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Title (Primary) A fluorescence-based bioassay for aquatic macrophytes and its suitability for effect analysis of non-photosystem II inhibitors
Author Küster, A.; Pohl, K.; Altenburger, R.;
Journal Environmental Science and Pollution Research
Year 2007
Department BIOTOX;
Volume 14
Issue 6
Language englisch;
Keywords aquatic macrophytes; bioassay; Chlorophyll fluorescence; high-throughput; Imaging-PAM; macrophytes; phytotoxicity; toxicity assessment
Abstract Background, Aim and Scope: During the last years the miniaturization of toxicity test systems for rapid and parallel measurements of high amounts of samples has often been discussed. For unicellular algae as well as for aquatic macrophytes fluorescence-based miniaturized test systems have been introduced for photosystem II (PSII) inhibitors. Nevertheless, in order to ensure a routinely applicable high-throughput measuring device experiments with a broad range of toxicants and modes of action others than PSII inhibition are necessary. Thus, the aim of this study was to establish a fast and reproducible measuring system for non-PSII inhibitors for aquatic macrophyte species to overcome those limitations for use.Materials and Methods: A newly developed pulse-amplitude modulated chlorophyll fluorometer (IPAM) was applied as an effect detector in short-term bioassays with the aquatic macrophyte species Lemna minor. This multiwell-plate based measuring device enabled the incubation and measurement of up to 24 samples in parallel. The chemicals paraquat-dichloride, alizarine and triclosan were chosen as representatives for the toxicant groups of non-PSII herbicides, PAHs and PPCPs which are often detected in environmental waters. The IPAM was used (i) to establish and validate the sensitivity of the test system to the three non-PSII inhibitors, (ii) to compare the test systems with standardised and established biotests for aquatic macrophytes and (iii) to define necessary time scales in aquatic macrophyte testing. For validation of the fluorescence-based with growth-based results the standard growth test with L. minor (ISO/DIS 20079) was performed in parallel for each chemical.Results: The results revealed that fluorescence-based measurements with the IPAM allow rapid and parallel analysis of large amounts of aquatic macrophyte samples and of toxicant effects of the three chemicals tested. The IPAM enabled the recording of concentration-effect-curves with L. minor samples on a 24-well plate with single measurements. Fluorescence-based concentration-effect-curves could been detected for all three chemicals after 30 min incubation only. Within 6 h incubation the maximum inhibition of fluorescence showed 80-100 % effect for the chemicals tested. The EC50 after 24 h incubation were 0.07 mg/L, 0.84 mg/L and 2.61 mg/L for paraquat-dichloride, alizarine and triclosan, respectively.Discussion: The results obtained with the IPAM after 24 h for the herbicide paraquat-dichloride and the polycyclic aromatic hydrocarbon alizarine could been shown to be in good accordance with median effective concentrations (EC50s) obtained by the standardised growth test for L. minor after 7 d incubation (0.09 mg/L and 0.79 mg/L for paraquat-dichloride and alizarine, respectively). Those results were in accordance with literature findings for the two chemicals. In contrast, fluorescence-based EC50 of the antimicrobial agent triclosan pointed out to be two orders of magnitude higher compared to the standard growth test with 7 d incubation time (0.026 mg/L) as well as to literature findings.Conclusions: Typically, aquatic macrophyte testing is very time consuming and relies on laborious experimental set-ups. The I-PAM measuring device enabled fast effect screening for the three chemicals tested. While established test systems for aquatic macrophytes need incubation times of ? 7 d, the I-PAM can detect inhibitory effects much earlier (24 h), even if inhibition of chemicals is not specifically associated with PSII. Thus, the fluorescence-based bioassay with the I-PAM offers a promising approach for the miniaturization and high-throughput testing of chemicals with aquatic macrophytes. For the chemical triclosan, however, the short-term effect prediction with the I-PAM has been shown to be less sensitive than with long-term bioassays, which might be due to physicochemical substance properties such as lipophilicity.Recommendations and Perspectives: The results of this study are promising steps to establish and validate a fast fluorescence-based bioassay for aquatic macrophytes. The chemicals paraquat-dichloride, alizarine and triclosan were chosen as representatives for the toxicant groups of non-PSII herbicides, PAHs and PPCPs which are often detected in environmental waters. Nevertheless, in order to ensure a routinely applicable measuring device experiments with an even broader range of toxicants and samples of superficial and/or waste waters are necessary.
ID 1982
Persistent UFZ Identifier https://www.ufz.de/index.php?en=20939&ufzPublicationIdentifier=1982
Küster, A., Pohl, K., Altenburger, R. (2007):
A fluorescence-based bioassay for aquatic macrophytes and its suitability for effect analysis of non-photosystem II inhibitors
Environ. Sci. Pollut. Res. 14 (6), 377 - 383