Determination of the anti-proliferative activity of A. precatorius

In document OF NATURAL KILLER (NK) CELLS BY Abrus precatorius LEAVES EXTRACT ON HUMAN (halaman 114-129)

CHAPTER 4 ANTI-PROLIFERATIVE ACTIVITY AND APOPTOSIS

4.3.2 Determination of the anti-proliferative activity of A. precatorius

A. precatorius aqueous leaves extract was prepared by decoction at 50°C.

Initially the cells were treated with concentration as listed in Table 4.1 (page 88).

However, no IC50 value was obtained even the cells were treated at the maximum concentration of 99µg/ml. Therefore, the aqueous extract concentration was increased to 990µg/ml. It was performed in the same manner as explained in section 4.2.2. A log concentration vs cell viability was also plotted. Figure 4.1 showed the representative points of each concentration, presented in means with ±SD of three independent experiments. A. precatorius aqueous extract demonstrated a null toxicity against all cells, according to the standard criteria used by The US National Cancer Institute and Geran Protocol.

Figure 4.1: Anti-proliferative activity of A. precatorius aqueous leaves extracts on selected cancer and normal cells.

The results were expressed as mean, ±SD of three independent experiments with three replicates

0 20 40 60 80 100 120

990 495

247.5 128.8

61.9 30.9

15.5 7.7

3.9

Cell viability (%)

Cocentration (µg/ml)

HeLa MCF-7 MDA-MB-231 SW 480 MCF10A NIH(3T3)

4.3.3 Determination of the anti-proliferative activity of A. precatorius solvents extract (Soxhlet) on selected normal and cancer cells

There were two types of extraction methods used for A. precatorius leaves.

Both methods applied solvents in a successive manner. Anti-proliferative activities of A. precatotius successive Soxhlet hexane-, ethyl acetate- and methanol- leaves extracts are shown in Figure 4.2 -4.7.

4.3.3(a) HeLa

Figure 4.2: Anti-proliferative activity of A. precatorius successive Soxhlet hexane-, ethyl acetate- and methanol- leaves extracts on HeLa cells.

The only IC50 values obtained were for methanol extract was 73.6µg/ml and 4.32 µg/ml for Tamoxifen, while the rest of the extracts has the IC50

values of >99 µg/ml. The results were expressed as mean, ±SD of three independent experiments with three replicates.

0

4.3.3(b) MCF7

Figure 4.3: Anti-proliferative activity of A. precatorius successive Soxhlet

hexane-, ethyl acetate- and methanol- leaves extracts on MCF7 cells.

The IC50 values obtained for hexane extract was 52.65µg/ml, ethyl acetate extract was 99µg/ml and methanol extract was 59.03µg/ml and 1.81µg/ml for Tamoxifen. The results were expressed as mean, ±SD of three independent experiments with three replicates.

0

4.3.3(c) MDA-MB-231

Figure 4.4: Anti-proliferative activity of A. precatorius successive Soxhlet hexane-, ethyl acetate- and methanol- leaves extracts on MDA MB-231 cells.

The IC50 obtained for hexane, ethyl acetate, methanol extract and tamoxifen were 45.60µg/ml, 54.5µg/ml, 26.4µg/ml and 2.27µg/ml, respectively. The results were expressed as mean, ±SD of three independent experiments with three replicates

0

4.3.3(d) SW 480

Figure 4.5: Anti-proliferative activity of A. precatorius successive Soxhlet

hexane-, ethyl acetate- and methanol- leaves extracts on SW 480 cells.

The IC50 obtained for methanol extract is 77.23µg/ml and 2.31µg/ml for Tamoxifen, while the rest of the extracts demonstrated the IC50 values of

>99µg/ml. The results were expressed as mean, ±SD of three independent experiments with three replicates

4.3.3(e) NIH(3T3)

Figure 4.6: Anti-proliferative activity of A. precatorius successive Soxhlet hexane-, ethyl acetate- and methanol- leaves extracts on NIH(3T3) cells.

4.3.3(f) MCF10A

Figure 4.7: Anti-proliferative activity of A. precatorius successive Soxhlet hexane-, ethyl acetate- and methanol- leaves extracts on MCF10A cells.

The IC50 obtained for Tamoxifen is 2.78µg/ml, while the rest of the extracts demonstrated the IC50 values of >99 µg/ml. The results were expressed as mean, ±SD of three independent experiments with three replicates.

0 20 40 60 80 100

99 49.5

24.8 12.4

6.19 3.09

1.55 0.73

0.39

Cell viability (%)

Concentration (µg/ml)

Hexane Ethyl acetate Methanol Tamoxifen

4.3.4 Determination of the anti-proliferative activity of A. precatorius solvents extracts (Maceration) on selected normal and cancer cells

Another method used for A. precatorius leaves extraction was by maceration.

The ground leaves were soaked with no heat successively with different solvents following their polarity. Initially the concentration of the extracts used were as stated in Table 4.1. However, no IC50 was recorded in all extracts even at the maximum concentration used. Therefore, the extracts concentration was increased to the maximum of 495µg/ml. Anti-proliferative activities of A. precatotius successive (maceration) hexane-, ethyl acetate- and methanol- leaves extracts are shown in Figure 4.8 – 4.13.

4.3.4(a) HeLa

Figure 4.8: Anti-proliferative activity of A. precatorius successive (maceration) hexane-, ethyl acetate- and methanol- leaves extracts on HeLa cells.

The IC50 obtained for hexane extract was 325µg/ml, ethyl acetate extract was 371µg/ml and methanol extract was 352 µg/ml. The results were expressed as mean, ±SD of three independent experiments with three replicates.

0 20 40 60 80 100 120

495 247.5

128.8 61.9

30.9 15.5

7.7 3.9

1.9

Cell Viability (%)

Concentration (µg/ml) Hexane Ethyl acetate Methanol

4.3.4(b) MCF-7

Figure 4.9: Anti-proliferative activity of A. precatorius successive (maceration) hexane-, ethyl acetate- and methanol- leaves extracts on MCF-7 cells.

The IC50 obtained for hexane extract was 672µg/ml and methanol extract was 423µg/ml. While ethyl acetate extract was >495µg/ml. The results were expressed as mean, ±SD of three independent experiments with three replicates.

0 20 40 60 80 100 120

495 247.5

128.8 61.9

30.9 15.5

7.7 3.9

1.9

Cell Viability (%)

Concentration (µg/ml) Hexane Ethyl acetate Methanol

4.3.4(c) MDA-MB-231

Figure 4.10: Anti-proliferative activity of A. precatorius successive (maceration) hexane-, ethyl acetate- and methanol- leaves extracts on MDA-MB-231 cells.

4.3.4(d) SW480

Figure 4.11: Anti-proliferative activity of A. precatorius successive (maceration) hexane-, ethyl acetate- and methanol- leaves extracts on SW480 cells.

The IC50 obtained for hexane extract was 301.3µg/ml, ethyl acetate extract was 447.5µg/ml and methanol 350.3was µg/ml. The results were expressed as mean, ±SD of three independent experiments with three replicates.

0 20 40 60 80 100 120

495 247.5

128.8 61.9

30.9 15.5

7.7 3.9

1.9

Cell Viability (%)

Concentration (µg/ml)

Hexane Ethyl acetate Methanol

4.3.4(e) NIH (3T3)

Figure 4.12: Anti-proliferative activity of A. precatorius successive (maceration) hexane-, ethyl acetate- and methanol- leaves extracts on NIH(3T3) cells.

No IC50 was obtained even at the maximum concentration of 495µg/ml.

The results were expressed as mean, ±SD of three independent

4.3.4(f) MCF10A

Figure 4.13: Anti-proliferative activity of A. precatorius successive (maceration) hexane-, ethyl acetate- and methanol- leaves extracts on MCF10A cells.

No IC50 was obtained even at the maximum concentration of 495µg/ml.

The results were expressed as mean, ±SD of three independent experiments with three replicates.

0 20 40 60 80 100 120

495 247.5

128.8 61.9

30.9 15.5

7.7 3.9

1.9

Cell viability (%)

Concentration (µg/ml) Hexane Ethyl acetate Methanol

In document OF NATURAL KILLER (NK) CELLS BY Abrus precatorius LEAVES EXTRACT ON HUMAN (halaman 114-129)