Algal growth inhibition test results of 425 organic chemical substances.

The toxicity towards the algal species Pseudokirchneriella subcapitata of 425 organic chemical substances was tested in a growth inhibition test. Precautions were taken to prevent loss of the compounds from the water phase and the test system (closed test system, low biomass, shorter test duration, silanized glass) and to keep pH constant by applying a higher alkalinity. Chemical phase distribution was modelled taking ionization, volatilisation, and adsorption to glass and biomass into consideration. If the modelled water concentration was below 90% of the nominal concentration the calculated EC values were corrected accordingly. The model helped to identify substances, where the calculated water concentration was too uncertain. Substances covering a wide range of physical-chemical properties and different modes of action were tested. Median effect concentrations (EC50) lower than 1000 mg/L were found for 310 substances; 216 of these were in the range from 1 to 1000 mg/L and 94 substances had EC50s below 1 mg/L and should be classified as "Very toxic". 36 substances fell in the group with EC50 > 1000 mg/L. An EC50 could not be established for 79 substances. These 425 different organic substances were tested under uniform conditions and thus considered a valuable source of information for administrators, industry, risk assessors and QSAR modellers.

Dose-response regressions for algal growth and similar continuous endpoints: calculation of effective concentrations.

We derive equations for the effective concentration giving 10% inhibition (EC10) with 95% confidence limits for probit (log-normal), Weibull, and logistic dose-response models on the basis of experimentally derived median effective concentrations (EC50s) and the curve slope at the central point (50% inhibition). For illustration, data from closed, freshwater algal assays are analyzed using the green alga Pseudokirchneriella subcapitata with growth rate as the response parameter. Dose-response regressions for four test chemicals (tetraethylammonium bromide, musculamine, benzonitrile, and 4-4-(trifluoromethyl)phenoxy-phenol) with ranges of representative slopes at 50% response (0.54-2.62) and EC50s (2.20-357 mg/L) were selected. Reference EC50s and EC10s with 95% confidence limits using probit or Weibull models are calculated by nonlinear regression on the whole dataset using a dose-response regression program with variance weighting and proper inverse estimation. The Weibull model provides the best fit to the data for all four chemicals. Predicted EC10s (95% confidence limits) from our derived equations are quite accurate; for example, with 4-4-(trifluoromethyl)phenoxy-phenol and the probit model, we obtain 1.40 (1.22-1.61) mg/L versus 1.40 (1.20- 1.64) mg/L obtained from the nonlinear regression program. The main advantage of the approach is that EC10 or ECx (where x = 1-99) can be predicted from well-determined responses around EC20 to EC80 without experimental data in the low- or high-response range. Problems with the estimation of confidence interval for EClow,x (concentration predicted to cause x% inhibition) from algal growth inhibition also are addressed. Large confidence intervals may be the result of experimental error and lack of a well-defined reference response value.

Biodegradation rates in adapted surface water can be assessed following a preadaptation period with semi-continuous operation.

The paper presents a semi-continuous preexposure procedure (SCEP) for use with surface water batch simulation biodegradability tests at low test substance concentrations (0.1-100 microg/l). Simple one step batch tests are normally used first of all for determining "initial rates" characteristic of the water as sampled, as by contrast to "adapted" rates obtained as a result of exposure of the microbial community to the test compound. The aim of the SCEP is to facilitate this adaptation and to become able to estimate reproducible "adapted rates" representing a steady state situation. This is accomplished by maintaining the microbial diversity and a supply of test substance and natural substrates. Conducting a SCEP involves regular renewal of part (e.g. one third) of the test suspension (e.g. every two weeks) adding freshly collected natural water with test compound of the initial concentration. An example study was performed with aniline, 4-nitrophenol, 2,4-dichlorophenoxyacetic acid, 4-chloroaniline, and water from the urban river Mølleå. Following preadaptation lag phases were considerably reduced and much more reproducible than obtained with simple batch tests. In tests at 100 microg/l lag phases for aniline decreased from 5.2 to <1 day, 4-nitrophenol from 10 to <1 day, 2,4-dichlorophenoxyacetic acid from 24 to <1 day, and 4-chloroaniline from 88 to 9 days, respectively. Adapted rates obtained with the SCECP were roughly the same as final rates in simple batch tests with successful adaptation. The adapted rate constant is perceived as an inherent characteristic of the test compound at a specific concentration and under environmental influence (temperature, natural substrates, etc.) but with no simple links back to the original microbial population. By contrast, the initial rates in one step batch tests are determined also by the microbial population initially present in the water sampled.

Shifts in biodegradation kinetics of the herbicides MCPP and 2,4-D at low concentrations in aerobic aquifer materials.

Biodegradation kinetics of two phenoxy acid herbicides, MCPP [(+/-)-2-(4-chloro-2-methylphenoxy)propanoic acid; mecoprop] and 2,4-D [2,4-dichlorophenoxyacetic acid] were studied in laboratory batch microcosms at low concentrations (0.025-100 microg/L) using 14C technique with sediments and groundwater from a shallow aerobic sandy aquifer. Below a certain threshold concentration of approximately 1 microg/L for 2,4-D and 10 microg/L for MCPP, the biodegradation followed first-order nongrowth kinetics, and no adaptation was observed within the experimental period of 341 d. Half-lifes for ultimate degradation were 500 d for 2,4-D and 1100 d for MCPP at 10 degrees C in unpolluted aquifer sediment in this environmentally relevant concentration regime. Above the threshold concentrations, the biodegradation rate accelerated gradually due to selective growth of specific biomass, which was ascertained from 14C most probable number enumerations of specific phenoxy acid degraders. Atthe highest concentration tested (100 microg/ L), specific degraders increased from 10(-1) to 10(5) cells/g during the experiment, and half-lifes after adaptation decreased to approximately 5 d. The enhanced rate of degradation by adapted systems was maintained during degradation of the last residuals measured to less than 0.1 microg/L. In situ long-term preexposure of the aquifer sediment also resulted in significant higher degradation rates of the phenoxy acids.

Laboratory shake flask batch tests can predict field biodegradation of aniline in the Rhine.

The aim of this study was to compare degradation rates of aniline in laboratory shake flask simulation tests with field rates in the river Rhine. The combined events of a low flow situation in the Rhine and residual aniline concentrations in the effluent from the BASF treatment plant in Ludwigshafen temporarily higher than normal, made it possible to monitor aniline at trace concentrations in the river water downstream the wastewater outlet by means of a sensitive GC headspace analytical method. Aniline was analyzed along a downstream gradient and the dilution along the gradient was calculated from measurements of conductivity, sulfate and a non-readily biodegradable substance, 1,4-dioxane. Compensating dilution, field first-order degradation rate constants downstream the discharge of BASF were estimated at 1.8 day(-1) for two different dates with water temperatures of 21.9 and 14.7 degrees C, respectively. This field rate estimate was compared with results from 38 laboratory shake flask batch tests with Rhine water which averaged 1.5 day(-1) at 15 degrees C and 2.0 day(-1) at 20 degrees C. These results indicate that laboratory shake flask batch tests with low concentrations of test substance can be good predictors of degradation rates in natural water bodies--at least as ascertained here for short duration tests with readily degradable compounds among which aniline is a commonly used reference.

Algal tests with soil suspensions and elutriates: a comparative evaluation for PAH-contaminated soils.

An algal growth inhibition test procedure with soil suspensions is proposed and evaluated for PAH-contaminated soil. The growth rate reduction of the standard freshwater green alga Pseudokirchneriella subcapitata (formerly known as Selenastrum capricornutum) was used as the toxicity endpoint, and was quantified by measuring the fluorescence of solvent-extracted algal pigments. No growth rate reduction was detected for soil contents up to 20 g/l testing five non-contaminated Danish soils. Comparative testing with PAH-contaminated soil elutriates and soil suspensions showed that the suspensions had toxicity endpoints 2.5-3000 times lower than tests with the corresponding elutriates. Algal growth inhibition tests with soil suspensions are recommended for screening purposes as a supplement to elutriate testing. Experiments with a phenanthrene-spiked soil, showed that the sorbed compound did not contribute to the toxicity. However, the soil did act as a reservoir for phenanthrene, allowing desorption to occur continuously during the algal test which maintained higher concentrations of phenanthrene in the dissolved phase. Phenanthrene-spiked soil incubated for 90 days before algal testing, resulted in a reduction of the toxicity to P. subcapitata by a factor of 76 (from EC10 = 0.3 to 23.6 g soil/l). However, during this 90-day period the total concentration of phenanthrene in the soil decreased by 38% (from 322 to 199 mg/kg) indicating that phenanthrene in the aged soil had become less bioavailable.