## 2 Materials and Methods
### Germination Index  
The potential phytotoxicity of EOMs was assessed by determining the Germination Index (GI) through water-soluble extracts using _Lactuca sativa_ (dicotyledon) and _Triticum aestivum_ (monocotyledon) seeds. Each EOM (20 g dry weight) was mixed with 240 mL of sterile deionized water using an orbital lab-shaker (Adolf Kuhner AG, Switzerland) at 125 rpm for 24 h. Subsequently, the extracts were centrifuged at 5000\(\times\)_g_ for 10 min and the supernatants were used to prepare test solutions at concentrations of 100% (pure EOM aqueous extract; only for biochar and sludge), 50%, 25%, 12.5%, 6.25%, and 0% (distilled water as negative control). The 100% aqueous extracts of digestate and manure were excluded due to their high-water adsorption capacity, making it challenging to obtain sufficient aqueous extract for testing at 100% concentration and preparing subsequent dilutions.  
Petri dishes (9 cm diameter) were prepared, each containing twenty _L. sativa_ or _T. aestivum_ seeds placed upon two sheets of Whatman N. 1 filter paper pre-treated with 10 mL of the test solution. Five replicates were prepared for a total of 100 seeds for each treatment. The dishes were transferred to a germination chamber under controlled temperature (20 \({}^{\circ}\)C) in the dark. The humidity within each Petri dish was regulated by the 10 mL of test solution added to moisten the Whatman sheets and no water was added during the incubation period to avoid diluting the test solution. After three days and one week of incubation, the number of germinated seeds in each Petri dish was counted. Seeding root elongation was measured after one week. The Germination Index (GI) was calculated by comparing the germination rate and main root length between the specific treatment and the negative control (water) [38; 39].  
\begin{table}
\begin{tabular}{c c c c c c}  & \multicolumn{1}{c}{**Cattle Manure**} & \multicolumn{1}{c}{**Digestate**} & \multicolumn{1}{c}{**Digestate**} & \multicolumn{1}{c}{**Biochar**} \\  & & **R1** & **R2** & **Biochar** \\ Total solid (TS) & \% & 23.9 & 12.5 & 12.9 & 98.3 \\ Volatile solid (VS) & \% (TS) & 91.3 & 83.5 & 85.4 & 29.8 \\ Carbon (C) & \% (TS) & 48.0 & 46.0 & 46.1 & 29.8 \\ Hydrogen (H) & \% (TS) & 5.8 & 4.8 & 5.3 & 0.3 \\ Nitrogen (N) & \% (TS) & 2.2 & 2.0 & 2.0 & 2.4 \\ Sulfur (S) & \% (TS) & 0.3 & 0.3 & 0.3 & 1.6 \\ Oxygen (O) & \% (TS) & 34.9 & 30.3 & 31.6 & \(-\)2.21 \\ Ash & \% (TS) & 8.7 & 16.5 & 14.6 & 68.1 \\ Phosphorous (P) & g/kg TS & 3.6 & 3.1 & 2.7 & 53.7 \\ Potassium (K) & g/kg TS & 15.5 & 38.7 & 38.7 & 2.2 \\ Calcium (Ca) & mg/kg TS & 6.5 & 7.7 & 8.5 & 31.3 \\ Iron (Fe) & mg/kg TS & 0.8 & 1.5 & 2.4 & 246.0 \\ Magnesium (Mg) & mg/kg TS & 4.7 & 5.3 & 5.3 & 3.6 \\ Sodium (Na) & mg/kg TS & 2.3 & 1.9 & 1.7 & 1.1 \\ Ammonium nitrogen (NH\({}_{4}\)-N) & g/kg TS & 23.0 & 8.0 & 6.20 & 0.4 \\ Total Kjeldahl nitrogen (TNK) & g/kg TS & n.d. & 23.9 & 21.69 & n.d. \\ \hline \end{tabular}  
* negative value indicates incomplete combustion (550 \({}^{\circ}\)C, 2 h) in the analysis.  
\end{table}
Table 1: Chemical characteristics of cattle manure, digestates [37] and biochar (n.d. = not determined).  
GI value equal to 100% indicates no differences in average seedling length and germination rate between a specific treatment and control. GI < 80% or >120% suggest inhibitory or bio-stimulatory effects of the specific treatment, respectively [6,38].