PRODUKSI SERBUK FITASE HASIL TEKNOLOGI REKOMBINAN DAN APLIKASINYA UNTUK MENINGKATKAN KUALITAS PAKAN DAN KINERJA AYAM BROILER
Adi Magna Patriadi Nuhriawangsa, Prof. Dr. Ir. Zaenal Bachruddin, M.Sc.
2012 | Disertasi | S3 Ilmu Peternakanber Penelitian tahap pertama tujuan memproduksi bakteri rekombinan dan fitase dari bakteri rekombinan yang mempunyai karakter optimum. Penelitian tahap kedua bertujuan mengetahui stabilitas fitase terhadap waktu penyimpanan dan kondisi organ pencernaan ayam dan aktivitas hidrolisis fitase secara in vitro. Penelitian tahap ketiga bertujuan mengetahui stabilitas aktivitas fitase dengan cara enkapsulasi, mencampur dengan bahan pembawa, memanfaatkan teknologi pengolahan enzim dan mengetahui umur waktu penyimpanan fitase. Penelitian tahap keempat bertujuan untuk mengetahui pengaruh serbuk fitase terhadap kinerja pertumbuhan dan kecernaan pakan ayam broiler secara in vivo. Plasmid pEAS1 ditransformasi dalam sel kompeten dan digunakan untuk produksi fitase. Produksi fitase menggunakan induktor 1,5 mM IPTG. Pemurnian fitase menggunakan kolom Ni-NTA agarosa dengan imidazole 200 mM. Bakteri rekombinan dioptimasi pH, tempertur dan IPTG. Karakterisasi fitase dengan melihat optimasi aktivitas relatif pada taraf pH, temperatur, waktu inkubasi, konsentrasi enzim dan kofaktor ion logam. Km dan Vm menggunakan metode Robyt dan White. BM fitase menggunakan poliakrilamid gel elektroforesis. Uji stabilitas fitase terhadap aras waktu simpan, temperatur dan pH berdasarkan aktivitas relatif. Hidrolisis fitase pada bekatul padi secara in vitro dengan melihat Ca, P, abu dan protease. Fitase diimobilisasi dan dipertahankan stabilitasnya menggunakan kitosan. Campuran fitase, kitosan dan media pembawa dikeringkan secara kering matahari, oven dan beku dan disimpan untuk diuji stabilitasnya. Serbuk fitase hasil teknologi rekombinan diuji secara in vivo pada ayam broiler fase grower menggunakan pakan R1: Ransum standar tanpa fitase, R2: Ransum dengan P rendah tanpa fitase dan R3: Ransum dengan P rendah ditambah serbuk fitase hasil teknologi rekombinan. Parameter yang diukur antara lain FC, FCR, ADG, kecernaan BK, Ca, P, protein kasar dan glukosa ekskretal semu, Ca, P, glukosa dan total protein darah, panjang, berat, kandungan P, Ca dan abu tulang tibia, bobot karkas, dada dan daging dada. Bakteri rekombinan dengan karakter optimum dapat diproduksi dan dapat menghasilkan fitase yang mempunyai karakter optimum. Aktivitas fitase mempunyai stabilitas tertentu dalam pencernaan ayam broiler secara in vitro. Fitase dapat dienkapsulasi dengan kitosan dan dihasilkan serbuk fitase yang siap diaplikasikan sebagai bahan tambahan pakan ayam broiler. Serbuk fitase dapat digunakan sebagai bahan tambahan untuk meningkatkan kecernaan pakan, profil darah, kinerja produksi fitase dan kinerja produksi.
The aims of the first phase of this research were to produce recombinant bacterium and the phytase from the respective recombinant bacteria, that had optimum character. The aims of the second phase of this research were to know the stability of the phytase to the storage time and conditions of the digestive organs of chickens, and to know the in vitro hydrolysis activity of phytase. The aims of third phase of this research were to know the stability of phytase activity after encapsulation, when mixed with a carrier material, using enzyme technology processing, and the length of storage time of phytase. The aims of fourth phase of this research were to determine the effect of the phytase powder on growth performance, and feed digestibility on broiler chickens in vivo. EAS1 plasmid was transformed in competent cells and used for the production of phytase. Phytase production was induced by 1.5 mM IPTG. Phytase purification was done by using Ni-NTA agarose column with 200 mM imidazole. Recombinant bacteria were optimized based on pH, temperture and IPTG values. Characterization of phytase by seeing relative activity to the degree of optimization of pH, temperature, incubation time, concentration of enzyme and cofactor metal ions. Km and Vm were evaluated using method of Robyt and White. The molecular weight of the phytase was evaluated using polyacrylamide gel electrophoresis. The phytase stability were tested on the levels of self life, temperature and pH on the relative activity. The in vitro hydrolysis activity of phytase in rice bran was done by determining Ca, P, ash and protease levels. Phytase was immobilized and maintained its stability using chitosan. The mixture of phytase, carrier, and chitosan were sun dried, oven and freeze dried and stored for stability testing. The phytase powder obtained from recombinant technology was tested in vivo on broiler chickens (grower phase) using experimental diets of R1: standard diets without phytase, R2: diets with low P without phytase and R3: diets with low P plus phytase powder. Parameters measured included FC, FCR, ADG, apparent excretal digestibility of DM, Ca, P, CP and glucose, blood Ca, P, glucose and total protein, length and weight, Ca, P, ash contents of tibia, carcasses, breast and breast meat. Recombinant bacteria and phytase could be produced with an optimum character. Phytase activity had certain stability in the digestive of broiler chickens by in vitro. Phytase can be encapsulated with chitosan and produced phytase powder that is ready to be applied as broiler chicken feed additives. Phytase powder could be used as feed additives to improve feed digestibility, blood profiles, the performance of phytase production and production performance.
Kata Kunci : Bakteri Rekombinan, Fitase dari bakteri rekombinan, Serbuk fitase hasil teknologi rekombinan, Karakterisasi, Stabilitas, Kinerja
