PENGARUH SORBET BUAH NAGA (Hylocereus polyrhizus) DENGAN PENAMBAHAN ISOLAT PROTEIN Spirulina platensis TERHADAP PERUBAHAN KADAR GULA DARAH DAN BERAT BADAN PADA TIKUS WISTAR

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45 7. LAMPIRAN

Lampiran 1. Isolasi Protein Spirulina platensis 7.1. Isolasi Protein Spirulina platensis

a b c d

g f e

Gambar 8. Penimbangan Spirulina platensis (a); Proses pemecahan dinding sel dengan menggunakan sonikator (b); Proses Salting out (c); Sentrifugasi untuk memisahkan dinding sel (d); Sentrifugasi dingin untuk mengendapkan protein (e); Pemisahan endapan protein (f); Isolat protein jadi setelah freeze drying (g).

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Efisiensi isolat protein dihitung berdasarkan perbandingan berat isolat protein yang telah dikeringkan dengan freeze dryer dengan berat Spirulina platensis kering sebelum dilakukan ekstraksi. Angka yang digunakan merupakan hasil berat isolat protein dari 1 batch percobaan dengan 3 kali ulangan.

 Jumlah sampel kering = 1 gram

 Berat isolat kering (gram) = 0,915 + 0,823 + 0,871 = 2,609  Rata-rata berat isolat protein = 2,609

3 = 0,869667  Efisiensi isolat protein = 0,869667₁ x 100% = 86,968%

Lampiran 2. Formulasi Penambahan Isolat Protein Spirulina platensis 7.3. Pembuatan Sorbet Buah Naga

a b c

d

Gambar 9. Pengupasan kulit buah naga (a); Penghalusan buah naga, disertai penambahan air dan isolat protein Spirulina platensis (b); Pembuatan Sorbet buah naga dengan menggunakan ice cream maker (c); Sorbet naga jadi (d).

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7.4. Perhitungan Penambahan Isolat Protein Spirulina platensis

Penambahan isolat protein Spirulina platensis menggunakan asumsi berat badan maksimal tikus pada saat pemilihan adalah 150gram dan akan disonde setiap hari masing-masing tikus 3,5ml. Selain itu asumsi 150gram buah adalah 150ml sehingga pembuatan sorbet akan menghasilkan 200ml sorbet buah naga merah.

7.4.1. Penambahan Isolat Protein Spirulina platensis 25 mg/KgBB Isolat yang ditambah = _{1000 𝑔𝑟𝑎𝑚}25𝑚𝑔 x 150gram x 200𝑚𝑙_3,5𝑚𝑙 = 0,214gram

7.4.2. Penambahan Isolat Protein Spirulina platensis 50 mg/KgBB Isolat yang ditambah = _{1000 𝑔𝑟𝑎𝑚}50𝑚𝑔 x 150gram x 200𝑚𝑙_3,5𝑚𝑙 = 0,429gram

Lampiran 3. Analisa Proksimat Sorbet Buah Naga Merah 7.5. Perhitungan proksimat sorbet buah naga merah

Analisa proksimat dihitung berdasarkan pengukuran kadar air, kadar abu, kadar lemak, kadar protein, dan kadar karbohidrat yang dilakukan dari 2 batch percobaan dengan 3 kali ulagan.

7.5.1. Sorbet Buah naga merah 7.5.1.1.Kadar Air  94,283−72,016_25,070 x100% = 88,819%  91,420−69,239_25,001 x 100% = 88,720%  90,097−67,949_25,001 x 100% = 88,588%  45,266−27,224_20,099 x 100% = 89,766%  42,424−24,181_20,060 x 100% = 90,942%  442,509−2,080_20,080 x 100% = 90,911%

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 29,783−29,296_0,509 x 100% = 4,322% → 100%−88,819%_100% 𝑥 4,322 % = 0,483%  31,693−31,171_0,534 x 100% = 2,247% → 100%−88,720%_100% 𝑥 3,770% = 0,253%  32,291−31,782_0,516 x 100% = 1,357% → 100%−88,588%_100% 𝑥 3,187 % = 0,155%  39,954−39,431_0,539 x 100% = 2,968% → 100%−89,766%_100% 𝑥 3,937 % = 0,304%  40,752−40,249_0,516 x 100% = 2,519% → 100%−90,942% 100% 𝑥 3,543 % = 0,228%  41,868−41,329_0,545 x 100% = 1,101% → 100%−90,911%_100% 𝑥 5,118% = 0,100% Rata-rata kadar abu = 0,483%+0,253%+0,155%+0,304%+0,228%+0,100%

6 = 0,254% 7.5.1.3.Kadar Lemak  29,824−29,817_0,5 x 100% = 1,4% →100%−88,819%_100% 𝑥 1,4 % = 0,157%  32,988−32,986_0,5 x 100% = 0,4% → 100%−88,720%_100% 𝑥 0,4% = 0,045%  28,598−28,589_0,5 x 100% = 1,8% → 100%−88,588%_100% 𝑥 1,8 % = 0,205%  40,241−40,237_0,5 x 100% = 0,8% → 100%−89,766%_100% 𝑥 0,8% = 0,082%  39,421−39,416_0,5 x 100% = 1% → 100%−90,942%_100% 𝑥 1% = 0,091%  41,328−41,326_0,5 x 100% = 0,4% → 100%−90,911%_100% 𝑥 0,4% = 0,036%

Rata-rata kadar lemak = 0,044%+0,087%+0,125%+0,036%+0,073%+0,143%₆ = 0,085%

7.5.1.4.Kadar Protein  %N = _0,5𝑥10001,7−0,2 x 0,1 x 14,007 x 100% = 0,476% %P = 0,476% x 6,25 = 2,976% → 100%−88,819%_100% 𝑥 2,976 % = 0,333%  %N = _0,5𝑥10001,9−0,2 x 0,1 x 14,007 x 100% = 0,532% %P = 0,532% x 6,25 = 3,327% → 100%−88,720%_100% 𝑥 3,327% = 0,375%  %N = 2,0−0,2 0,5𝑥1000 x 0,1 x 14,007 x 100% = 0,560% %P = 0,560% x 6,25 = 3,502% → 100%−88,588%_100% 𝑥 3,502 % = 0,400%

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 %N = 2,4−0,2 0,5𝑥1000 x 0,1 x 14,007 x 100% = 0,672% %P = 0,672% x 6,25 = 4,202% →100%−89,766%_100% 𝑥 4,202 % = 0,430%  %N = _0,5𝑥10003,4−0,2 x 0,1 x 14,007 x 100% = 0,952% %P = 0,952% x 6,25 = 5,953% → 100%−90,942%_100% 𝑥 5,953 % = 0,539%  %N = _0,5𝑥10002,5−0,2 x 0,1 x 14,007 x 100% = 0,700% %P = 0,700% x 6,25 = 4,377% → 100%−90,911%_100% 𝑥 4,377% = 0,398% Rata-rata kadar protein = 0,333%+0,375%+0,400%+0,430%+0,539%+0,398%₆ = 0,412%

7.5.1.5.Kadar Karbohidrat  By difference = 100% - (88,819%+0,483%+0,157%+0,333%) = 10,208%  By difference = 100% - (88,720%+0,253%+0,045%+0,375%) = 10,606%  By difference = 100% - (88,588%+0,155%+0,205%+0,400%) = 10,652%  By difference = 100% - (89,766%+0,304%+0,082%+0,430%) = 9,419%  By difference = 100% - (90,942 %+0,228%+0,091%+0,539%) = 8,200%  By difference = 100% - (90,911%+0,100%+0,036%+0,398%) = 8,554% Rata-rata kadar karbohidrat = 10,208%+10,606%+10,652%+9,419%+8,200%+8,554%₆ = 9,606%

7.5.2. Sorbet Buah naga merah + Isolat protein 25mg/KgBB 7.5.2.1.Kadar Air  99,854−77,302_25,097 x100% = 89,859%  84,420−61,844_25,060 x 100% = 90,088%  96,888−74,356_25,028 x 100% = 90,027%  65,362−42,943_25,127 x 100% = 89,223%  64,618−42,108_25,205 x 100% = 89,308%  66,336−44,052_25,016 x 100% = 89,079%

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 27,958−27,465_0,503 x 100% = 1,988% → 100%−89,859%_100% 𝑥 1,988 % = 0,202%  25,956−25,469_0,503 x 100% = 3,181% → 100%−90,088%_100% 𝑥 3,181% = 0,315%  27,914−27,428_0,506 x 100% = 3,953% → 100%−90,027%_100% 𝑥 3,953 % = 0,394%  31,736−31,261_0,503 x 100% = 5,567% → 100%−89,223%_100% 𝑥 5,567% = 0,600%  32,977−32,498_0,503 x 100% = 4,771% → 100%−89,308% 100% 𝑥 4,771% = 0,510%  34,659−34,181_0,503 x 100% = 4,970% → 100%−89,079%_100% 𝑥 4,970% = 0,543% Rata-rata kadar abu = 0,202%+0,315%+0,394%+0,600%+0,510%+0,543%

6 = 0,427% 7.5.2.3.Kadar Lemak  38,927−38,925_0,5 x 100% = 0,4% → 100%−89,859%_100% 𝑥 0,4 % = 0,041%  39,520−39,516_0,5 x 100% = 0,8% → 100%−90,088%_100% 𝑥 0,8% = 0,079%  40,864−40,858_0,5 x 100% = 1,2% → 100%−90,027%_100% 𝑥 1,2 % = 0,120%  42,075−42,072_0,5 x 100% = 0,6% → 100%−89,308%_100% 𝑥 0,6 % = 0,065%  42,015−42,012_0,5 x 100% = 0,6% →100%−89,308%_100% 𝑥 0,6 % = 0,064%  43,749−43,745_0,5 x 100% = 0,8% → 100%−89,079%_100% 𝑥 0,8% = 0,087%

7.5.2.4.Kadar Protein  %N = 3,5−0,2 0,5𝑥1000 x 0,1 x 14,007 x 100% = 0,980% %P = 0,980% x 6,25 = 6,128% → 100%−78,511%_100% 𝑥 6,128% = 0,621%  %N = _0,5𝑥10003,8−0,2 x 0,1 x 14,007 x 100% = 1,065% %P = 1,065% x 6,25 = 6,653% → 100%−78,060%_100% 𝑥 6,653% = 0,659%  %N = _0,5𝑥10003,6−0,2 x 0,1 x 14,007 x 100% = 1,009% %P = 1,009% x 6,25 = 6,303% → 100%−78,050%_100% 𝑥 6,303% = 0,629%

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 %N = _0,5𝑥10005,9−0,2 x 0,1 x 14,007 x 100% = 1,653% %P = 1,653% x 6,25 = 10,330% →100%−80,976%_100% 𝑥 10,330% = 1,113%  %N = 6,3−0,2 0,5𝑥1000 x 0,1 x 14,007 x 100% = 1,765% %P = 1,765% x 6,25 = 11,031% → 100%−81,055%_100% 𝑥 11,031% = 1,179%  %N = _0,5𝑥10006,3−0,2 x 0,1 x 14,007 x 100% = 1,765% %P = 1,765% x 6,25 = 11,031% → 100%−81,069%_100% 𝑥 11,031% = 1,205% Rata-rata kadar protein = 0,621%+0,659%+0,629%+1,113%+1,179%+1,205%₆ = 0,901%

7.5.2.5.Kadar Karbohidrat  By difference = 100% - (89,859%+0,202%+0,041%+0,621%) = 9,277%  By difference = 100% - (90,088%+0,315%+0,079%+0,659%) = 8,858%  By difference = 100% - (90,027%+0,394%+0,120%+0,629%) = 8,830%  By difference = 100% - (89,223%+0,600%+0,065%+1,113%) = 8,999%  By difference = 100% - (89,308%+0,510%+0,064%+1,179%) = 8,939%  By difference = 100% - (89,079%+0,543%+0,087%+1,205%) = 9,086% Rata-rata kadar karbohidrat = 9,277%+8,858%+8,830%+8,999%+8,939%+9,086%₆ = 8,998 %

7.5.3. Sorbet Buah naga merah + Isolat protein 50mg/KgBB 7.5.3.1.Kadar Air  94,055−71,630_25,033 x100% = 89,582%  85,911−63,453_25,019 x 100% = 89,764%  101,199−78,570_25,040 x 100% = 90,371%  99,957−77,375_25,218 x 100% = 89,547%  92,170−69,728_25,154 x 100% = 89,764%  97,010−74,520_25,152 x 100% = 89,416%

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 23,288−22,793_0,507 x 100% = 2,367% → 100%−89,582%_100% 𝑥 2,367% = 0,247%  23,998−23,497_0,508 x 100% = 1,378% → 100%−89,764%_100% 𝑥 1,378% = 0,141%  24,695−24,197_0,507 x 100% = 1,775% → 100%−90,371%_100% 𝑥 1,775% = 0,171%  30,316−29,830_0,502 x 100% = 3,187% → 100%−89,547%_100% 𝑥 3,187% = 0,333%  33,483−33,001_0,503 x 100% = 3,777% → 100%−89,764% 100% 𝑥 3,777% = 0,407%  29,088−28,601_0,504 x 100% = 3,373% → 100%−89,416%_100% 𝑥 3,373% = 0,357% Rata-rata kadar abu = 0,247%+0,141%+0,171%+0,333%+0,407%+0,357%

6 = 0,276% 7.5.3.3.Kadar Lemak  42,081−42,073_0,5 x 100% = 1,6% → 100%−89,582%_100% 𝑥 1,6 % = 0,167%  43,750−43,739_0,5 x 100% = 2,2% → 100%−89,764%_100% 𝑥 2,2% = 0,225%  42,038−42,020_0,5 x 100% = 3,6% → 100%−90,371%_100% 𝑥 3,6% = 0,347%  44,896−44,895_0,5 x 100% = 0,2% → 100%−89,547%_100% 𝑥 0,2 % = 0,021%  40,860−40,857_0,5 x 100% = 0,6% → 100%−89,764%_100% 𝑥 0,6% = 0,065%  38,917−38,914_0,5 x 100% = 0,6% → 100%−89,416%_100% 𝑥 0,6% = 0,064%

7.5.3.4.Kadar Protein  %N = _0,5𝑥10003,8−0,2 x 0,1 x 14,007 x 100% = 1,065% %P = 1,065% x 6,25 = 6,653% → 100%−89,582%_100% 𝑥 6,653 % = 0,693%  %N = _0,5𝑥10004,9−0,2 x 0,1 x 14,007 x 100% = 1,373% %P = 1,373% x 6,25 = 8,579% → 100%−89,764%_100% 𝑥 8,579% = 0,878%  %N = 4,3−0,2 0,5𝑥1000 x 0,1 x 14,007 x 100% = 1,205% %P = 1,205% x 6,25 = 7,529% → 100%−90,371%_100% 𝑥 7,529% = 0,725%

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 %N = 10,1−0,2 0,5𝑥1000 x 0,1 x 14,007 x 100% = 2,829% %P = 2,829% x 6,25 = 17,684% →100%−89,547%_100% 𝑥 17,684% = 1,848%  %N = _0,5𝑥100010,4−0,2 x 0,1 x 14,007 x 100% = 2,913% %P = 2,913% x 6,25 = 18,209% → 100%−89,764%_100% 𝑥 18,209% = 1,963%  %N = _0,5𝑥100010,2−0,2 x 0,1 x 14,007 x 100% = 2,857% %P = 2,857% x 6,25 = 17,859% → 100%−89,416%_100% 𝑥 17,859% = 1,890% Rata-rata kadar protein = 0,693%+0,878%+0,725%+1,848%+1,963%+1,890%₆ = 1,333 %

7.5.3.5.Kadar Karbohidrat  By difference = 100% - (89,582%+0,247%+0,167%+0,693%) = 9,312%  By difference = 100% - (89,764%+0,141%+0,225%+0,878%) = 8,992%  By difference = 100% - (90,906%+0,171%+0,347%+0,725%) = 8,386%  By difference = 100% - (89,547%+0,333%+0,021%+1,848%) = 8,250%  By difference = 100% - (79,961%+0,407%+0,065%+1,963%) = 8,346%  By difference = 100% - (80,647%+0,357%+0,064%+1,890%) = 8,273% Rata-rata kadar karbohidrat = 9,312%+8,992%+8,386%+8,250%+8,346%+8,273%₆ = 8,593%

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Hasil pengujian proksimat sorbet buah naga merah diuji normalitas dan one way anova untuk melihat perbedaan antar formulasi.

7.6. Analisis Statistik Proksimat Sorbet Buah Naga Merah 7.6.1. Uji Normalitas

6.1.1. Uji Anova

7.6.2. Uji One Way Anova

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Post Hoc Tests

Homogeneous Subsets

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Lampiran 5. Pengujian In Vivo 7.7. Pengujian In Vivo

7.7.1. Pelaksanaan Pengujian In Vivo

(a)

(b)

(d) (c)

(f) (e)

Gambar 10. Sortasi tikus (a); Induksi streptozotocin secara intraperitoneal (b); Pengukuran kadar gula darah (c); Penimbangan berat badan (d); Pemberian treatment secara sonde oral (e); Kondisi tikus dalam kandang selama pengujian (f).

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7.7.2. Dosis Induksi Streptozotocin

 Berat badan tikus maksimal pada saat pemilihan 150gram.  Dosis 45 mg/kgBB.

 Tikus dengan berat 150 gram akan diinjeksi streptozotocin yang dilarutkan dalam buffer sitrat-fosfat sebanyak 5 ml.

 Jumlah streptozotocin yang harus ada dalam 5 ml buffer sitrat-fosfat adalah 45𝑚𝑔

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7.7.3. Data Kadar Gula Darah dan Berat Badan Tikus

7.7.2.1. Data Kadar Gula Darah dan Berat Badan Tikus Dengan pakan standar

Tikus Awal

IP STZ

(pre-treatment) hari ke-7 hari ke-14 hari ke-21 hari ke-28

Post-Treatment BB KGD Dosis BB KGD BB KGD BB KGD BB KGD BB KGD BB KGD 1 133 97 0,44 133 119 155 80 177 141 192 103 224 131 240 116 2 112 91 0,37 147 86 151 85 154 89 157 91 182 89 194 103 3 123 99 0,41 113 66 116 572 139 511 160 551 178 337 187 94 4 133 100 0,44 160 154 169 149 200 118 206 108 237 124 259 114 5 142 103 0,47 194 95 190 97 186 94 201 155 232 108 239 88

7.7.3.2. Data Kadar Gula Darah dan Berat Badan Tikus Treatment Sonde Sorbet Buah Naga Merah

Tikus Awal

IP STZ

Post-Treatment BB KGD Dosis BB KGD BB KGD BB KGD BB KGD BB KGD BB KGD 1 126 95 0.42 152 308 190 134 218 125 239 127 255 101 264 94 2 105 107 0.35 133 131 175 99 208 101 220 75 235 94 245 68 3 113 103 0.37 141 188 171 105 190 106 212 89 221 89 230 72 4 134 100 0.44 100 110 188 116 212 96 234 80 247 70 261 70 5 121 98 0.4 136 193 157 110 170 114 183 108 187 84 195 72

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Isolat Protein Spirulina platensis 25 mg/kg BB.

Tikus Awal

IP STZ

Post-Treatment BB KGD Dosis BB KGD BB KGD BB KGD BB KGD BB KGD BB KGD 1 145 81 0.48 142 563 164 459 156 390 165 420 162 165 157 172 2 140 91 0.46 202 128 156 221 - - - - 3 135 99 0.45 190 140 200 105 219 112 230 112 232 101 239 87 4 124 97 0.41 99 382 149 464 161 284 170 118 183 185 195 106 5 126 105 0.42 158 496 178 131 185 128 230 140 247 69 263 130 Keterangan : (-) : tidak dapat diukur karena mati

7.7.3.4. Data Kadar Gula Darah dan Berat Badan Tikus Treatment Sonde Sorbet Buah Naga Merah dengan Penambahan Isolat Protein Spirulina platensis 50 mg/kg BB.

Tikus Awal

IP STZ

Post-Treatment BB KGD Dosis BB KGD BB KGD BB KGD BB KGD BB KGD BB KGD 1 149 88 0.49 176 191 224 119 251 118 263 121 264 62 289 106 2 107 123 0.35 156 163 169 165 188 114 190 73 203 97 214 89 3 142 92 0.47 162 392 178 131 186 106 201 108 203 59 222 94 4 124 88 0.41 171 110 163 85 162 91 158 58 177 73 190 68 5 140 108 0.46 175 300 187 122 203 84 206 70 218 85 226 94