Marianne Matzke

The influence of anion species on the toxicity of 1-alkyl-3-methylimidazolium ionic liquids observed in an (eco)toxicological test battery

To enlarge the restricted knowledge about the hazard potentials of ionic liquids to man and the environment we have concentrated on analysing systematically the anion effect of six different anion moieties (Cl–, BF4–, (CF3SO2)2N–, (CF3)2N–, octylsulfate and bis(1,2-benzenediolato)borate) and the influence of the side chain length at the cation on (eco)toxicity. For our investigations, we used the flexible (eco)toxicological test battery considering aquatical and terrestrial compartments as well as different trophic levels including enzymes (acetylcholinesterase), mammalian cells (IPC-81), luminescent marine bacteria (Vibrio fischeri), limnic unicellular green algae (Scenedesmus vacuolatus), wheat (Triticum aestivum), cress (Lepidium sativum), duckweed (Lemna minor) and a soil invertebrate (the spring tail Folsomia candida). In general, the side chain effect was found consistently in all used test systems from the molecular up to the organismic level. Such a consistent response of the different test systems could not be confirmed for the tested anion moieties. Furthermore, in most of the investigated test systems the anion effects are not as distinct as the demonstrated side chain length effect. Nevertheless, for (CF3SO2)2N– a clear (eco)toxicological hazard potential is evident. Thus, the strategy to check toxicities within a flexible (eco)toxicological test battery has been proven to be effective for uncovering hazard potentials of ionic liquids.

Anion effects on the cytotoxicity of ionic liquids

Most recent investigations concerning the toxicological and ecotoxicological risk potentials of ionic liquids are predominantly focusing on the cation moieties. In this study we elucidate, whether the anion species commonly used in ionic liquids are exhibiting intrinsic cytotoxic effects and if these effects can be rationalised by thinking in terms of structure–activity relationships (T-SAR). As test system to measure the cell viability as toxicologically relevant endpoint the IPC-81 rat leukemia cell line and the WST-1 assay were employed. Our results show an anion effect in ionic liquids on cytotoxicity for 10 of 27 tested anions. For the remaining 17 anions from our test kit no significant effect was found. With respect to structure–activity relationships, lipophilicity and/or vulnerability to hydrolytic cleavage seem to be the key structural features leading to the observed anion cytotoxicity. We also conclude that the model of concentration addition may be useful to estimate the EC50 values of ionic liquids that have not been tested or even synthesised yet. This can help to design not only task specific but also inherently safer ionic liquids.

Ecotoxicity evaluation of selected sulfonamides

Sulfonamides (SAs) are a group of antibiotic drugs widely used in veterinary medicine. The contamination of the environment by these pharmaceuticals has raised concern in recent years. However, knowledge of their (eco)toxicity is still very basic and is restricted to just a few of these substances. Even though their toxicological analysis has been thoroughly performed and ecotoxicological data are available in the literature, a systematic analysis of their ecotoxicological potential has yet to be carried out. To fill this gap, 12 different SAs were chosen for detailed analysis with the focus on different bacteria as well as non-target organisms (algae and plants). A flexible (eco)toxicological test battery was used, including enzymes (acetylcholinesterase and glutathione reductase), luminescent marine bacteria (Vibrio fischeri), soil bacteria (Arthrobacter globiformis), limnic unicellular green algae (Scenedesmus vacuolatus) and duckweed (Lemna minor), in order to take into account both the aquatic and terrestrial compartments of the environment, as well as different trophic levels. It was found that SAs are not only toxic towards green algae (EC₅₀=1.54-32.25 mg L⁻¹) but have even stronger adverse effect on duckweed (EC₅₀=0.02-4.89 mg L⁻¹) than atrazine - herbicide (EC₅₀=2.59 mg L⁻¹).

Imidazolium based ionic liquids in soils: effects of the side chain length on wheat (Triticum aestivum) and cress (Lepidium sativum) as affected by different clays and organic matter

This study provides data on the behaviour and toxicity of selected imidazolium based ionic liquids in the terrestrial environment with the aim to contribute to a prospective hazard assessment. Using the plant growth inhibition assay with wheat (Triticum aestivum) and cress (Lepidium sativum) we investigated the influence of two different clay minerals (kaolinite and smectite) in varying concentrations and clay mineral mixtures as well as the influence of organic matter in varying concentrations on the toxicity of three imidazolium based ionic liquids differing in the alkyl side chain length. The obtained results were compared to the German standard soil Lufa 2.2. Overall the influence of the 2:1 layer mineral smectite on toxicity was stronger than for the 1:1 layer mineral kaolinite resulting in lower toxicities when smectite was present. Comparable results were achieved in the tests with different clay mineral mixtures. The influence of the clay minerals was substance concentration dependent and the side chain effect could not consistently be confirmed for the different soil mixtures. The 1:1 clay mineral kaolinite caused in some cases an increase in toxicity. The obtained results for the influence of organic matter on the toxicity proved to be much more consistent than for the clay minerals: here an increase in organic matter concentration always resulted in a decrease of the toxicity. Differences in plant species sensitivity could be shown, but not in a consistent manner. A site specific hazard assessment of ionic liquids should therefore take into account organic matter content, quantity and especially quality of clay minerals.

Hydrolysis of sulphonamides in aqueous solutions.

Hydrolysis is one of the most common reactions controlling abiotic degradation and is one of the main paths by which substances are degraded in the environment. Nevertheless, the available information on this process for many compounds, including sulphonamides (a group of antibiotic drugs widely used in veterinary medicine), is very limited. This is the first study investigating the hydrolytic stabilities of 12 sulphonamides, which were determined according to OECD guideline 111 (1st category reliability data on the basis of regulatory demands on data quality for the environmental risk assessment of pharmaceuticals). Hydrolysis behaviour was examined at pH values normally found in the environment. This was prefaced by a discussion of the acid-base properties of sulphonamides. All the sulphonamides tested were hydrolytically stable at pH 9.0, nine (apart from sulphadiazine, sulphachloropyridazine and sulphamethoxypyridazine) were stable in this respect at pH 7.0 and two (sulphadiazine and sulphaguanidine) at pH 4.0 (hydrolysis rate≤10%; t(0.5 (25°C))>1 year). The degradation products were identified, indicating two independent mechanisms of this process. Our results show that under typical environmental conditions (pH and temperature) sulphonamides are hydrolytically stable with a long half-life; they thus contribute to the on-going assessment of their environmental fate.

Ionic liquids in soils: effects of different anion species of imidazolium based ionic liquids on wheat (Triticum aestivum) as affected by different clay minerals and clay concentrations.

This study contributes to a prospective hazard assessment of ionic liquids, focusing on the terrestrial environment. The influence of differently composed soils—varying contents of the clay minerals smectite and kaolinite—on the toxicity of different anion species of imidazolium based ionic liquids was studied for growth inhibition of wheat (Triticum aestivum). IM14 (CF3SO2)2N appeared the most toxic, independently of the investigated soil type. The toxicity of IM14 Cl, IM14 BF4 and IM14 HSO4 was mainly dominated by the cationic moiety. The observed effects varied in dependence of the added clay type and clay concentration. An increase of clay content resulted in less pronounced effects of these substances. In contrast, for IM14 (CF3SO2)2N the addition of clay minerals caused higher toxic effects in comparison to the reference soil. Our results give first hints for the assumption, that ionic liquids whose toxic action is based on the anionic moiety are especially hazardous for soils, particular for soils with high clay contents.

Mixture effects and predictability of combination effects of imidazolium based ionic liquids as well as imidazolium based ionic liquids and cadmium on terrestrial plants (Triticum aestivum) and limnic green algae (Scenedesmus vacuolatus)

Up to now the issues of mixture toxicity and combination effects were neglected within a prospective hazard assessment of ionic liquids (ILs). However when being released to the environment, mixtures of ILs or mixtures of ILs and other pollutants are likely to occur and therefore the impact of mixture toxicity should be taken into account. Thus this study investigates the effects of three differently composed mixtures containing ILs as well as ILs and the heavy metal cadmium on limnic green algae (Scenedesmus vacuolatus) and wheat (Triticum aestivum). For an evaluation of the results, two well established concepts of concentration addition (CA) and independent action (IA) were used. Both (CA and IA) underestimated the effects of the mixtures consisting exclusively of ILs leading to the assumption that interactions between the mixture components or between the mixture components and the environmental matrix occurred. The general applicability of CA and IA must be questioned. For the mixture of ILs and cadmium the deviations from the predictions were moderate (and less toxic than expected) so that CA can be recommended as a starting point for the analysis of combination effects of ILs and heavy metals. In general the presence of cadmium reduced the toxicity for both the aquatic as well as the terrestrial organisms. The green algae reacted approximately two orders of magnitude more sensitive to the mixture scenarios than wheat and proved to be a good reference test system for the evaluation of measured effects within a prospective hazard assessment of ionic liquids.

Sorption and desorption of imidazolium based ionic liquids in different soil types.

This study investigates the influence of the two different clay minerals kaolinite and smectite as well as of organic matter on the cation sorption and desorption behaviour of three imidazolium based ionic liquids -1-butyl-3-methyl-imidazolium tetrafluoroborate (IM14 BF(4)), 1-methyl-3-octyl-imidazolium tetrafluoroborate (IM18 BF(4)) and 1-butyl-3-methyl-imidazolium bis[(trifluoromethyl)sulfonyl]imide (IM14 (CF(3)SO(2))(2)N) - in soil. The German standard soil Lufa 2.2 - a natural soil classified as a loamy sand - was the basis substrate for the different soil compositions and also served as a reference soil. The addition of organic matter and clays increases the sorption of the substances and in particular smectite had striking effects on the sorption capacity for all three ionic liquids indicating that ionic interactions play an important role for sorption and desorption processes of ionic liquids in soil. One exception was for kaolinite-containing soils and the IM14 cation: with (CF(3)SO(2))(2)N(-) as an anion the sorption was identical at either 10 wt% or 15 wt% clay content, and with BF(4)(-) sorption was even lower at 15 wt% kaolinite than at 10 wt%. Desorption was weak for IM18 BF(4), presumably owing to the longer alkyl side chain. With regard to the influence of kaolinite on desorption, the same pattern was observed as it was found for the sorption of IM14 BF(4) and IM14 (CF(3)SO(2))(2)N.

Strategy to improve the characterization of chitosan for sustainable biomedical applications: SAR guided multi-dimensional analysis†

The biopolymer chitosan has shown great potential for a tremendous number of applications despite the fact that typical chitosan preparations are always mixtures of different chemical entities, natural impurities and process-induced impurities. However, chitosan preparations described in the literature or offered on the market are analytically highly undefined. Here we propose a T-SAR (thinking in terms of structure-activity-relationships) guided multi-dimensional analysis of distinct chitosan preparations with the aim a) to obtain the information needed for the production of reproducible chitosan preparations and b) to predict biological effects and technological properties of certain chitosan preparations. First, a physico-chemical description (molecular weight (MW), polydispersity (MW/MN), fraction of acetylation (FA), pattern of acetylation (PA), hydrodynamic radius (Rh), intrinsic viscosity ([η])) of six selected samples was done. Furthermore chitosan properties like solubility, crystallinity, conformation (Mark-Houwink-plot) and impurities of all the chitosan preparations from different origins were determined and biological effects were also analyzed using test systems with two different bacteria (Escherichia coli, Vibrio fischeri). It was found that the presence of HCl enabled the water solubility of chitosan, while chloride-free chitosan was only soluble in acetic acid. The pattern of acetylation PA showed no impact on this behavior. The analyzed biological effects revealed growth inhibition within 30 minutes for E. coli and a decreased bioluminescence for V. fisheri (IC50 = 0.035 w%). Thus, the strategy to check biological effects within a multi-dimensional analysis kit proved to be effective for detecting general structure-property-relationships of chitosan in relation to its biological effects.

Metalloproteins and phytochelatin synthase may confer protection against zinc oxide nanoparticle induced toxicity in Caenorhabditis elegans

Zinc oxide nanoparticles (ZnONPs) are used in large quantities by the cosmetic, food and textile industries. Here we exposed Caenorhabditis elegans wild-type and a metal sensitive triple knockout mutant (mtl-1;mtl-2;pcs-1) to ZnONPs (0-50mg/L) to study strain and exposure specific effects on transcription, reactive oxygen species generation, the biomolecular phenotype (measured by Raman microspectroscopy) and key endpoints of the nematode life cycle (growth, reproduction and lifespan). A significant dissolution effect was observed, where dissolved ZnO constituted over 50% of total Zn within a two day exposure to the test medium, suggesting that the nominal exposure to pure ZnONPs represents in vivo, at best, a mixture exposure of ionic zinc and nanoparticles. Nevertheless, the analyses provided evidence that the metallothioneins (mtl-1 and mtl-2), the phytochelatin synthase (pcs-1) and an apoptotic marker (cep-1) were transcriptionally activated. In addition, the DCFH-DA assay provided in vitro evidence of the oxidative potential of ZnONPs in the metal exposure sensitive triple mutant. Raman spectroscopy highlighted that the biomolecular phenotype changes significantly in the mtl-1;mtl-2;pcs-1 triple knockout worm upon ZnONP exposure, suggesting that these metalloproteins are instrumental in the protection against cytotoxic damage. Finally, ZnONP exposure was shown to decrease growth and development, reproductive capacity and lifespan, effects which were amplified in the triple knockout. By combining diverse toxicological strategies, we identified that individuals (genotypes) housing mutations in key metalloproteins and phytochelatin synthase are more susceptible to ZnONP exposure, which underlines their importance to minimize ZnONP induced toxicity.