## 2 Materials and Methods
### Sample Preparation and Analysis  
After 3 weeks growth, the plants were harvested and the above-ground (i.e., shoots) and below-ground (i.e., roots) parts were separated. The shoot and root samples were washed carefully with ultrapure water after tap water to remove any soil splash, were oven-dried at 80 \({}^{\circ}\)C for 24 h, and were then ground in a mortar. Dried plant samples (0.1 g of each sample) were digested with 10 cm\({}^{3}\) conc. HNO\({}_{3}\) (61%) and 3.0 cm\({}^{3}\) conc. HF (46%) in a microwave digestion system (Speedwave4, ACTAC. Co. LTD, Tokyo, Japan). The digested samples were diluted to 100 mL, then Cd and Zn were determined by Inductively Coupled Plasma Mass Spectrometry (ICP-MS, Thermo Fisher Scientific, X-series II, Massachusetts, USA) [29]. Replicate experiments were basically performed three times. The operating conditions of ICP-MS are shown in Supplementary Materials Table S1. The Cd and Zn standard solutions used for making the calibration curve were prepared by diluting the standard solutions (1000 mg/L in 6% HNO\({}_{3}\) solution). All of the chemical reagents used were of analytical grade. Water (>18.2 MJ), which was treated by an ultrapure water system (Advantec Aquarius: RFU 424 TA), was used throughout the work. The translocation factor from root to shoot was also calculated by dividing the metal concentration in the shoot by that in the root [30; 31].  
The surface morphology and element distribution of the roots and shoots of _Turacum officinal_ after cultivation were investigated using Scanning Electron Microscope and Energy Dispersive Spectrometer (SEM-EDS, JEOL, Akishima City, Tokyo, Japan: JCM-6000 with JED-2300).  
Extraction of Cd and Zn from Gazania after Phytoremediation: An Approach for Metal Recovery from Plant Tissues  
To realize the original purpose of our extraction studies mentioned in Introduction, extraction experiments of Cd and Zn from _Gazania_ after phytoremediation were conducted. In each extraction experiment, the spent plant was accurately weighed to 0.1 g into an Erlenmeyer flask containing 100 mL extractant solvent, and then heated in a water bath at  
Figure 1: The structure of each chelating agent: (**a**) EDDS; (**b**) HIDS; (**c**) GLDA.  
\begin{table}
\begin{tabular}{c c c c c c c c} \hline \hline
**Pot No.** & **1** & **2** & **3** & **4** & **5** & **6** & **7** & **8** \\ \hline Cd and Zn (mg/kg soil) & 6.5 & 6.5 & 6.5 & 6.5 & 6.5 & 6.5 & 6.5 & 6.5 \\ Al, Pb, Bi, and Ni & & 6.5 & 6.5 & 6.5 & 6.5 & 6.5 & 6.5 & 6.5 \\ (mg/kg soil) & & & & EDDS & EDDS & HIDS & HIDS & GLDA & GLDA \\ (mmol/kg soil) & & & 1 & 5 & 1 & 5 & 1 & 5 \\ \hline \hline \end{tabular}
\end{table}
Table 3: Conditions of the pot experiments using different kinds of chelating agents.  
60 \({}^{\circ}\)C for 1 h. Then, the mixed solution was filtered with a 0.45 \(\upmu\)m membrane filter, and the heavy metal (Cd or Zn) content in the filtrate was determined by ICP-MS.  
Effect of the Extractant Solvent on the Extraction of Cd and Zn from _Gazania_  
In order to investigate the effect of the extractant solvent on the extraction of Cd and Zn from _Gazania_, several kinds of solvents (i.e., CH\({}_{3}\)COOH, HNO\({}_{3}\), H\({}_{2}\)SO\({}_{4}\), H\({}_{3}\)PO\({}_{4}\), NH\({}_{3}\), and NaOH) at different concentrations i.e., (0.18, 0.36, 1.8, or 3.6 mol/L) were employed.  
Extraction Experiment Using Incineration Ash from _Gazania_  
The spent plant was incinerated using an electric furnace at 200, 400, and 600 \({}^{\circ}\)C. The above extraction experiment was performed using these incineration ashes. The experimental conditions were the same as that in Section 2.4.1.  
Statistics Analysis: Analysis of Variance (ANOVA)  
The one-way ANOVAs presented in Section 3.3 were performed in Excel [32] and used to determine significant differences among experimental treatments. To test data normality, we employed the chi-squared type goodness-of-fit test. In addition, [PERSON]'s test was used to check data homogeneity. Finally, Tukey-Kramer post hoc test in Excel was performed.