Total Nitrogen Determination by a Spectrophotometric Method.

Being able to measure total nitrogen (TN) is important for following the nitrogen budget. In this chapter, we present the spectrophotometric method we use for determining TN. The method relies on oxidation and reduction steps, involving persulfate digestion of nitrogen compounds into nitrate followed by spectrophotometric determination.

Determining Inorganic and Organic Nitrogen.

Nitrogen (N) is one of the key nutrients for algal growth and is an integral part of many cellular components, for example in proteins. Being able to determine the inorganic and organic pools of N is consequently critical for algal cultivation. In this chapter we present the methods we use for determining dissolved inorganic nitrogen (DIN), dissolved organic nitrogen (DON), and particulate organic nitrogen (PON). The methods presented here for DIN rely on colorimetric methods and those of DON and PON on filtration and high temperature catalytic oxidation.

Determining Dissolved and Biogenic Silica.

Most algae do not use silicon in any form with one notable exception, diatoms. Silicon is a major constituent of diatoms. Diatoms are characterized by high growth rates and are often one of the key groups in forming algal blooms in natural waters, and as such it is an interesting group for cultivation. In this chapter we present methods for determining dissolved silica (DSi) and biogenic silica (BSi), oxide forms of silicon, based on colorimetric methods. BSi is determined after filtration and alkaline digestion.

Determining Inorganic and Organic Phosphorus.

Phosphorus (P) is a macronutrient for all microalgal species, and the main form of uptake is orthophosphate (PO4). In this chapter we present a colorimetric method for determining the PO4 concentration and dissolved organic phosphorus (DOP) based on total phosphorus (TP) measurements. We also describe a method for determining particulate organic phosphorus (POP) based on the same principles.

Determining Inorganic and Organic Carbon.

Carbon is the element which makes up the major fraction of lipids and carbohydrates, which could be used for making biofuel. It is therefore important to provide enough carbon and also follow the flow into particulate organic carbon and potential loss to dissolved organic forms of carbon. Here we present methods for determining dissolved inorganic carbon, dissolved organic carbon, and particulate organic carbon.

Correction to: Determining Inorganic and Organic Phosphorus.

There are three corrections for this chapter. On page 90, 12th line from top should read as '(see step 6) with ultrapure water (e.g., 0.1 ml' instead of '(see step 6) with ultrapure water (e.g., 1 ml'.

Correction to: Determining Inorganic and Organic Nitrogen.

There are three corrections for this chapter. On page 74, 10th line from the bottom in Sect. 3.1, point 3 appears incorrect as '85 ml ultrapure water and 8.5 ml' instead of '85 ml ultrapure water and 8.4 ml'.

Organohalogen pollutants in herring from the northern Baltic Sea: concentrations, congener profiles and explanatory factors.

Organohalogen contaminants were investigated in Baltic herring caught from three catchment areas in the Baltic Sea, off the coasts of Finland. Pools of both small and large herring were analysed for polychlorinated biphenyls (PCB), dibenzo-p-dioxins, dibenzofurans, naphthalenes, camphenes (toxaphene), polybrominated diphenyl ethers and the pesticide DDT and its metabolites. PCB concentrations per fresh weight in small herring were at the same level in all catchment areas, i.e. the Bothnian Bay, the Bothnian Sea and the Gulf of Finland, revealing no hot spots and reflecting most likely long term emissions and atmospheric deposition. Differences in the levels and/or congener profiles of other contaminants between catchment areas may be explained by point sources. Similar concentrations in small and large herring in the Gulf of Finland were possibly due to their common nutrition. In the other areas, differences between small and large herring most likely reflected their different food sources.

Desorption of sediment-associated polychlorinated dibenzo-p-dioxins, dibenzofurans, diphenyl ethers and hydroxydiphenyl ethers from contaminated sediment.

The transport and bioavailability of sediment-associated contaminants are often controlled by the contaminants' desorbing behaviour. This study examines the desorption kinetics of polychlorinated dibenzo-p-dioxins (PCDDs), dibenzofurans (PCDFs), diphenyl ethers (PCDEs) and hydroxydiphenyl ethers (HO-PCDEs) from the highly contaminated River Kymijoki sediment in Finland. The desorption kinetics data were generated using Tenax((R)) extraction, and a first-order three-compartment kinetic model was fitted to the data. The desorption data was compared to the previously published accumulation data from this same location to investigate the relationship between the rapidly desorbing fraction (F(r)) and biota-sediment accumulation factors (BSAFs) as well as semipermeable membrane device sediment accumulation factors (SSAFs). The PCDDs, PCDFs, PCDEs and HO-PCDEs were tightly attached to sediment particles and formed a large very slowly desorbing fraction (F(vs)). Rapidly desorbing fractions (F(r)) varied between 0.8% and 8% of total amount in sediment. The size of the desorbing fraction was congener-specific and F(r) decreased with the increasing lipophilicity of congeners. The size of the F(r) was unable to explain the small variation in the BSAFs of Lumbriculus variegatus but may help to explain the observed variation in the SSAFs. To our best knowledge, this study is the first effort to investigate the desorption of PCDDs, PCDFs, PCDEs and HO-PCDEs in field-contaminated sediments. The major finding that the very slow desorption of these chemicals will continue years, provides essential information for the modern risk assessment process.

Bone-related effects of contaminants in seals may be associated with vitamin D and thyroid hormones.

The high levels of polychlorinated biphenyls (PCBs) and DDT in gray seal (Halichoerus grypus) and ringed seal (Phoca hispida botnica) in the Baltic Sea have been associated with pathological disruptions, including bone lesions and reproductive failures. The underlying environmental and toxicological mechanisms leading to these pathological changes are not yet fully understood. The present study investigated the relationship between the individual contaminant load and bone- and thyroid-related effects in adult gray seals (n=30) and ringed seals (n=46) in the highly contaminated Baltic Sea and in reference areas (Sable Island, Canada, and Svalbard, Norway). In the gray seals, multivariate and correlation analyses revealed a clear relationship between circulating 1,25-dihydroxyvitamin D(3) (1,25(OH)(2)D), calcium, phosphate, and thyroid hormone (TH) levels and hepatic PCB and DDT load, which suggests contaminant-mediated disruption of the bone and thyroid homeostasis. Contaminants may depress 1,25(OH)(2)D levels or lead to hyperthyroidism, which may cause bone resorption. In the ringed seals, associations between circulating 1,25(OH)(2)D, THs, and hepatic contaminants were less prominent. These results suggest that bone lesions observed in the Baltic gray seals may be associated with contaminant-mediated vitamin D and thyroid disruption.

Bioaccumulation, bioavailability and environmental fate of chlorophenol impurities, polychlorinated hydroxydiphenylethers and their methoxy analogues.

The bioaccumulation potential and environmental fate of polychlorinated hydroxydiphenyl ethers (HO-PCDEs; polychlorinated phenoxyphenols, PCPP), the major impurities of chlorophenol formulations and their methoxy analogues (MeO-PCDEs; polychlorinated methoxyanisoles, PCPAs) were investigated. Oligochaete worms (Lumbriculus variegatus) exposed to sediment spiked with a model substance of one HO-hexaCDE (4'-HO-PCDE 161) or its methoxy analogue (4'-MeO-PCDE 161) clearly accumulated the test compounds revealing the potential for environmental risk of HO-PCDEs and MeO-PCDEs. The HO-PCDE tested has earlier been reported as an abundant component in a Finnish chlorophenol formulation (Ky-5) and its methoxy analogue is recognized as an abundant MeO-PCDE in sawmill soil contaminated by the formulation. The occurrence of 4'-HO-PCDE 161 and its methoxy analogue among other HO-PCDEs and MeO-PCDEs in lake mussels (Anodonta piscinalis) incubated in a river contaminated via the manufacture of Ky-5 showed that these compounds are bioavailable and transported in the aquatic environment. Mussel comparison with sediment data pointed to a higher accumulation potential for MeO-PCDEs than for HO-PCDEs. The finding of HO-PCDEs in groundwater samples collected from a groundwater reservoir, which had been contaminated by chlorophenols, points to potential of HO-PCDEs for transport with water in soil.

Dioxin concentrations in sediments of the Baltic Sea--a survey of existing data.

Recent survey results for polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/Fs, dioxins) in Baltic Sea sediments from Finland, Sweden and Denmark were merged with previously published Baltic Sea data. Regional distribution of concentration levels, differences in congener patterns, and temporal changes in sediment profiles were examined. One of the main objectives was to study if any major point sources for different PCDD/F congeners could be identified on a regional scale, based on sediment records. The survey confirmed the impact of chlorophenol production derived highly chlorinated PCDF-congeners on the total toxicity in sediments in the Gulf of Finland near the Kymijoki river estuary. Signatures of other point sources or combined point sources pertinent to specific industry branches or particular production processes (such as pulp bleaching, vinyl chloride production, thermal processes) may be discerned. However, the findings did not support any of the known point sources significantly influencing those congeners that are most abundant in Baltic herring and salmon. Instead, regional distributions in the Baltic Sea indicate that atmospheric deposition may act as a major source for those congeners and especially for 2,3,4,7,8-PeCDF. There were clear indications of declines in levels in sediment in some areas, but generally the levels of highly chlorinated PCDD/Fs on the northern coast of the Gulf of Finland were still high when compared with other areas of the Baltic Sea. Major areas with data gaps cover the south-eastern and eastern coastal regions of the Baltic Proper and the southern Gulf of Finland.

Polychlorinated dibenzo-p-dioxins, dibenzofurans, biphenyls, naphthalenes and polybrominated diphenyl ethers in the edible fish caught from the Baltic Sea and lakes in Finland.

A total of 156 fish composite samples were collected from five areas of the Baltic Sea and from three lakes and analysed for polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/Fs), polychlorinated biphenyls (PCBs), polychlorinated naphthalenes (PCNs) and polybrominated diphenyl ethers (PBDEs). The European Union's maximum permissible level for PCDD/Fs, 4 pg WHO-TEQ/g fresh weight (fw), was exceeded in salmon, river lamprey and Baltic herring. In other species from the Baltic Sea, the 90th percentile was 3.42 pg WHO(PCDD/F)-TEQ/g fw. In the lake fish, the concentrations of PCDD/Fs, PCBs and PCNs were only 29-46% of those in the same species caught from the Baltic Sea, whereas the concentrations of PBDEs in the lake fish were as high as in the Baltic Sea fish. Dioxin-like PCBs contributed to the total dioxin-like toxicity of PCBs and PCDD/Fs by 49+/-12% in all the analysed samples.

The dependence of organohalogen compound concentrations on herring age and size in the Bothnian Sea, northern Baltic.

The concentrations and composition profiles of polychlorinated dibenzo-p-dioxins and dibenzofurans, polychlorinated biphenyls, polybrominated diphenyl ethers and polychlorinated naphthalenes were determined in herring individuals collected from the commercial catches of the Bothnian Sea, northern Baltic. The median age of herring was 5.0 years and in the muscle the median toxic equivalent concentration of PCDD/Fs was 5.6 WHO(PCDD/F)-TEQ pg/g fresh weight (fw) and that of PCBs 2.7 WHO(PCB)-TEQ pg/g fw. The median sum concentration of PBDEs was 1.4 ng/g fw and that of PCNs 0.1 ng/g fw. Differences in age-dependent accumulation between the organohalogen groups and individual congeners were major. In the Bothnian Sea the content of organohalogen compounds in herring is obviously elevated due to the availability and large proportion of Mysis crustaceans in their diet. More intensive fishing could reduce the concentrations of organohalogens, including the abundant, accumulative and toxic 2,3,4,7,8-PeCDF congener in the Baltic herring catch.

Suitability of passive sampling for the monitoring of pharmaceuticals in Finnish surface waters.

The occurrence of five pharmaceuticals, consisting of four anti-inflammatory and one antiepileptic drug, was studied by passive sampling and grab sampling in northern Lake Päijänne and River Vantaa. The passive sampling was performed by using Chemcatcher® sampler with a SDB-RPS Empore disk as a receiving phase. In Lake Päijänne, the sampling was conducted during summer 2013 at four locations near the discharge point of a wastewater treatment plant and in the years 2013 and 2015 at four locations along River Vantaa. The samples were analyzed by liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) in the multiple reaction monitoring mode. The concentrations of carbamazepine, diclofenac, ibuprofen, ketoprofen, and naproxen in Lake Päijänne determined by passive sampling ranged between 1.4-2.9 ng L(-1), 15-35 ng L(-1), 13-31 ng L(-1), 16-27 ng L(-1), and 3.3-32 ng L(-1), respectively. Similarly, the results in River Vantaa ranged between 1.2-40 ng L(-1), 15-65 ng L(-1), 13-33 ng L(-1), 16-31 ng L(-1), and 3.3-6.4 ng L(-1). The results suggest that the Chemcatcher passive samplers are suitable for detecting pharmaceuticals in lake and river waters.

Effects of in utero and lactational TCDD exposure on bone development in differentially sensitive rat lines.

2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) is a notorious model compound of highly toxic environmental pollutants, polychlorinated dibenzo-p-dioxins (PCDDs). Their toxic effects are mediated via cytosolic aryl hydrocarbon receptor (AHR). We studied the effects of several dose levels of TCDD on developing rat bone after maternal exposure at different times of gestation and lactation in three differentially sensitive rat lines. Rat lines A, B, and C differ in their sensitivity to TCDD due to mutated AHR (Ahr(hw)) in line A and another TCDD-resistance allele (B(hw)) in line B. Line C rats have no resistance alleles. Offspring were analyzed for bone mineral density and geometry by peripheral quantitative computed tomography (pQCT) and for bone biomechanics by three-point bending at mid-diaphysis of tibia and femur and by axial loading at femoral neck. TCDD treatment resulted in bone defects, mainly in offspring of the most sensitive line C at a maternal dose of 1 microg/kg. They included decreased bone length, cross-sectional area of cortex, and bone mineral density. Mechanical testing revealed significantly reduced bending breaking force and stiffness of tibia, femur, and femoral neck. The effects were exposure time-dependent, and earlier exposure caused more severe defects. Gestational exposure alone was not sufficient, but lactational exposure was required to cause the bone defects. Most of the defects were recovered at the age of 1 year. The results indicate that dioxins affect developing bone by interfering with bone growth and mechanical strength and that the effects are mainly reversible. The dioxin-resistance alleles, Ahr(hw) and B(hw) increase the resistance to these defects.

Accumulation of dietary organochlorines and vitamins in Baltic seals.

We studied the accumulation of polychlorinated biphenyls (PCBs), 1,1,1.trichloro-2,2-bis[p-chlorophenyl]ethane (DDT) and its metabolites, and vitamins A and E in the top levels of the Baltic Sea food web. The investigation focused on the transfer of contaminants and vitamins to the ringed seal (Phoca hispida baltica) and the grey seal (Halichoerus grypus) from their main prey species. The trophic level of the seals was investigated using stable isotopes of nitrogen and the results indicated that both species of Baltic seal feed at approximately the same level. PCBs accumulated to a greater extent in the grey seal than in the ringed seal. Biomagnification factors for DDT compounds were similar for both species of Baltic seal (85-140). Differences in observed DDT levels were due to different prey selection by the two species, while differences in PCB levels indicated a species-specific metabolic system. There was a clearly greater accumulation of DDT compounds than of PCBs in both species of seal. The supply of dietary vitamin A was normally above the recommended level in all the seal populations studied. Low levels of hepatic vitamin A in the Baltic seals could therefore indicate the toxic effects of a high level of persistent organic pollutants on vitamin A dynamics, at least in the ringed seal. In the grey seals, low hepatic vitamin A levels could also be explained by lower levels of dietary vitamin A, compared to the reference grey seals, as it is not known if seals can store unlimited amounts of vitamin A. The greater uptake of vitamin E by Baltic seals, compared to seals in the reference areas, demonstrated by elevated levels of vitamin E in the blubber, could be a response to oxidative stress caused by the high contaminant load. These results further support our previous hypothesis that the toxic effects of environmental contaminants could be causing the observed divergence in vitamin levels between the Baltic seals and the reference seal populations.

Development of supercritical fluid extraction with a solid-phase trapping for fast estimation of toxic load of polychlorinated dibenzo-p-dioxins-dibenzofurans in sawmill soil.

A method consisting of automated supercritical fluid extraction (SFE) with simultaneous cleanup by a solid-phase trap was developed for fast analysis of polychlorinated dibenzo-p-dioxins (PCDDs) and dibenzofurans (PCDFs) in soil. SFE was optimised to replace conventional liquid-based methods in routine analyses of PCDD/PCDFs in sawmill soil contaminated by a chlorophenol formulation. PCDD/PCDFs were quantitatively extracted in 60 min using CO2 at 400 atm and 100 degrees C without a modifier. A trap containing a small amount of activated carbon mixed with Celite efficiently collected PCDD/PCDFs after SFE. After SFE co-extracted impurities were eluted out from the trap with 4 ml of hexane and PCDD/PCDFs were eluted with 10 ml of toluene. The concentrations and TCDD-equivalent of PCDD/PCDFs corresponded to the results of traditional solvent extraction method (Soxhlet) in six sawmill soils tested. The performance of the trap was maintained over a long period of time (nearly 100 extractions).

Current levels of DDT, PCB and trace elements in the Baltic ringed seals (Phoca hispida baltica) and grey seals (Halichoerus grypus).

Residue levels of polychlorinated biphenyls (PCBs) and 1,1,1-trichloro-2,2-bis[p-chlorophenyl]ethane (DDT) were determined from liver samples of ringed seals (Phoca lispida) from the Baltic Sea and Svalbard, and of grey seals (Halichoerus grypus) from the Baltic Sea and Sable Island in Canada. Both Baltic seal populations showed clearly higher average sum PCB (SPCB) and sum DDT (SDDT) levels than the reference seal populations. Among the Baltic seals, SPCB levels were twice as high as SDDT levels, and both contaminants were higher in ringed seals than in grey seals. A difference in gender was observed only in the Sable Island grey seal population, in which males showed a higher level of contamination than females. A reduction of the SDDT levels, and to a lower extent of the SPCB levels can be observed in the Baltic seals since the peak contaminant levels during the 1970s. The decrease has been more rapid in the grey seals than in the ringed seals. The SPCB levels in the Baltic ringed seals are still high enough to cause a threat to their well being. Residue levels of the trace elements mercury, cadmium, lead and selenium were determined from liver, kidney and muscle samples of grey seals from the Baltic Sea and Sable Island. Only cadmium showed a geographic difference being higher in the seals from Sable Island than from the Baltic Sea. No clear reduction of the metal burden can be observed in the Baltic grey seal population since the 1970s.

PCDD/Fs and PCBs in Baltic herring during the 1990s.

Baltic herring samples caught from the Baltic Sea during the spring periods of 1993-1994 and 1999 were analysed for polychlorinated dibenzo-p-dioxins (PCDD), polychlorinated dibenzofurans (PCDF), and polychlorinated biphenyls (PCB). For analyses, 1570 individual herring were combined to 120 pools. Correlations between concentrations of congeners 1,2,3,7,8-PeCDD, 1,2,3,7,8-PeCDF, 2,3,4,7,8-PeCDF, and 2,3,4,6,7,8-HxCDF, and age of herring were the strongest (r>0.8) followed by correlations between PCB congeners PCB 105, 118, 126, 156, 169 and 180 (r>0.7), and age of herring. Due to higher fat percentage in herring in the Gulf of Bothnia the concentrations of PCDD/Fs and PCBs on fresh weight (fw) basis were higher than in herring in the Gulf of Finland. The concentrations of WHO(PCDD/F)-TEQs ranged from 1 to 27 pg/g fw, depending on the age and catchment area of herring, and concentrations of WHO(PCB)-TEQs reached 32 pg/g fw. Between the two studied time points no clear downward trend in concentrations was observed.