## 3 Results and Discussion  
### Antitumor Property  
The fluorescence images of the U2 OS cells after culturing in the different extracts for 5 days are shown in Figure 2a-f. No dead cells were observed in the control group (Figure 2a). Few dead cells were observed in the ZK60 extract (Figure 2b). Significantly, dead cells gradually increased and live cells decreased in ZK60-La extracts with La content increasing. In addition, the cells cultured in the ZK60-La extracts exhibited an unhealthy shrinkage shape. These results indicated that ZK60-La exhibited antitumor property, and higher La content exhibited higher antitumor efficiency. The cell viabilities of the U2 OS cells were studied by CCK assay, with results shown in Figure 2g. The cell viability of the U2 OS cells after culturing in ZK60 extract for 5 days was 94.2%. As a comparison, the cell viability of the U2 OS cells cultured in ZK60-0.5 La, ZK60-1.0 La, ZK60-1.5 La, and ZK60-2.0 La extracts were 50.8%, 38.1%, 31.4%, and 25.2%, respectively. It indicated that the alloying of La inhibited the growth of U2 OS cells, and the cell inhibition rate gradually increased with La content increasing. Among these, ZK60-1.0 La extract exhibited a cell inhibition rate of 61.9%. These results were in accordance with the florescence staining test. The fluorescence images of the HEK 293 cells after culturing in the different extracts for 5 days were exhibited in Figure 3a-f. From the results of live/dead staining, almost no dead cells were observed. The morphology of the cells was in spindle-like shape after culturing in different extracts, which indicated that the ZK60-La exhibited compatibility to normal cells. The CCK8 assay was used to assess the cytocompatibility for HEK 293 cells, with results shown in Figure 3g. The cell viability of HEK 293 cells after culturing in extract of ZK60, ZK60-0.5 La, ZK60-1.0 La, ZK60-1.5 La, and ZK60-2.0 La for 5 days was 80.3%, 88.9%, 91.9%, 90.3%, and 84.4%, respectively. The viabilities of the cells cultured in ZK60-La extract were higher than 80% (grade 1, according to the Standard [39]), indicating that ZK60-La showed no toxicity to HEK 293 cells. Figure 2: U2 OS cells after culturing in extracts for 5 days and stained with live/dead assay; red represented the dead cells and green represented the live cells: (**a**) Control (**b**) ZK60, (**c**) ZK60-0.5 La, (**d**) ZK60-1.0 La, (**e**) ZK60-1.5 La, (**f**) ZK60-2.0 La; (**g**) cell viability of U2 OS cells in different extracts after culturing for 1 day, 3 days, and 5 days. Compared to normal cells, tumor cells had a high ROS level, and were more vulnerable to oxidative insults under increased intrinsic oxidative stress. It was well known that the mitochondria were the main ROS factory in the cells. Thus, the level of mitochondrial membrane potential (MMP) and that of ROS in U2 OS cells were measured. As shown in Figure 4a, the level of MMP after incubating in ZK60 extract was 95.2%, and the level of MMP decreased in the U2 OS cells with the La content increasing. Specifically, the obtained MMP in the extract of ZK60-La was reduced to 88.6% for ZK60-0.5 La, 80.2% for ZK60-1.0 La, 76.7% for ZK60-1.5 La, and 73.2% for ZK60-2.0 La, respectively. Meanwhile, the ROS levels in U2 OS cells after incubating in the different extracts are shown in Figure 4b. The ROS level after incubating in ZK60 extract was 109.2%, and the ROS level increased in the U2 OS cells with the La content increasing. Specifically, the obtained ROS level in the extract of ZK60-La was increased to 122.3% for ZK60-0.5 La, 138.5% for ZK60-1.0 La, 145.8% for ZK60-1.5 La, and 152.1% for ZK60-2.0 La, respectively. Figure 3: HEK 293 cells after culturing in extracts for 5 days and stained with live/dead assay; red represented the dead cells and green represented the live cells: (**a**) Control (**b**) ZK60, (**c**) ZK60-0.5 La, (**d**) ZK60-1.0 La, (**e**) ZK60-1.5 La, (**f**) ZK60-2.0 La; (**g**) cell viability of HEK 293 cells in different extracts after culturing for 1 day, 3 days and 5 days.