Efek Kombinasi Ekstrak Aktif Daun Poguntano (Picria Fel-Terrae Lour.) Dengan Doxorubicin Terhadap Sel Kanker Payudara Secara In Vitro
Lampiran 1. Hasil identifikasi Poguntano (Picria fel-terrae Lour.)
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Lampiran 2. Gambar daun poguntano (Picria Fel-terrae Lour.)
a
b
Keterangan:
a. Gambar daun poguntano poguntano
b. Gambar simplisia daun poguntano
93
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Lampiran 3. Hasil Pemeriksaan Mikroskopik Serbuk Simplisia Daun
Poguntano
a
b
c
d
e
f
g
Keterangan :
Perbesaran 10 x 40
a. Trikoma
b. Sel epidermis
c. Kristal kalsium oksalat bentuk prisma
d. Stomata tipe diasitik
e. Stomata tipe anomositik
f. Berkas pembuluh angkut bentuk spiral
g. Sel tetangga
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Lampiran 4. Bagan skrining fitokimia, karakterisasi simplisia dan ekstrak
Daun poguntano
Dicuci
Ditiriskan
Dikeringkan (diangin-anginkan)
Simplisia
Dihaluskan
Serbuk Simplisia
Skrining
Fitokimia
Karakterisasi
Simplisia
Pembuatan
Ekstrak
Ekstrak
- Alkaloid
- Flavonoid
- Saponin
- Tanin
- Glikosida
- Antrakuinon
- Steroid/Triterpenoid
- Pemeriksaan mikroskopik
- Pemeriksaan makroskopik
- Penetapan kadar air
- Penetapan kadar sari yang larut dalam etanol
- Penetapan kadar sari yang larut dalam air
- Penetapan kadar abu total
- Penetapan kadar abu yang tidak larut dalam asam
95
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Lampiran 5. Bagan ekstraksi serbuk simplisia secara maserasi bertingkat
Serbuk simplisia
di maserasi dengan n-heksana
Ampas
Maserat
di maserasi dengan etilasetat
dipekatkan
Ampas
Maserat
di maserasi dengan etanol
Ekstrak kering n-heksana*
dipekatkan
di
skrining
Hasil
Maserat
Ampas
dipekatkan
Ekstrak kering etilasetat*
di skrining
Ekstrak kental etanol*
Hasil
di skrining
Hasil
96
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Keterangan : * = Uji Sitotoksik
Lampiran 6. Perhitungan kadar air simplisia daun poguntano (Picria Felterrae Lour.)
% Kadar air simplisia =
No.
1.
2.
3.
Berat sampel (g)
5,022
5,015
5,018
% Kadar air =
volume air (ml)
x 100%
berat sampel (g)
Volume awal (ml)
1,90
2,20
2,50
Volume akhir (ml)
2,20
2,50
2,80
volume akhir − volume awal
x 100%
berat sampel
1. Kadar air =
2,20 - 1,90
x 100% = 5,97%
5,022
2. Kadar air =
2,50 - 2,20
x 100% = 5,98%
5,015
3. Kadar air =
2,80 - 2,50
x 100% = 5,97%
5,018
% Rata-rata kadar air =
5,97% + 5,98% + 5,97%
= 5,97%
3
97
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Lampiran 7. Perhitungan kadar sari larut air simplisia daun poguntano
(Picria fel-terrae Lour.)
% Kadar sari larut dalam air =
No.
1.
2.
3.
100
berat sari (g)
x 100%
x
20
berat sampel (g)
Berat sampel (g)
4,008
4,002
4,010
Berat sari (g)
0,212
0,208
0,220
1. Kadar sari larut dalam air =
0,205 100
x
x 100% = 26,45%
4,008
20
2. Kadar sari larut dalam air =
0,208 100
x
x 100% = 25,99%
4,002
20
3. Kadar sari larut dalam air =
0,222 100
x
x 100% = 27,43%
4,010
20
% Rata-rata kadar sari larut dalam air =
26,45% + 25,99% + 27,43%
= 26,62%
3
98
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Lampiran 8. Perhitungan kadar sari larut etanol simplisia daun poguntano
(Picria fel-terrae Lour.)
% Kadar sari larut dalam etanol =
No.
1.
2.
3.
100
berat sari (g)
x 100%
x
20
berat sampel (g)
Berat sampel (g)
5,012
5,009
5,017
Berat sari (g)
0,198
0,182
0,205
1. Kadar sari larut dalam etanol
=
0,198 100
x
x 100% = 19,75%
5,012
20
2. Kadar sari larut dalam etanol
=
0,182 100
x
x 100% = 18,17%
5,009
20
3. Kadar sari larut dalam etanol
=
0,205 100
x
x 100% = 20,43%
5,017
20
% Rata-rata kadar sari larut dalam etanol =
19,75% + 20,43% + 18,17%
= 19,45%
3
99
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Lampiran 9. Perhitungan kadar abu total simplisia daun poguntano (Picria
fel-terrae Lour.)
% Kadar abu total =
berat abu (g)
x 100%
berat sampel (g)
No.
1.
2.
3.
Berat sampel (g)
2,0121
2,0118
2,0132
Berat abu (g)
0,0780
0,0791
0,0812
1. Kadar abu total =
0,0780
x 100% = 3,87%
2,0121
2. Kadar abu total =
0,0791
x 100% = 3,93%
2,0118
3. Kadar abu total =
0,0812
x 100% = 4,03%
2,0132
% Rata-rata kadar abu total =
3,87% + 3,93% + 4,03%
= 3,94%
3
100
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Lampiran 10. Perhitungan kadar abu tidak larut asam simplisia daun
poguntano (Picria fel-terrae Lour.)
% Kadar abu total =
berat abu (g)
x 100%
berat sampel (g)
No.
1.
2.
3.
Berat sampel (g)
1,9981
1,9935
2,0027
1. Kadar abu total =
0,0151
x 100% = 0,76%
1,9981
2. Kadar abu total =
0,0135
x 100% = 0,67%
1,9935
3. Kadar abu total =
0,0163
x 100% = 0,81%
2,0027
% Rata-rata kadar abu total =
Berat abu (g)
0,0151
0,0135
0,0163
0,76% + 0,67% + 0,81%
= 0,75%
3
101
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Lampiran 11. Perhitungan kadar air ekstrak n-heksana daun poguntano
(Picria fel-terrae Lour.)
% Kadar air simplisia =
No.
1.
2.
3.
volume air (ml)
x 100%
berat sampel (g)
Berat sampel (g)
2,005
2,001
2,002
% Kadar air =
Volume awal (ml)
1,90
2,00
2,10
Volume akhir (ml)
2,00
2,10
2,20
volume akhir − volume awal
x 100%
berat sampel
1. Kadar air
=
2,00 - 1,90
x 100% = 4,98%
2,005
2. Kadar air
=
2,10 - 2,00
x 100% = 4,99%
2,001
3. Kadar air
=
2,20 - 2,10
x 100% = 5,00%
2,002
% Rata-rata kadar air =
4,98% + 4,99% + 5,00%
= 4,99%
3
102
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Lampiran 12. Perhitungan kadar sari larut air ekstrak n-heksana daun
poguntano (Picria fel-terrae Lour.)
% Kadar sari larut dalam air =
No.
1.
2.
3.
berat sari (g)
100
x
x 100%
berat sampel (g)
20
Berat sampel (g)
2,011
2,105
2,025
Berat sari (g)
0,004
0,006
0,005
4. Kadar sari larut dalam air =
0,004 100
x
x 100% = 0,99%
2,011
20
5. Kadar sari larut dalam air =
0,002 100
x
x 100% = 1,42%
2,105
20
6. Kadar sari larut dalam air =
0,005 100
x
x 100% = 1,23%
2,025
20
% Rata-rata kadar sari larut dalam air =
0,99% + 1,42% + 1,23%
= 1,21%
3
103
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Lampiran 13. Perhitungan kadar sari larut etanol ekstrak n-heksana daun
poguntano (Picria fel-terrae Lour.)
% Kadar sari larut dalam etanol =
No.
1.
2.
3.
berat sari (g)
100
x
x 100%
berat sampel (g)
20
Berat sampel (g)
2,011
2,023
2,008
Berat sari (g)
0,012
0,013
0,009
4. Kadar sari larut dalam etanol
=
0,012 100
x
x 100% = 2,98%
2,011
20
5. Kadar sari larut dalam etanol
=
0,013 100
x
x 100% = 3,21%
2,023
20
6. Kadar sari larut dalam etanol
=
0,009 100
x
x 100% = 2,24%
2,008
20
% Rata-rata kadar sari larut dalam etanol =
2,98% + 3,21% + 2,24%
= 2,81%
3
104
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Lampiran 14. Perhitungan kadar abu total ekstrak n-heksana daun
poguntano (Picria fel-terrae Lour.)
% Kadar abu total =
berat abu (g)
x 100%
berat sampel (g)
No.
1.
2.
3.
Berat sampel (g)
2,0011
2,0052
2,0018
Berat abu (g)
0,0083
0,0091
0,0088
4. Kadar abu total =
0,0083
x 100% = 0,41%
2,0011
5. Kadar abu total =
0,0091
x 100% = 0,45%
2,0052
6. Kadar abu total =
0,0088
x 100% = 0,44%
2,0018
% Rata-rata kadar abu total =
0,41% + 0,45% + 0,44%
= 0,46%
3
105
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Lampiran 15. Perhitungan kadar abu tidak larut asam ekstrak n-heksana
daun poguntano (Picria fel-terrae Lour.)
berat abu (g)
x 100%
berat sampel (g)
% Kadar abu tidak larut dalam asam =
No. Berat sampel (g)
1.
2,0095
2.
2,0125
3.
2,0033
Berat abu (g)
0,0021
0,0017
0,0011
1. Kadar abu tidak larut dalam asam =
0,0021
x 100% = 0,10%
2,0095
2. Kadar abu tidak larut dalam asam =
0,0007
x 100% = 0,08%
1,0033
3. Kadar abu tidak larut dalam asam =
0,0011
x 100% = 0,05%
2,0033
% Rata-rata kadar abu tidak larut dalam asam =
0,10% + 0,08% + 0,05 %
3
= 0,08%
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Lampiran 16. Perhitungan kadar air ekstrak etilasetat daun poguntano
(Picria fel-terrae Lour.)
% Kadar air simplisia =
No.
1.
2.
3.
volume air (ml)
x 100%
berat sampel (g)
Berat sampel (g)
2,010
2,012
2,008
% Kadar air =
Volume awal (ml)
1,90
2,00
2,10
Volume akhir (ml)
2,00
2,10
2,20
volume akhir − volume awal
x 100%
berat sampel
1. Kadar air
=
2,00 - 1,90
x 100% = 4,97%
2,010
2. Kadar air
=
2,10 - 2,00
x 100% = 4,97%
2,012
3. Kadar air
=
2,20 - 2,10
x 100% = 4,98%
2,008
% Rata-rata kadar air =
4,97% + 4,97% + 4,98%
= 4,97%
3
107
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Lampiran 17. Perhitungan kadar sari larut air ekstrak etilasetat daun
poguntano (Picria fel-terrae Lour.)
% Kadar sari larut dalam air =
No.
1.
2.
3.
berat sari (g)
100
x
x 100%
berat sampel (g)
20
Berat sampel (g)
2,015
2,010
1,980
Berat sari (g)
0,053
0,048
0,050
1. Kadar sari larut dalam air =
0,053 100
x
x 100% = 13,15%
2,015
20
2. Kadar sari larut dalam air =
0,048 100
x
x 100% = 11,94%
2,010
20
3. Kadar sari larut dalam air =
0,050 100
x
x 100% = 12,62%
1,980
20
% Rata-rata kadar sari larut dalam air =
13,15% + 11,94% + 12,62%
= 12,57%
3
108
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Lampiran 18. Perhitungan kadar sari larut etanol ekstrak etilasetat daun
poguntano (Picria fel-terrae Lour.)
% Kadar sari larut dalam etanol =
No.
1.
2.
3.
berat sari (g)
100
x
x 100%
berat sampel (g)
20
Berat sampel (g)
2,018
2,006
2,021
Berat sari (g)
0,242
0,231
0,249
0,242 100
x
x 100% = 59,96%
2,018
20
1.
Kadar sari larut dalam etanol =
2.
Kadar sari larut dalam etanol=
0,231 100
x
x 100% = 57,58%
2,006
20
3.
Kadar sari larut dalam etanol=
0,249 100
x
x 100% = 61,60%
2,021
20
% Rata-rata kadar sari larut dalam etanol =
59,96% + 57,58 + 61,60%
= 59,71%
3
109
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Lampiran 19. Perhitungan kadar abu total ekstrak etilasetat daun poguntano
(Picria fel-terrae Lour.)
% Kadar abu total =
berat abu (g)
x 100%
berat sampel (g)
No.
1.
2.
3.
1. Kadar abu total =
2. Kadar abu total =
3. Kadar abu total =
Berat sampel (g)
2,0102
2,0094
2,0083
Berat abu (g)
0,0104
0,0097
0,0086
0,0104
x 100% = 0,52%
2,0102
0,0097
x 100% = 0,48%
2,0094
0,0086
x 100% = 0,43%
2,0083
% Rata-rata kadar abu total =
0,52% + 0,48% + 0,43%
= 0,47%
3
110
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Lampiran 20. Perhitungan kadar abu tidak larut asam ekstrak etilasetat
daun poguntano (Picria fel-terrae Lour.)
berat abu (g)
x 100%
berat sampel (g)
% Kadar abu tidak larut dalam asam =
No. Berat sampel (g)
1.
2,0106
2.
2,0092
3.
2,0118
Berat abu (g)
0,0017
0,0011
0,0021
4. Kadar abu tidak larut dalam asam =
0,0017
x 100% = 0,08%
2,0106
5. Kadar abu tidak larut dalam asam =
0,0011
x 100% = 0,05%
2,0092
6. Kadar abu tidak larut dalam asam =
0,0021
x 100% = 0,10%
2,0118
% Rata-rata kadar abu tidak larut dalam asam =
0,08% + 0,05 % + 0,10 %
3
= 0,08%
111
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Lampiran 21. Perhitungan kadar air ekstrak etanol daun poguntano (Picria
fel-terrae Lour.)
% Kadar air simplisia =
No.
1.
2.
3.
volume air (ml)
x 100%
berat sampel (g)
Berat sampel (g)
5,014
5,010
5,012
% Kadar air =
Volume awal (ml)
1,90
2,30
2,70
Volume akhir (ml)
2,30
2,70
3,10
volume akhir − volume awal
x 100%
berat sampel
1.
Kadar air =
2,30 - 1,90
x 100% = 8,00%
5,014
2.
Kadar air =
2,70 - 2,30
x 100% = 7,98%
5,010
3.
Kadar air =
3,10 - 2,70
x 100% = 7,98%
5,012
% Rata-rata kadar air =
8,00% + 7,98% + 7,98%
= 7,99%
3
112
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Lampiran 22. Perhitungan kadar sari larut air ekstrak etanol
poguntano (Picria fel-terrae Lour.)
% Kadar sari larut dalam air =
No.
1.
2.
3.
daun
berat sari (g)
100
x
x 100%
berat sampel (g)
20
Berat sampel (g)
2,008
2,006
2,017
Berat sari (g)
0,307
0,281
0,358
1.
Kadar sari larut dalam air =
0,307 100
x
x 100% = 76,44%
2,008
20
2.
Kadar sari larut dalam air =
0,281 100
x
x 100% = 70,03%
2,006
20
3. Kadar sari larut dalam air
=
% Rata-rata kadar sari larut dalam air =
0,319 100
x
x 100% = 78,30%
2,037
20
76,44% + 70,03% + 78,30%
= 74,92%
3
113
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Lampiran 23. Perhitungan kadar sari larut etanol ekstrak etanol daun
poguntano (Picria fel-terrae Lour.)
% Kadar sari larut dalam etanol =
No.
1.
2.
3.
berat sari (g)
100
x
x 100%
berat sampel (g)
20
Berat sampel (g)
2,072
2,056
2,044
1. Kadar sari larut dalam etanol
=
2. Kadar sari larut dalam etanol
=
3. Kadar sari larut dalam etanol
=
Berat sari (g)
0,348
0,339
0,328
0,348 100
x
x 100% = 83,98%
2,072
20
0,339 100
x
x 100% = 82,44%
2,056
20
0,328 100
x
x 100% = 80,23%
2,044
20
% Rata-rata kadar sari larut dalam etanol =
83,98% + 82,44% + 80,23
= 82,22%
3
114
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Lampiran 24. Perhitungan kadar abu total ekstrak etanol daun poguntano
(Picria fel-terrae Lour.)
% Kadar abu total =
berat abu (g)
x 100%
berat sampel (g)
No.
1.
2.
3.
Berat sampel (g)
1,9775
1,9950
2,0058
1. Kadar abu total =
0,0325
x 100% = 1,64%
1,9775
2. Kadar abu total =
0,0372
x 100% = 1,86%
1,9950
3. Kadar abu total =
0,0391
x 100% = 1,95%
2,0058
% Rata-rata kadar abu total =
Berat abu (g)
0,0325
0,0372
0,0391
1,64% + 1,86% + 1,95%
= 1,82%
3
115
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Lampiran 25. Perhitungan kadar abu tidak larut asam ekstrak etanol daun
poguntano (Picria fel-terrae Lour.)
% Kadar abu total =
berat abu (g)
x 100%
berat sampel (g)
No.
1.
2.
3.
Berat sampel (g)
1,8990
1,9923
1,8255
1. Kadar abu total =
0,0065
x 100% = 0,34%
1,8990
2. Kadar abu total =
0,0071
x 100% = 0,36%
1,9923
3. Kadar abu total =
0,0042
x 100% = 0,23%
1,8255
% Rata-rata kadar abu total =
Berat abu (g)
0,0065
0,0071
0,0042
0,34% + 0,36% + 0,23%
= 0,31%
3
116
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Lampiran 26. Perhitungan persen sel hidup sel MCF-7
a. Kontrol sel dan kontrol media
Absorbansi
kontrol sel
Rata-rata
absorbansi
kontrol sel
Absorbansi
kontrol media
Rata-rata
absorbansi
kontrol media
Absorbansi kontrol
sel dikurangi
kontrol media
0,154
0,907
0,150
1.232
1.02
1,061
0.931
0,147
0,164
b. Ekstrak n-heksana daun poguntano
Kadar
(µg/ml)
31.25
1.002
0.911
0.879
93.497 83.468 79.941
% sel hidup
rata-rata
85.636
62.5
0.947
0.929
0.705
87.436 85.452 60.764
77.884
125.0
0.682
0.655
0.602
58.229 55.253 49.412
54.298
250.0
0.363
0.342
0.345
23.071 20.757 21.087
21.639
500.0
0.201
0.215
0.241
5.217
7.201
Absorbansi
% sel hidup
6.760
9.625
c. Ekstrak etilasetat daun poguntano
Kadar
(µg/ml)
31.25
0.923
1.01
0.9
84.791 94.379 82.256
% sel hidup
rata-rata
87.142
62.5
0.9
0.904
0.759
82.256 82.697 66.716
77.223
125.0
0.557
0.639
0.59
44.453 53.490 48.090
48.677
250.0
0.309
0.341
0.397
17.120 20.647 26.819
21.528
500.0
0.232
0.248
0.243
8.633
9.625
Absorbansi
% sel hidup
10.397
9.846
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d. Ekstrak etanol daun poguntano
Kadar
(µg/ml)
Absorbansi
% sel hidup
rata-rata
% sel hidup
31.25
1.069
0.958
0.976
100.882
88.648
90.632
93.387
62.5
0.986
1.074
0.866
91.734
101.433 78.508
90.558
125.0
0.929
0.905
0.788
85.452
82.807
69.912
79.390
250.0
0.688
0.724
0.744
58.891
62.858
65.062
62.270
500.0
0.514
0.376
0.359
39.713
24.504
22.630
28.949
e. Doxorubicin
Absorbansi
kontrol sel
Rata-rata
absorbansi
kontrol sel
Absorbansi
kontrol media
Rata-rata
absorbansi
kontrol media
Absorbansi kontrol
sel dikurangi
kontrol media
0,159
0,694
0.159
0.865
0.851
0.853
0.843
0.161
0.157
Kadar
(µg/ml)
Absorbansi
% sel hidup
rata-rata
% sel hidup
25
0.836
0.749
0.982
97.550
85.014
118.588
100.384
50
0.706
0.809
0.782
78.818
93.660
89.769
87.416
100
0.767
0.789
0.719
87.608
90.778
80.692
86.359
200
0.694
0.728
0.723
77.089
81.988
81.268
80.115
400
0.404
0.425
0.43
35.303
38.329
39.049
37.560
118
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Lampiran 27. Perhitungan persen sel hidup sel T47D
a. Kontrol sel dan kontrol media
Absorbansi
kontrol sel
Rata-rata
absorbansi
kontrol sel
Absorbansi
kontrol media
Rata-rata
absorbansi
kontrol media
Absorbansi kontrol
sel dikurangi
kontrol media
0.126
0.934
0.13
1.059
1.078
1.061
1.045
0.115
0.134
b. Ekstrak n-heksana daun poguntano
Kadar
(µg/ml)
31.25
0.899
0.974
0.978
0.950
% sel hidup
rata-rata
88.191
62.5
0.785
0.888
0.898
0.857
78.202
125.0
0.751
0.804
0.916
0.824
74.634
250.0
0.681
0.714
0.712
0.702
61.648
Absorbansi
Absorbansi rata-rata
c. Ekstrak etilasetat daun poguntano
Kadar
(µg/ml)
31.25
0.917
0.959
0.932
84.624 89.119 86.229
% sel hidup
rata-rata
86.657
62.5
0.762
0.618
0.572
68.034 52.622 47.699
56.118
125.0
0.505
0.486
0.451
40.528 38.494 34.748
37.924
250.0
0.354
0.326
0.369
24.367 21.370 25.972
23.903
500.0
0.294
0.247
0.296
17.945 12.915 18.159
16.340
Absorbansi
% sel hidup
119
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d. Ekstrak etanol daun poguntano
Kadar
(µg/ml)
Absorbansi
% sel hidup
rata-rata
% sel hidup
31.25
1.02
1.087
1.091 95.648 102.818 103.247
100.571
62.5
1.021
0.997
1.037 95.755
93.186
97.467
95.469
125.0
0.938
0.978
0.966 86.871
91.152
89.868
89.297
250.0
0.761
0.825
0.795 67.927
74.777
71.566
71.423
500.0
0.316
0.277
0.307 20.300
16.126
19.336
18.587
120
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Lampiran 28. Perhitungan persen sel hidup sel Vero
a. Kontrol sel dan kontrol media
Absorbansi
kontrol sel
Rata-rata
absorbansi
kontrol sel
Absorbansi
kontrol media
Rata-rata
absorbansi
kontrol media
Absorbansi kontrol
sel dikurangi
kontrol media
0.065
0.437
0.071
0.479
0.498
0.502
0.528
0.063
0.06
b. Ekstrak n-heksana daun poguntano
Kadar
(µg/ml)
31.25
0.487
0.51
0.482
0.493
% sel hidup
rata-rata
98.017
62.5
0.486
0.498
0.498
0.494
98.246
125.0
0.475
0.449
0.478
0.467
92.143
250.0
0.108
0.096
0.122
0.109
10.069
500.0
0.077
0.079
0.083
0.080
3.432
Absorbansi
Absorbansi rata-rata
121
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Lampiran 29. Bagan pembuatan media DMEM
DMEM Sachet
2 g Hepes
2 g NaHCO3
Dimasukkan kedalam erlenmeyer
Ditambahkan 800 ml aquabidest steril
Dihomogenkan dengan menggunakan stirer magnet
Diatur pH 7,2 – 7,4 (HCl 1N atau NaOH 1N)
Ditambahkan aquabidest steril samapai 1 l
Larutan media
Dilakukan sterilisasi dengan penyaringan
Ditampung dalam botol steril
Diberi identitas pada botol media
Disimpan pada suhu 2 – 80 C
Media DMEM
122
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Lampiran 30. Bagan pembuatan media kultur lengkap (MK) DMEM
Fetal Bovine
Serum (FBS)
(10%)
PenisilinStreptomisin
(2%)
Fungizone
(amphotericin
B) (0,5%)
DMEM ad
100%
Dicampur
Diberi identitas pada botol MK
Disimpan pada suhu 2 – 80C
Media Kultur Lengkap (MK)
123
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Lampiran 31. Bagan penumbuhan sel
Sel dalam freezer
Diambil diambil dari freezer
Diambil beberapa tetes
Konikel
Dimasukkan kedalam konikel yg berisi DMEM
Disentrifuge 6000 rpm selama 5 menit
Dibuang supernatan
Ditambahkan 4 ml MK DMEM
Di resuspensi hingga homogen
Flask
Dimasukkan ke dalam flask
Ditambahkan 5 ml MK kedalam setiap flask
Dihomogenkan
Diamati kondisi sel dengan mikroskop inverted
Diberi identitas pada flask
Disimpan dalam inkubator CO2
Sel
124
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Lampiran 32. Bagan panen sel
Sel, alat, dan bahan
Dipersiapkan dan dikondisikan
Diamati apakah sel telah konfluen 80%
Dibuang MK dari flask dengan mikropipet
Sel yang melekat
Dicuci sel 2x dengan PBS
Ditambahkan 400 µl trypsine-EDTA 0,25%
Diinkubasi dalam inkubator CO2 selama 5 menit
Ditambahkan 4 ml MK
Di resuspensi dengan mikropipet
Diamati sel dibawah mikroskop inverted
Di resuspensi kembali jika masih ada sel yang menggerombol
Ditransfer sel kedalam tabung konikel
Panen Sel
125
Universitas Sumatera Utara
Lampiran 33. Bagan penghitungan sel
Kultur Sel
Diambil 10µl panenan sel
Dipipetkan kedalam hemositometer
Dihitung jumlah sel dibawah mikroskop
Jumlah Sel
126
Universitas Sumatera Utara
Lampiran 34. Bagan pembuatan larutan uji
Ekstrak n-
heksan
Ekstrak
Etilasetat
Ekstrak
Etanol
Ditimbang sebanyak 50 mg
Dimasukkan kedalam polytube
Dilarutkan dalam 1.000 µl DMSO
Di vortex
Dibuat pengenceran sampai diperoleh
konsentrasi 500 µg/ml, 250 µg/ml,
125 µg/ml, 62,5 µg/ml, dan 31,25 µg/ml
Larutan Uji
127
Universitas Sumatera Utara
Lampiran 35. Bagan pengujian sitotoksik
Sel
Ditanam pada microplate 96 sumuran dengan
kepadatan 1 x 104
Diinkubasi selama 24 jam
Dibuang medium
Ditambahkan medium baru
Ditambahkan larutan uji
Diinkubasi selama 24 jam
Dibuang media dan larutan uji setelah 24 jam
Dicuci dengan PBS
Ditambahkan 100 µl MK dan 10 µl MTT (5 mg/ml)
Diinkubasi selama 4-6 jam
Ditambahkan SDS (sebagai stopper)
Dibungkus dengan aluminium foil
Dibiarkan selama 1 malam
Dibaca serapan dengan ELISA reader pada λ 595 nm
Absorbansi
Dihitung % sel hidup
Dihitung IC50 dengan analisa probit menggunakan
SPSS 17
IC50
128
Universitas Sumatera Utara
Lampiran 36. Sel MCF-7 dan T47D di bawah mikroskop
a
b
Keterangan: a. Sel MCF-7 sebelum diberi larutan uji (perbesaran 10 x 10)
b. Sel MCF-7 setelah diberi larutan uji (sel mengalami
perubahan bentuk morfologi) (perbesaran 10 x 10)
Keterangan: a. Sel T47D sebelum diberi larutan uji (perbesaran 10 x 10)
b. Sel T47D setelah diberi larutan uji (sel mengalami perubahan
bentuk morfologi) (perbesaran 10 x 10)
129
Universitas Sumatera Utara
Lampiran 37. Sel Vero di bawah mikroskop
a
b
Keterangan: a. Sel Vero sebelum diberi larutan uji (perbesaran 10 x 10)
b. Sel Vero setelah diberi larutan uji (sel mengalami
perubahan bentuk morfologi) (perbesaran 10 x 10)
130
Universitas Sumatera Utara
Lampiran 38. Microplate -96 sumuran dan kristal formazan
ekstrak
n-heksana
500 µg/ml
250 µg/ml
125 µg/ml
62,5 µg/ml
31,25 µg/ml
Kontrol
media
Kontrol
sel
ekstrak
etilasetat
ekstrak
etanol
Keterangan: microplate-96 sumuran yang berisi sel dan larutan uji
Keterangan: a. Kristal Formazan (perbesaran 10 x 10)
a
131
Universitas Sumatera Utara
Lampiran 39. Hasil penentuan IC50 sel MCF7 dengan analisa probit SPSS 17
a. Ekstrak n-heksana daun poguntano
Confidence Limits
95% Confidence Limits for
konsentrasi ekstrak
Probabil Estimat
ity
e
Lower
Bound
Upper
Bound
95% Confidence Limits for
log(konsentrasi ekstrak)b
Estimat
e
Lower
Bound
Upper
Bound
PROBI .010
Ta
.020
9.001
.012
29.397
.954
-1.905
1.468
12.192
.036
35.579
1.086
-1.449
1.551
.030
14.780
.069
40.240
1.170
-1.160
1.605
.040
17.083
.114
44.204
1.233
-.943
1.645
.050
19.219
.171
47.763
1.284
-.767
1.679
.060
21.246
.241
51.062
1.327
-.618
1.708
.070
23.198
.326
54.182
1.365
-.487
1.734
.080
25.098
.426
57.178
1.400
-.371
1.757
.090
26.961
.544
60.085
1.431
-.265
1.779
.100
28.797
.680
62.930
1.459
-.168
1.799
.150
37.830
1.701
76.840
1.578
.231
1.886
.200
46.990
3.481
91.282
1.672
.542
1.960
.250
56.598
6.334
107.446
1.753
.802
2.031
.300
66.888
10.646
126.718
1.825
1.027
2.103
.350
78.087
16.824
151.148
1.893
1.226
2.179
.400
90.443
25.209
184.086
1.956
1.402
2.265
.450
104.255
35.923
231.186
2.018
1.555
2.364
.500
119.906
48.755
302.040
2.079
1.688
2.480
.550
137.907
63.232
412.953
2.140
1.801
2.616
.600
158.968
78.898
592.288
2.201
1.897
2.773
.650
184.121
95.595
892.096
2.265
1.980
2.950
.700
214.948
113.580
1415.331
2.332
2.055
3.151
.750
254.031
133.568
2385.435
2.405
2.126
3.378
.800
305.968
156.889
4350.330
2.486
2.196
3.639
.850
380.056
186.082
8913.542
2.580
2.270
3.950
.900
499.271
226.935 22339.662
2.698
2.356
4.349
.910
533.279
237.630 27942.929
2.727
2.376
4.446
.920
572.855
249.660 35656.796
2.758
2.397
4.552
.930
619.769
263.410 46650.018
2.792
2.421
4.669
132
Universitas Sumatera Utara
.940
676.721
279.454 63026.633
2.830
2.446
4.800
.950
748.090
298.696 88904.222
2.874
2.475
4.949
.960
841.613
322.684 133313.51
4
2.925
2.509
5.125
.970
972.760
354.395 219643.65
1
2.988
2.549
5.342
.980
1179.26
4
400.736 427288.05
6
3.072
2.603
5.631
1597.28
484.898 1223353.1
9
97
a. A heterogeneity factor is used.
3.203
2.686
6.088
.990
b. Logarithm base = 10.
b. Ekstrak etilasetat daun poguntano
Confidence Limits
95% Confidence Limits for
konsentrasi ekstrak
Probabilit
y
Estimate
Lower
Bound
Upper
Bound
95% Confidence Limits for log
(konsentrasi ekstrak)a
Estimate
Lower
Bound
Upper
Bound
PROBI .010
T
.020
9.748
5.989
14.006
.989
.777
1.146
13.082
8.460
18.119
1.117
.927
1.258
.030
15.766
10.528
21.342
1.198
1.022
1.329
.040
18.143
12.409
24.146
1.259
1.094
1.383
.050
20.338
14.180
26.701
1.308
1.152
1.427
.060
22.414
15.884
29.092
1.351
1.201
1.464
.070
24.408
17.542
31.369
1.388
1.244
1.496
.080
26.343
19.172
33.562
1.421
1.283
1.526
.090
28.237
20.782
35.693
1.451
1.318
1.553
.100
30.099
22.382
37.778
1.479
1.350
1.577
.150
39.213
30.384
47.851
1.593
1.483
1.680
.200
48.387
38.662
57.857
1.685
1.587
1.762
.250
57.950
47.442
68.233
1.763
1.676
1.834
.300
68.138
56.888
79.303
1.833
1.755
1.899
.350
79.171
67.147
91.381
1.899
1.827
1.961
.400
91.289
78.373
104.825
1.960
1.894
2.020
.450
104.775
90.745
120.072
2.020
1.958
2.079
133
Universitas Sumatera Utara
.500
119.990
104.487
137.684
2.079
2.019
2.139
.550
137.416
119.909
158.410
2.138
2.079
2.200
.600
157.716
137.452
183.279
2.198
2.138
2.263
.650
181.854
157.780
213.777
2.260
2.198
2.330
.700
211.302
181.918
252.181
2.325
2.260
2.402
.750
248.451
211.536
302.240
2.395
2.325
2.480
.800
297.554
249.575
370.723
2.474
2.397
2.569
.850
367.167
301.865
471.561
2.565
2.480
2.674
.900
478.339
382.467
639.987
2.680
2.583
2.806
.910
509.896
404.832
689.207
2.707
2.607
2.838
.920
546.541
430.562
747.066
2.738
2.634
2.873
.930
589.882
460.687
816.411
2.771
2.663
2.912
.940
642.362
496.760
901.622
2.808
2.696
2.955
.950
707.938
541.274
1009.875
2.850
2.733
3.004
.960
793.583
598.581
1153.997
2.900
2.777
3.062
.970
913.201
677.250
1359.989
2.961
2.831
3.134
.980
1100.57
8
797.787
1692.410
3.042
2.902
3.229
.990
1476.97
2
1032.146
2390.336
3.169
3.014
3.378
a. Logarithm base = 10.
c. Ekstrak etanol daun poguntano
Confidence Limits
95% Confidence Limits for
konsentrasi ekstrak
Probabili
ty
Estimate
Lower
Bound
Upper
Bound
95% Confidence Limits for log
(konsentrasi ekstrak)b
Estimate
Lower
Bound
Upper
Bound
PROBI .010
Ta
.020
15.532
1.856
35.622
1.191
.269
1.552
22.039
3.510
45.793
1.343
.545
1.661
.030
27.518
5.248
53.813
1.440
.720
1.731
.040
32.520
7.092
60.848
1.512
.851
1.784
.050
37.252
9.050
67.322
1.571
.957
1.828
.060
41.819
11.126
73.447
1.621
1.046
1.866
.070
46.281
13.322
79.347
1.665
1.125
1.900
.080
50.679
15.640
85.106
1.705
1.194
1.930
134
Universitas Sumatera Utara
.090
55.041
18.081
90.782
1.741
1.257
1.958
.100
59.387
20.647
96.419
1.774
1.315
1.984
.150
81.347
35.365
125.134
1.910
1.549
2.097
.200
104.460
53.187
156.988
2.019
1.726
2.196
.250
129.458
73.834
194.970
2.112
1.868
2.290
.300
156.965
96.766
242.620
2.196
1.986
2.385
.350
187.646
121.385
304.322
2.273
2.084
2.483
.400
222.287
147.321
385.495
2.347
2.168
2.586
.450
261.878
174.606
493.111
2.418
2.242
2.693
.500
307.719
203.639
636.789
2.488
2.309
2.804
.550
361.586
235.122
830.650
2.558
2.371
2.919
.600
425.986
270.039
1096.489
2.629
2.431
3.040
.650
504.627
309.759
1469.585
2.703
2.491
3.167
.700
603.265
356.273
2010.395
2.781
2.552
3.303
.750
731.446
412.705
2830.436
2.864
2.616
3.452
.800
906.480
484.431
4157.286
2.957
2.685
3.619
.850
1164.034
581.981
6529.187
3.066
2.765
3.815
.900
1594.480
730.507 11562.985
3.203
2.864
4.063
.910
1720.382
771.390 13280.488
3.236
2.887
4.123
.920
1868.449
818.267 15439.124
3.271
2.913
4.189
.930
2046.008
872.953 18222.991
3.311
2.941
4.261
.940
2264.332
938.173 21933.847
3.355
2.972
4.341
.950
2541.907
1018.294 27103.093
3.405
3.008
4.433
.960
2911.797
1120.903 34762.692
3.464
3.050
4.541
.970
3441.090
1260.878 47223.194
3.537
3.101
4.674
.980
4296.536
1473.623 70995.912
3.633
3.168
4.851
.990
6096.662
1882.339 135127.39
3
3.785
3.275
5.131
a. A heterogeneity factor is used.
b. Logarithm base = 10.
d. Doxorubicin
Confidence Limits
Probab
ility
95% Confidence Limits for
konsentrasi ekstrak
95% Confidence Limits for
log(konsentrasi ekstrak)b
135
Universitas Sumatera Utara
Estimate
a
Lower
Bound
Upper
Bound
Estimate
Lower
Bound
Upper
Bound
PROBIT .010
8878.18
0
.
.
3.948
.
.
.020
6115.25
0
.
.
3.786
.
.
.030
4827.11
5
.
.
3.684
.
.
.040
4040.27
1
.
.
3.606
.
.
.050
3495.83
9
.
.
3.544
.
.
.060
3090.64
1
.
.
3.490
.
.
.070
2774.19
5
.
.
3.443
.
.
.080
2518.45
0
.
.
3.401
.
.
.090
2306.38
9
.
.
3.363
.
.
.100
2127.00
6
.
.
3.328
.
.
.150
1521.18
2
.
.
3.182
.
.
.200
1165.39
4
.
.
3.066
.
.
.250
927.268
.
.
2.967
.
.
.300
755.197
.
.
2.878
.
.
.350
624.387
.
.
2.795
.
.
.400
521.277
.
.
2.717
.
.
.450
437.753
.
.
2.641
.
.
.500
368.632
.
.
2.567
.
.
.550
310.425
.
.
2.492
.
.
.600
260.686
.
.
2.416
.
.
.650
217.637
.
.
2.338
.
.
.700
179.939
.
.
2.255
.
.
.750
146.548
.
.
2.166
.
.
.800
116.604
.
.
2.067
.
.
.850
89.332
.
.
1.951
.
.
136
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.900
63.888
.
.
1.805
.
.
.910
58.919
.
.
1.770
.
.
.920
53.958
.
.
1.732
.
.
.930
48.983
.
.
1.690
.
.
.940
43.968
.
.
1.643
.
.
.950
38.872
.
.
1.590
.
.
.960
33.634
.
.
1.527
.
.
.970
28.151
.
.
1.449
.
.
.980
22.221
.
.
1.347
.
.
.
.
1.185
.
.
.990
15.306
a. A heterogeneity factor is used.
b. Logarithm base = 10.
Lampiran 40. Hasil penentuan IC50 sel T47D dengan analisa probit SPSS 17
a. Ekstrak n-heksan daun poguntano
Confidence Limits
95% Confidence Limits for
konsentrasi ekstrak
Probab
ility
Estimate
PROBI .010
T
Lower
Bound
Upper
Bound
125061.18 14273.985
7
95% Confidence Limits for
log(konsentrasi ekstrak)a
Estimate
Lower
Bound
Upper
Bound
2.918E7
5.097
4.155
7.465
.020
64746.770
8999.177 9161910.65
1
4.811
3.954
6.962
.030
42640.364
6714.397 4394429.53
9
4.630
3.827
6.643
.040
31143.173
5386.029 2528845.22
1
4.493
3.731
6.403
.050
24119.303
4501.454 1613455.49
1
4.382
3.653
6.208
.060
19403.852
3863.684 1100710.89
5
4.288
3.587
6.042
.070
16034.310
3379.066 787187.406
4.205
3.529
5.896
.080
13516.959
2996.792 583095.471
4.131
3.477
5.766
.090
11572.394
2686.674 443837.897
4.063
3.429
5.647
.100
10030.672
2429.540 345264.200
4.001
3.386
5.538
.150
5549.421
1600.943 122128.624
3.744
3.204
5.087
137
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.200
3466.784
1148.188
53518.436
3.540
3.060
4.729
.250
2315.462
862.505
26393.848
3.365
2.936
4.422
.300
1611.472
666.394
14004.117
3.207
2.824
4.146
.350
1151.739
524.056
7793.605
3.061
2.719
3.892
.400
837.412
416.548
4476.293
2.923
2.620
3.651
.450
615.207
332.847
2623.428
2.789
2.522
3.419
.500
454.178
266.059
1555.572
2.657
2.425
3.192
.550
335.298
211.587
927.115
2.525
2.325
2.967
.600
246.328
166.137
552.949
2.392
2.220
2.743
.650
179.101
127.090
329.996
2.253
2.104
2.519
.700
128.006
92.119
199.250
2.107
1.964
2.299
.750
89.087
60.040
125.323
1.950
1.778
2.098
.800
59.501
33.462
83.304
1.775
1.525
1.921
.850
37.171
15.615
56.106
1.570
1.194
1.749
.900
20.565
5.715
35.732
1.313
.757
1.553
.910
17.825
4.468
32.152
1.251
.650
1.507
.920
15.261
3.416
28.694
1.184
.534
1.458
.930
12.865
2.541
25.342
1.109
.405
1.404
.940
10.631
1.824
22.077
1.027
.261
1.344
.950
8.552
1.249
18.880
.932
.097
1.276
.960
6.624
.800
15.724
.821
-.097
1.197
.970
4.838
.462
12.570
.685
-.336
1.099
.980
3.186
.222
9.346
.503
-.653
.971
.990
1.649
a. Logarithm base = 10.
.070
5.869
.217
-1.154
.769
b. Ekstrak etilasetat daun poguntano
Confidence Limits
95% Confidence Limits for
konsentrasi ekstrak
Proba
bility
Estimate
Lower
Bound
Upper
Bound
95% Confidence Limits for
log(konsentrasi ekstrak)b
Estimate
Lower
Bound
Upper
Bound
PROBI .010
Ta
.020
2480.677
863.709 44772.484
3.395
2.936
4.651
1698.688
662.987 21764.332
3.230
2.822
4.338
.030
1335.891
559.821 13790.129
3.126
2.748
4.140
138
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.040
1115.009
492.491
9792.180
3.047
2.692
3.991
.050
962.566
443.429
7417.199
2.983
2.647
3.870
.060
849.349
405.296
5858.919
2.929
2.608
3.768
.070
761.091
374.368
4766.997
2.881
2.573
3.678
.080
689.877
348.514
3965.119
2.839
2.542
3.598
.090
630.914
326.403
3355.135
2.800
2.514
3.526
.100
581.103
307.157
2878.212
2.764
2.487
3.459
.150
413.405
237.424
1534.579
2.616
2.376
3.186
.200
315.389
191.566
940.235
2.499
2.282
2.973
.250
250.045
157.537
624.719
2.398
2.197
2.796
.300
202.988
130.337
438.813
2.307
2.115
2.642
.350
167.326
107.483
321.791
2.224
2.031
2.508
.400
139.298
87.688
244.740
2.144
1.943
2.389
.450
116.658
70.362
192.209
2.067
1.847
2.284
.500
97.972
55.328
155.169
1.991
1.743
2.191
.550
82.280
42.567
128.034
1.915
1.629
2.107
.600
68.907
32.014
107.282
1.838
1.505
2.031
.650
57.365
23.496
90.697
1.759
1.371
1.958
.700
47.287
16.762
76.883
1.675
1.224
1.886
.750
38.388
11.533
64.935
1.584
1.062
1.812
.800
30.434
7.546
54.227
1.483
.878
1.734
.850
23.218
4.570
44.262
1.366
.660
1.646
.900
16.518
2.415
34.522
1.218
.383
1.538
.910
15.214
2.068
32.538
1.182
.316
1.512
.920
13.913
1.748
30.520
1.143
.242
1.485
.930
12.612
1.451
28.454
1.101
.162
1.454
.940
11.301
1.179
26.321
1.053
.072
1.420
.950
9.972
.930
24.092
.999
-.031
1.382
.960
8.609
.704
21.723
.935
-.153
1.337
.970
7.185
.499
19.139
.856
-.302
1.282
.980
5.651
.316
16.186
.752
-.501
1.209
.153
12.447
.588
-.815
1.095
.990
3.869
a. A heterogeneity factor is used.
b. Logarithm base = 10.
c. Ekstrak etanol daun poguntano
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Confidence Limits
95% Confidence Limits for
konsentrasi ekstrak
Probab
ility
Estimate
Lower
Bound
Upper
Bound
95% Confidence Limits for
log(konsentrasi ekstrak)b
Estimate
Lower
Bound
Upper
Bound
PROBI .010
Ta
.020
1741.777
790.001 27458.872
3.241
2.898
4.439
1420.898
689.497 17132.511
3.153
2.839
4.234
.030
1248.698
631.963 12711.285
3.096
2.801
4.104
.040
1133.055
591.567 10160.007
3.054
2.772
4.007
.050
1046.935
560.407
8470.663
3.020
2.749
3.928
.060
978.810
535.015
7258.231
2.991
2.728
3.861
.070
922.735
513.563
6340.513
2.965
2.711
3.802
.080
875.257
494.969
5619.039
2.942
2.695
3.750
.090
834.203
478.542
5035.501
2.921
2.680
3.702
.100
798.117
463.811
4552.974
2.902
2.666
3.658
.150
664.587
406.492
3007.714
2.823
2.609
3.478
.200
574.587
364.669
2171.467
2.759
2.562
3.337
.250
507.153
330.992
1648.170
2.705
2.520
3.217
.300
453.368
302.175
1291.900
2.656
2.480
3.111
.350
408.634
276.429
1035.697
2.611
2.442
3.015
.400
370.274
252.635
844.372
2.569
2.402
2.927
.450
336.591
230.020
697.630
2.527
2.362
2.844
.500
306.435
208.013
582.936
2.486
2.318
2.766
.550
278.981
186.200
492.102
2.446
2.270
2.692
.600
253.603
164.320
419.476
2.404
2.216
2.623
.650
229.796
142.303
360.906
2.361
2.153
2.557
.700
207.122
120.294
313.107
2.316
2.080
2.496
.750
185.156
98.616
273.312
2.268
1.994
2.437
.800
163.426
77.666
239.080
2.213
1.890
2.379
.850
141.295
57.782
208.135
2.150
1.762
2.318
.900
117.655
39.124
177.968
2.071
1.592
2.250
.910
112.565
35.533
171.724
2.051
1.551
2.235
.920
107.286
31.981
165.307
2.031
1.505
2.218
.930
101.765
28.462
158.648
2.008
1.454
2.200
.940
95.935
24.968
151.653
1.982
1.397
2.181
.950
89.693
21.483
144.182
1.953
1.332
2.159
140
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.960
82.875
17.986
136.018
1.918
1.255
2.134
.970
75.200
14.438
126.776
1.876
1.160
2.103
.980
66.087
10.762
115.662
1.820
1.032
2.063
.990
53.912
6.751
a. A heterogeneity factor is used.
b. Logarithm base = 10.
100.398
1.732
.829
2.002
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Lampiran 41. Combination Index
I= (D)1/(Dx)1 + (D)2/(Dx)2
I = CI
Dx = konsentrasi dari satu senyawa tunggal (IC50)
(D)1 dan (D)2 = besarnya konsentrasi kedua senyawa untuk memberikan efek yang sama
ug/ml
15.0
30.0
45.0
60.0
ug/ml
15.0
30.0
45.0
60.0
0.731
0.681
0.663
0.559
62.80
57.40
55.46
44.22
ENDP
0.825
0.781
0.728
0.588
absorbansi tunggal
uM
0.783
25.0
0.74
50.0
0.708
100.0
0.592
200.0
Doxorubicin tunggal
0.836
0.749
0.982 kontrol sel
0.767
0.799
0.819
1.189
1.025
0.694
0.728
0.723 kontrol media
0.625
0.702
0.617
0.14
0.15
ENDP
72.96
68.20
62.48
47.35
cell viability tunggal
rata-rata uM
68.42
68.06 25.0
63.77
63.13 50.0
60.32
59.42 100.0
47.79
46.45 200.0
Doxorubicin tunggal
rata-rata
74.14
64.75
89.92
76.27
66.69
70.15
72.31
69.72
58.80
62.48
61.94
61.07
51.35
59.67
50.49
53.84
1.012 1.075333
0.159
0.150
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ENDP
ug/ml
ENDP
ug/ml
EADP
ENDP
0
15.0
30.0
45.0
15.0
30.0
45.0
60.0
15.0
30.0
45.0
60.0
0.918
0.834
0.807
0.708
83.00
73.93
71.01
60.32
doxo 25
0.88
0.855
0.759
0.735
dox 25
78.90
76.20
65.83
63.23
y = -0.1682x +
81.93
R² = 0.8752
IC 50 = 120 µg/ml
0
100
68.06
63.13
59.42
25
76.27
80.45
75.51
66.83
0.885
0.857
0.739
0.747
79.44
76.41
63.67
64.53
0.769
0.765
0.803
0.739
absorbansi kombinasi
doxo
50
doxo100
0.856
0.857
0.726
0.807
0.824
0.826
0.757
0.728
0.788
0.767
0.75
0.723
0.772
0.757
0.69
0.652
66.91
66.47
70.58
63.67
cell viability kombinasi
dox 50
dox 100
76.31
76.41
62.26
71.01
72.85
73.06
65.61
62.48
68.96
66.69
64.85
61.94
67.23
65.61
58.37
54.27
Doxorubicin
Doxo
50
69.72
73.21
70.80
68.74
100
61.07
68.78
65.36
63.45
0.826
0.779
0.738
0.715
73.06
67.99
63.56
61.07
0.706
0.701
0.632
0.443
doxo 200
0.733
0.754
0.632
0.473
0.744
0.665
0.662
0.443
60.10
59.56
52.11
31.69
dox 200
63.02
65.29
52.11
34.93
64.21
55.67
55.35
31.69
y = -0.1524x + 101.99
R² = 0.9393
IC50 = 369 nM
200
53.84
62.44
60.17
53.19
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60.0
doxo tunggal
15.0
30.0
45.0
60.0
ENDP
15.0
30.0
45.0
60.0
46.45
25
141.361
173.733
230.678
257.851
62.69
65.50
y = -0.1524x + 101.99
50
100
188.855
217.918
204.686
240.365
218.155
252.889
239.420
289.277
y = -0.1682x + 81.93
25
50
100
8.820
51.852
78.185
38.150
66.196
98.524
89.746
78.399
109.871
114.367
97.667
142.841
57.90
32.77
200
259.505
274.391
320.230
454.205
200
115.865
129.353
170.886
292.276
CI
Doxo
ENDP
15.0
30.0
45.0
60.0
25
1.88
0.93
0.61
0.62
50
0.55
0.70
0.80
0.82
100
0.65
0.72
0.81
0.77
200
0.90
0.96
0.89
0.65
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Lampiran 42. Hasil penentuan IC50 sel vero dengan analisa probit SPSS 17
Confidence Limits
95% Confidence Limits for
konsentrasi ekstrak
Probab
ility
Estimate
Lower
Bound
95% Confidence Limits for
log(konsentrasi ekstrak)b
Upper
Bound
Estimate
Lower
Bound
Upper
Bound
PROBI .010
Ta
.020
553.113
.
.
2.743
.
.
484.041
.
.
2.685
.
.
.030
444.760
.
.
2.648
.
.
.040
417.326
.
.
2.620
.
.
.050
396.264
.
.
2.598
.
.
.060
379.176
.
.
2.579
.
.
.070
364.801
.
.
2.562
.
.
.080
352.392
.
.
2.547
.
.
.090
341.474
.
.
2.533
.
.
.100
331.723
.
.
2.521
.
.
.150
294.228
.
.
2.469
.
.
.200
267.475
.
.
2.427
.
.
.250
246.470
.
.
2.392
.
.
.300
229.017
.
.
2.360
.
.
.350
213.949
.
.
2.330
.
.
.400
200.569
.
.
2.302
.
.
.450
188.421
.
.
2.275
.
.
.500
177.184
.
.
2.248
.
.
.550
166.616
.
.
2.222
.
.
.600
156.525
.
.
2.195
.
.
.650
146.736
.
.
2.167
.
.
.700
137.081
.
.
2.137
.
.
.750
127.374
.
.
2.105
.
.
.800
117.372
.
.
2.070
.
.
.850
106.700
.
.
2.028
.
.
.900
94.639
.
.
1.976
.
.
.910
91.937
.
.
1.963
.
.
.920
89.088
.
.
1.950
.
.
.930
86.058
.
.
1.935
.
.
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.940
82.795
.
.
1.918
.
.
.950
79.225
.
.
1.899
.
.
.960
75.227
.
.
1.876
.
.
.970
70.586
.
.
1.849
.
.
.980
64.858
.
.
1.812
.
.
.
.
1.754
.
.
.990
56.759
a. A heterogeneity factor is used.
b. Logarithm base = 10.
Lampiran 43. Mikroskop inverted, elisa reader dan inkubator CO2
Keterangan : Elisa reader
Keterangan : Mikroskop inverted
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Keterangan : Inkubator CO2
Keterangan: Flowsitometri
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Keterangan: Laminar Air Flow
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Lampiran 2. Gambar daun poguntano (Picria Fel-terrae Lour.)
a
b
Keterangan:
a. Gambar daun poguntano poguntano
b. Gambar simplisia daun poguntano
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Lampiran 3. Hasil Pemeriksaan Mikroskopik Serbuk Simplisia Daun
Poguntano
a
b
c
d
e
f
g
Keterangan :
Perbesaran 10 x 40
a. Trikoma
b. Sel epidermis
c. Kristal kalsium oksalat bentuk prisma
d. Stomata tipe diasitik
e. Stomata tipe anomositik
f. Berkas pembuluh angkut bentuk spiral
g. Sel tetangga
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Lampiran 4. Bagan skrining fitokimia, karakterisasi simplisia dan ekstrak
Daun poguntano
Dicuci
Ditiriskan
Dikeringkan (diangin-anginkan)
Simplisia
Dihaluskan
Serbuk Simplisia
Skrining
Fitokimia
Karakterisasi
Simplisia
Pembuatan
Ekstrak
Ekstrak
- Alkaloid
- Flavonoid
- Saponin
- Tanin
- Glikosida
- Antrakuinon
- Steroid/Triterpenoid
- Pemeriksaan mikroskopik
- Pemeriksaan makroskopik
- Penetapan kadar air
- Penetapan kadar sari yang larut dalam etanol
- Penetapan kadar sari yang larut dalam air
- Penetapan kadar abu total
- Penetapan kadar abu yang tidak larut dalam asam
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Lampiran 5. Bagan ekstraksi serbuk simplisia secara maserasi bertingkat
Serbuk simplisia
di maserasi dengan n-heksana
Ampas
Maserat
di maserasi dengan etilasetat
dipekatkan
Ampas
Maserat
di maserasi dengan etanol
Ekstrak kering n-heksana*
dipekatkan
di
skrining
Hasil
Maserat
Ampas
dipekatkan
Ekstrak kering etilasetat*
di skrining
Ekstrak kental etanol*
Hasil
di skrining
Hasil
96
Universitas Sumatera Utara
Keterangan : * = Uji Sitotoksik
Lampiran 6. Perhitungan kadar air simplisia daun poguntano (Picria Felterrae Lour.)
% Kadar air simplisia =
No.
1.
2.
3.
Berat sampel (g)
5,022
5,015
5,018
% Kadar air =
volume air (ml)
x 100%
berat sampel (g)
Volume awal (ml)
1,90
2,20
2,50
Volume akhir (ml)
2,20
2,50
2,80
volume akhir − volume awal
x 100%
berat sampel
1. Kadar air =
2,20 - 1,90
x 100% = 5,97%
5,022
2. Kadar air =
2,50 - 2,20
x 100% = 5,98%
5,015
3. Kadar air =
2,80 - 2,50
x 100% = 5,97%
5,018
% Rata-rata kadar air =
5,97% + 5,98% + 5,97%
= 5,97%
3
97
Universitas Sumatera Utara
Lampiran 7. Perhitungan kadar sari larut air simplisia daun poguntano
(Picria fel-terrae Lour.)
% Kadar sari larut dalam air =
No.
1.
2.
3.
100
berat sari (g)
x 100%
x
20
berat sampel (g)
Berat sampel (g)
4,008
4,002
4,010
Berat sari (g)
0,212
0,208
0,220
1. Kadar sari larut dalam air =
0,205 100
x
x 100% = 26,45%
4,008
20
2. Kadar sari larut dalam air =
0,208 100
x
x 100% = 25,99%
4,002
20
3. Kadar sari larut dalam air =
0,222 100
x
x 100% = 27,43%
4,010
20
% Rata-rata kadar sari larut dalam air =
26,45% + 25,99% + 27,43%
= 26,62%
3
98
Universitas Sumatera Utara
Lampiran 8. Perhitungan kadar sari larut etanol simplisia daun poguntano
(Picria fel-terrae Lour.)
% Kadar sari larut dalam etanol =
No.
1.
2.
3.
100
berat sari (g)
x 100%
x
20
berat sampel (g)
Berat sampel (g)
5,012
5,009
5,017
Berat sari (g)
0,198
0,182
0,205
1. Kadar sari larut dalam etanol
=
0,198 100
x
x 100% = 19,75%
5,012
20
2. Kadar sari larut dalam etanol
=
0,182 100
x
x 100% = 18,17%
5,009
20
3. Kadar sari larut dalam etanol
=
0,205 100
x
x 100% = 20,43%
5,017
20
% Rata-rata kadar sari larut dalam etanol =
19,75% + 20,43% + 18,17%
= 19,45%
3
99
Universitas Sumatera Utara
Lampiran 9. Perhitungan kadar abu total simplisia daun poguntano (Picria
fel-terrae Lour.)
% Kadar abu total =
berat abu (g)
x 100%
berat sampel (g)
No.
1.
2.
3.
Berat sampel (g)
2,0121
2,0118
2,0132
Berat abu (g)
0,0780
0,0791
0,0812
1. Kadar abu total =
0,0780
x 100% = 3,87%
2,0121
2. Kadar abu total =
0,0791
x 100% = 3,93%
2,0118
3. Kadar abu total =
0,0812
x 100% = 4,03%
2,0132
% Rata-rata kadar abu total =
3,87% + 3,93% + 4,03%
= 3,94%
3
100
Universitas Sumatera Utara
Lampiran 10. Perhitungan kadar abu tidak larut asam simplisia daun
poguntano (Picria fel-terrae Lour.)
% Kadar abu total =
berat abu (g)
x 100%
berat sampel (g)
No.
1.
2.
3.
Berat sampel (g)
1,9981
1,9935
2,0027
1. Kadar abu total =
0,0151
x 100% = 0,76%
1,9981
2. Kadar abu total =
0,0135
x 100% = 0,67%
1,9935
3. Kadar abu total =
0,0163
x 100% = 0,81%
2,0027
% Rata-rata kadar abu total =
Berat abu (g)
0,0151
0,0135
0,0163
0,76% + 0,67% + 0,81%
= 0,75%
3
101
Universitas Sumatera Utara
Lampiran 11. Perhitungan kadar air ekstrak n-heksana daun poguntano
(Picria fel-terrae Lour.)
% Kadar air simplisia =
No.
1.
2.
3.
volume air (ml)
x 100%
berat sampel (g)
Berat sampel (g)
2,005
2,001
2,002
% Kadar air =
Volume awal (ml)
1,90
2,00
2,10
Volume akhir (ml)
2,00
2,10
2,20
volume akhir − volume awal
x 100%
berat sampel
1. Kadar air
=
2,00 - 1,90
x 100% = 4,98%
2,005
2. Kadar air
=
2,10 - 2,00
x 100% = 4,99%
2,001
3. Kadar air
=
2,20 - 2,10
x 100% = 5,00%
2,002
% Rata-rata kadar air =
4,98% + 4,99% + 5,00%
= 4,99%
3
102
Universitas Sumatera Utara
Lampiran 12. Perhitungan kadar sari larut air ekstrak n-heksana daun
poguntano (Picria fel-terrae Lour.)
% Kadar sari larut dalam air =
No.
1.
2.
3.
berat sari (g)
100
x
x 100%
berat sampel (g)
20
Berat sampel (g)
2,011
2,105
2,025
Berat sari (g)
0,004
0,006
0,005
4. Kadar sari larut dalam air =
0,004 100
x
x 100% = 0,99%
2,011
20
5. Kadar sari larut dalam air =
0,002 100
x
x 100% = 1,42%
2,105
20
6. Kadar sari larut dalam air =
0,005 100
x
x 100% = 1,23%
2,025
20
% Rata-rata kadar sari larut dalam air =
0,99% + 1,42% + 1,23%
= 1,21%
3
103
Universitas Sumatera Utara
Lampiran 13. Perhitungan kadar sari larut etanol ekstrak n-heksana daun
poguntano (Picria fel-terrae Lour.)
% Kadar sari larut dalam etanol =
No.
1.
2.
3.
berat sari (g)
100
x
x 100%
berat sampel (g)
20
Berat sampel (g)
2,011
2,023
2,008
Berat sari (g)
0,012
0,013
0,009
4. Kadar sari larut dalam etanol
=
0,012 100
x
x 100% = 2,98%
2,011
20
5. Kadar sari larut dalam etanol
=
0,013 100
x
x 100% = 3,21%
2,023
20
6. Kadar sari larut dalam etanol
=
0,009 100
x
x 100% = 2,24%
2,008
20
% Rata-rata kadar sari larut dalam etanol =
2,98% + 3,21% + 2,24%
= 2,81%
3
104
Universitas Sumatera Utara
Lampiran 14. Perhitungan kadar abu total ekstrak n-heksana daun
poguntano (Picria fel-terrae Lour.)
% Kadar abu total =
berat abu (g)
x 100%
berat sampel (g)
No.
1.
2.
3.
Berat sampel (g)
2,0011
2,0052
2,0018
Berat abu (g)
0,0083
0,0091
0,0088
4. Kadar abu total =
0,0083
x 100% = 0,41%
2,0011
5. Kadar abu total =
0,0091
x 100% = 0,45%
2,0052
6. Kadar abu total =
0,0088
x 100% = 0,44%
2,0018
% Rata-rata kadar abu total =
0,41% + 0,45% + 0,44%
= 0,46%
3
105
Universitas Sumatera Utara
Lampiran 15. Perhitungan kadar abu tidak larut asam ekstrak n-heksana
daun poguntano (Picria fel-terrae Lour.)
berat abu (g)
x 100%
berat sampel (g)
% Kadar abu tidak larut dalam asam =
No. Berat sampel (g)
1.
2,0095
2.
2,0125
3.
2,0033
Berat abu (g)
0,0021
0,0017
0,0011
1. Kadar abu tidak larut dalam asam =
0,0021
x 100% = 0,10%
2,0095
2. Kadar abu tidak larut dalam asam =
0,0007
x 100% = 0,08%
1,0033
3. Kadar abu tidak larut dalam asam =
0,0011
x 100% = 0,05%
2,0033
% Rata-rata kadar abu tidak larut dalam asam =
0,10% + 0,08% + 0,05 %
3
= 0,08%
106
Universitas Sumatera Utara
Lampiran 16. Perhitungan kadar air ekstrak etilasetat daun poguntano
(Picria fel-terrae Lour.)
% Kadar air simplisia =
No.
1.
2.
3.
volume air (ml)
x 100%
berat sampel (g)
Berat sampel (g)
2,010
2,012
2,008
% Kadar air =
Volume awal (ml)
1,90
2,00
2,10
Volume akhir (ml)
2,00
2,10
2,20
volume akhir − volume awal
x 100%
berat sampel
1. Kadar air
=
2,00 - 1,90
x 100% = 4,97%
2,010
2. Kadar air
=
2,10 - 2,00
x 100% = 4,97%
2,012
3. Kadar air
=
2,20 - 2,10
x 100% = 4,98%
2,008
% Rata-rata kadar air =
4,97% + 4,97% + 4,98%
= 4,97%
3
107
Universitas Sumatera Utara
Lampiran 17. Perhitungan kadar sari larut air ekstrak etilasetat daun
poguntano (Picria fel-terrae Lour.)
% Kadar sari larut dalam air =
No.
1.
2.
3.
berat sari (g)
100
x
x 100%
berat sampel (g)
20
Berat sampel (g)
2,015
2,010
1,980
Berat sari (g)
0,053
0,048
0,050
1. Kadar sari larut dalam air =
0,053 100
x
x 100% = 13,15%
2,015
20
2. Kadar sari larut dalam air =
0,048 100
x
x 100% = 11,94%
2,010
20
3. Kadar sari larut dalam air =
0,050 100
x
x 100% = 12,62%
1,980
20
% Rata-rata kadar sari larut dalam air =
13,15% + 11,94% + 12,62%
= 12,57%
3
108
Universitas Sumatera Utara
Lampiran 18. Perhitungan kadar sari larut etanol ekstrak etilasetat daun
poguntano (Picria fel-terrae Lour.)
% Kadar sari larut dalam etanol =
No.
1.
2.
3.
berat sari (g)
100
x
x 100%
berat sampel (g)
20
Berat sampel (g)
2,018
2,006
2,021
Berat sari (g)
0,242
0,231
0,249
0,242 100
x
x 100% = 59,96%
2,018
20
1.
Kadar sari larut dalam etanol =
2.
Kadar sari larut dalam etanol=
0,231 100
x
x 100% = 57,58%
2,006
20
3.
Kadar sari larut dalam etanol=
0,249 100
x
x 100% = 61,60%
2,021
20
% Rata-rata kadar sari larut dalam etanol =
59,96% + 57,58 + 61,60%
= 59,71%
3
109
Universitas Sumatera Utara
Lampiran 19. Perhitungan kadar abu total ekstrak etilasetat daun poguntano
(Picria fel-terrae Lour.)
% Kadar abu total =
berat abu (g)
x 100%
berat sampel (g)
No.
1.
2.
3.
1. Kadar abu total =
2. Kadar abu total =
3. Kadar abu total =
Berat sampel (g)
2,0102
2,0094
2,0083
Berat abu (g)
0,0104
0,0097
0,0086
0,0104
x 100% = 0,52%
2,0102
0,0097
x 100% = 0,48%
2,0094
0,0086
x 100% = 0,43%
2,0083
% Rata-rata kadar abu total =
0,52% + 0,48% + 0,43%
= 0,47%
3
110
Universitas Sumatera Utara
Lampiran 20. Perhitungan kadar abu tidak larut asam ekstrak etilasetat
daun poguntano (Picria fel-terrae Lour.)
berat abu (g)
x 100%
berat sampel (g)
% Kadar abu tidak larut dalam asam =
No. Berat sampel (g)
1.
2,0106
2.
2,0092
3.
2,0118
Berat abu (g)
0,0017
0,0011
0,0021
4. Kadar abu tidak larut dalam asam =
0,0017
x 100% = 0,08%
2,0106
5. Kadar abu tidak larut dalam asam =
0,0011
x 100% = 0,05%
2,0092
6. Kadar abu tidak larut dalam asam =
0,0021
x 100% = 0,10%
2,0118
% Rata-rata kadar abu tidak larut dalam asam =
0,08% + 0,05 % + 0,10 %
3
= 0,08%
111
Universitas Sumatera Utara
Lampiran 21. Perhitungan kadar air ekstrak etanol daun poguntano (Picria
fel-terrae Lour.)
% Kadar air simplisia =
No.
1.
2.
3.
volume air (ml)
x 100%
berat sampel (g)
Berat sampel (g)
5,014
5,010
5,012
% Kadar air =
Volume awal (ml)
1,90
2,30
2,70
Volume akhir (ml)
2,30
2,70
3,10
volume akhir − volume awal
x 100%
berat sampel
1.
Kadar air =
2,30 - 1,90
x 100% = 8,00%
5,014
2.
Kadar air =
2,70 - 2,30
x 100% = 7,98%
5,010
3.
Kadar air =
3,10 - 2,70
x 100% = 7,98%
5,012
% Rata-rata kadar air =
8,00% + 7,98% + 7,98%
= 7,99%
3
112
Universitas Sumatera Utara
Lampiran 22. Perhitungan kadar sari larut air ekstrak etanol
poguntano (Picria fel-terrae Lour.)
% Kadar sari larut dalam air =
No.
1.
2.
3.
daun
berat sari (g)
100
x
x 100%
berat sampel (g)
20
Berat sampel (g)
2,008
2,006
2,017
Berat sari (g)
0,307
0,281
0,358
1.
Kadar sari larut dalam air =
0,307 100
x
x 100% = 76,44%
2,008
20
2.
Kadar sari larut dalam air =
0,281 100
x
x 100% = 70,03%
2,006
20
3. Kadar sari larut dalam air
=
% Rata-rata kadar sari larut dalam air =
0,319 100
x
x 100% = 78,30%
2,037
20
76,44% + 70,03% + 78,30%
= 74,92%
3
113
Universitas Sumatera Utara
Lampiran 23. Perhitungan kadar sari larut etanol ekstrak etanol daun
poguntano (Picria fel-terrae Lour.)
% Kadar sari larut dalam etanol =
No.
1.
2.
3.
berat sari (g)
100
x
x 100%
berat sampel (g)
20
Berat sampel (g)
2,072
2,056
2,044
1. Kadar sari larut dalam etanol
=
2. Kadar sari larut dalam etanol
=
3. Kadar sari larut dalam etanol
=
Berat sari (g)
0,348
0,339
0,328
0,348 100
x
x 100% = 83,98%
2,072
20
0,339 100
x
x 100% = 82,44%
2,056
20
0,328 100
x
x 100% = 80,23%
2,044
20
% Rata-rata kadar sari larut dalam etanol =
83,98% + 82,44% + 80,23
= 82,22%
3
114
Universitas Sumatera Utara
Lampiran 24. Perhitungan kadar abu total ekstrak etanol daun poguntano
(Picria fel-terrae Lour.)
% Kadar abu total =
berat abu (g)
x 100%
berat sampel (g)
No.
1.
2.
3.
Berat sampel (g)
1,9775
1,9950
2,0058
1. Kadar abu total =
0,0325
x 100% = 1,64%
1,9775
2. Kadar abu total =
0,0372
x 100% = 1,86%
1,9950
3. Kadar abu total =
0,0391
x 100% = 1,95%
2,0058
% Rata-rata kadar abu total =
Berat abu (g)
0,0325
0,0372
0,0391
1,64% + 1,86% + 1,95%
= 1,82%
3
115
Universitas Sumatera Utara
Lampiran 25. Perhitungan kadar abu tidak larut asam ekstrak etanol daun
poguntano (Picria fel-terrae Lour.)
% Kadar abu total =
berat abu (g)
x 100%
berat sampel (g)
No.
1.
2.
3.
Berat sampel (g)
1,8990
1,9923
1,8255
1. Kadar abu total =
0,0065
x 100% = 0,34%
1,8990
2. Kadar abu total =
0,0071
x 100% = 0,36%
1,9923
3. Kadar abu total =
0,0042
x 100% = 0,23%
1,8255
% Rata-rata kadar abu total =
Berat abu (g)
0,0065
0,0071
0,0042
0,34% + 0,36% + 0,23%
= 0,31%
3
116
Universitas Sumatera Utara
Lampiran 26. Perhitungan persen sel hidup sel MCF-7
a. Kontrol sel dan kontrol media
Absorbansi
kontrol sel
Rata-rata
absorbansi
kontrol sel
Absorbansi
kontrol media
Rata-rata
absorbansi
kontrol media
Absorbansi kontrol
sel dikurangi
kontrol media
0,154
0,907
0,150
1.232
1.02
1,061
0.931
0,147
0,164
b. Ekstrak n-heksana daun poguntano
Kadar
(µg/ml)
31.25
1.002
0.911
0.879
93.497 83.468 79.941
% sel hidup
rata-rata
85.636
62.5
0.947
0.929
0.705
87.436 85.452 60.764
77.884
125.0
0.682
0.655
0.602
58.229 55.253 49.412
54.298
250.0
0.363
0.342
0.345
23.071 20.757 21.087
21.639
500.0
0.201
0.215
0.241
5.217
7.201
Absorbansi
% sel hidup
6.760
9.625
c. Ekstrak etilasetat daun poguntano
Kadar
(µg/ml)
31.25
0.923
1.01
0.9
84.791 94.379 82.256
% sel hidup
rata-rata
87.142
62.5
0.9
0.904
0.759
82.256 82.697 66.716
77.223
125.0
0.557
0.639
0.59
44.453 53.490 48.090
48.677
250.0
0.309
0.341
0.397
17.120 20.647 26.819
21.528
500.0
0.232
0.248
0.243
8.633
9.625
Absorbansi
% sel hidup
10.397
9.846
117
Universitas Sumatera Utara
d. Ekstrak etanol daun poguntano
Kadar
(µg/ml)
Absorbansi
% sel hidup
rata-rata
% sel hidup
31.25
1.069
0.958
0.976
100.882
88.648
90.632
93.387
62.5
0.986
1.074
0.866
91.734
101.433 78.508
90.558
125.0
0.929
0.905
0.788
85.452
82.807
69.912
79.390
250.0
0.688
0.724
0.744
58.891
62.858
65.062
62.270
500.0
0.514
0.376
0.359
39.713
24.504
22.630
28.949
e. Doxorubicin
Absorbansi
kontrol sel
Rata-rata
absorbansi
kontrol sel
Absorbansi
kontrol media
Rata-rata
absorbansi
kontrol media
Absorbansi kontrol
sel dikurangi
kontrol media
0,159
0,694
0.159
0.865
0.851
0.853
0.843
0.161
0.157
Kadar
(µg/ml)
Absorbansi
% sel hidup
rata-rata
% sel hidup
25
0.836
0.749
0.982
97.550
85.014
118.588
100.384
50
0.706
0.809
0.782
78.818
93.660
89.769
87.416
100
0.767
0.789
0.719
87.608
90.778
80.692
86.359
200
0.694
0.728
0.723
77.089
81.988
81.268
80.115
400
0.404
0.425
0.43
35.303
38.329
39.049
37.560
118
Universitas Sumatera Utara
Lampiran 27. Perhitungan persen sel hidup sel T47D
a. Kontrol sel dan kontrol media
Absorbansi
kontrol sel
Rata-rata
absorbansi
kontrol sel
Absorbansi
kontrol media
Rata-rata
absorbansi
kontrol media
Absorbansi kontrol
sel dikurangi
kontrol media
0.126
0.934
0.13
1.059
1.078
1.061
1.045
0.115
0.134
b. Ekstrak n-heksana daun poguntano
Kadar
(µg/ml)
31.25
0.899
0.974
0.978
0.950
% sel hidup
rata-rata
88.191
62.5
0.785
0.888
0.898
0.857
78.202
125.0
0.751
0.804
0.916
0.824
74.634
250.0
0.681
0.714
0.712
0.702
61.648
Absorbansi
Absorbansi rata-rata
c. Ekstrak etilasetat daun poguntano
Kadar
(µg/ml)
31.25
0.917
0.959
0.932
84.624 89.119 86.229
% sel hidup
rata-rata
86.657
62.5
0.762
0.618
0.572
68.034 52.622 47.699
56.118
125.0
0.505
0.486
0.451
40.528 38.494 34.748
37.924
250.0
0.354
0.326
0.369
24.367 21.370 25.972
23.903
500.0
0.294
0.247
0.296
17.945 12.915 18.159
16.340
Absorbansi
% sel hidup
119
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d. Ekstrak etanol daun poguntano
Kadar
(µg/ml)
Absorbansi
% sel hidup
rata-rata
% sel hidup
31.25
1.02
1.087
1.091 95.648 102.818 103.247
100.571
62.5
1.021
0.997
1.037 95.755
93.186
97.467
95.469
125.0
0.938
0.978
0.966 86.871
91.152
89.868
89.297
250.0
0.761
0.825
0.795 67.927
74.777
71.566
71.423
500.0
0.316
0.277
0.307 20.300
16.126
19.336
18.587
120
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Lampiran 28. Perhitungan persen sel hidup sel Vero
a. Kontrol sel dan kontrol media
Absorbansi
kontrol sel
Rata-rata
absorbansi
kontrol sel
Absorbansi
kontrol media
Rata-rata
absorbansi
kontrol media
Absorbansi kontrol
sel dikurangi
kontrol media
0.065
0.437
0.071
0.479
0.498
0.502
0.528
0.063
0.06
b. Ekstrak n-heksana daun poguntano
Kadar
(µg/ml)
31.25
0.487
0.51
0.482
0.493
% sel hidup
rata-rata
98.017
62.5
0.486
0.498
0.498
0.494
98.246
125.0
0.475
0.449
0.478
0.467
92.143
250.0
0.108
0.096
0.122
0.109
10.069
500.0
0.077
0.079
0.083
0.080
3.432
Absorbansi
Absorbansi rata-rata
121
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Lampiran 29. Bagan pembuatan media DMEM
DMEM Sachet
2 g Hepes
2 g NaHCO3
Dimasukkan kedalam erlenmeyer
Ditambahkan 800 ml aquabidest steril
Dihomogenkan dengan menggunakan stirer magnet
Diatur pH 7,2 – 7,4 (HCl 1N atau NaOH 1N)
Ditambahkan aquabidest steril samapai 1 l
Larutan media
Dilakukan sterilisasi dengan penyaringan
Ditampung dalam botol steril
Diberi identitas pada botol media
Disimpan pada suhu 2 – 80 C
Media DMEM
122
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Lampiran 30. Bagan pembuatan media kultur lengkap (MK) DMEM
Fetal Bovine
Serum (FBS)
(10%)
PenisilinStreptomisin
(2%)
Fungizone
(amphotericin
B) (0,5%)
DMEM ad
100%
Dicampur
Diberi identitas pada botol MK
Disimpan pada suhu 2 – 80C
Media Kultur Lengkap (MK)
123
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Lampiran 31. Bagan penumbuhan sel
Sel dalam freezer
Diambil diambil dari freezer
Diambil beberapa tetes
Konikel
Dimasukkan kedalam konikel yg berisi DMEM
Disentrifuge 6000 rpm selama 5 menit
Dibuang supernatan
Ditambahkan 4 ml MK DMEM
Di resuspensi hingga homogen
Flask
Dimasukkan ke dalam flask
Ditambahkan 5 ml MK kedalam setiap flask
Dihomogenkan
Diamati kondisi sel dengan mikroskop inverted
Diberi identitas pada flask
Disimpan dalam inkubator CO2
Sel
124
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Lampiran 32. Bagan panen sel
Sel, alat, dan bahan
Dipersiapkan dan dikondisikan
Diamati apakah sel telah konfluen 80%
Dibuang MK dari flask dengan mikropipet
Sel yang melekat
Dicuci sel 2x dengan PBS
Ditambahkan 400 µl trypsine-EDTA 0,25%
Diinkubasi dalam inkubator CO2 selama 5 menit
Ditambahkan 4 ml MK
Di resuspensi dengan mikropipet
Diamati sel dibawah mikroskop inverted
Di resuspensi kembali jika masih ada sel yang menggerombol
Ditransfer sel kedalam tabung konikel
Panen Sel
125
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Lampiran 33. Bagan penghitungan sel
Kultur Sel
Diambil 10µl panenan sel
Dipipetkan kedalam hemositometer
Dihitung jumlah sel dibawah mikroskop
Jumlah Sel
126
Universitas Sumatera Utara
Lampiran 34. Bagan pembuatan larutan uji
Ekstrak n-
heksan
Ekstrak
Etilasetat
Ekstrak
Etanol
Ditimbang sebanyak 50 mg
Dimasukkan kedalam polytube
Dilarutkan dalam 1.000 µl DMSO
Di vortex
Dibuat pengenceran sampai diperoleh
konsentrasi 500 µg/ml, 250 µg/ml,
125 µg/ml, 62,5 µg/ml, dan 31,25 µg/ml
Larutan Uji
127
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Lampiran 35. Bagan pengujian sitotoksik
Sel
Ditanam pada microplate 96 sumuran dengan
kepadatan 1 x 104
Diinkubasi selama 24 jam
Dibuang medium
Ditambahkan medium baru
Ditambahkan larutan uji
Diinkubasi selama 24 jam
Dibuang media dan larutan uji setelah 24 jam
Dicuci dengan PBS
Ditambahkan 100 µl MK dan 10 µl MTT (5 mg/ml)
Diinkubasi selama 4-6 jam
Ditambahkan SDS (sebagai stopper)
Dibungkus dengan aluminium foil
Dibiarkan selama 1 malam
Dibaca serapan dengan ELISA reader pada λ 595 nm
Absorbansi
Dihitung % sel hidup
Dihitung IC50 dengan analisa probit menggunakan
SPSS 17
IC50
128
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Lampiran 36. Sel MCF-7 dan T47D di bawah mikroskop
a
b
Keterangan: a. Sel MCF-7 sebelum diberi larutan uji (perbesaran 10 x 10)
b. Sel MCF-7 setelah diberi larutan uji (sel mengalami
perubahan bentuk morfologi) (perbesaran 10 x 10)
Keterangan: a. Sel T47D sebelum diberi larutan uji (perbesaran 10 x 10)
b. Sel T47D setelah diberi larutan uji (sel mengalami perubahan
bentuk morfologi) (perbesaran 10 x 10)
129
Universitas Sumatera Utara
Lampiran 37. Sel Vero di bawah mikroskop
a
b
Keterangan: a. Sel Vero sebelum diberi larutan uji (perbesaran 10 x 10)
b. Sel Vero setelah diberi larutan uji (sel mengalami
perubahan bentuk morfologi) (perbesaran 10 x 10)
130
Universitas Sumatera Utara
Lampiran 38. Microplate -96 sumuran dan kristal formazan
ekstrak
n-heksana
500 µg/ml
250 µg/ml
125 µg/ml
62,5 µg/ml
31,25 µg/ml
Kontrol
media
Kontrol
sel
ekstrak
etilasetat
ekstrak
etanol
Keterangan: microplate-96 sumuran yang berisi sel dan larutan uji
Keterangan: a. Kristal Formazan (perbesaran 10 x 10)
a
131
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Lampiran 39. Hasil penentuan IC50 sel MCF7 dengan analisa probit SPSS 17
a. Ekstrak n-heksana daun poguntano
Confidence Limits
95% Confidence Limits for
konsentrasi ekstrak
Probabil Estimat
ity
e
Lower
Bound
Upper
Bound
95% Confidence Limits for
log(konsentrasi ekstrak)b
Estimat
e
Lower
Bound
Upper
Bound
PROBI .010
Ta
.020
9.001
.012
29.397
.954
-1.905
1.468
12.192
.036
35.579
1.086
-1.449
1.551
.030
14.780
.069
40.240
1.170
-1.160
1.605
.040
17.083
.114
44.204
1.233
-.943
1.645
.050
19.219
.171
47.763
1.284
-.767
1.679
.060
21.246
.241
51.062
1.327
-.618
1.708
.070
23.198
.326
54.182
1.365
-.487
1.734
.080
25.098
.426
57.178
1.400
-.371
1.757
.090
26.961
.544
60.085
1.431
-.265
1.779
.100
28.797
.680
62.930
1.459
-.168
1.799
.150
37.830
1.701
76.840
1.578
.231
1.886
.200
46.990
3.481
91.282
1.672
.542
1.960
.250
56.598
6.334
107.446
1.753
.802
2.031
.300
66.888
10.646
126.718
1.825
1.027
2.103
.350
78.087
16.824
151.148
1.893
1.226
2.179
.400
90.443
25.209
184.086
1.956
1.402
2.265
.450
104.255
35.923
231.186
2.018
1.555
2.364
.500
119.906
48.755
302.040
2.079
1.688
2.480
.550
137.907
63.232
412.953
2.140
1.801
2.616
.600
158.968
78.898
592.288
2.201
1.897
2.773
.650
184.121
95.595
892.096
2.265
1.980
2.950
.700
214.948
113.580
1415.331
2.332
2.055
3.151
.750
254.031
133.568
2385.435
2.405
2.126
3.378
.800
305.968
156.889
4350.330
2.486
2.196
3.639
.850
380.056
186.082
8913.542
2.580
2.270
3.950
.900
499.271
226.935 22339.662
2.698
2.356
4.349
.910
533.279
237.630 27942.929
2.727
2.376
4.446
.920
572.855
249.660 35656.796
2.758
2.397
4.552
.930
619.769
263.410 46650.018
2.792
2.421
4.669
132
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.940
676.721
279.454 63026.633
2.830
2.446
4.800
.950
748.090
298.696 88904.222
2.874
2.475
4.949
.960
841.613
322.684 133313.51
4
2.925
2.509
5.125
.970
972.760
354.395 219643.65
1
2.988
2.549
5.342
.980
1179.26
4
400.736 427288.05
6
3.072
2.603
5.631
1597.28
484.898 1223353.1
9
97
a. A heterogeneity factor is used.
3.203
2.686
6.088
.990
b. Logarithm base = 10.
b. Ekstrak etilasetat daun poguntano
Confidence Limits
95% Confidence Limits for
konsentrasi ekstrak
Probabilit
y
Estimate
Lower
Bound
Upper
Bound
95% Confidence Limits for log
(konsentrasi ekstrak)a
Estimate
Lower
Bound
Upper
Bound
PROBI .010
T
.020
9.748
5.989
14.006
.989
.777
1.146
13.082
8.460
18.119
1.117
.927
1.258
.030
15.766
10.528
21.342
1.198
1.022
1.329
.040
18.143
12.409
24.146
1.259
1.094
1.383
.050
20.338
14.180
26.701
1.308
1.152
1.427
.060
22.414
15.884
29.092
1.351
1.201
1.464
.070
24.408
17.542
31.369
1.388
1.244
1.496
.080
26.343
19.172
33.562
1.421
1.283
1.526
.090
28.237
20.782
35.693
1.451
1.318
1.553
.100
30.099
22.382
37.778
1.479
1.350
1.577
.150
39.213
30.384
47.851
1.593
1.483
1.680
.200
48.387
38.662
57.857
1.685
1.587
1.762
.250
57.950
47.442
68.233
1.763
1.676
1.834
.300
68.138
56.888
79.303
1.833
1.755
1.899
.350
79.171
67.147
91.381
1.899
1.827
1.961
.400
91.289
78.373
104.825
1.960
1.894
2.020
.450
104.775
90.745
120.072
2.020
1.958
2.079
133
Universitas Sumatera Utara
.500
119.990
104.487
137.684
2.079
2.019
2.139
.550
137.416
119.909
158.410
2.138
2.079
2.200
.600
157.716
137.452
183.279
2.198
2.138
2.263
.650
181.854
157.780
213.777
2.260
2.198
2.330
.700
211.302
181.918
252.181
2.325
2.260
2.402
.750
248.451
211.536
302.240
2.395
2.325
2.480
.800
297.554
249.575
370.723
2.474
2.397
2.569
.850
367.167
301.865
471.561
2.565
2.480
2.674
.900
478.339
382.467
639.987
2.680
2.583
2.806
.910
509.896
404.832
689.207
2.707
2.607
2.838
.920
546.541
430.562
747.066
2.738
2.634
2.873
.930
589.882
460.687
816.411
2.771
2.663
2.912
.940
642.362
496.760
901.622
2.808
2.696
2.955
.950
707.938
541.274
1009.875
2.850
2.733
3.004
.960
793.583
598.581
1153.997
2.900
2.777
3.062
.970
913.201
677.250
1359.989
2.961
2.831
3.134
.980
1100.57
8
797.787
1692.410
3.042
2.902
3.229
.990
1476.97
2
1032.146
2390.336
3.169
3.014
3.378
a. Logarithm base = 10.
c. Ekstrak etanol daun poguntano
Confidence Limits
95% Confidence Limits for
konsentrasi ekstrak
Probabili
ty
Estimate
Lower
Bound
Upper
Bound
95% Confidence Limits for log
(konsentrasi ekstrak)b
Estimate
Lower
Bound
Upper
Bound
PROBI .010
Ta
.020
15.532
1.856
35.622
1.191
.269
1.552
22.039
3.510
45.793
1.343
.545
1.661
.030
27.518
5.248
53.813
1.440
.720
1.731
.040
32.520
7.092
60.848
1.512
.851
1.784
.050
37.252
9.050
67.322
1.571
.957
1.828
.060
41.819
11.126
73.447
1.621
1.046
1.866
.070
46.281
13.322
79.347
1.665
1.125
1.900
.080
50.679
15.640
85.106
1.705
1.194
1.930
134
Universitas Sumatera Utara
.090
55.041
18.081
90.782
1.741
1.257
1.958
.100
59.387
20.647
96.419
1.774
1.315
1.984
.150
81.347
35.365
125.134
1.910
1.549
2.097
.200
104.460
53.187
156.988
2.019
1.726
2.196
.250
129.458
73.834
194.970
2.112
1.868
2.290
.300
156.965
96.766
242.620
2.196
1.986
2.385
.350
187.646
121.385
304.322
2.273
2.084
2.483
.400
222.287
147.321
385.495
2.347
2.168
2.586
.450
261.878
174.606
493.111
2.418
2.242
2.693
.500
307.719
203.639
636.789
2.488
2.309
2.804
.550
361.586
235.122
830.650
2.558
2.371
2.919
.600
425.986
270.039
1096.489
2.629
2.431
3.040
.650
504.627
309.759
1469.585
2.703
2.491
3.167
.700
603.265
356.273
2010.395
2.781
2.552
3.303
.750
731.446
412.705
2830.436
2.864
2.616
3.452
.800
906.480
484.431
4157.286
2.957
2.685
3.619
.850
1164.034
581.981
6529.187
3.066
2.765
3.815
.900
1594.480
730.507 11562.985
3.203
2.864
4.063
.910
1720.382
771.390 13280.488
3.236
2.887
4.123
.920
1868.449
818.267 15439.124
3.271
2.913
4.189
.930
2046.008
872.953 18222.991
3.311
2.941
4.261
.940
2264.332
938.173 21933.847
3.355
2.972
4.341
.950
2541.907
1018.294 27103.093
3.405
3.008
4.433
.960
2911.797
1120.903 34762.692
3.464
3.050
4.541
.970
3441.090
1260.878 47223.194
3.537
3.101
4.674
.980
4296.536
1473.623 70995.912
3.633
3.168
4.851
.990
6096.662
1882.339 135127.39
3
3.785
3.275
5.131
a. A heterogeneity factor is used.
b. Logarithm base = 10.
d. Doxorubicin
Confidence Limits
Probab
ility
95% Confidence Limits for
konsentrasi ekstrak
95% Confidence Limits for
log(konsentrasi ekstrak)b
135
Universitas Sumatera Utara
Estimate
a
Lower
Bound
Upper
Bound
Estimate
Lower
Bound
Upper
Bound
PROBIT .010
8878.18
0
.
.
3.948
.
.
.020
6115.25
0
.
.
3.786
.
.
.030
4827.11
5
.
.
3.684
.
.
.040
4040.27
1
.
.
3.606
.
.
.050
3495.83
9
.
.
3.544
.
.
.060
3090.64
1
.
.
3.490
.
.
.070
2774.19
5
.
.
3.443
.
.
.080
2518.45
0
.
.
3.401
.
.
.090
2306.38
9
.
.
3.363
.
.
.100
2127.00
6
.
.
3.328
.
.
.150
1521.18
2
.
.
3.182
.
.
.200
1165.39
4
.
.
3.066
.
.
.250
927.268
.
.
2.967
.
.
.300
755.197
.
.
2.878
.
.
.350
624.387
.
.
2.795
.
.
.400
521.277
.
.
2.717
.
.
.450
437.753
.
.
2.641
.
.
.500
368.632
.
.
2.567
.
.
.550
310.425
.
.
2.492
.
.
.600
260.686
.
.
2.416
.
.
.650
217.637
.
.
2.338
.
.
.700
179.939
.
.
2.255
.
.
.750
146.548
.
.
2.166
.
.
.800
116.604
.
.
2.067
.
.
.850
89.332
.
.
1.951
.
.
136
Universitas Sumatera Utara
.900
63.888
.
.
1.805
.
.
.910
58.919
.
.
1.770
.
.
.920
53.958
.
.
1.732
.
.
.930
48.983
.
.
1.690
.
.
.940
43.968
.
.
1.643
.
.
.950
38.872
.
.
1.590
.
.
.960
33.634
.
.
1.527
.
.
.970
28.151
.
.
1.449
.
.
.980
22.221
.
.
1.347
.
.
.
.
1.185
.
.
.990
15.306
a. A heterogeneity factor is used.
b. Logarithm base = 10.
Lampiran 40. Hasil penentuan IC50 sel T47D dengan analisa probit SPSS 17
a. Ekstrak n-heksan daun poguntano
Confidence Limits
95% Confidence Limits for
konsentrasi ekstrak
Probab
ility
Estimate
PROBI .010
T
Lower
Bound
Upper
Bound
125061.18 14273.985
7
95% Confidence Limits for
log(konsentrasi ekstrak)a
Estimate
Lower
Bound
Upper
Bound
2.918E7
5.097
4.155
7.465
.020
64746.770
8999.177 9161910.65
1
4.811
3.954
6.962
.030
42640.364
6714.397 4394429.53
9
4.630
3.827
6.643
.040
31143.173
5386.029 2528845.22
1
4.493
3.731
6.403
.050
24119.303
4501.454 1613455.49
1
4.382
3.653
6.208
.060
19403.852
3863.684 1100710.89
5
4.288
3.587
6.042
.070
16034.310
3379.066 787187.406
4.205
3.529
5.896
.080
13516.959
2996.792 583095.471
4.131
3.477
5.766
.090
11572.394
2686.674 443837.897
4.063
3.429
5.647
.100
10030.672
2429.540 345264.200
4.001
3.386
5.538
.150
5549.421
1600.943 122128.624
3.744
3.204
5.087
137
Universitas Sumatera Utara
.200
3466.784
1148.188
53518.436
3.540
3.060
4.729
.250
2315.462
862.505
26393.848
3.365
2.936
4.422
.300
1611.472
666.394
14004.117
3.207
2.824
4.146
.350
1151.739
524.056
7793.605
3.061
2.719
3.892
.400
837.412
416.548
4476.293
2.923
2.620
3.651
.450
615.207
332.847
2623.428
2.789
2.522
3.419
.500
454.178
266.059
1555.572
2.657
2.425
3.192
.550
335.298
211.587
927.115
2.525
2.325
2.967
.600
246.328
166.137
552.949
2.392
2.220
2.743
.650
179.101
127.090
329.996
2.253
2.104
2.519
.700
128.006
92.119
199.250
2.107
1.964
2.299
.750
89.087
60.040
125.323
1.950
1.778
2.098
.800
59.501
33.462
83.304
1.775
1.525
1.921
.850
37.171
15.615
56.106
1.570
1.194
1.749
.900
20.565
5.715
35.732
1.313
.757
1.553
.910
17.825
4.468
32.152
1.251
.650
1.507
.920
15.261
3.416
28.694
1.184
.534
1.458
.930
12.865
2.541
25.342
1.109
.405
1.404
.940
10.631
1.824
22.077
1.027
.261
1.344
.950
8.552
1.249
18.880
.932
.097
1.276
.960
6.624
.800
15.724
.821
-.097
1.197
.970
4.838
.462
12.570
.685
-.336
1.099
.980
3.186
.222
9.346
.503
-.653
.971
.990
1.649
a. Logarithm base = 10.
.070
5.869
.217
-1.154
.769
b. Ekstrak etilasetat daun poguntano
Confidence Limits
95% Confidence Limits for
konsentrasi ekstrak
Proba
bility
Estimate
Lower
Bound
Upper
Bound
95% Confidence Limits for
log(konsentrasi ekstrak)b
Estimate
Lower
Bound
Upper
Bound
PROBI .010
Ta
.020
2480.677
863.709 44772.484
3.395
2.936
4.651
1698.688
662.987 21764.332
3.230
2.822
4.338
.030
1335.891
559.821 13790.129
3.126
2.748
4.140
138
Universitas Sumatera Utara
.040
1115.009
492.491
9792.180
3.047
2.692
3.991
.050
962.566
443.429
7417.199
2.983
2.647
3.870
.060
849.349
405.296
5858.919
2.929
2.608
3.768
.070
761.091
374.368
4766.997
2.881
2.573
3.678
.080
689.877
348.514
3965.119
2.839
2.542
3.598
.090
630.914
326.403
3355.135
2.800
2.514
3.526
.100
581.103
307.157
2878.212
2.764
2.487
3.459
.150
413.405
237.424
1534.579
2.616
2.376
3.186
.200
315.389
191.566
940.235
2.499
2.282
2.973
.250
250.045
157.537
624.719
2.398
2.197
2.796
.300
202.988
130.337
438.813
2.307
2.115
2.642
.350
167.326
107.483
321.791
2.224
2.031
2.508
.400
139.298
87.688
244.740
2.144
1.943
2.389
.450
116.658
70.362
192.209
2.067
1.847
2.284
.500
97.972
55.328
155.169
1.991
1.743
2.191
.550
82.280
42.567
128.034
1.915
1.629
2.107
.600
68.907
32.014
107.282
1.838
1.505
2.031
.650
57.365
23.496
90.697
1.759
1.371
1.958
.700
47.287
16.762
76.883
1.675
1.224
1.886
.750
38.388
11.533
64.935
1.584
1.062
1.812
.800
30.434
7.546
54.227
1.483
.878
1.734
.850
23.218
4.570
44.262
1.366
.660
1.646
.900
16.518
2.415
34.522
1.218
.383
1.538
.910
15.214
2.068
32.538
1.182
.316
1.512
.920
13.913
1.748
30.520
1.143
.242
1.485
.930
12.612
1.451
28.454
1.101
.162
1.454
.940
11.301
1.179
26.321
1.053
.072
1.420
.950
9.972
.930
24.092
.999
-.031
1.382
.960
8.609
.704
21.723
.935
-.153
1.337
.970
7.185
.499
19.139
.856
-.302
1.282
.980
5.651
.316
16.186
.752
-.501
1.209
.153
12.447
.588
-.815
1.095
.990
3.869
a. A heterogeneity factor is used.
b. Logarithm base = 10.
c. Ekstrak etanol daun poguntano
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Confidence Limits
95% Confidence Limits for
konsentrasi ekstrak
Probab
ility
Estimate
Lower
Bound
Upper
Bound
95% Confidence Limits for
log(konsentrasi ekstrak)b
Estimate
Lower
Bound
Upper
Bound
PROBI .010
Ta
.020
1741.777
790.001 27458.872
3.241
2.898
4.439
1420.898
689.497 17132.511
3.153
2.839
4.234
.030
1248.698
631.963 12711.285
3.096
2.801
4.104
.040
1133.055
591.567 10160.007
3.054
2.772
4.007
.050
1046.935
560.407
8470.663
3.020
2.749
3.928
.060
978.810
535.015
7258.231
2.991
2.728
3.861
.070
922.735
513.563
6340.513
2.965
2.711
3.802
.080
875.257
494.969
5619.039
2.942
2.695
3.750
.090
834.203
478.542
5035.501
2.921
2.680
3.702
.100
798.117
463.811
4552.974
2.902
2.666
3.658
.150
664.587
406.492
3007.714
2.823
2.609
3.478
.200
574.587
364.669
2171.467
2.759
2.562
3.337
.250
507.153
330.992
1648.170
2.705
2.520
3.217
.300
453.368
302.175
1291.900
2.656
2.480
3.111
.350
408.634
276.429
1035.697
2.611
2.442
3.015
.400
370.274
252.635
844.372
2.569
2.402
2.927
.450
336.591
230.020
697.630
2.527
2.362
2.844
.500
306.435
208.013
582.936
2.486
2.318
2.766
.550
278.981
186.200
492.102
2.446
2.270
2.692
.600
253.603
164.320
419.476
2.404
2.216
2.623
.650
229.796
142.303
360.906
2.361
2.153
2.557
.700
207.122
120.294
313.107
2.316
2.080
2.496
.750
185.156
98.616
273.312
2.268
1.994
2.437
.800
163.426
77.666
239.080
2.213
1.890
2.379
.850
141.295
57.782
208.135
2.150
1.762
2.318
.900
117.655
39.124
177.968
2.071
1.592
2.250
.910
112.565
35.533
171.724
2.051
1.551
2.235
.920
107.286
31.981
165.307
2.031
1.505
2.218
.930
101.765
28.462
158.648
2.008
1.454
2.200
.940
95.935
24.968
151.653
1.982
1.397
2.181
.950
89.693
21.483
144.182
1.953
1.332
2.159
140
Universitas Sumatera Utara
.960
82.875
17.986
136.018
1.918
1.255
2.134
.970
75.200
14.438
126.776
1.876
1.160
2.103
.980
66.087
10.762
115.662
1.820
1.032
2.063
.990
53.912
6.751
a. A heterogeneity factor is used.
b. Logarithm base = 10.
100.398
1.732
.829
2.002
141
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Lampiran 41. Combination Index
I= (D)1/(Dx)1 + (D)2/(Dx)2
I = CI
Dx = konsentrasi dari satu senyawa tunggal (IC50)
(D)1 dan (D)2 = besarnya konsentrasi kedua senyawa untuk memberikan efek yang sama
ug/ml
15.0
30.0
45.0
60.0
ug/ml
15.0
30.0
45.0
60.0
0.731
0.681
0.663
0.559
62.80
57.40
55.46
44.22
ENDP
0.825
0.781
0.728
0.588
absorbansi tunggal
uM
0.783
25.0
0.74
50.0
0.708
100.0
0.592
200.0
Doxorubicin tunggal
0.836
0.749
0.982 kontrol sel
0.767
0.799
0.819
1.189
1.025
0.694
0.728
0.723 kontrol media
0.625
0.702
0.617
0.14
0.15
ENDP
72.96
68.20
62.48
47.35
cell viability tunggal
rata-rata uM
68.42
68.06 25.0
63.77
63.13 50.0
60.32
59.42 100.0
47.79
46.45 200.0
Doxorubicin tunggal
rata-rata
74.14
64.75
89.92
76.27
66.69
70.15
72.31
69.72
58.80
62.48
61.94
61.07
51.35
59.67
50.49
53.84
1.012 1.075333
0.159
0.150
142
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ENDP
ug/ml
ENDP
ug/ml
EADP
ENDP
0
15.0
30.0
45.0
15.0
30.0
45.0
60.0
15.0
30.0
45.0
60.0
0.918
0.834
0.807
0.708
83.00
73.93
71.01
60.32
doxo 25
0.88
0.855
0.759
0.735
dox 25
78.90
76.20
65.83
63.23
y = -0.1682x +
81.93
R² = 0.8752
IC 50 = 120 µg/ml
0
100
68.06
63.13
59.42
25
76.27
80.45
75.51
66.83
0.885
0.857
0.739
0.747
79.44
76.41
63.67
64.53
0.769
0.765
0.803
0.739
absorbansi kombinasi
doxo
50
doxo100
0.856
0.857
0.726
0.807
0.824
0.826
0.757
0.728
0.788
0.767
0.75
0.723
0.772
0.757
0.69
0.652
66.91
66.47
70.58
63.67
cell viability kombinasi
dox 50
dox 100
76.31
76.41
62.26
71.01
72.85
73.06
65.61
62.48
68.96
66.69
64.85
61.94
67.23
65.61
58.37
54.27
Doxorubicin
Doxo
50
69.72
73.21
70.80
68.74
100
61.07
68.78
65.36
63.45
0.826
0.779
0.738
0.715
73.06
67.99
63.56
61.07
0.706
0.701
0.632
0.443
doxo 200
0.733
0.754
0.632
0.473
0.744
0.665
0.662
0.443
60.10
59.56
52.11
31.69
dox 200
63.02
65.29
52.11
34.93
64.21
55.67
55.35
31.69
y = -0.1524x + 101.99
R² = 0.9393
IC50 = 369 nM
200
53.84
62.44
60.17
53.19
143
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60.0
doxo tunggal
15.0
30.0
45.0
60.0
ENDP
15.0
30.0
45.0
60.0
46.45
25
141.361
173.733
230.678
257.851
62.69
65.50
y = -0.1524x + 101.99
50
100
188.855
217.918
204.686
240.365
218.155
252.889
239.420
289.277
y = -0.1682x + 81.93
25
50
100
8.820
51.852
78.185
38.150
66.196
98.524
89.746
78.399
109.871
114.367
97.667
142.841
57.90
32.77
200
259.505
274.391
320.230
454.205
200
115.865
129.353
170.886
292.276
CI
Doxo
ENDP
15.0
30.0
45.0
60.0
25
1.88
0.93
0.61
0.62
50
0.55
0.70
0.80
0.82
100
0.65
0.72
0.81
0.77
200
0.90
0.96
0.89
0.65
144
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Lampiran 42. Hasil penentuan IC50 sel vero dengan analisa probit SPSS 17
Confidence Limits
95% Confidence Limits for
konsentrasi ekstrak
Probab
ility
Estimate
Lower
Bound
95% Confidence Limits for
log(konsentrasi ekstrak)b
Upper
Bound
Estimate
Lower
Bound
Upper
Bound
PROBI .010
Ta
.020
553.113
.
.
2.743
.
.
484.041
.
.
2.685
.
.
.030
444.760
.
.
2.648
.
.
.040
417.326
.
.
2.620
.
.
.050
396.264
.
.
2.598
.
.
.060
379.176
.
.
2.579
.
.
.070
364.801
.
.
2.562
.
.
.080
352.392
.
.
2.547
.
.
.090
341.474
.
.
2.533
.
.
.100
331.723
.
.
2.521
.
.
.150
294.228
.
.
2.469
.
.
.200
267.475
.
.
2.427
.
.
.250
246.470
.
.
2.392
.
.
.300
229.017
.
.
2.360
.
.
.350
213.949
.
.
2.330
.
.
.400
200.569
.
.
2.302
.
.
.450
188.421
.
.
2.275
.
.
.500
177.184
.
.
2.248
.
.
.550
166.616
.
.
2.222
.
.
.600
156.525
.
.
2.195
.
.
.650
146.736
.
.
2.167
.
.
.700
137.081
.
.
2.137
.
.
.750
127.374
.
.
2.105
.
.
.800
117.372
.
.
2.070
.
.
.850
106.700
.
.
2.028
.
.
.900
94.639
.
.
1.976
.
.
.910
91.937
.
.
1.963
.
.
.920
89.088
.
.
1.950
.
.
.930
86.058
.
.
1.935
.
.
145
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.940
82.795
.
.
1.918
.
.
.950
79.225
.
.
1.899
.
.
.960
75.227
.
.
1.876
.
.
.970
70.586
.
.
1.849
.
.
.980
64.858
.
.
1.812
.
.
.
.
1.754
.
.
.990
56.759
a. A heterogeneity factor is used.
b. Logarithm base = 10.
Lampiran 43. Mikroskop inverted, elisa reader dan inkubator CO2
Keterangan : Elisa reader
Keterangan : Mikroskop inverted
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Keterangan : Inkubator CO2
Keterangan: Flowsitometri
147
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Keterangan: Laminar Air Flow
148
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