SKRIPSI Jurusan Kimia - Fakultas MIPA UM, 2015

Ukuran Huruf:  Kecil  Sedang  Besar

OPTIMASI PEMURNIAN ENZIM PROTEASE ALKALI Bacillus brevis MENGGUNAKAN METODE FRAKSINASI AMONIUM SULFAT

APRIANA WIKE NUR MAHARIYANTI

Abstrak


ABSTRAK

 

Mahariyanti, Apriana Wike Nur. 2015. Optimasi Pemurnian Enzim Protease Alkali Bacillus brevis Menggunakan Metode Fraksinasi Amonium Sulfat. Skripsi, Jurusan Kimia, FMIPA, Universitas Negeri Malang. Pembimbing: (I) Dr. Suharti, S.Pd., M.Si., (II) Eli Hendrik, S.Si., M.Si.

 

Kata kunci : protease alkali, Bacillus brevis, pemurnian, fraksinasi amonium sulfat, aktivitas spesifik

 

Protease merupakan kelompok enzim hidrolase yang mengkatalisis reaksi hidrolisis ikatan peptida pada protein.Enzim ini banyak dimanfaatkan dalam bidang industri, antara lain industri deterjen, penyamakan kulit, dan tes diagnostik. Pada penelitian sebelumnya,protease dari Bacillus brevis telah dicoba dimurnikan tetapi memiliki aktivitas spesifik yang masih rendah sehingga perlu dilakukan perbaikan metode pemurnian. Penelitian ini bertujuan untuk mengoptimalkan pemurnian protease alkali dari Bacillus brevisdengan metode fraksinasi amonium sulfat.

Penelitian ini adalah penelitian eksperimen laboratoris. Penelitian dilakukan dalam empat tahap, yaitu (1) regenerasi bakteri Bacillus brevisdari media padat NA + susu skim; (2) produksi ekstrak kasar protease dari Bacillus brevis;(3) pemurnian enzim dengan metode fraksinasi amonium sulfat, dan (4)penentuan aktivitas dan kadar protein enzim. Penentuan aktifitas protease dilakukan dengan metode Anson pada pH 9 dan penentuan kadar protein ditentukan dengan metode Lowry. Pengendapan amonium sulfat dilakukan pada fraksi 0-20%, 20-40%, 40-60%, 60-80%, dan 80-100%.

Hasil penelitian menunjukkan bahwa aktivitas protease alkali tertinggi diperoleh pada  konsentrasi amonium sulfat 60-80% dengan aktivitas spesifik enzim sebesar 2,81 U/mg, tingkat kemurnian sebesar 3,28 kali ekstrak kasar, dan yield sebesar 7,79%. Namun, nilai aktivitas spesifik dan tingkat kemurnian tersebut tidak jauh berbeda dengan nilai aktivitas spesifik dan tingkat kemurnian pada fraksi 40-80% yaitu berturut-turut sebesar 2,48 U/mg dan 2,806  kali ekstrak kasar.Sedangkan pada penelitian sebelumnya diperoleh aktivitas spesifik dan tingkat kemurnian berturut-turut sebesar 0,245 U/mg dan 1,26 kali ekstrak kasar.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

ABSTRACT

 

Mahariyanti, Apriana Wike Nur. 2015. Optimization of Purification Bacillus brevis’s Alkaline ProteaseusingAmmoniumSulfateFractionationMethods. Undergraduate, Department of Chemistry, Mathematics and Science Faculty, State University of Malang.Advisors: (I) Dr. Suharti, S.Pd., M.Si., (II) Eli Hendrik Sanjaya, S.Si., M.Si.

 

Key words: alkaline protease, Bacillus brevis, purification, ammoniumsulfatefractionation

 

Protease is an enzyme whichcatalyzes  hydrolysis of peptide bonds in proteins. Theenzyme is widely used in industry, for example  as additve agent in detergent, dehairing, and diagnostic test. Previous research showed thatBacillus brevisproduced  alkaline protease. However, its specific activity is relatively low. Therefore, the enzyme needs to be purified. This study aims to purify the enzyme by ammonium sulfate fractination.

This study is an experimental research laboratory. The study was conducted in four stages: (1) regeneration ofBacillus brevisbacteria on solid media NA containing skim milk; (2) production of crude extract protease from Bacillus brevis; (3) purification of the enzyme by ammonium sulfate fractionation method, and (4) enzyme activity and protein concentration measurements. Protease activity was measured using Anson method at pH 9 and total  protein concentration was determined usingLowry method.Ammonium sulfate precipitation was performed at concentration of  0-20%, 20-40%, 40-60%, 60-80% and 80-100%.

The results showed that the highest alkaline protease activity was achieved at ammonium sulfateconcentration of 60-80 % which yieldedenzyme specific activity of 2,81 U/mg, enzyme purity of 3.28 times the crude extract, and the yield of 7.79%.The value of specific activity and purity levels are not much different from its compare to purified enzyme at ammonium sulfate concentration of  of 40-80%, which has specific activity and purity  the 2,48 U/mg and 2.806 times the crude extract, respectively. However, overall results are better compare to previous purification results of which only purified the enzyme at 1.26 times the crude extract with specific activity of alkaline protease 0,245 U/mg.