicting population-level impacts through the adverse outcome pathway methodology. Environ Toxicol Chem 2017;36:1547-1555. © 2016 SETAC [ABSTRACT FROM AUTHOR]

The open pit oil sands mining operations north of Fort McMurray, Alberta, Canada, are accumulating tailings waste at a rate approximately equal to 4.9 million m3/d. Naphthenic acids are among the most toxic components within tailings to aquatic life, but structural components have largely remained unidentified. In the present study, electrospray ionization high-resolution mass spectrometry (ESI-HRMS) and synchronous fluorescence spectroscopy (SFS) were used to characterize fractions derived from the distillation of an acid-extractable organics (AEO) mixture isolated from oil sands process-affected water (OSPW). Mean molecular weights of each fraction, and their relative proportions to the whole AEO extract, were as follows: fraction 1: 237 Da, 8.3%; fraction 2: 240 Da, 23.8%; fraction 3: 257 Da, 26.7%; fraction 4: 308 Da, 18.9%; fraction 5: 355 Da, 10.0%. With increasing mean molecular weight of the AEO fractions, a concurrent increase occurred in the relative abundance of nitrogen-, sulfur-, and oxygen-containing ions, double-bond equivalents, and degree of aromaticity. Structures present in the higher-molecular-weight fractions (fraction 4 and fraction 5) suggested the presence of heteroatoms, dicarboxyl and dihydroxy groups, and organic acid compounds with the potential to function as estrogens. Because organic acid compositions become dominated by more recalcitrant, higher-molecular-weight acids during natural degradation, these findings are important in the context of oil sands tailings pond water remediation. Environ Toxicol Chem 2015;34:1001-1008. © 2015 SETAC [ABSTRACT FROM AUTHOR]

ABSTRACT Neurogenic orthostatic hypotension (nOH) results from failure of norepinephrine responses to postural change to maintain standing systolic blood pressure (s-SBP). Droxidopa is an oral prodrug of norepinephrine. Study nOH306 enrolled patients with Parkinson's disease (PD) and symptomatic nOH. Subjects underwent up to 2 weeks of double-blind titration of droxidopa or placebo, followed by 8 weeks of double-blind maintenance treatment (100-600 mg thrice-daily). For the initial 51 subjects (study nOH306A, previously reported), the primary efficacy measure, Orthostatic Hypotension Questionnaire (OHQ) composite score, did not demonstrate significant change versus placebo at maintenance week 8. For the subsequent 171 subjects (study nOH306B, reported here), the primary efficacy measure was change versus placebo on item 1 ('dizziness, lightheadedness, feeling faint, or feeling like you might black out') of the Orthostatic Hypotension Symptom Assessment (OHSA) subsection of the OHQ at maintenance week 1. At week 1, mean (standard deviation) improvement on OHSA item 1 was 2.3 (2.95) for droxidopa versus 1.3 (3.16) for placebo ( P = 0.018). In addition, mean increase in s-SBP at week 1 was 6.4 (18.85) for droxidopa versus 0.7 (20.18) mmHg for placebo (nominal P value: 0.032). Differences in change in OHSA item 1 scores from baseline to maintenance weeks 2, 4, and 8 were not statistically significant. Adverse-event (AE) incidence was similar across groups, but 12.4% of droxidopa and 6.1% of placebo subjects withdrew because of AEs. The most common AEs on droxidopa (vs. placebo) were headache (13.5% vs. 7.3%) and dizziness (10.1% vs. 4.9%). Study nOH306B demonstrated subjective (OHSA item 1) and objective (s-SBP) evidence of short-term droxidopa efficacy (vs. placebo) for symptomatic nOH in PD. © 2014 The Authors. Movement Disorders published by Wiley Periodicals, Inc. on behalf of International Parkinson and Movement Disorder Society. [ABSTRACT FROM AUTHOR]

RATIONALE: Characterising complex mixtures of organic compounds in polar fractions of heavy petroleum is challenging, but is important for pollution studies and for exploration and production geochemistry. Oil sands process-affected water (OSPW) stored in large tailings ponds by Canadian oil sands industries contains such mixtures. METHODS: A polar OSPW fraction was obtained by silver ion solid-phase extraction with methanol elution. This was examined by numerous methods, including electrospray ionisation (ESI) Fourier transform ion cyclotron resonance mass spectrometry (FTICRMS) and ultra-high-pressure liquid chromatography (uHPLC)/Orbitrap MS, in multiple ionisation and MS/MS modes. Compounds were also synthesised for comparison. RESULTS: The major ESI ionisable compounds detected (+ion mode) were C15-28 SO3 species with 3-7 double bond equivalents (DBE) and C27-28 SO5 species with 5 DBE. ESI-MS/MS collision-induced losses were due to water, methanol, water plus methanol and water plus methyl formate, typical of methyl esters of hydroxy acids. Once the fraction was re-saponified, species originally detected by positive ion MS, could be detected only by negative ion MS, consistent with their assignment as sulphur-containing hydroxy carboxylic acids. The free acid of a keto dibenzothiophene alkanoic acid was added to an unesterified acid extract of OSPW in known concentrations as a putative internal standard, but attempted quantification in this way proved unreliable. CONCLUSIONS: The results suggest the more polar acidic organic SO3 constituents of OSPW include C15-28 S-containing, alicyclic and aromatic hydroxy carboxylic acids. SO5 species are possibly sulphone analogues of these. The origin of such compounds is probably via further biotransformation (hydroxylation) of the related S-containing carboxylic acids identified previously in a less polar OSPW fraction. The environmental risks, corrosivity and oil flow assurance effects should be easier to assess, given that partial structures are now known, although further identification is still needed. [ABSTRACT FROM AUTHOR]

ABSTRACT Estuarine eutrophication as a result of agricultural land use, including the use of chemical fertilizers, is increasing worldwide. Prince Edward Island (PEI), Canada has very high agricultural intensity by international standards with approximately 44% of the land area under production, and some watersheds in excess of 75% agricultural land-use. The type of agriculture is also intensive with primarily row crops that have high chemical fertilizer and pesticide usage. In light of these stressors, the hypothesis of this study was that mummichog ( Fundulus heteroclitus) population parameters would change with point and nonpoint source pollution, and that multivariate statistics could be used to draw associations with specific stressors. Fish were sampled on a monthly basis from May through August at 7 estuaries spanning a range of land use, nutrient, and contaminant loadings. A suite of environmental variables were simplified into 3 principal components: PC1 representing agricultural land use, N loading, and plant habitat, PC2 being dominated by sediment sand and silt distribution, and PC3 largely reflecting P loading and sediment organic matter. There were significant differences in abundance of both adult and young-of-the-year mummichog, and these changes associated most strongly with PC1, the largely N-driven agricultural influences. In contrast, somatic variables such as liver and gonad size did not show strong association with the environmental quality principal component scores. The sand and silt PC2 appeared to have the opposite association with the biological data, with siltier environments correlating to older, larger, less dense populations of mummichog. Although pesticide residues were detected in estuarine sediment, there was no clear relationship between these and watershed agricultural intensity or biochemical indicators. There was, however, a strong relationship between agricultural environmental variables (PC1) and in vitro steroid production that is suggestive of a potential chemical effect. Eutrophication appeared to be a primary stressor affecting mummichog populations, as nutrient enrichment was associated with changes in habitat variables and these in turn were associated with high mummichog density. Thus, mummichog population demographics appear to have use as an indicator of adverse or worsening conditions in estuaries. We concluded that, based on the subset of environmental factors evaluated, the nonpoint-source inputs of sediments and nutrients exerted the greatest influence on mummichog populations in PEI estuaries. Integr Environ Assess Manag 2013;9:496-507. © 2013 SETAC [ABSTRACT FROM AUTHOR]

The overall health and endocrine function of wild brown bullhead ( Ameiurus nebulosus) and goldfish ( Carassius auratus) from the Wheatley Harbour Area of Concern (Lake Erie, Ontario, Canada) was assessed using a suite of physiological and biochemical endpoints. Smaller gonads were detected in female brown bullhead and goldfish from Wheatley Harbour compared with Hillman Marsh (Ontario, Canada) reference fish. Female brown bullhead exhibited decreased in vitro synthesis of 17β-estradiol. Female goldfish had decreased plasma vitellogenin concentrations. Plasma testosterone and 11-ketotestosterone were significantly depressed in males of both species. Perturbations in the thyroid status were detected, but varied between sexes and species. Observed differences included lower plasma concentrations of thyroid hormones and/or elevated liver deiodinase activity. Histological evaluation of the thyroid tissue indicated that in the case of female goldfish, those perturbations stimulated the thyroid (as indicated by increased thyroid epithelial cell height) and partially depleted the thyroxine reserves, as indicated by decreased colloid and elevated thyroid activation index. Increased mixed-function oxygenase activity in brown bullhead from Wheatley Harbour was consistent with exposure to planar aromatic contaminants. A principal component analysis of selected variables showed the separation of fish by collection site. The endpoints most strongly associated with the separation were generally those exhibiting significant differences between sites. The results of the present study indicate that the health of fish populations within Wheatley Harbour warrants continued attention. Environ. Toxicol. Chem. 2012;31:2798-2811. © 2012 Crown in the Right of Canada [ABSTRACT FROM AUTHOR]

Pulp and paper mill effluents have been linked to effects on fish reproduction for more than 25 years. To date, the causes of these effects and remedial strategies have eluded investigators. Recent work has shown that the degree of reproductive effect caused by a mill effluent is related to the overall organic content. If verified, this could lead to breakthroughs for best management practices (BMPs). For this study, the effluents from seven kraft mills were assessed for their ability to reduce egg production in fathead minnow ( Pimephales promelas) in the laboratory. At the same time, the effluents were analyzed for three parameters thought to be good indicators of organic losses: biochemical oxygen demand (BOD), methyl-substituted 2-cyclopentenones (cellulose degradation products), and a gas chromatographic (GC) profile index, which integrates the total area of the chromatographic peaks of solvent-extracted effluents using low-resolution mass spectrometry. The results showed that the degree to which the effluents reduced egg production increased with increased organic losses as characterized by BOD and the GC profile index. Therefore, these parameters could be used to guide BMPs at kraft mills according to specific targets: BOD < 20 mg/L and GC profile index equivalent to effluent with BOD of 20 mg/L. Such targets should be achievable by good in-plant control of organic losses and optimized effluent biotreatment systems. Environ. Toxicol. Chem. 2011; 30:1421-1429. © 2011 SETAC [ABSTRACT FROM AUTHOR]

For the last 20 years, studies conducted in North America, Scandinavia, and New Zealand have shown that pulp and paper mill effluents affect fish reproduction. Despite the level of effort applied, few leads are available regarding the factors responsible. Effluents affect reproduction in multiple fish species, as evidenced by decreased gonad size, decreased circulating and gonadal production of reproductive steroids, altered expression of secondary sex characteristics, and decreased egg production. Several studies also have shown that effluent constituents are capable of accumulating in fish and binding to sex steroid receptors/binding proteins. Studies aimed at isolating biologically active substances within the pulping and papermaking process have provided clues about their source, and work has progressed in identifying opportunities for in-mill treatment technologies. Following comparisons of manufacturing processes and fish responses before and after process changes, it can be concluded that effluent from all types of mill processes are capable of affecting fish reproduction and that any improvements could not be attributed to a specific process modification (because mills normally performed multiple modifications simultaneously). Improved reproductive performance in fish generally was associated with reduced use of molecular chlorine, improved condensate handling, and liquor spill control. Effluent biotreatment has been effective in reducing some effects, but biotreated effluents also have shown no difference or an exacerbation of effects. The role of biotreatment in relation to effects on fish reproduction remains unclear and needs to be resolved. [ABSTRACT FROM AUTHOR]

The results of the present study have demonstrated the utility of an estrogen receptor (ER) competitive ligand-binding assay, a hepatocyte vitellogenin (VTG) induction bioassay, and an ER reporter gene bioassay in characterizing the activity of model estrogen agonists (17β-estradiol [E2], ethynylestradiol, and nonylphenol) and antagonists (tamoxifen and ZM 189154) in rainbow trout (Oncorhynchus mykiss). The in vitro results were validated against in vivo trout waterborne exposures to E2 and tamoxifen. The results showed that all three in vitro assays were capable of detecting the hormonal activities of the selected model compounds in a dose-dependent manner, with the exception of nonylphenol in the ER reporter gene bioassay. However, the relative potency rankings of the model compounds were not consistent between these assays, which suggests that the relative potencies obtained within assays may have limited predictive value between assays. Discrepancies in potencies most likely can be attributed to the different levels of cellular organization in each assay. In addition to model compounds, we also evaluated the responses of the ER-binding assay and the hepatocyte VTG induction bioassay to complex mixtures associated with endocrine effects in fish, specifically extracts of pulp mill effluent. Of the 14 pulp mill effluent extracts tested, only six showed activity in both assays, whereas the remaining eight samples showed activity in only one of the two assays. The hepatocyte VTG induction bioassay consistently showed that the pulp mill effluent extracts were antiestrogenic, which to our knowledge has not been reported in previous studies. Collectively, these results suggest that a combination of in vitro assays that depend on differing endpoints is required to identify potential xenoestrogens and to characterize their modes of action. [ABSTRACT FROM AUTHOR]

In previous experiments, bleached kraft chemical recovery condensates generated during softwood pulp production have been identified as a primary source of substances causing testosterone depressions in mummichog (Fundulus heteroclitus). Recent toxicity identification evaluations have encountered inconsistencies in the potential of condensates and fractions to reduce steroid concentrations between three repeated exposures. To assess the sources of these inconsistencies, temporal assessments of condensate extractives and analytical methodologies were evaluated. Condensates were collected during a six-month period, extracted by solid-phase extraction (SPE), and analyzed by gas chromatography-mass spectrometry and gas chromatography-flame photometric detection. All unique extractives were cataloged by mass spectra, peak area, and gas chromatographic retention time, and nine were confirmed and quantified against authentic standards. Confirmed components included phenolic guaiacyl-based lignin degradation products, sulfur (S8), three diterpenoids, and a dimethoxy stilbene. Concentrations of confirmed condensate extractives were consistent in all samples collected. Spiking experiments of confirmed extractives revealed substantial losses following high-pressure liquid chromatographic fractionation and three different methods of fraction preparation. In an effort to propose chemical classes associated with biological activity, all unique condensate extractives in previously established bioactive and inactive fractions were classified based on their potential to depress testosterone in mummichog. Of 39 unique components in bioactive SPE extracts of condensates, six were associated with hormonal activity. Mass spectral interpretation indicated hydroxylated diterpenoids, sesquiterpenoids, and a lignin-derived stilbene as classes of chemicals associated with steroid depressions. [ABSTRACT FROM AUTHOR]

Reproductive effects have been recorded in wild fish in waters receiving pulp mill effluent (PME) since the mid to late 1980s. Laboratory assays with fathead minnow (FHM; Pimephales promelas) have been developed to better understand fish responses to PME. However, observations from laboratory studies have been variable, making it difficult to establish cause/effect relationships. A lack of environmental relevance in these laboratory studies may have contributed to the variability observed. The objectives of the present study were, first, to determine the effects of bleached kraft PME (BKME) on FHM under environmentally realistic conditions (i.e., ambient water and effluent quality) and, second, to determine the suitability of pair-breeding FHM to better link BKME-induced changes in indicators at the biochemical, individual, and population levels. A mobile bioassay trailer was situated on-site at a bleached kraft mill for 60 d, allowing supply of both ambient water (Lake Superior, Canada) and final BKME. The reproductive output of FHM was initially assessed for 21 d to obtain baseline data pre-exposure. At the end of the pre-exposure period, selected breeding pairs were exposed to final BKME (100% v/v and 1% v/v) for 21 d. Results demonstrated a stimulatory response pattern at 1% BKME (e.g., increased egg production) compared to control. In the 100% treatment, spawning events were reduced and fewer eggs were produced during the first two weeks of exposure. Exposure to 100% (v/v) BKME also resulted in ovipositor development in males and development of male secondary sex characteristics in females. Obtaining pre-exposure data and use of pair-breeding FHM in this assay gave a sensitive indication of effluent effects and allowed accurate comparisons of endpoints to be made. [ABSTRACT FROM AUTHOR]

Reproductive effects of pulp mill effluents on fish continue to be reported in Canadian waters. Spawning delays, reduced gonad size, and altered levels of sex steroid hormones have been found in both sexes of various fish species exposed to effluents. We initiated a project to identify the source/cause of such effects. In part 1 of this two-part series, we exposed mature adult fathead minnow (FHM; Pimephales promelas) for 21 d to final treated effluent (1% and 100% v/v) from a bleached kraft pulp mill in Terrace Bay (ON, Canada). Results suggested pulp mill effluent from this mill affected reproductive indicators in FHM and effects were dependent on effluent concentration, duration of exposure, and method of data analysis. The main objective of this paper was to use the FHM assay to identify waste stream sources within the mill that affect reproductive indicators. Various process streams were selected, characterized with respect to effluent chemistry and acute toxicity, and a subset was tested on-site with the 21-d FHM bioassay. Results showed that both the combined mill effluent (before secondary treatment) and the combined alkaline stream (CALK) caused both decreased spawning events (∼55% for both streams) and decreased egg production (28 and 74%, respectively), and the CALK stream resulted in significant male ovipositor development. By comparing response patterns we were able to identify the CALK stream as a source of compounds at this mill affecting reproductive indicators in FHM and highlight this stream for further toxicity identification evaluation. [ABSTRACT FROM AUTHOR]

The accumulation of ligands for the aryl hydrocarbon receptor (AhR) and fish sex steroid receptors was investigated using two separate controlled fish exposures to final effluent from a bleached sulfite/groundwood mill in New Brunswick, Canada. In the first experiment, hepatic tissue extracts from exposed fish were fractionated according to lipophilicity. Fractions with different octanol-water (Kow) partition coefficients were tested for the presence of bioavailable chemicals that function as ligands for the AhR in H4IIE cells, rainbow trout hepatic estrogen receptors (ER), goldfish testicular androgen receptors (AR), and goldfish sex steroid binding protein (SSBP). Fish accumulated ligands for each receptor after 4-d exposure to effluent. Single fractions contained ligands for the AhR and the ER, while multiple fractions competed for the AR and SSBP. Fish also accumulated ligands for the AhR and SSBP from Saint John River dilution water, indicating upstream sources of bioactive substances. Semipermeable-membrane devices deployed concurrently with fish accumulated ligands from effluent for all receptors except the ER. In the second experiment, accumulated ligands were evaluated after exposure of fish to effluent for two different durations and following a depuration period. Hepatic mixed function oxygenase activity and whole-liver hormonal activity, measured as binding to SSBP, returned to background following 6 d depuration and were reduced but still significant after 12-d exposure to effluent. Whole-liver extract affinities for the AR were maintained after extended exposure and depuration, indicating the potential for AR ligands to bioaccumulate. The accumulation of AhR ligands and ligands for sex steroid receptors provides a mechanistic linkage to effects on growth, development, and performance of fish exposed to effluent from this and other mills. [ABSTRACT FROM AUTHOR]

Reports on the development of a solid-phase extraction (SPE) method to isolate chemical recovery concentrate extractives for evaluation of hormonal activity. Potential of condensates generated during softwood and hardwood pulp production to affect circulating and gonadal production of sex steroids in mummichog.

The transport of farm animals has been the subject of much research in recent years; however, the information available on ostrich transportation is limited. This paper draws together past and recent scientific literature and practical experience. The current state of knowledge is summarised and areas which require further research are identified. [ABSTRACT FROM PUBLISHER]

ABSTRACT- Ketones can potentiate the hepatotoxicity of haloalkanes in animals. This may be due, in part, to changes in organelle susceptibility. Male Sprague-Dawley rats were administered 15 mmol/kg (po) acetone, 2-butanone, 2-hexanone or 50 mg/kg (po) chlordecone or mirex (a nonketonic analog of chlordecone). Eighteen hours later, tests of organelle structure/function were performed (osmotic stress, respiration, and calcium pump activity). Other rats were given 14CHCl3 (0.5 or 1.0 ml/kg, po) 18 h after chlordecone or mirex administration. Three hours later, the organelle distribution of 14C was evaluated. In a final experiment, ketone-pretreated (chlordecone or 2-hexanone) animals were killed 6 h after CHCl3 administration and evaluated morphologically for evidence of modified organelle response. Acetone and chlordecone, when given alone, enhanced lysosomal fragility to osmotic stress; no changes in functional capacity of mitochondria or microsomes were observed. CHCl3-derived 14C in the mitochondrial fraction increased 2-fold in chlordecone-treated rats. Morphological evaluation suggested mitochondria respond differently to CHCl3 in ketone-pretreated (chlordecone or 2-hexanone) animals compared to corn oil-pretreated controls. These results support the concept that modifications of organelles contribute to the mechanism of ketone-potentiation of CHCl3-induced hepatotoxicity. [ABSTRACT FROM AUTHOR]

The contamination of field formulations of the lampricide 3-trifluoromethyl-4-nitrophenol (TFM) by a dibenzo-p-dioxin containing -chloro,-nitro, and -trifluoromethyl substituents was suspected from chemical fractionations of a TFM formulation that were directed by mixed function oxidase (MFO) induction in rainbow trout. Three dioxin isomers containing these substituents in field formulations were identified by gas chromatography-mass spectrometry (GC-MS). Short-term waterborneexposures to a mixture of two isomers, 2-trifluoromethyl-3-nitro-7-(and 8)-chloro-dibenzo-p-dioxin, elevated MFO activity in trout, with a threshold between 0.143 and 0.745 nM (4.1--20.5 ng/L). Synthetic preparations of other dioxins related to formulation impurities enabledcharacterizations of this previously unknown family of dioxin congeners by CC-MS. The elution order of the isomers followed those established for halogenated dioxins except where there was a lone -nitro substitution on one ring. The average concentration of these compounds in TFM formulations spanning more than a decade was 288 1 47 mu g/L, which translates into an annual loading of approximately 40 g to the Great Lakes Basin. Using mammalian (H4IIE) and fish (PLHC-1) in vitro assays, a 2,3,7-substituted chloro-nitro-trifluoromethyl isomer was afour to five times more potent inducer of MFO activity than 2,3.7-trichlorodibenzo-p-dioxin. Characterizations of the synthetic isomers indicate that the isomers present in the formulation are not substituted in the 2,3,7- positions and are relatively weak inducers of MFO activity. [ABSTRACT FROM AUTHOR]

Lampricide formulations containing 3-trifluoromethyl-4-nitrophenol (TFM) have been associated with induction of hepatic mixed function oxygenase (MFO) activity and altered levels of circulating steroids in fish. Bioassay-directed chemical fractionations have shown MFO induction to be associated with impurities in TFM field formulations. A similar toxicity identification/evaluation approach coupled to competitive binding to rainbow trout estrogen receptors (ER) was used to identify compounds associated with estrogenic responses in fish. In contrast to MFO induction, nearly all binding to the ER was associated withfractions containing TFM, TFM isomers, and p-chlorophenol. p-Chlorophenol did not show any affinity for the receptor when tested separately. Diphenyl ether impurities identified in the fractions causing MFOinduction as well as analogues to dibenzo-p-dioxins suspected in these fractions also showed no affinity for the receptor when tested individually. Relative to estradiol, TFM demonstrated an affinity of 5.03 x 10-5, compared to 2.47 x 10-4 for p-nonylphenol, a reported estradiol agonist. Vitellogenin induction in primarycultures of rainbow trout hepatocytes indicated that TFM acts as an estradiol agonist. [ABSTRACT FROM AUTHOR]

Aims: This study describes the pharmacokinetic (PK)/target engagement (TE) relationship of tozorakimab, an anti-interleukin (IL)-33 antibody, by building a mechanistic population PK/TE model using phase 1 biomarker data., Methods: The analysis included tozorakimab PK and TE in serum assessed in 60 tozorakimab-treated participants, including healthy adults and patients with mild chronic obstructive pulmonary disease. Scenarios evaluated three dose frequencies (once every 2, 4 or 6 weeks) administered subcutaneously at seven doses of tozorakimab (30, 60, 90, 120, 150, 300 or 600 mg). For each dose, simulations were performed with 5000 virtual individuals to predict systemic TE. Inhibition of IL-33/soluble ST2 (sST2) complex levels at trough PK at steady state was assessed in each dosing scenario. The PK/TE modelling analyses were performed using a nonlinear mixed-effect modelling approach., Results: The final two-compartment PK model with tozorakimab binding IL-33 in the central compartment adequately described the systemic PK and TE of tozorakimab at population and individual levels. The mean PK parameter estimates of absorption rate, central volume of distribution and clearance were 0.48 (90% confidence interval [CI]: 0.40-0.59, 1/day), 12.64 (90% CI: 8.60-18.62, L) and 0.87 (90% CI: 0.65-1.16, L/day), respectively. Consistent with the observed value, tozorakimab bioavailability was 45%. For all three dose frequencies, predicted inhibition of systemic IL-33/sST2 levels was more than 95% at doses greater than 90 mg., Conclusions: The PK/TE model reliably quantified the relationship between PK and systemic TE of tozorakimab, with potential utility for predicting clinical dose-response relationships and supporting clinical dose selection., (© 2024 AstraZeneca. British Journal of Clinical Pharmacology published by John Wiley & Sons Ltd on behalf of British Pharmacological Society.)

The X-ray crystal structure of d(TGCGCA)2 has been determined at 120 K to a resolution of 1.3 A. Hexamer duplexes, in the Z-DNA conformation, pack in an arrangement similar to the 'pure spermine form' [Egli et al. (1991). Biochemistry, 30, 11388-11402] but with significantly different cell dimensions. The phosphate backbone exists in two equally populated discrete conformations at one nucleotide step, around phosphate 11. The structure contains two ordered cobalt hexammine molecules which have roles in stabilization of both the Z-DNA conformation of the duplex and in crystal packing. A comparison of d(TGCGCA)2 with other Z-DNA hexamer structures available in the Nucleic Acid Database illustrates the elusive nature of crystal packing. A review of the interactions with the metal cations Na+, Mg2+ and Co3+ reveals a relatively small proportion of phosphate binding and that close contacts between metal ions are common. A prediction of the water structure is compared with the observed pattern in the reported structure.