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Lampiran 1. Bagan alir proses ekstraksi senyawa antimikroba biji atung secara bertingkat (Adawiyah, 1998).
Biji atung
Penepungan
Pemisahan lemak
lemak
dengan metoda soxhlet
Ekstraksi senyawa antimikroba dengan metoda refluks
Evaporasi dengan rotavapor
Pemekatan dengan gas N cair sampai berat konstan
Ekstrak /senyawa antimikroba biji atung
ampas
122
Lampiran 2. Pertumbuhan bakteri S. aureus selama 24 jam pada medium cair pada dosis ekstrak atung 0 sampai dengan 0.6 % (v/v), dengan pengukuran absorbansi (λ= 660 nm) dan metoda hitungan cawan. Dosis ekstrak biji atung (%v/v)
Jam ke Absorbansi
TPC
Absorbansi
TPC
0 0.4 0.45 0.5 0.55 0.6
0.017 0.000 0.064 0.047 0.002 0.048
4.0 x 107 3.2 x 107 2.5 x 107 4.0 x 107 3.6 x 107 2.0 x 107
0.389 0.031 0.017 0.037 -0.003 -0.003
1.09 x 108 -
0
24
Lampiran 3. Pertumbuhan bakteri S. aureus selama 24 jam pada medium cair pada dosis ekstrak atung 0 sampai dengan 0.4 % (v/v), dengan pengukuran absorbansi (λ= 660 nm) dan metoda hitungan cawan. Dosis ekstrak biji atung (%v/v)
Jam ke Absorbansi
TPC
Absorbansi
TPC
0 0.1 0.2 0.3 0.35 0.4
0.017 0.052 0.046 0.044 0.003 0.046
2.8 x 107 2.3 x 107 1.7 x 107 1.5 x 107 1.6 x 107 0.8 x 107
0.736 0.278 0.033 -0.032 0.039 0.017
7.9 x 109 5.6 x 108 5.9 x 103 -
0
24
Lampiran 4. Pertumbuhan bakteri P. fluorescens selama 24 jam pada medium cair pada dosis ekstrak atung 0 sampai dengan 0.6 % v/v, dengan pengukuran absorbansi (λ= 660 nm) dan metoda hitungan cawan. Dosis ekstrak biji atung (%v/v)
Jam ke Absorbansi
TPC
0 0.4 0.45 0.5 0.55 0.6
0.017 0.013 0.020 0.039 0.003 0.007
2.0 x 107 1.0 x 107 1.7 x 107 1.6 x 107 1.8 x 107 1.0 x 107
0
Keterangan: - = tidak tumbuh
24 Absorbansi 0.764 0.042 0.032 0.026 0.032 0.026
TPC 2.9 x 109 -
123
Lampiran 5. Pertumbuhan bakteri P. fluorescens selama 24 jam pada medium cair pada dosis ekstrak atung 0 sampai dengan 0.4 % v/v, dengan pengukuran absorbansi (λ= 660 nm) dan metoda hitungan cawan. Dosis ekstrak biji atung (%v/v)
Jam ke Absorbansi
TPC
Absorbansi
TPC
0 0.1 0.2 0.3 0.4
0.015 0.030 0.011 0.065 0.035
3.1 x 107 3.0 x 107 3.1 x 107 3.3 x 107 4.1 x 107
0.772 -0.105 -0.218 0.340 0.020
1.3 x1010 9.8 x 109 7.4 x 105 -
0
24
Lampiran 6. Pertumbuhan bakteri L. plantarum selama 24 jam pada medium cair pada dosis ekstrak atung 0, 0.45, 0.5, 0.55 dan 0.6 % v/v, dengan pengukuran absorbansi (λ= 660 nm) dan dengan metoda hitungan cawan. Dosis ekstrak biji atung (%v/v)
Jam ke Absorbansi
TPC
Absorbansi
TPC
0 0.45 0.50 0.55 0.60
0.293 0.312 0.311 0.294 0.285
1.8 x 108 2.3 x 108 2.2 x 108 2.6 x 108 1.3 x 108
1.700 1.650 1.630 1.530 1.480
3.8 x 109 2.5 x 109 1.8 x 109 9.0 x 108 9.6 x 108
0
24
Lampiran 7. Pertumbuhan bakteri L. plantarum selama 24 jam pada medium cair pada dosis ekstrak atung 0, 0.65, 0.70, 0.75 dan 0.8 % v/v, dengan pengukuran absorbansi (λ= 660 nm) dan metoda hitungan cawan. Dosis ekstrak biji atung (%v/v)
Jam ke Absorbansi
TPC
Absorbansi
TPC
0 0.65 0.70 0.75 0.80
0.295 0.284 0.225 0.338 0.306
1.4 x 108 1.6 x 108 1.6 x 108 1.5 x 108 1.6 x 108
1.650 1.315 1.215 1.345 1.265
2.7 x 109 1.1 x 109 1.1 x 109 1.1 x 108 7.9 x 108
0
Keterangan: - = tidak tumbuh
24
124
Lampiran 8. Pertumbuhan bakteri L. plantarum selama 24 jam pada medium cair pada dosis ekstrak atung 0, 1.0, 1.5, 2.0 dan 2.5 % (v/v), dengan pengukuran absorbansi (λ= 660 nm) Dosis ekstrak biji atung (%v/v)
Absorbansi jam ke
0 1.0 1.5 2.0 2.5
0
24
0.312 0.490 0.561 0.419 0.512
1.530 1.988 1.876 1.844 1.746
Lampiran 9. Pertumbuhan bakteri B. subtilis selama 24 jam pada medium cair pada dosis ekstrak atung 0, 0.45, 0.5, 0.55 dan 0.6 % v/v, dengan pengukuran absorbansi (λ= 660 nm) dan metoda hitungan cawan. Dosis ekstrak biji atung (%v/v)
Jam ke Absorbansi
TPC
Absorbansi
TPC
0 0.45 0.50 0.55 0.60
0.100 0.030 0.145 0.145 0.183
1.8 x 107 3.6 x 107 3.1 x 107 1.4 x 107 2.0 x 107
1.105 0.258 -0.016 -0.121 -0.100
1.0 x 108 1.1 x 102 8.7 x 101 7.8 x 101 7.5 x 101
0
24
Lampiran 10. Pertumbuhan bakteri E. coli selama 24 jam pada medium cair pada dosis ekstrak atung 0, s/d 0.6 % (v/v), dengan pengukuran absorbansi (λ= 660 nm) dan metoda hitungan cawan. Dosis ekstrak biji atung (%v/v)
Jam ke Absorbansi
TPC
Absorbansi
TPC
0 0.1 0.2 0.3 0.4 0.5 0.6
0.017 0.018 0.006 0.057 0.033 0.009 0.023
1.1 x 107 1.3 x 107 1.3 x 107 1.2 x 107 1.3 x 107 1.3 x 107 1.0 x 107
0.623 0.563 0.609 0.063 0.031 0.016 0.025
5.0 x 108 2.1 x 109 1.6 x 109 2.2 x 106 9.0 x 102 -
0
Keterangan: - = tidak tumbuh
24
125
Lampiran 11. Pengaruh ekstrak etil asetat biji atung pada dosis di bawah MIC pada pertumbuhan bakteri S. aureus pada medium cair dengan metoda hitungan cawan. Waktu Jam Ke-
Dosis Ekstrak Etil Asetat Biji Atung (MIC) 0
0,3
0,35
0,66
0,83
0,9
0,93
0,96
1,0
0
1,8x107
1,8x107
2,3x107
1,6x107
1,4x107
1,3 x107
1,7x107
1,6x107
1,3x107
2
5,2x107
1,3x107
6,6x106
8,5x106
1,1x107
7,1x106
4
4,0x108
1,8x107
8,5x107
7,2x106
2,1x106
6,8x106
2,0x106
2,6x106
6
9,7x108
4,2x107
1,5x107
4,4x106
6,2x105
1,7x106
3,9x105
7,2x105
8
4,5x109
7,1x107
3,2x107
5,0x106
3,7x105
4,2x105
2,4x105
4,5x105
12
4,5x109
5,0x107
6,6x106
3,3x105
2,5x105
7,9x105
7,9x104
16
2,8x109
4,7x107
1,0x107
1,4x106
4,8x105
1,1x105
3,1x104
20
4,5x108
3,7x107
1,2x106
7,2x105
7,8x104
8,1x103
24
7,8x109
4,8x107
3,3x106
3,7x105
3,0x104
2,4x103
`
9,5x107
1,6x108
4,1x107
1,7x105
1,1x104
< 30
126
Lampiran 12.
Pengaruh ekstrak etil asetat biji atung pada dosis di bawah MIC pada pertumbuhan bakteri P. flourescens pada medium cair dengan metoda hitungan cawan.
Waktu Jam Ke-
Dosis Ekstrak Etil Asetat Biji Atung (MIC) 0
0,1
0,2
0.5
0.64
0.71
0.86
1.0
0 2 3 4 6
4,1x107
2,5x107
3,0x107
2,0x107
2,2x107
1,9 x107
1,6x107
1,3x107
1,3x107
1,2x107
8
8
7
1,8x107
1,1x107
8 11 12 13 15 16 18 19 20 21 24 28 30 36 42
2,9x109
7,1x108
3,4x108 3,6x106
1,6x106
9
9
9
1,4x106
1,0x105
1,6x106
9,1x105
4,2x103
1,0x105 1,4x104
4,8x103 -
8,5x10
1,3x10
6,8x10
1,2x107
6,0x10
4,6x10
1,0x10
9,3x10
5,0x10
9
3,98x10
1,1x107
8,7x10 1,4x107
9
8,0x106
6
5,1x106
9
8,0x10 8,1x109
1,7x109
6,6x109
6,8x109 9,6x109
6,3x109 5,6x109
6,7x109 6,85x109
6
4,5x10
6
8,3x106
2,4x106
6,5x105
1,0x107 9,3x106 9,6x106
1,3x106 7,7x105 4,3x105
4,5x105
127
Lampiran 13. Pengaruh ekstrak etil asetat biji atung pada dosis stres pertumbuhan bakteri S. aureus pada medium cair dengan metoda hitungan cawan. Waktu Jam Ke-
Dosis Ekstrak Etil Asetat Biji Atung (MIC) 0,66 MIC
0,83 MIC
0,9 MIC
0,93 MIC
0
1,6x107
1,4x107
1,3x107
1,7x107
2
1,3x107
6,6x106
8,5x106
1,1x107
4
7,2x106
2,0x106
6,8x106
2,0x106
6
4,4x106
6,2x105
1,7x106
4,0x105
8
5,0x106
3,7x105
4,2x105
2,4x105
12
6,6x106
3,3x105
2,5x105
1,2x105
16
1,0x107
6,0x105
4,8x105
1,1x105
20
3,7x107
6,0x105
7,2x104
7,7x104
24
7
6
5
3,0x104 4,8x10
3,3x10
3,7x10
Lampiran 14. Pengaruh ekstrak etil asetat biji atung pada dosis stres pertumbuhan bakteri P. fluorescens pada medium cair dengan metoda hitungan cawan. Waktu Jam Ke-
Dosis Ekstrak Etil Asetat Biji Atung (MIC) 0
0.53
0.56
0,58
0,61
0
9,8x107
1,2x108
1,3x108
1,3x108
1,1x108
2
1,4x108
1,2x108
8,4x107
1,3x108
1,8x108
4
2,0x108
8,1x107
5,4x107
9,9x107
6,0x107
6
7,9x109
8,1x106
7,9x106
7,6x106
6,3x106
8
7,8x109
7,8x106
6,7x106
5,8x106
4,9x106
10
1,0x1010
7,7x107
6,3x106
6,2x106
3,2x107
12
1,3x1010
5,5x107
5,2x107
4,3x107
3,2x107
128
Lampiran 15. Pengaruh ekstrak etil asetat biji atung pada dosis di atas MIC pada pertumbuhan bakteri S. aureus pada medium cair dengan metoda hitungan cawan.
Waktu Jam Ke-
Dosis Ekstrak Etil Asetat Biji Atung (MIC) 1,0
1,03
1,1
1,17
1,24
1,31
1,38
1,6
0 1 2 3 4 5 6 7 8 12
5,2x107 3,8x107 2,7x107 2,0x107 7,1x106 2,5x106 8,9x105 4,3x105 2,1x105 1,1x104
5,1x107 3,1x107 2,9x107 6,3x106
4,2x107 3,6x107
4,4x107 3,6x107
4,2x107
2,7x106 2,5x105 5,4x104 8,1x103 2,5x103 -
2,8x106 1,2x105 2,7x104 4,0x103 1,4x103 -
3,9x107 4,3x106 1,9x105 1,6x104 1,4x103 6,5x102 3,1x102 1,7x102 9,8x101 -
3,5x107 2,4x106 1,7x105 1,3x104 1,0x103 3,9x102 1,5x102 6,0x101 -
3,6x107 1,4x105 5,4x102 2,6x102 1,2x102 6,0x101 2,9x101 -
20
6,3x101
1,4x105 1,9x104 4,9x103 2,2x103
7,1x105 4,5x104 6,0x103 1,1x103 4,2x102 -
129
Lampiran 16. Pengaruh ekstrak etil asetat biji atung pada dosis di atas MIC pada pertumbuhan bakteri P. fluorescens pada medium cair dengan metoda hitungan cawan.
Waktu Jam Ke-
0 2 4 6 8 10 12 16 20 24
Dosis Ekstrak Etil Asetat Biji Atung (MIC) 1,0
1,03
1,1
1,17
1,24
1,33
1,5
1,67
2,6x107 7,8x106 7,4x106 1,9x106 5,1x105 1,3x105 3,5x104 5,6x103 3,6x102 4,8x101
3,6x107 1,6x107 1,1x106 2,2x105 3,5x104 6,5x103 6,9x103 -
2,5x107 9,0x106 1,0x106 5,0x104 7,9x104 1,9x103 1,5x102 -
2,5x107 1,2x107 6,3x105 2,5x104 1,0x103 7,9x101 1,2x102 -
3,2x107 7,9x106 1,2x105 5,0x103 6,3x102 5,0x101 5,0x101 -
1,1x107 1,1x106 1,9x103 -
1,0x107 2,1x103 -
9,3x106 1,0x101 -
Catatan; Jam ke 10 dan 12 sel kecil-kecil
130
Lampiran 17. Pengaruh ekstrak etil asetat biji atung pada dosis di atas MIC pada pertumbuhan bakteri E. coli pada medium cair dengan metoda hitungan cawan.
Waktu Jam Ke-
Dosis Ekstrak Etil Asetat Biji Atung (MIC) 1,0
1,1
1,2
1,3
1,4
0
2,2x107
2,0x107
2,1x107
2,0x107
2,0x107
2
1,8x107
1,7x107
1,7x107
1,4x107
3
1,6x107
1,5x107
1,5x107
1,2x107
1,0x107
4
1,5x107
1,3x107
1,1x107
9,3x106
7,1x106
5
9,3x106
5,5x106
5,2x106
5,5x106
4,6x106
6
5,8x106
2,3x106
2,6x106
9,1x105
5,1x105
7
1,7x106
2,8x105
4,2x105
4,5x104
2,2x104
8
5,0x105
3,5x104
2,2x104
10
3,5x104
8,1x102
6,2x102
12
1,3x103
-
-
14
-
Keterangan : - = tidak tumbuh
1,2x104 -
7,9x101 -
131
Lampiran 18: Skema proses pembuatan dekstrin tapioka (Mukodiningsih, (1991).
Tepung Tapioka Dicampur dengan larutan HCl
Dikeringkan dengan oven
Digiling dan diayak
disangrai
Dekstrin Tapioka
132
Lampiran 19. Skema proses pembuatan tepung kedele (Koeswara, 1995).
Biji Kedele
sortasi
Perendaman dalam air bersih
Perebusan
dikuliti
ditiriskan
Dikeringkan dengan oven
Digiling dan diayak
Tepung Kedele
133
Lampiran 20. Pengaruh ekstrak etil asetat biji atung pada pertumbuhan bakteri S. aureus pada medium padat ( pangan model ) dengan metoda hitungan cawan.
Waktu Hari Ke-
Dosis Ekstrak Etil Asetat Biji Atung (MIC) 0
0,5
1,0
1,5
2,0
3,0
3,6x106
3,6x106
5,4x106
4,8x106
4,0x106
5,1x106
1
3,6x105
4,5x104
2,3x104
7,9x103
7,9x103
2
8,7x104
1,4x104
9,8x103
5,5x103
3,9x103
5,6x104
8,3x103
7,6x103
2,7x103
1,4x103
4
4,1x104
6,4x103
2,5x103
7,2x102
3,8x102
5
3,1x104
2,9x103
7,8x102
2,9x102
1,0x102
7,6x101
tt
tt
tt
0
3
8,1x105
6
3,7x105
2,6x104
9
5
2,6x10
8,3x10
3
13
3,6x105
2,4x102
16
9,7x104
tt
tt
Keterangan : tt = tidak tumbuh Lampiran 21 Data absorbansi dari bahan yang dilepaskan oleh sel S. aureus dan P. fluorescens pengaruh ekstrak etil asetat biji atung pada OD 260 dan 280 nm. Absorbansi Dosis (MIC)
0 0,3 0,5 0,7 0,9
S. aureus
P. fluorescens
OD260
OD280
OD260
OD280
0,283 1,926 2,181 2,353 1,429
0,142 1,627 1,877 1,955 1,96
0,380 1,482 1,908 2,437 2,505
0,208 1,326 1,881 2,491 2,491
134
Lampiran 22. Data peningkatan jumlah ion Ca++ dan K+ (% total) yang dilepaskan oleh sel S. aureus dan P. fluorescens pengaruh ekstrak etil asetat biji atung. Dosis (MIC) 0 0,3 0,5 0,7 0,9
S. aureus
P. fluorescens
Ca++
K+
Ca++
K+
0 2,16 5,81 29,17 58,44
0 4,93 13,71 36,76 38,72
0 64,0 77,5 79,0 82 ,0
0 1,53 4,40 12,28 21,33
