Sains Malaysiana 40(9)(2011): 965–972

Properties of Immobilized Candida antarctica Lipase B on Highly  Macroporous Copolymer

(Sifat Pegun Candida antarctica Lipase B ke atas Kopolimer yang Sangat Mikroporos)

Nurrahmi Handayani*, Sadijah Achmad & Deana Wahyuningrum

Department of Chemistry, Bandung Institute of Technology, Indonesia

Nemanja Miletic & Katja Loos

Zernike Institute for Advanced Materials, University of Groningen, Netherland

Received: 16 June 2010 / Accepted: 3 January 2011

ABSTRACT

In spite of their excellent catalytic properties, enzymes should be improved before their implementation both in industrial and laboratorium scales. Immobilization of enzyme is one of the ways to improve their properties. Candida antarctica lipase B (Cal-B) has been reported in numerous publications to be a particularly useful enzyme catalizing in many type of reaction including regio- and enantio- synthesis. For this case, cross-linking of immobilized Cal-B with 1,2,7,8 diepoxy octane is one of methods that proved significantly more stable from denaturation by heat, organic solvents, and proteolysis than lyophilized powder or soluble enzymes. More over, the aim of this procedure is to improve the activity and reusability of lipase. Enzyme kinetics test was carried out by transesterification reaction between 4-nitrophenyl acetate (pNPA) and methanol by varying substrate concentrations, and the result is immobilized enzymes follows the Michaelis-Menten models and their activity is match with previous experiment. Based on the Vmax values, the immobilized enzymes showed higher activity than the free enzyme. Cross-linking of immobilized lipase indicate that cross-linking by lower concentration of cross-linker, FIC (immobilized lipase that was incubated for 24 h) gave the highest activity and cross-linking by higher concentration of cross-linker, PIC (immobilized lipase that was incubated for 2 h) gives the highest activity. However, pore size and saturation level influenced their activity.

Keywords: Candida antarctica lipase B (Cal-B); cross-linking; enzyme immobilization

ABSTRAK

Walaupun sifat katalitisis yang sangat baik, enzim perlu ditingkatkan keupayaannya sebelum diimplementasikan dalam skala industri maupun makmal. Imobilisasi enzim merupakan salah satu cara untuk memperbaiki sifat enzim. Candida antarctica lipase B (Cal-B) telah dilaporkan dalam sejumlah penerbitan sebagai enzim yang dapat digunakan dalam mengkatalisis enzim dalam berbagai tindak balas termasuk sintesis yang bersifat regio- dan enantio-selektif. Dalam kajian ini didapati taut-silang dalam Cal-B terimobilisasi oleh 1,2,7,8 diepoksi oktana merupakan salah satu kaedah yang terbukti secara signifikan lebih stabil terhadap penyahaslian oleh pemanasan, pelarut organik, dan proteolisis dibandingkan dengan enzim dalam bentuk larutan maupun serbuk terliofilik. Selain itu, tujuan kajian ini adalah meningkatkan keaktifan dan kebolehulangan pemakaian lipase. Ujian kinetik enzim dilakukan melalui tindak balas transesterifikasi antara 4-nitrofenil asetat (pNPA) dan metanol dengan memvariasikan kepekatan substrat, dan hasilnya adalah bahawa enzim terimobilisasi mengikuti model Michaelis-Menten serta keaktifannya sesuai dengan hasil kajian sebelumnya. Berdasarkan nilai Vmax, enzim yang terimobilisasi menunjukkan keaktifan yang lebih tinggi daripada enzim bebas. Taut-silang pada lipase yang terimobilisasi menunjukkan bahawa taut-silang oleh adanya rangkaian silang FIC (lipase terimobilisasi yang diinkubasi selama 24 jam) berkepekatan rendah menunjukkan keaktifan tertinggi, sedangkan taut-silang oleh adanya rangkaian silang PIC (lipase terimobilisasi yang diinkubasi selama 2 jam) berkepekatan lebih tinggi menunjukkan keaktifan tertinggi. Di samping itu, saiz liang dan aras ketepuan memberikan pengaruh terhadap keaktfian enzim tersebut.

Kata kunci: Candida antarctica lipase B (Cal-B); imobilisasi enzim; taut-silang

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*Corresponding author; email: ami_chemie2003@yahoo.com