The antiproliferation effects of different broccoli parts were tested against A549 - lung cancer cell lines. Percentage of viable cells (estimated cell growth and cell death along three days of treatment) was obtained by trypan blue dye exclusion assay. The cytotoxicity activity was characterized by conducting cell viability assay carried out by using MTT assay. As can be seen from Figure 27 (a) that the extracts of broccoli seed and sprout exhibited high antiproliferative capacity against the tested cell lines when compared with the extracts of broccoli floret and leave.
The data also indicated the effect of broccoli extracts on the A549 cells at different concentrations (50 - 500 mg/L) showed the decrease of cell viability from nearly 70% - 40%. The increasing of the concentration of extracts almost completely blocked the growth A549 cells. Therefore, an IC50 values were calculated as the concentrations that show 50% inhibition of proliferation on any tested cell line. IC50 values were reported as the average of three replicates in the Figure 27 (b), ranged at approximately 380, 300, 100 and 70 mg/L for four group samples including floret, leave, seed and sprout, respectively.
As can be considered from the data that broccoli sample had the antiproliferative effects on the A549 lung tumor cell line. It was consistent with previous researches which demonstrated the anti-cancer activity of broccoli extracts (Chaudhary et al. 2012; Gawlik-Dziki et al. 2012).
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(a)
(b)
Figure 27. Different concentrations of dried different broccoli part‟s extracts on A549 cell viability by MTT method (a) and IC 50 value (b). Data with different superscripts are significantly different at P<0.05 (n=3) analyzed by ANOVA with Duncan‟s post hoc using SPSS 17. In which, a-b were done in different parts - same concentration; and v-x-y were done in same part - different concentrations.
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4.3.2. Proliferation on HepG2 cell line
Similar to A549 cell line, the antiproliferation activities were also investigated in vitro using HepG2 – human liver hepatocellular carcinoma cell line. Under those experimental conditions, a dose-dependent decrease in cell proliferation was observed with treatment of broccoli extract (Figure 28).
Percentage of viable cells was obtained by MTT test. The Figure 28 (a) presented clear difference was detected between the extracts of broccoli samples. Seed and sprout groups expressed high antiproliferation against the HepG2 cell line in the comparison with the extracts of broccoli floret and leave samples. The results also determined the effect of broccoli extracts on the HepG2 cells at different concentrations (50 - 500 mg/L) showed the decrease of viable cell from approximately 80% - 40%. The increasing of the concentration of extracts prevented the rising of HepG2 cells. For instance, at the highest concentration (500 mg/L) induced cell death at nearly 50, 40, 55 and 60% for floret, leave, seed and sprout, respectively.
An IC50 value denoted the concentration of sample that show 50%
inhibition of proliferation on any tested cell line, was calculated and displayed in the Figure 28 (b) with a rage of values around 480, 330 and 300 mg/L for three group samples including floret, seed and sprout, respectively. In this analysis, IC50 value of leave group could not calculate due to the number of viable cells after treatment by leave extract made up to approximately 60 – 80%. The IC50 data presented that to inhibit 50% of proliferation on HepG2 cell line required higher extract concentration than several previous studies which used different tumor cell lines such as Caco-2, AT-2 or MAT-Lylu (Gawlik-Dziki et al. 2012; Domínguez-Perles et al. 2012).
Likewise for last analysis on A549 cell line, the result of this examination also estimated the effects of broccoli samples on the proliferation of the HepG2 - liver tumor cell line. It was appropriate to several previous studies which exhibited the anti-cancer capacity of broccoli extracts (Gawlik- Dziki et al. 2012; Chaudhary et al. 2012; Guo et al. 2011).
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Figure 28. Different concentrations of dried different broccoli part‟s extracts on HepG2 cell viability by MTT method (a) and IC 50 value (b). Data with different superscripts are significantly different at P<0.05 (n=3) analyzed by ANOVA with Duncan‟s post hoc using SPSS 17. In which, a-b were done in different parts - same concentration; and v-x-y were done in same part - different concentrations.
(a)
(b)
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4.3.3. Proliferation on FL83B cell line
As shown in Figure 29, dried broccoli extracts caused increase in the number of cell viability up to from around 120% - 130% cell viability at the concentration 50 and 150 àg/L extracts (from 100% cell viability in control) in FL83B mouse liver cell. Nevertheless, their effects were found to be relatively reduced (under 100 – 110 % of viable cell) at higher concentrations (250 and 500 àg/L). It also can be seen from the figure that among four different broccoli parts, seed and sprout groups had the inconsiderable higher number of cell viability than floret and leave groups. In the 100-àg/L concentration, the highest cell viability of broccoli parts were expressed with a range of 118.04 ± 6.7, 117.9 ± 5.1, 127.2 ± 6.2 and 119 ± 8.5% for floret, leave, seed and sprout, respectively. The obtained results determined that cell viability of broccoli extracts were significant higher than that of control (100%) at all of concentrations except 500-àg/L concentration. From these results, it can be considered that broccoli extracts demonstrated no toxicity to FL83B normal liver cell line even at high dose in vitro.
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Figure 29. Different concentrations of dried different broccoli part‟s extracts on FL83B cell viability by MTT method (a) and IC 50 value (b). Data with different superscripts are significantly different at P<0.05 (n=3) analyzed by ANOVA with Duncan‟s post hoc using SPSS 17. In which, a-b-c were done in different parts - same concentration; and v-x were done in same part - different concentrations.
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