FRAKSINASI SELULASE MIKROBA SELULOLITIK  DENGAN AMONIUM SULFAT DAN AMOBILISASI  PADA AGAR-AGAR KOMERSIAL

Oka Ratnayani; Putu Elvira Yulianthi; I Nengah Wirajana

Oka Ratnayani Program Studi Kimia, Fakultas Matematika dan Ilmu Pengetahuan Alam, Universitas Udayana, Jimbaran, Bali, Indonesia
Putu Elvira Yulianthi Program Studi Kimia FMIPA Universitas Udayana, Bukit Jimbaran, Bali-Indonesia
I Nengah Wirajana Program Studi Kimia FMIPA Universitas Udayana, Bukit Jimbaran, Bali-Indonesia

Abstract

ABSTRAK: Penggunaan selulase yang semakin meningkat telah mendorong berbagai studi tentang eksplorasi, pemurnian, dan amobilisasi selulase. Penelitian ini bertujuan untuk fraksinasi ekstrak kasar selulase yang diperoleh dari mikroba selulolitik serta amobilisasinya pada agar-agar komersial. Produksi selulase dari mikroba selulolitik U3.1 dilakukan pada media yang mengandung substrat spesifik yaitu Carboxymethyl cellulose (CMC). Ekstrak kasar selulase dimurnikan dengan metode fraksinasi amonium sulfat dan dialisis. Ekstrak kasar selulase dan selulase murni diamobilisasi pada matrik agar-agar. Aktivitas selulase ditentukan berdasarkan analisis gula pereduksi yang dihasilkan dari reaksi enzimatis dengan reagen DNS. Kadar protein total diukur dengan metode Biuret. Aktivitas spesifik ekstrak kasar selulase dan selulase murni diperoleh berdasarkan rasio antara aktivitas selulase dengan kadar protein total tiap fraksi selulase. Fraksi 2 (selulase yang dimurnikan dengan garam amonium sulfat dengan tingkat kejenuhan 20-40%) memiliki aktivitas spesifik selulase tertinggi sebesar 3,3977 x 10^-3U/µg, dengan tingkat kemurnian 21,0317 kali dibandingkan ekstrak kasar selulase. Konsentrasi agar-agar optimum untuk amobilisasi ekstrak kasar selulase dan selulase murni adalah masing- masing 2% (b/v) dan 3% (b/v) dengan efisiensi amobil sebesar 13,99% dan 51,26%.

ABSTRACT: The increasing use of cellulase has encouraged various studies on exploration, purification and immobilization of the enzyme. This study aimed to fractionate the crude extract of cellulase obtained from cellulolytic microbe as well as immobilize it into commercial agar. The cellulase production from cellulolytic microbe U3.1 was carried out on the media containing a specific substrate of Carboxymethyl cellulose (CMC). The crude extract of cellulase was purified by fractionation of ammonium sulfate and dialysis. The crude extract and the pure cellulase were immobilized into the agar matrix. Cellulase activity was determined based on the analysis of reducing sugars produced by enzymatic reaction with DNS reagent. The total protein content was measured by using the Biuret method. The specific activity of the crude extract and the pure cellulase was obtained from the ratio of the cellulase activity to the total protein of each fraction. Fraction 2 (cellulase purified with 20-40% saturation of ammonium sulfate) had the highest specific activity of 3.3977 x 10^-3 U/µg with the purity level increased by 21.0317 times compared to the crude extract of cellulase. The optimum agar concentration for immobilization of the crude extract and the pure cellulase was 2% (b/v) and 3% (b/v) with the immobilization efficiency of 13.99% and 51.26%, respectively.

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