Nonylphenol induced changes in trophic web structure of plankton analysed by multivariate statistical approaches.

Microcosm studies unveil relevant information in ecotoxicology. However, statistical evaluation of data gained from the different parts of the ecosystem often remain on unsatisfying, discretely level, e.g. separately for different species communities. A combination of different multivariate statistical methods enables an ecological interpretation of the found changes in zoo- and phytoplankton abundances in connection with the physico-chemical variables. To investigate the effects of nonylphenol (NP) on plankton communities, NP was continuously applicated in microcosms by controlled release. Plankton taxa were identified and quantified, various physico-chemical variables were measured. Maximum NP concentration ranged between 29 and 129 microg/L. The most important abiotic variable explaining both zoo- and phytoplankton abundances was NP concentration. NP primarily inhibited Crustaceae, especially juveniles. Cladocera and Copepoda abundances decreased. Rotatoria tend to increase, probably due to a decline of competition and predator pressure. Shifts in phytoplankton structure occurred with a time lag. Aggregating phytoplankton on class level did not show NP effects, whereas aggregating them according to feeding protection strategies did. Thus NP influences phytoplankton at least prevailingly indirectly via zooplankton. Zooplankton shifts led to species composition with different feeding preferences and strategies. Thereby the feeding pressure on phytoplankton changed. NP reorganized the plankton interrelation. Whereas without NP phytoplankton composition was dominated by Rotatoria, in the NP treated microcosms the Crustaceae unveiled highest explanatory power. The found effects were reversible in all but the highest treated microcosm. There are indications that the found effects were caused by endocrine activity of the chemical. Iteratively combining multivariate statistical methods with ecological and toxicological knowledge proved to provide a deeper insight into the mode of ecosystem disturbance by toxic substances.

Influence of 4-nonylphenol on the structure of nematode communities in freshwater microcosms.

We investigated the effect of 4-nonylphenol (NP) on nematode communities in the sediment of freshwater microcosms. Seven treatments were dosed with various concentrations of NP over a period of six weeks by using a controlled-release method (NPI-NP7; maximum sediment concentrations: 0.29-3.37 mg/kg dry wt). Four microcosms were not exposed to NP and served as controls. Nematode communities were analyzed over a period of 15 weeks, including sampling dates before, within, and after the NP application. Communities were characterized in terms of total nematode abundance and species diversity (Shannon index and evenness), as well as composition of species, feeding types, and different life-history strategists (maturity index [MI]). Species composition was analyzed by using a multivariate method (principal response curves). Total nematode abundance and species diversity were not affected in any of the NP-treated microcosms. However, in the highest dosed treatment, NP-induced changes in the nematode communities occurred. Species and feeding types composition, as well as the MI, were affected in the postapplication period, with species composition being altered most clearly. In the highest dosed treatment, deposit-feeding species, classified as colonizers (Eumonhystera), increased in dominance, whereas epistrate feeders and chewers (Prodesmodora and Tobrilus) decreased in relative abundance compared to the control.

Effects of 17 alpha-ethinylestradiol on zoo- and phytoplankton in lentic microcosms.

The effects of 17 alpha-ethinylestradiol (EE), an endocrine disruptor, on zoo- and phytoplankton were studied in outdoor 230-L still-water microcosms. Cell density and biomass, diversity, and community composition were analyzed. Five microcosms were treated by controlled release for six weeks, three by direct application of EE. To investigate recovery, sampling was continued for four weeks after treatment. Most characteristics of the zooplankton were not unambiguously affected by EE. Only the relative density of copepods, especially of their larvae, decreased significantly after EE application. For phytoplankton, no unambiguous concentration- or toxodose-correlated effects on any biotic characteristics could be found. However, most properties of the phytoplankton deviated from those of controls, i.e. tended to be smaller (number of species per microcosm, biomass, cell density) or covered a wider range (diversity, evenness). PCA indicated a shift of species structure in the treated microcosms. This was supported by the species scores calculated by the principal response curve method, although the principal response curve itself showed no clear EE-correlated shifts. High variability within the biocenosis between microcosms and over time, probably because of disturbance of the ecosystem before starting of the test, might have superimposed EE-dependent effects.

Effects of 4-nonylphenol on phytoplankton and periphyton in aquatic microcosms.

The effects of nonylphenol (NP) on phytoplankton and periphyton were studied in 230 L outdoor microcosms. Phytoplankton cell density and biomass, phytoplankton and periphyton diversity, and assemblage composition were analyzed during a four-week preapplication period, followed by six weeks of NP treatment via controlled release and a six weeks postapplication period. Changes in species richness and diversity were not correlated with NP concentrations. However, changes in phytoplankton cell densities during the first week of the postapplication period were related to previous exposure. In the controls and the lowest-dosed microcosm. Conjugatophyceae constituted most of the biomass during the dosing and the postapplication period. In contrast, Dinophyceae dominated the biomass during the dosing and the postapplication period at higher NP concentrations. Principal response curves revealed changes in phytoplankton assemblage composition during the dosing and the postapplication period. Dinophyceae and most Cyanophyceae were more abundant at intermediate and higher NP concentrations, whereas Conjugatophyceae were less abundant compared to controls. Assemblages only partly recovered during the postapplication period. Periphyton taxon richness, diversity, and assemblage change was not related to NP concentrations. At the lowest and intermediate concentration, assemblages were significantly different from the controls and the higher concentrations, which were similar during the treatment period.

The bioaccumulation and fate of a branched 14C-p-nonylphenol isomer in Lymnaea stagnalis L.

A single branched isomer of p-nonylphenol, 4(3',6'-dimethyl-3'-heptyl)-phenol, previously identified by gas chromatography-mass spectrometry as one of the major constituent isomers in p-nonylphenol (constituting approximately 10% of all its isomers), was synthesized and used in studies of its bioaccumulation and excretion in the hermophroditic pond snail Lymnaea stagnalis L. Branched isomers of nonylphenol are perceived to have more estrogenlike toxicity than the straight-chain isomers in addition to being more resistant to biodegradation in the environment. With an average static exposure concentration of 104 microg/L (range: 92-116 microg/L) in water at 19 degrees C for 8 d, the uptake of the compound was found to be fairly rapid, reaching a peak concentration of 23,548 microg/kg of whole tissue wet weight after 5 d and a peak bioaccumulation factor (BAFw) of 242 (5,562, based on lipid weight) after 3 d. The uptake data fitted into a logarithmic expression C(t) = 5,231 ln(t) + 11,956, where C(t) is the amount of residues accumulated in whole tissue in micrograms per kilogram tissue wet weight after a period of time, t, and t is the period of exposure in days. By determination of the excretion of 14C-residues released in water and in feces, an average loss of 96% of the accumulated residues was achieved after 22 d of continuous exposure to clean water. By first-order kinetics analysis of the excretion data, an average half-life of excretion of 4.9 d was obtained. By high-performance liquid chromatography and gas-liquid chromatography-mass spectrometry, a catechol metabolite, 4(3',6'-dimethyl-3'-heptyl)-catechol, was detected in tissue extracts (after hydrolysis with beta-glucuronidase) and in feces, in addition to the parent isomer, suggesting that the isomer may have been metabolized by glucuronic acid conjugation and hydroxylation at the ortho position of its phenolic ring.

Controlled release experiments with nonylphenol in aquatic microcosms.

A method of controlled release of technical nonylphenol (tNP) was developed to simulate realistic exposure in ecotoxicological studies on aquatic organisms. The direct addition of tNP from an aqueous stock solution into 50 ml of water led to a concentration decrease of 80 to 90% weight/volume (w/v) from nominal values within 48 h. The inclusion of tNP in semipermeable low-density polyethylene (LDPE) lay-flat tubing (controlled-release devices [CRDs]) of different length allowed a continuous release into pure water at a rate of about 30 microg/cm2/d. Using CRDs in aquaria containing 15 L of 63-microm-filtered lake water, eight different concentrations with maxima between 38.1 and 326.7 microg/L were maintained for 11 d. During a second experiment in 15-L aquaria, five replicates of three concentrations were maintained using CRDs of the same length. Concentrations after 38 d varied between 0.1 and 6.7, 26.1 and 41.9, and 49.9 and 76.0 microg/L. In aquatic microcosms containing 230 L of lake water, a natural plankton community, 50 L of sediment, and macrophytes, seven different tNP concentrations (maxima 11-120.1 microg/L) were maintained over 45 d using CRDs of different length. They were replaced after 14 and 25 d because release of tNP was slower than predicted from laboratory experiments. Concentrations in the top 1-cm sediment layer were on average 19 times higher during the dosing period than concentrations in the water at the same time. In the sediments, different levels of applications led to concentrations that differed less distinctly than in the water. This method is suitable for exposing aquatic organisms continuously to constant, ecologically relevant concentrations of NP and represents an improvement over previous dosing methods in which exposure varied.

Effects of 17alpha-ethinylestradiol on the reproduction of the cladoceran species Ceriodaphnia reticulata and Sida crystallina.

Single-species tests allow the assessment of chronical effects of endocrine disruptors on organisms under laboratory conditions. In the current study, three-generation tests with Ceriodaphnia reticulata and Sida crystallina were carried out to examine the influence of the synthetic hormone 17alpha-ethinylestradiol (EE) on the reproduction of these cladoceran species. For each species, six different concentrations (10-500 microg/l EE) and two controls were tested with eight replicates for a duration of 4 weeks. The test was initiated by transferring one neonate individual into a test vessel which was incubated under standardized conditions. Every 2 days, the medium was renewed and life history parameters such as survivorship of the adults and juveniles, clutch size, first appearance and number of produced offspring were investigated. Acute toxicity tests showed that C. reticulata (EC50 (24 h) 1814 microg/l) was more sensitive towards the substance compared to S. crystallina (EC50 (24 h) >4100 microg/l). The juvenile phase of S. crystallina was significantly shorter at concentrations above 100 microg/l EE. For C. reticulata, 17alpha-ethinylestradiol caused a higher mortality of the newly hatched juveniles at EE concentrations above 200 microg/l. No effects were found for mortality of adult animals, birth rate, number of juveniles per female and net reproduction rate of S. crystallina and C. reticulata. Thus, sublethal effects on parental generation exposed to EE lead to disturbances in reproduction and to affection of their offspring. Negative consequences for the population dynamic cannot be excluded, e.g. the decrease of a population.