BIOREAKTOR

Medium Pertumbuhan Preculture

3%
27%
Untuk meningkatkan
Produk Bioreakor
Untuk meningkatkan pertumbuhan
97% P. Pastoris
73%
produksi Pre-Insulin
Glyserol Methanol
Nutrisi umum lain
Nutrisi Umum Lain
Udara
BIOSTAT-C BIOREACTOR
Fermentation
• Highly reproducible
• Intracellular
• E. Coli W3110 with a pB322-derived plasmid coding
• Antibiotic resistance of geneticaly modified E.coli strain
• Typically tetracycline and ampicillin resistant
Fermentation
• Glucose and mineral salt medium
• Exponentially growing cells in batch mode
oTransferred from seed fermentor to production bioreactor
o10% of final volume
• High cell density fermentation
Fermentation
• Fed-batch bioreactor
oGlucose feed ~ 500-600 g/L
oMay use peptone for amino acid source
oTemperature near maximum tolerance of E. Coli (37 C)
oAgitation generally ranges 100-1500 rpm
oSlightly positive pressure
PURIFIKASI
Centrifuge
 Protein Campuran

Pemisahan didasarkan
pada tingkat affinitas
protein terhadap
logam
Pre-insulin Kasar

Immobilized Metal Ion Affinity Chromatography

(IMAC)
Pre-insulin Kasar

Pemisahan didasarkan
pada muatan Protein

Pre-insulin

Cation Exchange Chromatography

Transpeptidation
 Pre-Insulin TFA
(Trifluoroacetic acid)
Transpeptidation
Sistem Batch selama 24 Jam
Pada pH 6
Suhu 12 C Insulin
Deprotection
Insulin Amino Acids L-Glutamine
dan L-tryptophan

Insulin dengan
Struktur lebih kuat
Purifikasi Akhir
Insulin Kadar Sedang

Insulin Kadar Tinggi (99%)

Semi Preparative Chromatography