0000000000016547
AUTHOR
Aimo Oikari
The use of fish bile metabolite analyses as exposure biomarkers to pulp and paper mill effluents
The exposure of caged one-year-old whitefish (Coregonus lavaretus) to wood extractives and chlorophenolics from pulp and paper mill effluents in Lake Saimaa (SE Finland) was investigated. Whitefish were exposed at 17 locations affected by effluents from pulp and paper mills and at 5 reference areas. Resin and fatty acids and chlorophenolics in bile were analysed by GC and GC-MS. Cholesterol concentration in bile was also measured. Results show that despite of process changes to ECF and waste water treatment improvements employed by the pulp and paper industry in the study area, it is still possible to detect elevated concentrations of these substances in the bile of fish exposed in the vici…
Vertical distribution of AhR-activating compounds in sediments contaminated by modernized pulp and paper industry
Increased ethoxyresorufin-O-deethylase (EROD) activity is a sensitive biomarker of exposure to the chemicals which activate the aryl hydrocarbon receptor (AhR) and induce the cytochrome P450 system, such as many polychlorinated dibenzo-p-dioxins (PCDDs), polychlorinated dibenzofurans (PCDFs) and polychlorinated biphenyls (PCBs). Pulp bleaching was one of the main sources of PCDDs and PCDFs until elemental chlorine free (ECF) and total chlorine free bleaching processes since 1990s have remarkably decreased but not completely eliminate discharges of these chemicals. In addition, historically contaminated sediments may act as a source of these persistent contaminants. In this study, the contam…
Vertical distribution of pharmaceuticals in lake sediments-citalopram as potential chemomarker
The use of pharmaceuticals has increased enormously over the last few decades and serious concerns about their environmental fate and effects have arisen. Thus far, there is little knowledge about the historical pollution of the environment by pharmaceuticals. In the present study, sediment columns from three lake sites adjacent to wastewater treatment plants were collected, further divided in 2- or 2.5-cm slices, and analyzed for 15 pharmaceuticals by liquid chromatography-tandem mass spectrometry. In addition, sediment columns were historically interpreted by analyzing fecal sterols (coprostanol and cholesterol) as well as organic material and organic carbon. Several pharmaceuticals were …
Identification and dose dependency of ibuprofen biliary metabolites in rainbow trout
Abstract The biotransformation of the anti-inflammatory drug ibuprofen (IBF) was studied by exposing rainbow trout ( Oncorhynchus mykiss ) to IBF via intraperitoneal (i.p.) injection, and via water at four (0.17, 1.9, 13 and 145 μg L −1 ) exposure levels for 4 d. Following exposure, the bile was collected and analyzed by LC–MS/MS methods. The identification of the formed metabolites in i.p. injected fish bile was based on the exact mass determinations by a time-of-flight mass analyzer (Q–TOF–MS) and on the studies of fragments and fragmentation patterns of precursor ions by ion trap mass analyzer (IT-MS). In addition to unmetabolized IBF, several phase I and phase II metabolites were found …
Anaerobically digested poultry slaughterhouse wastes as fertiliser in agriculture
Chemical and physical analysis, 27-d plant growth assays with carrot (Daucus carota) and Chinese cabbage (Brassica campestris var. chinensis), and 5-d phytotoxicity assays with Chinese cabbage and perennial ryegrass (Lolium perenne) were used to investigate the suitability of anaerobically digested poultry slaughterhouse waste for fertiliser in agriculture and the effect of aerobic post-treatment on the properties of the digested material. The digested material appeared to be rich in nitrogen. In 27-d assays with digested material as nitrogen source, carrots grew almost as well as those fertilised with a commercial mineral fertiliser used as reference, whereas, the growth of Chinese cabbage…
Pharmaceuticals in settleable particulate material in urban and non-urban waters
Abstract Wastewater treatment plants (WWTP) are important sources of settleable particulate material (SPM), heading to sediments with natural suspended solids. To date, there is little information about the fate of pharmaceuticals in sediment systems. In this study, the objective was to determine if pharmaceuticals are detected in SPM at locations near WWTPs or even in rural areas, thus being susceptible for sedimentation. SPM samples were collected from 10 sites in Finland, grouped as reference, rural and wastewater effluent sites. SPM collectors were placed about 35 cm above bottom for about 2 months during summer. After extraction, a set of 17 pharmaceuticals was analyzed. Several pharma…
A field methodology to study effects of UV radiation on fish larvae.
There is a considerable lack of in situ specific information about the effects of UV-B radiation on limnic animals studied in the field. We exposed larval pike (Esox lucius L.) in two types of cuvettes (glass and quartz) placed at different depths (5 or 15 cm) to natural solar UV or to artificially enhanced UV-B (lamps on 3 h per day), simulating the scenarios for coming decades. Dose realism and comparability with earlier laboratory experiments was the main purpose, and therefore UV-B irradiances to the surface as well as underwater irradiances were directly measured. Result showed that UV-B dose rates in natural waters are low even though DOC concentration was low (4.8 mg/l) in our study …
Bioaccumulation and subchronic physiological effects of waterborne iron overload on whitefish exposed in humic and nonhumic water.
One-year-old whitefish, Coregonus lavaretus, were exposed to three types of iron-rich water, two dilutions for each, in a subchronic (30-day) experiment. In natural iron-rich humic water, both the bioaccumulation and physiological effects of iron exposure were negligible. In humic-free water with high amount of additional inorganic iron (nominally 8 mg Fe/L), Fe accumulated in gills, liver, and gut. This accumulation was accompanied by decreased glycogen phosphorylase activities and microsomal EROD activity in the liver as well as decreased plasma sodium and potassium concentrations. The third group of whitefish were exposed by adding inorganic iron (nominally 2 and 8 mg Fe/L) to natural ir…
Metabolites of the aquatic pollutant diclofenac in fish bile.
The uptake and metabolism of anti-inflammatory drug diclofenac (DCF) was studied by exposing rainbow trout (Oncorhynchus mykiss) to DCF intraperitoneally, and via water at concentration of 1.7 μg L(-1). The bile was collected and the formed metabolites were identified. The identification was based on the exact mass determinations by a time-of-flight mass analyzer and on the studies of fragments and fragmentation patterns of precursor ions by an ion trap mass analyzer. The main metabolites found were acyl glucuronides of hydroxylated DCFs. In addition, one ether glucuronide of hydroxylated DCF was found. Also, unmetabolized DCF was detected in the bile. The total bioconcentration factors (BC…
Fluence Rate or Cumulative Dose? Vulnerability of Larval Northern Pike (Esox lucius) to Ultraviolet Radiation
Newly hatched larvae of northern pike were exposed in the laboratory to four fluence rates of ultraviolet radiation (UVR; 290-400 nm) over three different time periods, resulting in total doses ranging from 3.0 +/- 0.2 to 63.0 +/- 4.4 kJ.m(-2). Mortality and behavior of the larvae were followed for 8-12 days, and growth measured at the end of the experiment. Also, the principle of reciprocity-that the UVR-induced mortality depends on the cumulative dose, independent of fluence rate-was tested. Fluence rates higher than 1480 +/- 150 mW.m(-2) caused mortality and growth retardation. The highest fluence rate (3040 +/- 210 mW.m(-2)) caused 100% mortality in 5 days. All fluence rates caused beha…
Retene causes multifunctional transcriptomic changes in the heart of rainbow trout (Oncorhynchus mykiss) embryos
Fish are particularly sensitive to aryl hydrocarbon receptor (AhR)-mediated developmental toxicity. The molecular mechanisms behind these adverse effects have remained largely unresolved in salmonids, and for AhR-agonistic polycyclic aromatic hydrocarbons (PAHs). This study explored the cardiac transcriptome of rainbow trout (Oncorhynchus mykiss) eleuteroembryos exposed to retene, an AhR-agonistic PAH. The embryos were exposed to retene (nominal concentration 32 μg/L) and control, their hearts were collected before, at and after the onset of the visible signs of developmental toxicity, and transcriptomic changes were studied by microarray analysis. Retene up- or down-regulated 122 genes. Th…
Importance of ligand speciation in environmental research:A case study
The speciations of EDTA and DTPA in process, waste and river waters are modelled and simulated, specifically to the mode of occurrence in the pulp and paper mill effluents and subsequently in receiving waters. Due to relatively short residence times in bleaching process and waste water treatment and slow exchange kinetics, it is expected that the thermodynamic equilibrium is not necessarily reached. Therefore, the initial speciation plays a key role. As such, the simulations have been extended to the process waters of the pulp and paper industry taking into account estimated average conditions. The results reveal that the main species are; Mn and Ca complexes of EDTA and DTPA in pulp mill p…
Use of Early Juvenile Zebrafish Danio Rerio for In-Vivo Assessment of Endocrine Modulation by Xenoestrogens
Reliable and cost-effective early-life stage (ELS) bioassays incorporating practical experimentation without compromising scientific relevance are crucial in chemical risk assessment. This study investigated the use of 20 days- post-fertilization life stage ( 20dpfZF ) of zebrafish Danio rerio to screen environmental chemicals known to be estrogenic in adult fish. Firstly, studies with key genes in steroidogenesis were conducted; the brain isoform of aromatase gene ( cyp19a1b ) being the most prominently expressed biomarker. Regulation of mRNA levels of molecular biomarkers, vitellogenin 1 gene ( vtg1) and cyp19a1b were selected to assess the endocrine modulation by xenoestrogens, 17α-ethin…
Time- and concentration-dependent metabolic and genomic responses to exposure to resin acids in brown trout (Salmo trutta m. lacustris)
The presence of metabolically conjugated resin acids (RAs) in the bile is considered to be a sensitive indicator for exposure of fish to pulp and paper industry effluents; however, to our knowledge, no comprehensive kinetic study of this response has been made. Juvenile brown trout (Salmo trutta m. lacustris) were exposed to a waterborne mixture of seven RAs (wood rosin) in time (0.1-192.0 h; average concentration, 8 microg/L) and dose (average concentrations, 0, 0.6, 4, 14, and 78 microg/L; 10 d) series, and total RAs were analyzed in bile. In time-dependent exposure, total RAs in bile increased up to 24 h. In concentration-dependent exposure, RAs increased along with the concentration of …
Confirmation of in situ exposure of fish to secondary treated bleached-kraft mill effluent using a laboratory simulation
To corroborate the responses in whitefish (Coregonus Iavaretus L.) exposed to elemental chlorine free (ECF) bleached-kraft pulp mill effluent (BKME) in situ, a 30-d laboratory exposure was carried out at concentrations simulating the field conditions. The flow-through exposures were conducted at four secondary (activated sludge) treated effluent (STE) concentrations: 1.3, 2.3, 3.5, and 7%. To evaluate the role of the secondary treatment, fish were also exposed to one concentration (3.5%) of pretreated effluent (PTE) from the mill. Compared to the control, whitefish liver 7-ethoxyresorufin-O-deethylase (EROD) activity was twofold in fish exposed to 3.5% STE, which was similar to monooxygenas…
Photoinduced lethal and sublethal toxicity of retene, a polycyclic aromatic hydrocarbon derived from resin acid, to coregonid larvae
A comparative investigation on the acute phototoxicity of retene to vendace (Coregonus albula) and whitefish (C. lavaretus), both having pelagial larvae in spring, was conducted. To test the concept of early warning of sublethal biomarkers in relation to lethality to posthatch stages, we examined the effects of ultraviolet-B (UV-B) and retene on the levels of cytochrome P4501A (CYP1A) and heat shock protein 70 (HSP70) by exposing the animals to elevated levels of these factors for 48 and 72 h, respectively. Whereas UV-B and retene on their own were not lethal, simultaneous retene and UV-B exposure caused very high mortality to both species. The median lethal concentration (LC50; i.e., the c…
High sensitivity of northern pike larvae to UV-B but no UV-photoinduced toxicity of retene
In order to investigate whether increased UV-B radiation is a risk factor, a series of acute laboratory experiments was conducted with larval stages of the northern pike (Esox lucius L.), hatching in Nordic waters in May. Further, a comparative investigation on the acute phototoxicity of retene (7-isopropyl-1-methylphenanthrene), a PAH compound recently revealed to posses UV-B-induced phototoxicity in larval coregonids, was conducted with pike larvae. In semi-static experiment, larvae were pre-exposed to retene (3, 9, 30 and 82 microg/g), with relevant controls, for 24 h and then irradiated for 3 h once a day (two consecutive days) with three UV-B doses (CIE-weighted 1.0, 1.8 or 2.7 kJ/m2 p…
Occurrence of retene and resin acids in sediments and fish bile from a lake receiving pulp and paper mill effluents
Retene (7-isopropyl−1-methylphenanthrene) is a dialkyl-substituted PAH derived thermally and microbially from dehydroabietic acid. We have analyzed for retene and several resin acids in sediments at five depths at several sites in a lake receiving effluents from three pulp and paper mills, and two upstream reference sites. The highest concentration of retene was 1,600 μg/g dry weight (11,700 μg/g organic carbon [OC]) and of total resin acids was 1,500 μg/g dry weight (9,300 μg/g OC). Twelve kilometers downstream from the point of bleached-kraft mill effluent (BKME) discharge, the concentration of retene at a depth of 5 to 10 cm was 16 μg/g dry weight (650 μg/g OC) and of resin acids was 139…
Transcriptome responses to carbon tetrachloride and pyrene in the kidney and liver of juvenile rainbow trout (Oncorhynchus mykiss)
Abstract We report the effects of the hepatotoxic compound carbon tetrachloride (CCl 4 ) and pyrene, a model polycyclic aromatic hydrocarbon, on the transcriptomes of juvenile rainbow trout kidneys and livers. Fish were exposed to sublethal doses for 4 days and expression of 1273 genes was measured using a cDNA microarray. Efforts were focused on differentiating between unspecific responses and those that can be regarded as molecular signatures of CCl 4 and pyrene toxicities. Expression profiles were analyzed in terms of Gene Ontology categories. Universal reactions to chemical toxicity were observed in metallothionein, HSP90 and mitochondrial proteins of oxidative phosphorylation, which we…
How to preserve and handle fish liver samples to conserve RNA integrity
As transcriptomic studies are becoming more and more common, it is important to ensure that the RNA used in the analyses is of good quality. The RNA integrity may be compromised by storage temperature or freeze-thaw cycles, but these have not been well studied in poikilothermic fishes. This work studied the effects of tissue storage time and temperature, and freeze-thaw cycles of tissue and extracted RNA on RNA integrity in brown trout (Salmo trutta L.) liver. The storage time and temperature had an effect on RNA integrity, but RNA suitable for quantitative reverse transcription PCR (RT-qPCR) (RIN > 7) was still obtained from samples preserved at − 20 °C for 6 months. Freeze-thaw cycles of …
Hepatic responses of gene expression in juvenile brown trout (Salmo trutta lacustris) exposed to three model contaminants applied singly and in combination.
Chemical pollution of the aquatic environment is almost always the result of multiple rather than single toxic compounds. The possibility of separating the effects of key risk chemicals from those of others, including their joint effects, is of clear theoretical interest and high technical importance. We addressed this goal using multiple gene expression profiling in the liver of juvenile brown trout (Salmo trutta lacustris) exposed to three model chemicals (cadmium, carbon tetrachloride [CCl4], and pyrene) administered singly, in binary and trinary combinations at low acutely sublethal concentrations, and in the partial dose–response manner. Differentially expressed genes were grouped by c…
Different sensitivities of whitefish (Coregonus lavaretus) and northern pike (Esox lucius) eleutheroembryos to photoinduced toxicity of polycyclic aromatic hydrocarbons.
The photoinduced toxicity of two polycyclic aromatic hydrocarbons (PAHs), retene (RET) and pyrene (PYR), to the eleutheroembryos of whitefish and northern pike was studied. Fish were exposed to three concentrations of RET and PYR, and irradiated with ultraviolet radiation (UVR) or visible light for 3 h on two consecutive days. UVR covered the absorption maxima of RET and PYR at UVB and UVA, the daily UVR doses were 30 and 28 kJ m−2, respectively. After 72 h, mortality and behavioral abnormalities were observed. Survivors were analyzed for the proteins of CYP1A, CYP3A, Hsp70, and the activity of glutathione reductase (GR). While neither PAHs nor UVR on their own were lethal, simultaneous exp…
Aerobic composting and anaerobic digestion of pulp and paper mill sludges
The feasibility of aerobic vessel composting and anaerobic digestion for the treatment of pulp and paper mill sludges were studied. The composting studies made use of primary and secondary sludge from a de-inking and paper mill. In six parallely aerated 500 1 vessels with various carbon : nitrogen (C:N) -ratios, the most optimal performance was obtained with C:N -ratios of c. 22–35, while higher and lower ratios delayed the temperature increase. With the optimal ratios, the thermophilic stage was reached within 36 h, and the stage lasted for about seven days. In the scale-up study (18 m3 compost vessel), the thermophilic stage was reached within 24 h. An effective dehydratation of the mass …
Use of Early Life-Stages of Zebrafish to Assess Toxicity of Sediments Contaminated by Organotin Compounds
ABSTRACTThis study examined the response of early life-stages (ELS) of zebrafish to organotin-contaminated sediment from Lake Huruslahti (HL) in Central Finland. A dilution series (0, 10, 33, and 100%) of the native (HL) and the sediment spiked with tributyltin (TBT) determined a dose-response of zebrafish ELS to organotin-contaminated sediment. Sediment elutriates were assessed by bacterial bioluminescence assay and microscopical pathologies of 1–3 days post-fertilization zebrafish (1–3dpfZF). Brain aromatase (cyp19a1b) and tissue vitellogenin (vtg1) were assayed from early-juvenile zebrafish (20dpfZF) exposed to intact sediment. In vivo modulation of cyp19a1b and vtg1 transcripts in 20dpf…
Assessment of the bioactivity of creosote-contaminated sediment by liver biotransformation system of rainbow trout.
A sediment site in the Lake Jamsanvesi (municipality of Petajavesi, Finland) contaminated by creosote was investigated to assess the possible ecotoxicological risks it may cause to benthic animals, including ones which may arise due to physical measures in remediating the site. It is suggested that polycyclic aromatic hydrocarbons (PAHs) are bioavailable to fish and other aquatic animals during exposure to contaminated water, sediment, and food. In order to assess toxicological risks of sediment contents to fish, juvenile rainbow trout (Onchorhynchus mykiss) were intraperitoneally dosed with extracts of the creosote-contaminated sediments and their elutriates. This was compared to pristine …
BIOAVAILABILITY TO JUVENILE RAINBOW TROUT (ONCORYNCHUS MYKISS) OF RETENE AND OTHER MIXED-FUNCTION OXYGENASE-ACTIVE COMPOUNDS FROM SEDIMENTS
Retene (7-isopropyl-1-methylphenanthrene) is a naturally formed polycyclic aromatic hydrocarbon (PAH) that causes teratogenicity in fish larvae and induction of cytochrome P450 (CYP1A) enzymes. Retene occurs at high concentrations (or =3,300 microg/g dry wt) in surface sediments contaminated by resin acids from pulp mill effluents. To assess the environmental risks of retene, it is important to evaluate conditions affecting its bioavailability and accumulation by fish. Fingerling rainbow trout were exposed to retene-spiked or naturally contaminated sediments and sampled after 4 d to determine liver CYP1A activity and concentrations of retene metabolites in bile as indicators of retene accum…
Biomarker responses of the earthworm Aporrectodea tuberculata to copper and zinc exposure: differences between populations with and without earlier metal exposure.
Biomarkers in the earthworm Aporrectodea tuberculata (Eisen) were measured to find out their possible induction under Cu and Zn exposure and differences in the responses between two populations with different exposure history. The biomarkers applied were concentration of metallothioneins (MT), and cytochrome P4501A (CYP1A) monooxygenase and glutathione-S-transferase (GST) activities. These were measured from earthworms sampled at three distances from a steel smelter in Finland and from the individuals from two populations, one with and another without earlier metal exposure, exposed to three combined Cu/Zn concentrations in the laboratory. In the field, MT concentration, and cytochrome CYP1…
Phytotoxicity of low-weight carboxylic acids.
Abstract Presence of low-weight carboxylic acids (LWCAs) can be the reason for phytotoxicity of green manures, treated bio-waste or digestates from biogas production applied to soils. As the phytotoxic concentrations of LWCA are poorly known, this work presents data on six acids (C 1 C 6 : formic, acetic, propionic, butyric, valeric, and caproic). Phytotoxicity was measured in acute (72 or 120 h) and subchronic (21 d) assays for seed germination, seedling elongation, and plant growth for garden cress Lepidium sativum and ryegrass Lolium multiflorum . The dose–response relationship was modeled using Weibull model. Results showed a trend that toxicity of LWCA increases with the length of the …
Uptake of organic xenobiotics by benthic invertebrates from sediment contaminated by the pulp and paper industry
Abstract Uptake of pulp and paper mill-derived pollutants by benthic invertebrates from sediment in Southern Lake Saimaa, eastern Finland, was studied. Two groups of benthic invertebrates (Diptera and Oligochaeta) were analyzed for their concentrations of resin acids (RAs), chlorophenolics (CPs) and β -sitosterol. The samples were collected 1 and 3 km downstream from the mill. In laboratory experiments Chironomus plumosus (a dipteran) and Lumbriculus variegatus (oligochaete) were exposed for 14 d to sediments collected from the same locations. The concentrations of RAs, CPs and β -sitosterol were higher in the areas downstream from the mill than those in the upstream reference area in both …
Induction of erod activity in HEPA-1 mouse hepatoma cells and estrogenicity in mcf-7 human breast cancer cells by extracts of pulp mill effluents, sludge, and sediment exposed to effluents
Extracts of effluents and sludges from the primary and secondary clarifiers of an activated sludge treatment plant at a Finnish bleached kraft pulp and paper mill were analyzed in two cell bioassays. Total dioxin-like activities were determined by measuring the induction of ethoxyresorufin-O-deethylase (EROD) activity in Hepa-1 mouse hepatoma cells. Estrogenicity was studied by measuring luciferase activity in MCF-7 ERE-luc, which are MCF-7 human breast cancer cells stably transfected with an estrogen-responsive element linked to a luciferase promoter. Sediments collected near the pulp mill and from other sites in Lake Saimaa as well as fillets of whitefish exposed to effluents were examine…
Reproductive, biochemical, physiological, and population responses in perch (Perca fluviatilisL.) and roach (Rutilus rutilusL.) downstream of two elemental chlorine-free pulp and paper mills
Perch (Perca fluviatilis L.) and roach (Rutilus rutilus L.) populations in a state of late vitellogenesis were studied downstream of two pulp and paper mills and at upstream references in southern Lake Saimaa, Finland. The mills used elemental chlorine-free bleaching and activated sludge effluent treatment technologies. The exposure of fish to pulp mill effluents, as measured by concentrations of chlorophenolics in the bile and liver ethoxyresorufin-O-deethylase (EROD) activity, was low and almost similar to the references. Nevertheless, bile resin acid concentrations in exposed perch and roach (260-320 micrograms/ml) and bile beta-sitosterol concentrations in exposed roach (1.5-3.5 microgr…
Wood-derived estrogens: studies in vitro with breast cancer cell lines and in vivo in trout.
The wood-derived compound, beta-sitosterol (purity > 90%), was shown to be estrogenic in fish. It induced the expression of the vitellogenin gene in the liver of juvenile and methyltestosterone-treated rainbow trout. Structural similarities to beta-sitosterol notwithstanding, cholesterol, citrostadienol, beta-sitostanol, and 5-androstene-3 beta,17 beta-diol, an estrogenic member of the androstenic steroid group, were inactive. An abietic acid mixture (37% abietic acid, 6% dehydroabietic acid, and a remainder of unknown compounds) showed slight hormonal activity in feed, but it was completely inactive when given intraperitoneally in implants. The estrogenic component of the abietic acid prep…
Assessment of bioavailability and effects of chemicals due to remediation actions with caging mussels (Anodonta anatina) at a creosote-contaminated lake sediment site
A study was conducted at Lake Jämsänvesi in Central Finland, to identify the potential ecotoxicological risks of the remediation operation of a creosote-/PAH-contaminated lake sediment, made by capping during the years 1998-1999. Mussels (Anodonta anatina) were deployed to the lake at the same time as the remediation operation was started in November 1998. The contaminated area (0.5 ha) was covered by a filter geotextile (polypropylene), gravel and sand (1-1.5m) which were spread out on the ice and let to sink onto the bottom of the lake when the ice melted in May 1999. The possible impacts of capping to the adjacent environment were assessed from mussels exposed and particulate material se…
More accuracy to the EROD measurements—The resorufin fluorescence differs between species and individuals
Ethoxyresorufin-O-deethylase (EROD) activity is a biomarker of exposure to planar aromatic hydrocarbons, and it is often measured from the S9 fraction. The effect of the liver S9 fraction of seven boreal freshwater fish species on the fluorescence of resorufin was studied. The S9 fractions diminished resorufin fluorescence by 40–80%, and there were large differences between species. Thus, using a resorufin standard curve without the S9 fraction leads to a large underestimation of the EROD activity. Therefore a microwell plate EROD method was developed that takes into account the effect of each sample on resorufin fluorescence. At least two mechanisms were involved in the decrease of the flu…
Photoinduced toxicity of retene to Daphnia magna under enhanced UV-B radiation.
Abstract The effects of UV radiation on the acute toxicity of retene (7-isopropyl-1-methylphenanthrene) to Daphnia magna Straus were studied. Dehydroabietic acid (DHAA) from which retene is formed in the vicinity of pulp and paper industry was also studied. Pyrene, anthracene, and phenanthrene were used as model PAH compounds. The time taken for immobilization (ET50) was monitored under biologically effective UV-B dose rates of 240, 365, 565, and 650 mW m−2 (UV-A and visible light also present). Median effective concentrations (EC50) were determined after a 15-min UV exposure (565 mW m−2) followed by 24 h in the dark. Retene ( 10–320 μg l −1 ) was not acutely toxic in the dark. The inductio…
The occurrence and bioavailability of retene and resin acids in sediments of a lake receiving BKME (bleached kraft mill effluent)
Retene (7-isopropyl-1-methylphenantrene) is an alkyl substituted PAH derived primarily from bacterial aromatization of abietic type resin acids. Retene has been shown to induce cytochrome P450 1A in rainbow trout whereas e.g. dehydroabietic acid does not induce it. We analysed resin acids and retene in sediments from seven sites receiving pulp and paper mill effluents from 4 mills, and from two reference sites. All mills have employed treatment of waste waters by activated sludge. The highest concentration of retene measured in sediment was 1600 μg/g d.w. (11 700 μg/g organic carbon, OC) while the highest concentration of resin acids was 1500 μg/g d.w. (9 300 μg/g OC). Downstream from the p…
Uptake from water, biotransformation, and biliary excretion of pharmaceuticals by rainbow trout.
An urgent need exists to assess the exposure of fish to pharmaceuticals. The aim of the present study was to assess the uptake and metabolism of waterborne pharmaceuticals in rainbow trout (Oncorhynchus mykiss). A further objective was to determine the possibility of monitoring exposure to low levels of pharmaceuticals by bile assays. Rainbow trout were exposed for 10 d under flow-through conditions to mixtures of five pharmaceuticals (diclofenac, naproxen, ibuprofen, bisoprolol, and carbamazepine) at high and low concentrations. The low concentration was used to mimic the conditions prevailing in the vicinity of the discharge points of wastewater treatment plants. The uptake and the biocon…
Pentachlorophenol accumulation in the freshwater mussels Anodonta anatina and Pseudanodonta complanata, and some physiological consequences of laboratory maintenance
Freshwater mussels Anodanta anatina and Pseudanodonta complanata were exposed to (14C)-pentachlorophenol. The wet weight based bioconcentration factor (BCF = activity in animal per activity in water) at steady state varied from 80 to 120 for A. anatina and from 61 to 85 for P. complanata. The species did not differ significantly in their wet weight or lipid based BCFs but dry weight based values were significantly higher (40-50%) for A. anatina. The soft tissue dry weight and dry weight based condition index of A. anatina (Cl4 = soft tissue dry weight per shell length) differed significantly between natural mussel populations. In animals kept from 4 to 8 months in laboratory conditions, the…
The anti-inflammatory drugs diclofenac, naproxen and ibuprofen are found in the bile of wild fish caught downstream of a wastewater treatment plant.
Pharmaceutical residues are ubiquitous in rivers, lakes, and at coastal waters affected by discharges from municipal wastewater treatment plants. In this study, the presence of 17 different pharmaceuticals and six different phase I metabolites was determined in the bile of two wild fish species, bream (Abramis brama) and roach (Rutilus rutilus). The fish were caught from a lake that receives treated municipal wastewater via a small river. Prior to analyses, the bile content was enzymatically hydrolyzed to convert the glucuronide metabolites into the original pharmaceuticals or phase I metabolites. The solid phase extracts of hydrolyzates were analyzed by liquid chromatography-tandem mass sp…
Caging techniques for field exposures of fish to chemical contaminants
The article reviews current state-of-the-art to conduct fish caging as an assessment and monitoring technique in aquatic toxicology. A multitude of scientific, practical and management factors influence and may restrict how field research is carried out. For many purposes the advantages of transplant fish caging outweigh the alternative methodologies of impact studies of xenobiotics. However, besides mortality, virtually no study has evaluated the physiological consequences of caging fish. It is not known how caging itself affects the responses of fishes to chemical pollution. The used caging densities are commonly too high. However, with the aid of variables describing physiological stress…
UV-B exposure causes DNA damage and changes in protein expression in northern pike (Esox lucius) posthatched embryos.
The ongoing anthropogenically caused ozone depletion and climate change has increased the amount of biologically harmful UV-B radiation, which is detrimental to fish in embryonal stages. The effects of UV-B radiation on the levels and locations of DNA damage manifested as cyclobutane pyrimidine dimers (CPDs), heat shock protein 70 (HSP70) and p53 protein in newly hatched embryos of pike were examined. Pike larvae were exposed in the laboratory to current and enhanced doses of UV-B radiation. UV-B exposure caused the formation of CPDs in a fluence rate-dependent manner, and the CPDs were found deeper in the tissues with increasing fluence rates. UV-B radiation induced HSP70 in epidermis, and…
PCA and PLS methods applied to ecotoxicological data: Ecobalance project
Within a case study ‘Ecobalance’, the fate and effects of various chlorinated and non-chlorinated organic compounds and some heavy metals discharged from pulp and paper mills into water, sediment and aquatic animals were studied in a recipient area of southern Lake Saimaa, SE Finland. The main aim of the project was to find an empirical link between chemical emission parameters and ecotoxicological effects expressed in the ecosystem. As part of the \‘Ecobalance’ project, principal component analysis (PCA) and partial least squares (PLS) methods were used to interpret the data of the lake area. The PLS method was used to estimate the lake area affected by mill effluents and to calculate the …
Do laboratory exposures represent field exposures? Effects of sediments contaminated by wood industry on yolk-sac fry of rainbow trout (Oncorhynchus mykiss)
Purpose. Risk assessment of contaminated sediments is routinely based on laboratory exposures. The purpose of this work was to study if sediments contaminated by the chemical wood industry cause developmental defects in fish fry and how well a laboratory exposure correlates with a field exposure. Materials and methods. Newly hatched yolk-sac fry of rainbow trout (Oncorhynchus mykiss) were exposed in the laboratory and in situ. In the laboratory, the fish were placed in contact with either clean or contaminated sediment in aquaria. In the field, half of the fish were placed in contact with the lake sediment and the other half were similarly caged 2 m above it, to discern the effects of the s…
Bioaccumulation of PAHs from creosote-contaminated sediment in a laboratory-exposed freshwater oligochaete, Lumbriculus variegatus
The oligochaete, Lumbriculus variegatus, was used for a bioaccumulation assay in the creosote-contaminated sediment of Lake Jämsänvesi in a 28-day experiment. The PAH concentrations of the whole body tissue of worms, sediments and water samples were determinated by GC-MS. Chemical analyses showed that benzo(k)fluoranthene, anthracene and fluorene were the main PAH compounds present in the tissue of oligochaetes, just as in the sediment. The biota-sediment accumulation factors (BSAFs) of the individual PAHs varied from 1.2 to 5.7. It is concluded that oligochaetes have a marked ability to accumulate and retain PAHs from creosote-contaminated sediment.
Do historical sediments of pulp and paper industry contribute to the exposure of fish caged in receiving waters?
Field experiments were conducted in early summer 2004 by caging brown trout (Salmo trutta m. lacustris) in Southern Lake Saimaa (SE Finland) for 10 days. Cages were positioned into two depths at four distances (5 to 17 km) from the pulp and paper mill; one set with direct contact to the sediment and other locating at the three meters' depth from the water surface. Resin acids were measured from sediment, water, and fish bile. Possible toxicities were investigated by measuring liver EROD-activity, HSP70-protein, and gene expression profiles by microarray. The concentration of resin acids in the sediment showed clear gradient along the distance from the mill, and their concentrations in water…
Fluence rate or cumulative dose? : Vulnerability of larval northern pike (Esox lucius) to ultraviolet radiation
Newly hatched larvae of northern pike were exposed in the laboratory to four fluence rates of ultraviolet radiation (UVR; 290–400 nm) over three different time periods, resulting in total doses ranging from 3.0 ± 0.2 to 63.0 ± 4.4 kJ·m−2. Mortality and behavior of the larvae were followed for 8–12 days, and growth measured at the end of the experiment. Also, the principle of reciprocity—that the UVR-induced mortality depends on the cumulative dose, independent of fluence rate—was tested. Fluence rates higher than 1480 ± 150 mW·m−2 caused mortality and growth retardation. The highest fluence rate (3040 ± 210 mW·m−2) caused 100% mortality in 5 days. All fluence rates caused behavioral disorde…
Expression of the Vitellogenin Gene in the Liver of Juvenile Whitefish (Coregonus lavaretus L. s.l.) Exposed to Effluents from Pulp and Paper Mills
Juvenile whitefish (Coregonus lavaretus L. s.l.) were exposed by caging in the field to diluted effluents from three operating pulp, paper, and paperboard mills in Southern Lake Saimaa, Finland. The expression of the vitellogenin gene, used as a biomarker of estrogenic contamination of effluents, was measured using a Northern blotting method. Increased mRNA levels, the most specific and reliable evidence for estrogen receptor-mediated actions in vivo, were found in fish caged in the vicinity of one of three mills studied. This mill was found to discharge wood-derived compounds, such as sterols and resin acids, into Lake Saimaa in amounts considerably exceeding those from the other two mills…
Removal of toxic pollutants from pulp mill effluents by electrocoagulation
This study investigated the effect of electrocoagulation treatment on toxic pollutant removal from pulp mill effluents. Synthetic wastewaters containing wood rosin and copper or pure resin acids were used to investigate the removal of resin acids and copper by electrocoagulation. Removal of pollutants by electrocoagulation was also tested with real debarking effluent. In this study, statistical experimental design and partial least squares modeling were used to investigate the effect of initial pH, current (current density) and treatment time. Electrocoagulation and subsequent filtration removed resin acids and copper from the synthetic wastewaters with high efficiency. Toxicity to algae (P…
Histopathological responses of newly hatched larvae of whitefish (Coregonus lavaretus s.l.) to UV-B induced toxicity of retene
Positively phototactic fish larvae may be exposed to increased ultraviolet-B (UV-B) radiation alone or, potentially and in addition, to polycyclic aromatic hydrocarbons (PAHs) such as retene (7-isopropyl-1-methylphenanthrene) at the egg or larval stages. Suspended and sedimental particulate material near pulp and paper mills may act as sources of retene in chemically contaminated lake areas. In laboratory conditions whitefish larvae were pre-exposed to retene (10, 32 and 100 microg/l), with relevant controls, and irradiated in semi-static tests for 3 h once a day (2 consecutive days) with two UV-B doses (CIE-weighted 2.8 or 5.4 kJ per m(2) per day) or with visible light only. These UV-B dos…
The capacity of liver microsomes to form benzo[a]pyrene-diolepoxide-DNA adducts and induction of cytochrome P450 1A in feral fish exposed to pulp mill effluents.
An investigation was made of cytochrome P4501A (CYP1A) induction, determined by the activity of EROD (7-ethoxyresorufin O-deethylase), and formation of benzo[a]pyrene-diolepoxide-DNA (BPDE-DNA) adducts, measured by synchronous fluorescence spectrophotometry, in liver microsomes of perch (Perca fluviatilis), bream (Abramis brama), and roach (Rutilus rutilus). Fish were collected from the southern part of Lake Saimaa (Finland), an area polluted by effluents from the pulp and paper industry. In addition, two conjugation enzymes (UDP-glucuronosyltransferase and glutathione S-transferase) were determined. Overall, when compared to an upstream reference, EROD activity was higher in fish at waters…
Concentration of retene and resin acids in sedimenting particles collected from a bleached kraft mill effluent receiving lake
Abstract The concentration of retene, a di-alkyl substituted PAH derived microbially and thermally from dehydroabietic acid, eight resin acids and their derivatives were analysed from sedimenting particles collected from five lake sites receiving pulp and paper mill effluents and from two reference sites. At five of the sites, samples were collected prior and after a conventional aerated lagoon wastewater treatment system of the mill was modernized with an activated sludge plant, and elemental chlorine free bleaching (ECF) had replaced chlorine bleaching. The highest level of retene was 30.0–53.8 μ g/g dry weight (d.w.), observed 3 km downstream from the mill. Nine kilometers downstream fr…
Assessment of toxicity hazards of dredged lake sediment contaminated by creosote.
In order to predict the potential toxicity hazards of sediment remediation by dredging, an experimental laboratory simulation was made by investigating seven ratios of creosote-contaminated sediment (Lake Jamsanvesi, central Finland) and artificial lake water mixtures. Sediment was suspended in water at the ratios of 1:1, 1:2, 1:4, 1:8, 1:16, 1:32, 1:64, 1:128 v/v. The elutriates were analysed for the acute toxicity by photoluminescence bacterial and waterflea (Daphnia magna Straus) tests. The concentrations of polycyclic aromatic hydrocarbons (PAHs) are determined by gas chromatography (GC/FID). The elutriate of ratio 1:2 was most toxic to bacteria (EC50 = 4.5%), whereas the ratio 1:4 was …
Sediment-associated retene bioavailability of sediment-associated retene to an oligochaete wormlumbriculus variegatus
The bioavailability of retene spiked to two sediment concentrations (50 and 200 µg/g dry weight) was measured in long-term (28-day) bioaccumulation tests performed on the oligochaeteLumbriculus variegatus Muller. Three sediment organic carbon (SOC) concentrations (1.1, 3.0, 23.0% of dry weight) were used. Growth and reproduction, used as endpoints, indicated that retene was chronically nontoxic at the concentrations employed. Retene was clearly bioavailable, but bioaccumulated about 100 times less in the SOC-rich (23.0%) sediment than in the other two (SOC <3.0%). At the lower retene concentration, retene in the worms was undetectable in the high-SOC sediment. Based on final retene present …
Bioavailability of pharmaceuticals in waters close to wastewater treatment plants: Use of fish bile for exposure assessment
Pharmaceuticals are ubiquitous in surface waters as a consequence of discharges from municipal wastewater treatment plants. However, few studies have assessed the bioavailability of pharmaceuticals to fish in natural waters. In the present study, passive samplers and rainbow trout were experimentally deployed next to three municipal wastewater treatment plants in Finland to evaluate the degree of animal exposure. Pharmaceuticals from several therapeutic classes (in total 15) were analyzed by liquid chromatography-tandem mass spectrometry in extracts of passive samplers and in bile and blood plasma of rainbow trout held at polluted sites for 10 d. Each approach indicated the highest exposure…
Agricultural potential of anaerobically digested industrial orange waste with and without aerobic posttreatment
The potential of anaerobically digested orange waste with (AAD) and without (AD) aerobic post-treatment for use in agriculture was evaluated through chemical analyses, short-term phytotoxicity and long-term plant assays. Chemical analyses showed that AD contained ammonia and organic acids, and aerobic post-treatment did not significantly remove these phytotoxins. The N:P2O5:K2O ratio in AD was 1:0.26:0.96 and aerobic post-treatment did not change the composition in AAD except for K2O (1:0.26:1.24). Heavy metal contents in AD and AAD were more or less the same and were below the upper limit recommended for non-sewage sludge application on agricultural soils. Short-term phytotoxicity tests sh…
Caddis larvae (Trichoptera, Hydropsychidae) indicate delaying recovery of a watercourse polluted by pulp and paper industry
Abstract We studied ecotoxicological responses of aquatic insects to evaluate the recovery of a watercourse with a history of over 100 years as a recipient of pulp and paper mill effluents. The recovery of the water quality began in 1985 due to the improved technology, but ecotoxicological effects on biota have still been evident two decades later, as indicated by morphological abnormalities in tracheal gills of caddis larvae. We hypothesized that these abnormalities are related to organic contaminants, such as resin acids, alkylated PAHs and chlorinated organic compounds, released historically from the mills, and more recently via resuspension from the sediment. Samples were collected betw…
Biomarker responses along a pollution gradient: Effects of pulp and paper mill effluents on caged whitefish
Abstract Lake Saimaa (SE Finland) is a large oligotrophic lake receiving biologically treated effluent from a bleached kraft pulp and paper mill. Hatchery-reared juvenile whitefish ( Coregonus lavaretus L. s.l.), a species known to feed on plankton and other particulates, were caged using a technique developed and optimized for this species and were exposed for one month in five downstream stations (3.3–16 km). Two reference stations (4.5–8.5 km) upstream from the effluent outlet were used. An exposure gradient of chlorinated organics was assessed by determining concentrations of conjugated chlorophenolics (CPs) in bile and CPs as well as extractable organic halogens (EOX) in gut lipids. Ac…
Response of rainbow trout transcriptome to model chemical contaminants.
We used high-density cDNA microarray in studies of responses of rainbow trout fry at sublethal ranges of beta-naphthoflavone, cadmium, carbon tetrachloride, and pyrene. The differentially expressed genes were grouped by the functional categories of Gene Ontology. Significantly different response to the studied compounds was shown by a number of classes, such as cell cycle, apoptosis, signal transduction, oxidative stress, subcellular and extracellular structures, protein biosynthesis, and modification. Cluster analysis separated responses to the contaminants at low and medium doses, whereas at high levels the adaptive reactions were masked with general unspecific response to toxicity. We fo…
Altered Challenge Response in Whitefish Subchronically Exposed in Areas Polluted by Bleached Kraft Mill Effluents
Exposure to impaired water quality, as in bleached kraft mill effluents (BKMEs), has recently been demonstrated to impair the ability of fish to elicit an acute stress response. Acute stress caused by catching is, in most field studies, an unavoidable incident that may markedly affect physiological functions. Consistently, dissimilar stress responses to catching procedures in exposed and reference animals may lead to altered results. In this study, juvenile whitefish (Coregonus levaretus) were caged for 30 days in two reference areas and three areas affected by different BKMEs, and the immediate effects of low-level handling on physiological functions in four periods within an hour of the s…
Toxicity of waste gasification bottom ash leachate
Abstract Toxicity of waste gasification bottom ash leachate from landfill lysimeters (112 m3) was studied over three years. The leachate of grate incineration bottom ash from a parallel setup was used as reference material. Three aquatic organisms (bioluminescent bacteria, green algae and water flea) were used to study acute toxicity. In addition, an ethoxyresorufin-O-deethylase (EROD) assay was performed with mouse hepatoma cells to indicate the presence of organic contaminants. Concentrations of 14 elements and 15 PAH compounds were determined to characterise leachate. Gasification ash leachate had a high pH (9.2–12.4) and assays with and without pH adjustment to neutral were used. Gasifi…
Recovery of Historically Contaminated Watercourse Polluted by the Chemical Wood Industry: EROD Activity in Fish as Biomarker
Despite outstanding process alterations over decades, pulp- and paper-mill-contaminated sediments and continuing exposure by the effluents may still have effects on biota. In this study, ecotoxicological impacts in the boreal watercourse were analyzed by measuring ethoxyresorufin-O-deethylase (EROD) induction from wild fish populations and from experimentally exposed fish. In order to assess the role of sediment-borne chemicals, juvenile rainbow trout (Oncorhynchus mykiss) were exposed in the laboratory to the surface sediments of Lake Vatianjarvi and Southern Lake Saimaa, both watercourses impacted by the chemical wood industry for approximately a century. Hepatic EROD activity was also me…
Dissolution of resin acids, retene and wood sterols from contaminated lake sediments.
The dissolution potency of hydrophobic resin acids (RAs), retene and wood sterols from sediments was studied. These wood extractives and their metabolites are sorbed from pulp and paper mill effluents to downstream sediments. With harmful components like these, sediments can pose a hazard to the aquatic environment. Therefore, sediment elutriates with water were produced under variable conditions (agitation rate and efficiency, time), and concentrations of the dissoluted compounds were analyzed. Both naturally contaminated field sediments and artificially spiked sediments were studied. By vigorous agitation RAs can be released fast from the sediment matrix and equilibrium reached within 3 d…
Identification of lipophilic pollutants discharged from a Finnish pulp and paper mill.
Lipophilic organic compounds originating from kraft pulping and papermaking were identified by straight gas chromatography/mass spectrometry (GC/MS) analyses. Samples analyzed included a primary clarifier effluent (PE) and a secondary clarifier effluent (SE) from the activated sludge treatment plant of a Finnish ECF-bleached (Elementally Chlorine Free) kraft pulp and paper mill. Liquid-liquid extraction composition of PE was compared with solid phase extraction (SPE) and dialysis of a semipermeable membrane device (SPMD) exposed to PE. Dichloromethane extracts of particulate material isolated from PE, sludges from both the primary and secondary clarifier, a sediment collected in the vicinit…
Use of Early Life-Stages of Zebrafish to Assess Toxicity of Sediments Contaminated by Organotin Compounds
This study examined the response of early life-stages (ELS) of zebrafish to organotin-contaminated sediment from Lake Huruslahti (HL) in Central Finland. A dilution series (0, 10, 33, and 100%) of the native (HL) and the sediment spiked with tributyltin (TBT) determined a dose-response of zebrafish ELS to organotin-contaminated sediment. Sediment elutriates were assessed by bacterial bioluminescence assay and microscopical pathologies of 1–3 days post-fertilization zebrafish (1–3dpfZF). Brain aromatase (cyp19a1b) and tissue vitellogenin (vtg1) were assayed from early-juvenile zebrafish (20dpfZF) exposed to intact sediment. In vivo modulation of cyp19a1b and vtg1 transcripts in 20dpfZF was t…