## 3 Results and Discussion
### Acute Toxicity Assays
#### 3.3.1 Growth Inhibition  
EC\({}_{50}\) (72 h) values were significantly lower for _R. subcapitata_ than for _T. obliquus_: the values range from 0.28 to 0.47 mg L\({}^{-1}\) for the former and 1.76-2.68 mg L\({}^{-1}\) for the latter (Table 4). This agrees with an observation by [PERSON] et al. [13], that thioglycolate-based ILs [N\({}_{1888}\)][C\({}_{6}\)SAc] and [P\({}_{1888}\)][C\({}_{6}\)SAc] as well as commercially available ILs [N\({}_{1888}\)][Cl] and [P\({}_{66614}\)][Cl] exhibited higher values for _T. obliquus_. This was attributed to the inert character of the biological polymer sporopolenin, covering the cell walls of _T. obliquus_, thus reducing diffusion into the cells. In further comparing our results to [13], our EC\({}_{50}\) values were significantly higher for _R. subcapitata_ and, additionally, the chosen anion had a clear effect on acute toxicity. Moreover, a comparison of three different anions is possible for cation [N\({}_{1888}\)], with acute toxicity decreasing in the order [C\({}_{6}\)SAc]\({}^{-}>\)[Cl]\({}^{-}>\)[HNA]\({}^{-}\). For the latter, a value of 0.28 \(\pm\) 0.01 mg L\({}^{-1}\) signifies a decreased toxicity by a factor of 4 compared with [Cl]\({}^{-}\) and by a factor of 5.6 for [C\({}_{6}\)SAc]. The same holds true for [P\({}_{66614}\)][Cl] and [P\({}_{66614}\)][HNA] and is highly pronounced for [P\({}_{1888}\)][C\({}_{6}\)SAc] and [P\({}_{1888}\)][HNA], with EC\({}_{50}\) values of 0.04 \(\pm\) 0.01 and 0.39 \(\pm\) 0.05 mg L\({}^{-1}\), respectively. The chloride anion was proposed as a reference point in determining anion toxicity because no effect of chloride on the overall toxicity of the compound is assumed [45]. Accordingly, [HNA]\({}^{-}\) has a positive effect on IL acute toxicity. The exact interaction with cells is unclear, but recent work revealed a positive influence of 3-hydroxy-2-naphthoate on cell activity [46], which indicates a generally lower toxicity of [HNA]\({}^{-}\) compared to [C\({}_{6}\)SAc]\({}^{-}\). Whereas no effect of alkyl chain length could be detected for growth of _R. subcapitata_, [P\({}_{66614}\)][HNA] exhibited a slightly higher toxicity compared to the less hydrophobic cations [P\({}_{1888}\)]\({}^{+}\) and [N\({}_{1888}\)]\({}^{+}\) for _T. obliquus_. This agrees with the literature on the alkyl-length-dependent toxicity of imidazolium-based ILs on _T. obliquus_[47]. EC\({}_{50}\) values (72 h) for ILs with alkyl chains \(\geq\) C\({}_{10}\) and chloride anion were below 0.06 mg L\({}^{-1}\)[48] and below 0.11 mg L\({}^{-1}\) in the case of bromide [47], which indicates a higher acute toxicity compared to the ILs we tested. Likewise, [49] reported an EC\({}_{50}\) value (72 h) of 0.06 mg L\({}^{-1}\) for 1-decylpyridinium bromide. Comparing cations with the same alkyl chains revealed no influence of the central atom on toxicity, yielding similar results for [P\({}_{1888}\)][HNA] and [N\({}_{1888}\)][HNA] towards both _T. obliquus_ and _R. subcapitata_. As is frequently suggested, predicting the toxicity of ILs based on chemical structures is complicated by the distinct reactions of different test organisms. Moreover, [PERSON] et al. [45] described overadditive effects on toxicity by specific cation-anion combinations. Although the three ILs we tested here displayed a lower acute toxicity than similar compounds in previous studies, it is important to emphasize that they nevertheless are classified as \"acute toxic 1\" (EC\({}_{50}\)\(\leq\) 1 mg L\({}^{-1}\)) towards _R. subcapitata_ and as \"acute toxic 2\" (1 mg L\({}^{-1}\)\(\leq\) EC\({}_{50}\)\(\leq\) 10 mg L\({}^{-1}\)) towards _T. obliquus_ for an exposure time of 72 h in the _Globally Harmonized System of the Classification and Labelling of Chemicals_ (GHS) [50]. As such, their potential application must be further reviewed in greater detail, e.g., by assessing their toxicity on additional organisms or conducting studies on biodegradability.