## 2 Materials and Methods
### Course of the Experiment
#### 2.2.3 Soil Toxicity Analysis  
The toxicity of soils at baseline and after completion of the bioaugmentation-assisted phytoremediation process carried out in the studied systems was evaluated using 4 commercially available microbiostets: Phytotoxkit(tm) (MicroBioTests Inc., Gent Belgium), Ostracodotoxit(tm) (MicroBioTests Inc., Gent, Belgium), Microtox(r) Solid Phase Test (Modern Water Inc., New Castle, DE, USA) and MARA (NCIMB Ltd., Aberdeen, UK) [65]. Phytotoxkit(tm) and Ostracodotoxit(tm) tests are among the chronic toxicity assessment tests. The Phytotoxkit(tm) test is based on evaluating the germination and early growth of test plants, which include _Sorghum saccharatum_, _Lepidium sativum_ and _Sinapis alba_. The result of the test is inhibition of germination and early root growth after three days of incubation at 25 \({}^{\circ}\)C. The Ostracodotoxit(tm) test is based on assessing the percentage mortality and growth inhibition of crustaceans of the species _Heterocypris incongruruens_ after 6 days of contact with the test soil. The Microtox(r) Solid Phase Test is an acute toxicity test and involves evaluating the decrease in fluorescence of _Vibrio fischeri_ bacteria following con \begin{table}
\begin{tabular}{c c c c c} \hline \multicolumn{3}{c}{**Chromatographic Analysis of Aliphatic Hydrocarbons—TPH, nC\({}_{\text{c}}\)=nC\({}_{\text{4}}\), Pristane, Phytane (Clarus 500 GC Perkin Elmer)**} \\ \multicolumn{3}{c}{**Parameter of Chromatography**} \\ \hline \multicolumn{3}{c}{**Temperature (\({}^{\circ}\)C)**} \\ \hline
**Injector** & **Carrier Gas** & **Temperature Program** & **Column** & **Detector** & **Calibration Standards** \\ \hline \multicolumn{3}{c}{} & & & Reference soil: \\  & & & BAM—K010 (Tusnovic Instruments, Poland) \\  & & 30 \({}^{\circ}\)C—isothermal run & & Instruments, Poland \\  & for 2 min & & Standard mixture \\  & & 30–105 \({}^{\circ}\)C—temp. & RTX-1 & & hydrocarbons (nC\({}_{\text{c}}\)=nC\({}_{\text{4}}\)) \\ \multicolumn{3}{c}{} & He & increase rate 10 \({}^{\circ}\)C min\({}^{-1}\) & 30 m \(\times\) 0.53 mm & FID \\
290 \({}^{\circ}\)C & 20 mL min\({}^{-1}\) & 105–285 \({}^{\circ}\)C—temp. & (Restek, Bellefonte, & 300 \({}^{\circ}\)C \\  & & increase rate 5 \({}^{\circ}\)C min\({}^{-1}\) & PA, USA) & Fuel Oil Degradation Mix \\  & & 285 \({}^{\circ}\)C—isothermal run & & nc\({}_{\text{17}}\), pristane, nC\({}_{\text{18}}\), \\  & & for 5 min\({}^{-1}\) & & phytane No. A029668 (Restek, Bellefonte, PA, USA) \\ \hline \multicolumn{3}{c}{**Chromatographic Analysis of Polycyclic Aromatic Hydrocarbons—PAH (Vanquish Core Thermo Scientific)**} \\ \multicolumn{3}{c}{**Parameter of Chromatography**} \\ \hline
**Eluents** & **Flow** & **Gradient** & **Column** & **Detector** & **Calibration Standards** \\ \hline \multicolumn{3}{c}{} & 20\% B—for 1.5 min & NUCLEODUR C\({}_{\text{18}}\) & Reference soil: PAHs by HPLC No. SQC017-40G \\ \multicolumn{3}{c}{} & 20–50\% B—for 1.5 min & PAH column & (Sigma-Aldrich, Saint \\ \multicolumn{3}{c}{} & \multicolumn{3}{c}{} & 50–100\% B—for 1 min & 125 mm \(\times\) 4 mm, & UV-ViS \\ \multicolumn{3}{c}{} & \multicolumn{3}{c}{} & 100\% B—for 1 min & 3 μm & FLD \\ \multicolumn{3}{c}{} & \multicolumn{3}{c}{} & 100\% B—for 3 min & (Marchey-Nagel, \\ \multicolumn{3}{c}{} & \multicolumn{3}{c}{} & 100\% A—for 3 min & Dueren, Germany) & \\ \hline \end{tabular}
\end{table} 
Table 2: Operating parameters of chromatographs during the determination of TPHs and PAHs. tact with the test sample. The result of the Microtox(r) Solid Phase test is EC\({}_{50}\), which is the concentration of the test sample, which produces 50% of the test reaction.