Significance of an In Vivo Genotoxicity Test for Surface Waters Is Increased when the Physiological Characteristics of the Test Organism (Bivalvia: Mollusca) Are Taken into Account?

The alkaline filter elution assay using the gills of the freshwater clam species Corbicula fluminea detects breaks in single‐stranded DNA and is thus a good method for determining the genotoxic potential of surface waters. In attempting to standardize the procedure, a wide range of factors which could have an influence on the uptake of genotoxic substances by the exposed clams were studied. The most important parameters of the static exposure in relation to the rate of filtration by the animals turned out to be the temperature, the volume of the water, and the exposure time. Differences in body size and in the amount of suspended particles in surface waters did not play a significant role. The results demonstrate that the in vivo test system can be quite sensitive and its results reproducible when the relevant species‐specific characteristics of the test organisms are brought into consideration, even if the test organism belongs to a biologically more advanced group. A clear dose‐response relationship to the reference substances 4‐nitroquinoline‐1‐oxide (NQO) and N‐methyl‐N‐nitro‐N‐nitrosoguanidine (MNNG) can be observed even after a short exposure time of between four and twenty hours. No genotoxic effects were observed when using this method on surface waters from the area of Cologne (including water from the Rhine River and within the protection zone 2 of the Cologne waterworks).     

Use of Chemotaxonomy To Study the Influence of Benzalkonium Chloride on Bacterial Populations in Biodegradation Testing

Contradictory results are reported for the behaviour of quaternary ammonium compounds (QACs) in sewage treatment plants (STPs). QACs may sorb onto activated sludge. Only little information is available with respect to effects of QACs against bacteria in STPs. Only 5 to 15 % of bacteria present in sewage sludge can be detected by means of culture dependent microbiological methods. The shift of the bacterial populations due to effects of test compounds have not been studied up to now with culture independent methods. The microbial populations shift was studied in situ using culture independent chemotaxonomy profiling ubiquinones and polyamines. Additionally, toxic effects of QACs against bacteria present in the test vessels of the Zahn‐Wellens test (OECD 302 B) were assessed with a toxicity control in the test. The ubiquinone profiles representing changes in Gram‐negative populations mainly showed that the activated sludge was affected only in test vessels containing benzalkonium chloride. According to chemotaxonomy Acinetobacter or/and some members of Pseudomonas spp. have been selected by benzalkonium chloride after some adaptation period (8 to 12 days).     

Eignung mikrobieller Biotests für gebundene Schadstoffe Suitability of Microbial Bioassays for Bound Contaminants

Toxicity of soil spiked with water soluble and insoluble contaminants was assessed with bacteria. Three bioassays were used with common soil bacteria. Water soluble fractions from the soils were estimated in microtiterplates via dehydrogenase activity of B. cereus and growth of P. putida. A new method was developed using B. cereus to include effects of soil particles with bound contaminants. As contaminants p-nitrophenol and 2-naphthol were tested in these systems. Results indicated that the concentration of the contaminants in the soil solution is determined by soil properties. Toxicity of water soluble fractions was not only a result of the toxicant's concentration but further modified by soluble components of the soils. It is not possible to assess soil pollution from toxicity of the water soluble fraction in general, because unknown factors of the soluble soil fraction can influence toxicity greatly. The newly developed contact bioassay with B. cereus showed clearly higher toxicity of 2-naphthol estimated in soil suspension than did the comparable test system with the same bacteria using water soluble fractions only. The results stress the importance of including soil particles into bioassay procedures. It was shown that also water insoluble, bound contaminants can act as toxicants. For an estimation of toxicity under conditions more relevant for the environment, the suggested test system can provide useful information to bioavailability of bound contaminants.     

Use of Plant Bioassays for the Detection of Genotoxins in the Aquatic Environment

The present article describes the use of plant bioassays for the detection of genotoxins in the aquatic environment and gives an overview of test methods, their detection spectrum for environmental mutagens and carcinogens and their limitations and pitfalls. The most widely used test systems are micronucleus assays with meiotic cells of Tradescantia and with meristematic root tip cells of Allium and Vicia. In the last years, protocols for single cell gel electrophoresis assays have been developed, which can be conducted with a variety of species. Also various gene mutation test procedures have been developed with plants but they have hardly ever been used in environmental studies. Plants detect a broad variety of environmentally relevant genotoxins, in particular directly acting compounds. Many pesticides and industrial chemicals caused positive results; plant assays are also a unique tool for the detection of DNA‐reactive carcinogenic heavy metals in the environment. In many studies with complex mixtures, positive results were obtained which indicates that plants are sufficiently sensitive to detect effects without concentration of water samples. One of the shortcomings of the use of plants as indicators is their lack of sensitivity towards certain classes of promutagens such as nitrosamines, heterocyclic amines, and polycyclic aromatic hydrocarbons (PAHs). However, the former compounds are hardly encountered in the environment and PAHs can easily be detected chemically and in other mutagenicity tests. Taken together, the currently available data show that plant bioassays are a useful component of test batteries for environmental monitoring.     

Characterization of Estuarine and Fluvial Dissolved Organic Matter by Thermochemolysis using Tetramethylammonium Hydroxide

Selected samples of dissolved organic matter (DOM) isolated by ultrafiltration (UDOM) have been analyzed by thermochemolysis in the presence of tetramethylammonium hydroxide (TMAH). This technique cleaves ester and ether bonds of bio‐ and geological macromolecules and releases monomer subunits and methylates them in situ as their methyl ethers and methyl esters. Compared with conventional pyrolysis, TMAH thermochemolysis avoids decarboxylation of preexisting carboxylic moieties and produces aromatic acids as their methyl esters. Various phenolic derivatives, which might originate from incorporated lignin‐derived structures, from the highly aliphatic and resistant biopolymer cutan and also from proteinaceous materials, were identified among the products produced from UDOM upon thermochemolysis. The presence of lignin derivatives in UDOM indicates input of organic matter derived from terrestrial sources. Various aromatic acids, perhaps representing the final steps in the oxidation of the side‐chain during microbial oxidation of lignin, were released upon TMAH thermochemolysis, suggesting they are structural constituents of the UDOM. Different ratios of lignin‐derived materials, commonly determined using the CuO oxidation method, such as the Δ value, indicative of the amount of lignin present, the acid/aldehyde ratio (Ad/Al)_G, indicative of the extent of oxidative degradation of the lignin component, and the syringyl/guaiacyl (S/G) and p‐hydroxyphenyl/guaiacyl (P/G) ratios, indicative of the contribution for the different types of lignin, were determined.     

Scum in Nutrient Removal Plants: The Role of Carbon Sources in “Microthrix parvicella” Growth

Scum formation is a widespread problem in activated sludge nutrient removal plants. It often comes along with an excessive development of the filamentous bacterium “Microthrix parvicella” stabilizing the flotation process. As “M. parvicella” was found to depend on long‐chain fatty acids (LCFA) as sole carbon source not only in vitro but also in situ, some options of in‐situ substrate supply are discussed. Wastewater concentrations of fatty acids in the range of 2 to 15 mg L^‐1 and homologue concentrations from synthetic surfactant degradation below 10 mg L^‐1 rule out these substrates as source for excessive biomass production. They might, however, well be suitable for start‐up of a “M. parvicella” population. Build‐up of excessive biomass might rely on fatty acid supply originating in cell walls of lysed stationary phase bacteria of long residence time sludge fractions such as scum layers. Moreover, biogenic surfactants such as rhamnolipids have been proved to be an excellent carbon source for excessive biomass production in vitro.     

Review Paper: A semi‐empirical model of strain sensitivity for 4D seismic interpretation

We reformulate the original model of Hatchell and Bourne and Røste, Stovas and Landrø that couples fractional velocity change to subsurface strain via a fundamental constant R. The new model combines elastic compressibility of a dual‐porosity system for a sand–shale mixture with horizontal planes of inter‐granular weakness. The majority of observed R‐factor magnitudes from post‐stack 4D seismic data in both the reservoir and overburden can thus be explained. R is predicted to depend strongly on lithology and also initial strain state. The model is also extended to predict the observed angle‐dependence of time‐lapse time‐shifts from pre‐stack data. An expression for the gradient of time‐shift with incidence angle is obtained in terms of the background V_P/V_S, and also the ratio of tangential to normal compliances B_T/B_N representing loss or creation of inter‐granular coupling. If accurately estimated from data, this compliance ratio can be used as an additional parameter to assess the post‐production state of the overburden. It is concluded that whilst R remains the over‐arching parameter controlling the magnitude of time‐shifts measured from 4D seismic data, B_T/B_N is a subtler parameter that may also prove of future value.     

Separation and Preconcentration of Cobalt Using a New Schiff Base Derivative on Amberlite XAD‐7

In this study, a new solid‐phase extraction procedure has been developed for preconcentration and determination of Co ions in different water samples by flame atomic absorption spectrometry (FAAS). Cobalt was preconcentrated as N,N′‐bis(pyridine‐2‐yl‐methyl)benzene‐1,4‐diamine (Co‐BPMBDA) from sample solutions using a column containing Amberlite XAD‐7 and was determined. In order to achieve the best performance for the method, effects of several parameters such as pH, concentrations of ligand, sample flow rate, eluent, and matrix ions on the method efficiency were investigated. Under optimum conditions, the preconcentration factor was found to be 200 for 1000 mL waters samples. Detection limit based on the 3S_b criterion was calculated as 0.24 µg/L for 100 mL of sample solution and relative standard deviation was found to be 1.8%. The method was applied to determine the trace amounts of cobalt in water samples.     

Simultaneous and Sensitive HPLC Determination of Mono‐ and Disaccharides,Uronic Acids,and Amino Sugars after Derivatization by Reductive Amination

The reductive amination of low‐molecular‐weight saccharides, uronic acids, and amino sugars, followed by a separation of the derivates by means of ion‐pair chromatography or RP‐HPLC, offers an interesting alternative to HPAEC‐PAD for the environmental analysis of these compounds. Under this aspect various potential amination reagents, i.e., p‐amino‐benzoic acid (p‐AMBA), p‐AMBA propyl ester, 1‐aminopyrene, 2‐(2‐aminophenyl)indole, and 4‐aminoazobenzene, were tested with regard to the formation of derivates and to the chromatographic properties of the formed derivates. p‐AMBA, p‐AMBA propyl ester and 4‐aminoazobenzene proved to be especially suited, because they facilitate the amination of all carbohydrate reference components together with a complete separation and sensible detection (detection limits < 0.5 mg/L) of the derivates. Mainly the following elution sequence was ascertained: amino sugars (hexosamines) / disaccharide(s) / monosaccharides (hexoses) / hexuronic acid(s) / N‐acetyl‐D‐glucosamine. Detection limits down to 0.1 μmol/L were realized using p‐AMBA as reagent, facilitating the determination of the target compounds in landfill leachates and lysimeter percolates. Applying the p‐AMBA propyl ester for derivatization, chromatographic interferences with weakly retained derivates and the coelution of the reagent with its galactosamine derivate can be avoided, since the ester elutes after its derivates unlike p‐AMBA itself.     

Inorganic nitrogen retention in acid‐sensitive lakes in southern Norway and southern Ontario,Canada—a comparison of mass balance data with an empirical N retention model

In‐lake retention of inorganic nitrogen species (nitrate and ammonium) was estimated from mass balances in five acid‐sensitive lakes in southern Norway and eight in southern Ontario, Canada, to evaluate an empirical in‐lake N retention (R_N) model. This model is included in the First‐order Acidity Balance (FAB) model, which currently is used for calculation of critical acid loads and exceedances in many countries. To estimate in‐lake R_N, the FAB model uses a recommended mass transfer coefficient (S_N) of 5 m year⁻¹, which mainly is derived from NO₃⁻ mass balances in Canadian lakes. To date, the in‐lake R_N model has not been evaluated for large parts of Europe. At the Norwegian study sites receiving the highest N deposition (>120 meq m⁻² year⁻¹) the net in‐lake retention of inorganic N (TIN) exceeded the corresponding terrestrial retention by a factor of 1·1–2·6. Despite differences in N loading and hydrology at the Norwegian and Canadian sites, both the mean mass transfer coefficients for NO₃⁻ (S_NO3; 6·5 versus 5·6 m year⁻¹) and TIN (S_TIN; 7·9 versus 7·0 m year⁻¹) were of comparable magnitude. Both mean values and ranges of S_NO3 suggest that the default S_N value presently recommended for FAB model applications seems valid over a large range in N inputs and areal water loads (q_s). However, owing to the relatively few data available for lakes with high q_s values (15–150 m year⁻¹), it is recommended that more lakes within this range be included in future studies to obtain a more precise prediction of in‐lake N retention over a wide q_s gradient. Also, when considering that the FAB model treats all inorganic N leaching from a catchment as NO₃⁻, it seems reasonable to use a default S_TIN value instead of just S_NO3 when estimating in‐lake R_N. In that case, the in‐lake R_N presently calculated by the FAB model might be slightly underestimated. Copyright © 2003 John Wiley & Sons, Ltd.     

Natural and experimental structural evolution of dispersed organic matter in mudstones: The Shimanto accretionary complex,southwest Japan

Structural changes induced by thermal maturation of dispersed organic matter (OM) in the Shimanto accretionary complex, southwest Japan, were investigated using micro‐Fourier‐transform infrared spectroscopy and micro‐Raman spectroscopy. Natural dispersed OM exhibits systematic structural changes inferred from D1‐ and G‐band FWHM values, Raman band separation (RBS), and intensity ratios of the D1‐ and G‐bands (I_D1/I_G ratio) from diagenetic zone to anchizone (IC values: 0.75–0.30). Infrared spectra indicate a loss of aliphatic CH _x, aromatic CH _x, and oxygen‐containing structures as temperature increases. These changes are consistent with discontinuities in thermal structures bounded by out‐of‐sequence thrusts. Kinetic pyrolysis experiments indicate that the I_D1/I_G ratio of synthesized OM has a power law relationship with heat treatment time. Kinetic models of temperature dependence were fitted using the I_D1/I_G ratio, and an effective activation energy of 106 ±17 kJ/mol was estimated using an Arrhenius equation. The activation energies estimated by power law rate and Avrami models have a least‐square correlation coefficient of 0.93, indicating the temperature dependence of carbonization. The estimated effective activation energy is consistent with that of coal, lignin, cellulose, and hemicellulose during thermal degradation. On the other hand, RBS, and D1‐ and G‐band FWHM values of OM display more complex changes with increasing heating temperature and time, and it is difficult to constrain rate parameters during pyrolysis experiments. Our data indicate that the I_D1/I_G ratio is controlled by a simple thermally activated process, whereas RBS and D1‐ and G‐band FWHM values can be affected by lithostatic pressure, fluid activity, hydrogen index, and host lithology, as well as temperature. Structural evolution of dispersed OM in mudstones differs between natural and anhydrous closed experimental systems. Natural carbonization based on micro‐Raman spectroscopy should be applied for a limited indicator of thermal maturation, especially for dispersed OM in diagenetic zone.     

Caustics in a periodically layered transversely isotropic medium with vertical symmetry axis

Wave propagation in a finely layered medium is a very important topic in seismic modelling and inversion. Here we analyse non‐vertical wave propagation in a periodically layered transversely isotropic (VTI) medium and show that the evanescent (attenuation) zones in the frequency‐horizontal slowness domain result in caustics in the group velocity domain. These caustics, which may appear for both the quasi‐compressional (qP) and quasi‐shear (qSV) wave surfaces are frequency dependent but display weak dependence at low frequencies. The caustics computed for a specific frequency differ from those observed at the low‐ and high‐frequency limits. We illustrate these caustics with a few numerical examples and snapshots computed for both qP‐ and qSV‐wave types.     

New Strategy to Reduce the Harmful Effects of Smoking: Reducing the Level of N‐nitrosamines in Mainstream Smoke by NaA Zeolite and In vitro and In vivo Investigations

A new effort to control the pollution caused by smoking by use of zeolite sieving the N‐nitrosamines in smoke is depicted and assessed in this paper, which is a first preliminary approach to demonstrate the performance characteristics of zeolite for the reduction of the concentration of N‐nitrosamines in the cigarette smoke. Smoking causes global pollution and is difficult to control because smoke is a complex system containing thousands of components. A new strategy is presented here for the reduction of the harmful effects of smoking through dispersion of zeolite into the tobacco rod of cigarettes to eliminate the N‐nitrosamines once they form in the smoke, and both in vitro and in vivo tests were employed to check the efficiency of this strategy. The impact of zeolite on the combustion of tobacco and the total toxicity of smoke are evaluated for the first time. Lower mutagenic activity and higher CHO cell livability were found for the test cigarette containing zeolite in comparison with the control, and no significant damage was detected in 30‐day animal experiments when exposed to the cigarette smoke. This confirms the feasibility of the new strategy and promotes the potential application of zeolite for protecting public health.     

On the Limnology of Deep and Saline Lake Burdur in Turkey

Limnological characteristics of Lake Burdur in Lake District in south‐western Turkey are presented. It is a deep, tectonic (estimated max. depth 100 m), athalassic, highly alkaline, and saline lake. A set of physical and chemical variables was monitored, phyto‐ and zooplankton was sampled from surface layer of the lake during 1997. Physico‐chemical variables indicated that the lake is hyposaline and composed of some hydrochemically different water layers formed by groundwater sources located on the bottom of the lake. The phytoplankton composition of Lake Burdur consisted of Cyanophyta, Chlorophyta, Bacillariophyta, Dinophyta, and Chrysophyta. The abundance and number of species of Chlorophyta and Cyanophyta were higher than the other taxa. The zooplankton composition of the Lake consisted of Rhizopoda, Rotifera, and Crustacea. Number of species of Rotifera was higher than the other taxa. The diversities of the phyto‐ and zooplankton were calculated according to the Shannon‐Weaver diversity index. The diversity of each group was found to be low in the lake.     

Beeinflussung der Enzymaktivität einer Protease durch Huminstoffe Influence of Humic Substances on the Activity of a Protease

This paper deals with the influence of different humic substances (HS) on the enzyme activity (EA) of the proteolytic enzyme pronase E. The EA was determined by analyzing the concentration of the amino acid valine hydrolyzed from caseine. For amino acid analysis, a flourescence detector was used after precolumn derivatization and HPLC-separation of the hydrolysis products. The data show that the presence of HS has no adverse effects on amino acid analysis. Based on this result, a decrease of EA was observed in the presence of HS acting as enzyme inhibitors. Clear inhibition effects could be seen for different fulvic acids (HO3 FA, FBR FA and FBR 1.0 FA) at concentrations of 150 mg/L. In case of FBI-2 FA, higher concentrations were necessary to decrease the EA significantly. At an inhibitor concentration of 1500 mg/L the EA was mostly found to be less than 50%. The standard deviations of the results don't allow a differentiation between the HS of different origin and treatment. Specifying HS according to their influence on EA has to remain uncertain. As shown in a kinetic study of the hydrolysis of L-Leucine-2-naphthylamide-hydrochloride, the inhibitor HO3 FA decreases the maximum velocity of the reaction but has no effect on the Michaelis constant. These results indicate that this HS acts as a noncompetitive inhibitor.     

Biological Test for the Detection of Low Concentrations of Infectious Cryptosporidium parvum Oocysts in Water

An in vivo SCID mouse infectivity assay was used to determine its capacity to detect the infectivity of low concentrations of Cryptosporidium parvum oocysts in water. This biological test can be applied to demonstrate oocysts infectivity in water samples derived from drinking water supply and/or environmental sources.     

Simultaneous Analysis of Natural Free Estrogens and Their Conjugates in Wastewater by GC‐MS

A solid‐phase extraction (SPE)‐gas chromatography (GC)‐mass spectrometry (MS) analytical method was developed for the simultaneous analysis of natural free estrogens and their conjugates in wastewater samples. Natural free estrogens and their conjugates in wastewater were successfully separated by the oasis hydrophilic‐lipophilic balance solid phase extraction (Oasis HLB SPE) method, and the conjugates were initially enzyme hydrolyzed by β‐glucuronidase or arylsulfatase from Helix pomatia prior to derivatization. N‐methyl‐N‐(tert‐butyldimethylsilyl)trifluoroacetamide (MTBSTFA) plus 1% tert‐butyldimetheylchlorosilane (TBDMCS) was chosen as the derivatization reagent, and the most appropriate conditions of derivatization were determined to be at 95°C for 90 min. The recovery ratios of nine target chemicals were determined by spiking them in 1 L of ultra‐purified water or the influent of a wastewater treatment plant (WWTP). The recovery ratios of six out of nine for the analytes ranged from 73.3–114.9% with relative standard deviations (RSD) from 1.6–19.9%. The established method was successfully applied to environmental wastewater samples which were collected from one municipal wastewater treatment plant (WWTP) in Osaka, Japan, for the determination of natural free estrogens and their conjugates. In the influent sample, E1, E2, E1‐3S, E3‐3S, and E1‐3G were detected at concentrations of 16.6, 9.6, 8.2, 21.9, and 3.2 ng L^–1, respectively. However, only E1 was detected at a high concentration of 44 ng L^–1 in the effluent sample, suggesting that it is the dominant natural free estrogen in the effluent.     

Preconcentration by Coprecipitation of Copper and Nickel with Mo(VI)/Triazole Derivative System and Their Determinations by Flame Atomic Absorption Spectrometry in Food and Water Samples

A new separation and preconcentration technique based on coprecipitation of Cu(II) and Ni(II) ions by the aid of Mo(VI)/di‐tert‐butyl{methylenebis[5‐(chlorobenzyl)‐4H‐1,2,4‐triazol‐3,4‐diyl]}biscarbamate (BUMECTAC) precipitate has been established. The Mo(VI)/BUMECTAC precipitate was dissolved by concentrated HNO₃ and the solution was completed to 5.0 mL with distilled/deionized water. The levels of the analyte ions were determined by flame atomic absorption spectrometer. The effects of experimental conditions like HNO₃ concentration, amount of BUMECTAC and Mo(VI), sample volume, etc. and also the influences of some foreign ions were investigated in detail on the quantitative recoveries of analyte ions. The preconcentration factors were found to be 40 for Cu(II) and 100 for Ni(II) ions. The detection limits for Cu(II) and Ni(II) ions based on 3σ (N:10) were 0.43 and 0.70 µg L^?1, respectively. The relative standard deviations were found to be lower than 4.0% for both analyte ions. The accuracy of the method was checked by spiked/recovery tests and the analysis of two certified reference materials (Environment Canada TM‐25.3 and CRM‐SA‐C Sandy Soil C). The procedure was successfully applied to sea water and stream water as liquid samples and baby food as solid sample in order to determine the levels of Cu(II) and Ni(II) ions.     

New Attempt to Reduce the Harm of Smoking: Reducing the Nitrosamines Level in Tobacco Smoke by Microwave Irradiation

In order to protect the environment and public health, microwave irradiation was investigated as a means of reducing the nitrosamines level of tobacco smoke under mild conditions where the microwave energy was 1 kW and the irradiation time was shorter than 2 min. The microwave‐induced elimination of nitrosamines such as N‐nitrosopyrrolidine (NPYR) and N‐nitrosodiphenylamine (NDPA) was investigated and the impact of the presence of water and salt in the medium on the reduction was assessed in detail. The existence of water in the medium was crucial for the microwave‐induced reduction of NDPA, because the rapid movement of water molecules under the influence of microwaves strongly affects the adjacent nitrosamines leading to their decomposition. This was the first time that microwaves have been used to directly irradiate cigarettes sealed in their packaging, and the nitrosamines content of tobacco and smoke of these irradiated cigarettes were analyzed with care. Irradiation for 90 s with microwave energy of 1kW was seen to reduce 25 to 30% of the nitrosamines in tobacco and 50 to 60% of the levels present in the mainstream smoke of the cigarette. Moreover, lower mutagenic activity and higher CHO cell livability were found for the irradiated cigarettes in comparison with the control according to the results of in vitro tests.     

Effect of Nickel and Cadmium on Ammonium Uptake Kinetics of Free and Immobilized Cells of Anacystis nidulans

The stress effect of Ni and Cd on the ammonium uptake varied significantly (ANOVA test) in free and immobilized state of the test organism. The effect due to the interaction between different variables (cell state type, metal type and metal dose) was studied to depict the significant or non‐significant variation in the ammonium uptake by free and immobilized cells in the presence of metal ions. Ammonium uptake exhibited a competitive mode of inhibition in the presence of Ni in both free and immobilized state of the organism. However, Cd exhibited non‐competitive and competitive inhibition in free and immobilized cells, respectively. The study demonstrates that there is a considerable influence of metal ions on the ammonium uptake. Cd was found to be more toxic compared to Ni in both free and immobilized state.