## 3 Results  
### Cytotoxicity of Hydrolyzed Riceberry Rice Bran Extract on Colon Cancer Cell Lines  
For the aqueous extraction of hydrolyzed riceberry rice barn extract (HRBE), the percentage yield was reached at 85.94%. To ensure the cytotoxicity levels on cancer and normal cells, an MTT assay was performed on a primary human dermal fibroblast neonatal cell line (PCS-201-010), a non-metastatic human colon cancer cell line (HT-29) and a metastatic human colon cancer cell line (SW-620) after they were treated with the different concentrations of HRBE (0-10,000 \(\upmu\)g/mL) for 24 h, 48 h and 72 h. For the positive control, different concentrations of mitomycin C (0-100 \(\upmu\)g/mL) were used (Table 1).  
According to the MTT assay results on cancer cell lines, a significant decrease in cell viability with a dose- and time-dependent manner was observed after they were treated with HRBE. The half-maximal inhibitory concentrations (IC\({}_{50}\)) of HRBE on HT-29 of 43,677, 11,164 and 6053 \(\upmu\)g/mL, and on SW-620 of 9880, 6870 and 5468 \(\upmu\)g/mL were indicated at 24 h, 48 h and 72 h, respectively. For the effect of HRBE on the fibroblast normal human cell line (PCS-201-010), the IC\({}_{50}\) concentrations were 15,352, 8007 and 6745 \(\upmu\)g/mL, respectively, (Table 1 and Figure 1), which were significantly higher than those of cancer cell lines (the responsive curve patterns can be seen in Figure S1 (Supplementary Materials)).  
\begin{table}
\begin{tabular}{c c c c c c} \hline
**Time/Treatment** & & & & **IC\({}_{50}\) (ug/mL)** & \\ \hline \multicolumn{5}{c}{**HT-29**} & **SW-620** & **PCS-201-010** \\ \hline \multicolumn{5}{c}{HRBE} & Mitomycin C & HRBE & Mitomycin C & HRBE & Mitomycin C \\
24 h & 43,677.08 \(\pm\) 30,191.29 & 762.91 \(\pm\) 650.87 & 9879.66 \(\pm\) 1855.29 & 192.93 \(\pm\) 90.21 & 15,351.45 \(\pm\) 7954.70 & 209.78 \(\pm\) 78.02 \\
48 h & 11,164.35 \(\pm\) 1642.52 & 141.26 \  
Figure 1: Cytotoxicity test by MTT assay on normal human fibroblast cell line (PCS-201-010), nonmetastatic colon cancer cell line (HT-29) and metastatic colon cancer cell line (SW-620) after being treated with HRBE concentrations (0–10,000 \(\mathrm{\SIUnitSymbolMicro m}\)/mL) for 24 h (**A**), 48 h (**B**) and 72 h (**C**). The untreated cells (0 mg/mL) were considered as negative control. The percentage of cell viability was determined as compared to an untreated condition. Data are represented by the mean \(\pm\) SD. Statistical analysis was performed using one-way ANOVA (*, \(p\leq 0.05\) significant; **, \(p\leq 0.01\) significant).  
For the positive control treated with mitomycin C on HT-29, SW-620 and PCS-201-010, IC\({}_{50}\) concentrations were indicated for HT-29 as 763, 141 and 57 \(\upmu\)g/mL, for SW-620 as 193, 58 and 40 \(\upmu\)g/mL and for PCS-201-010 as 210, 177 and 145 \(\upmu\)g/mL (Figure 2) at 24 h, 48 h and 72 h, respectively. The result showed that the HRBE had less effect on normal cells than the two types of cancer cells and the IC\({}_{50}\) concentrations decreased in a dose-time-dependent manner (the responsive curve patterns can be seen in Figure S2). Since the lowest IC\({}_{50}\) value was observed with the treated SW-620 cell line at 72 h, this metastatic human colon cancer cell line was chosen for the apoptotic inductive efficiency determination of HRBE in the next experiment.  
Figure 2: Cytotoxicity test by MTT assay on normal human fibroblast cell line (PCS-201-010), non-metastatic colon cancer cell line (HT-29) and metastatic colon cancer cell line (SW-620) after being treated with mitomycin C (as the positive control) concentrations (0–100 \(\upmu\)g/mL) for 24 h (**A**), 48 h (**B**) and 72 h (**C**). The percentage of cell viability was determined as compared to an untreated condition. Data are represented by the mean \(\pm\) SD. Statistical analysis was performed using one-way ANOVA (*, \(p\leq\) 0.05 significant; *, \(p\leq\) 0.01 significant).