p-value 0,05 selama hang time hingga jam ke-8, yang menunjukkan BAL L. rhamnosus R21 dan R25 tidak terganggu oleh keberadaan C. sakazakii YRC3a.
5.2 Saran
Perlu dilakukan kajian mengenai penggunaan isolat L. rhamnosus R21 asal ASI yang ditambahkan pada susu bubuk formula untuk berkompetisi dan
menghambat pertumbuhan bakteri C. sakazakii isolat lainnya yang berasal dari susu formula. Perlu dilakukan kajian terhadap aktivitas senyawa antibakteri isolat
L. rhamnosus R21 lebih mendalam untuk mengetahui jenis senyawa antibakteri yang berperan utama untuk menghambatannya pertumbuhan C. sakazakii yang
mungkin terjadi selama hang time.
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LAMPIRAN
Lampiran 1 . Pengaruh suhu rekonstitusi 50 °C terhadap isolat BAL
Uji F: ANOVA Single Factor
Perubahan jumlah BAL Sumber Keragaman
Jumlah Kuadrat
Derajat Bebas
Kuadrat Tengah
F-hit F-tab
Between Groups 3,12
10 0,31
34,35 2,85
Within Groups 0,09
11 0,01
Total 3,22
21 Hipotesis
: Ho: µ1= µ2= µ3= µ4= µ5=µ6= µ7= µ8= µ9= µ10= µ11 Ho: Minimal Terdapat sepasang µi≠ µi’
Keputusan : Berdasarkan Hasil uji-F dapat dilihat minimal terdapat sepasang
perlakuan yang memberikan pengaruh yang berbeda nyata pada
taraf α=0,05 dimana FhitFtab
Kode Isolat
Genusgalur Penghambatan
EPEC Log CFUmL
Jumlah Awal No
CFUmL Rekonstitusi
suhu 27 °C Jumlah Akhir
Nt CFUmL Rekonstitusi
suhu 50 °C Perubahan
Jumlah Log
CFUmL Rata-
rata SB
A22 L. acidophilus
1,26 5,1 x 10
6
1,9 x 10
5
1,43 1,44
0,01 6,0 x 10
6
2,2 x 10
5
1,44 A23
L. rhamnosus 1,95
1,3 x 10
7
1,2 x 10
7
0,04 0,18
0,19 1,8 x 10
7
8,8 x 10
6
0,31 A24
L. rhamnosus 1,00
3,6 x 10
7
3,5 x 10
7
0,01 0,06
0,07 5,6 x 10
7
4,4 x 10
7
0,11 A27
L. rhamnosus 1,16
1,9 x 10
7
1,6 x 10
7
0,09 0,06
0,04 2,1 x 10
7
1,9 x 10
7
0,03 A29
L. rhamnosus 1,13
3,2 x 10
7
2,2 x 10
7
0,17 0,11
0,08 2,9 x 10
7
2,6 x 10
7
0,05 R14
L. rhamnosus 2,05
5,4 x 10
7
3,3 x 10
7
0,21 0,32
0,15 5,7 x 10
7
2,1 x 10
7
0,42 R21
L. rhamnosus 2,12
6,9 x 10
7
3,8 x 10
7
0,27 0,26
0,01 6,7 x 10
7
3,7 x 10
7
0,25 R23
L. rhamnosus 2,06
4,5 x 10
7
3,2 x 10
7
0,14 0,08
0,09 2,4 x 10
7
2,4 x 10
7
0,01 R25
Lactobacillus 2,06
6,9 x 10
7
4,3 x 10
7
0,21 0,18
0,05 6,3 x 10
7
4,6 x 10
7
0,14 R27
Lactobacillus 1,06
9,7 x 10
7
6,6 x 10
7
0,17 0,10
0,11 10,1 x 10
7
9,6 x 10
7
0,02 R32
L. rhamnosus 1,00
5,1 x 10
7
2,3 x 10
7
0,34 0,29
0,07 3,1 x 10
7
1,8 x 10
7
0,24 Keterangan: Air Susu Ibu; Hartanti 2010; SB= Simpangan baku
Uji Lanjut: Uji pembanding berpasangan uji Duncan Perubahan jumlah BAL
Duncan
a b
c
Isolat Keputusan
N Subset for alpha = 0.05
1 2
3 A24
a 2
0,06 A27
a 2
0,06 R23
a 2
0,08 R27
ab 2
0,09 0,09
A29 ab
2 0,11
0,11 A23
ab 2
0,18 0,18
R25 ab
2 0,18
0,18 R21
ab 2
0,26 0,26
R32 ab
2 0,29
0,29 R14
b 2
0,32 A22
c 2
1,44 Sig.
0,05 0,06
1,00 Means for groups in homogeneous subsets are displayed.
Lampiran 2 . Kompetisi BAL asal ASI dengan C. sakazakii YRC3a dalam susu
formula rekonstitusi pada suhu 27 °C selama 24 jam
•
Lampiran 2a .
Perubahan jumlah C. sakazakii YRC3a
YRC3a
pada media TSAYE-SC
Keterangan Jumlah Log CFUmL
Δ Log CFUmL Rata-
rata SB
U 1 U 2
Δ U1 24-0
jam Δ U2
24-0 jam
0 jam 24 jam
0 jam 24 jam
R14 vs YRC3a 2,69
7,11 2,69
7,04 4,41
4,36 4,38
0,01 R21 vs YRC3a
2,89 4,13
2,96 4,31
1,24 1,34
1,29 0,04
R23 vs YRC3a 3,06
9,89 3,02
9,91 6,83
6,89 6,86
0,02 R25 vs YRC3a
3,06 6,63
3,02 6,45
3,57 3,43
3,50 0,05
•
Lampiran 2b .
Perubahan jumlah pada media MRSA-AA
Keterangan Jumlah Log CFUmL
Δ Log CFUmL Rata-
rata SB
U 1 U 2
Δ U1 24-0
jam Δ U2
24-0 jam
0 jam 24 jam
0 jam 24 jam
R14 vs YRC3a 7,42
10,04 7,39
10,03 2,61
2,64 2,63
0,01 R21 vs YRC3a
7,79 9,39
7,67 9,40
1,59 1,73
1,66 0,05
R23 vs YRC3a 7,41
9,89 7,30
9,92 2,48
2,61 2,54
0,05 R25 vs YRC3a
7,48 9,86
7,60 9,89
2,38 2,29
2,34 0,03
Lampiran 2c .
Penghambatan C. sakazakii YRC3a Keterangan
Rata-rata STDEV
Penghambatan R14 vs YRC3a
4,38 0,02
-0,27 R21 vs YRC3a
1,29 0,04
2,82 R23 vs YRC3a
6,86 0,02
-2,75 R25 vs YRC3a
3,50 0,05
0,61 YRC3a
4,11 0,07
-
Uji F: ANOVA Single Factor
Penghambatan jumlah C. sakazakii YRC3a
Sumber Keragaman Jumlah
Kuadrat Derajat
Bebas Kuadrat
Tengah F-hit
F-tab
Between Groups 31,89
4 7,97
1,04E3 5,19
Within Groups 0,04
5 0,01
Total 31,93
9
Hipotesis : Ho: µ1= µ2= µ3= µ4= µ5
Ho: Minimal Terdapat sepasang µi≠ µi’ Keputusan
: Berdasarkan Hasil uji-F dapat dilihat minimal terdapat sepasang perlakuan yang memberikan pengaruh yang berbeda nyata pada
taraf α=0,05 dimana FhitFtab
Uji Lanjut: Uji pembanding berpasangan uji Duncan
Penghambatan C. sakazakii YRC3a Duncan
a
Kompetisi BAL vs Cronobacter
N Subset for alpha = 0.05
Keputusan 1
2 3
4 5
R21 vs YRC3a a
2 1.29E0
R25 vs YRC3a b
2 3.50E0
Kontrol YRC3a c
2 4.11E0
R14 vs YRC3a d
2 4.38E0
R23 vs YRC3a e
2 6.86E0
Sig. 1.00
1.00 1.00
1.00 1.00
Means for groups in homogeneous subsets are displayed.
Lampiran 3. Perubahan nilai pH pada kompetisi BAL asal ASI dengan C. sakazakii
YRC3a dalam susu formula rekonstitusi
Perlakuan pH 0 jam
pH 24 jam YRC3a vs R14
7,7 4,5
YRC3a vs R21 7,6
4,5 YRC3a vs R23
7,6 4,4
YRC3a vs R25 7,6
4,4 Kontrol R14
7,6 4,9
Kontrol R21 7,6
4,8 Kontrol R23
7,7 4,8
Kontrol R25 7,7
4,8 Kontrol YRC3a
7,8 5,7
Lampiran 4. Pengeringan beku dan uji viabilitas BAL
• KODE ISOLAT: L. rhamnosus R21
Ʃ No biomassa sel +kriogenik CFUmL
Ʃ Nb setelah pembekuan CFUmL
Ʃ Nt setelah pengeringan beku CFUg
Pengenceran U1
U2 Pengenceran
U1 U2
Pengenceran U1
U2 10
-8
TBUD TBUD
10
-7
TBUD TBUD
10
-7
TBUD TBUD
TBUD TBUD
TBUD TBUD
TBUD TBUD
10
-9
156 135
10
-8
TBUD TBUD
10
-8
TBUD TBUD
127 100
TBUD TBUD
TBUD TBUD
10
-10
17 13
10
-9
80 73
10
-9
112 185
15 14
80 68
140 145
10
-11
2 3
10
-10
8 10
10
-10
25 46
2 7
5 39
35 Ʃ
1,42.10
11
1,18.10
11
Ʃ 8,00.10
10
7,05.10
10
Ʃ 1,43.10
11
1,86.10
11
Log 11,15
11,07 Log
10,90 10,45
Log 11,16
11,27
• KODE ISOLAT: L. rhamnosus R25
Ʃ No biomassa sel +Kriogenik CFUmL
Ʃ Nb setelah pembekuan CFUmL
Ʃ Nt setelah pengeringan beku CFUg
Pengenceran U1
U2 Pengenceran
U1 U2
Pengenceran U1
U2 10
-8
TBUD TBUD
10
-7
TBUD TBUD
10
-7
TBUD TBUD
TBUD TBUD
TBUD TBUD
TBUD TBUD
10
-9
111 140
10
-8
TBUD TBUD
10
-8
TBUD TBUD
159 110
TBUD TBUD
TBUD TBUD
10
-10
11 28
10
-9
80 67
10
-9
202 212
13 24
73 76
183 226
10
-11
2 1
10
-10
8 2
10
-10
50 53
7 8
7 47
56 Ʃ
1,31.10
11
1,32.10
11
Ʃ 7,65.10
10
7,15.10
10
Ʃ 2,19.10
11
2,48.10
11
Log 11,12
11,12 Log
10,88 10,85
Log 11,34
11,39 Keterangan: Jumlah bakteri merupakan hasil rata-rata 2 kali ulangan dan duplo; No: Jumlah mikroba awal+laktosa;
Nb: Jumlah mikroba setelah pembekuan; Nt: Jumlah mikroba setelah freeze driying
Lampiran 5. Gambar persiapan dan proses pembuatan BAL kering beku
Lampiran 5a: Proses penyalutan sel BAL Lampiran 5b: Produksi sel BAL dengan kriogenik laktosa pada medium MRSB
Lampiran 5c : Proses pengeringan beku BAL
Lampiran 5d : Kondisi pengaturan alat pada proses pengeringan beku BAL
Lampiran 5e : Uji viabilitas BAL setelah pengeringan beku
Lampiran 6
: Uji Kompetisi BAL kering beku R21 dan R25 dengan C.sakazakii YRC3a pada T= 50;60;70 °C; dan selama Hang time t= 0;2;4;6;8 jam
Lampiran 6a . Data kompetisi R21 vs YRC3a pada media MRSAA-AA pada suhu rekonstitusi 50
o
C
Jam ke-
Suhu 50
o
C
Rata-rata Ulangan 1
Ulangan 2 Pengenceran
∑Koloni CFUmL Log
CFUmL Pengenceran
∑Koloni CFUmL Log
CFUmL CFUmL
Log CFUmL
10
-5
TBUD
1.43x10
8
8.16
10
-4
TBUD
1.47x10
8
8.17 1.45x10