Metakrilik asit aşılanmış kitosan kürelerin karakterizasyonu ve lizozim enziminin saflaştırılmasında kullanılması
Date
2007
Authors
Journal Title
Journal ISSN
Volume Title
Publisher
Uludağ Üniversitesi
Abstract
Bu çalısmada, yumurta beyazından doğrudan lizozim enziminin saflastırılması için yeni bir kromatografik matriks gelistirildi. Destek materyali olarak, kitosan-g-pMAA küreleri kullanıldı. Kesikli sistemde bu afinite adsorbentleri üzerine farklı pH'larda farklı miktarda lizozim içeren sulu ortamdan lizozim adsorpsiyonu davranısı incelendi. Lizozimim iyon değistirici kürelere adsorpsiyonu için optimal pH değeri 4.0- 8.0 değerleri arasında arastırıldı. Sıcaklık ve iyonik siddetin, adsorbentlerin, adsorpsiyon kapasiteleri üzerindeki etkisi belirlendi. Kitosan-g-pMAA iyon değistirici kürelerin lizozim adsorpsiyon kapasitesi, sırasıyla 65.7 mg/g olarak belirlendi. ?mmobilize lizozim enziminin baslangıçtaki aktifliğini koruyarak 7 kez tekrar kulanılabildiği belirlendi. Yumurta beyazından doğrudan lizozim enzimi saflastırması sonuçları YBSK ile tayin edildi. Kitosan-g-pMAA iyon değistirici kürelerin, lizozim enziminin yumurta akından saflastırılmasında kullanılmasından sonra, lizozim enziminin 1.0 M KSCN ile elüsyonları yapıldı. Kitosan-g-pMAA iyon değistirici kürelerden elüe edilen lizozimin saflık derecesi ve geri kazanımları sırasıyla %94.16 ve %87.30 olarak belirlendi. Kitosan-g-pMAA küreler ile saflastırılan lizozimin spesifik aktivitesi 40486 U mg-1 olarak belirlendi.Ayrıstırma ve saflastırma islemlerinde lizozim aktivitesi, substrat olarak Micrococcus lysodeikticus bakterisi kullanılarak belirlendi.
In this study, for direct purification of lysozyme enzyme from egg white, a new chromatography matrixs was developed. Chitosan-g-pMAA ion-exchange beads were used as support material.Behaviour of lysozyme adsorption onto these ion-exchange beads, from Aqueous solutions containing different amounts of lysozyme at different pH, investigated in a batch system. The optimal pH values for adsorption of lysozyme on the affinity adsorbents, were investigated in the pH range of 4.0-8.0. The effects of temperature and ionic strenght,on the adsorption capacities of the adsorbents, were determined. The lysozyme adsorption capacity of the chitosan-g-pMAA ion-exchange beads was found as 65.7 mg/g, respectively. It was determined that the orginial activity was preserved when immobilized lysozyme was used repeatedly 7 times. The results of direct lysozyme enzyme purification from egg white were determined by HPLC. After usage of chitosan-g-pMAA ion-exchange beads the lysozyme enzyme from the egg white, lysozyme enzyme was eluted by 1.0 M KSCN solution. The purity and the recovery of the eluted lysozyme, were 94.16% and 87.30 % for chitosan-g-pMAA ion-exchange beads, respectively. The specific activity of the lysozyme purified with Kitosan-g-pMAA beads 40486 Umg-1. Separation and purification was monitored by determination of lysozyme activity using Micrococcus lysodeikticus as substrate.
In this study, for direct purification of lysozyme enzyme from egg white, a new chromatography matrixs was developed. Chitosan-g-pMAA ion-exchange beads were used as support material.Behaviour of lysozyme adsorption onto these ion-exchange beads, from Aqueous solutions containing different amounts of lysozyme at different pH, investigated in a batch system. The optimal pH values for adsorption of lysozyme on the affinity adsorbents, were investigated in the pH range of 4.0-8.0. The effects of temperature and ionic strenght,on the adsorption capacities of the adsorbents, were determined. The lysozyme adsorption capacity of the chitosan-g-pMAA ion-exchange beads was found as 65.7 mg/g, respectively. It was determined that the orginial activity was preserved when immobilized lysozyme was used repeatedly 7 times. The results of direct lysozyme enzyme purification from egg white were determined by HPLC. After usage of chitosan-g-pMAA ion-exchange beads the lysozyme enzyme from the egg white, lysozyme enzyme was eluted by 1.0 M KSCN solution. The purity and the recovery of the eluted lysozyme, were 94.16% and 87.30 % for chitosan-g-pMAA ion-exchange beads, respectively. The specific activity of the lysozyme purified with Kitosan-g-pMAA beads 40486 Umg-1. Separation and purification was monitored by determination of lysozyme activity using Micrococcus lysodeikticus as substrate.
Description
Keywords
Kitosan küre, Lizozim, İmmobilizasyon, İyon değişim, Yumurta beyazı, Aşı kopolimer, Metakrilik asit, Chitosan bead, Lysozyme, Immobilization, Ion-exchange, Metacrylic acid, Graft copolymer, Egg white
Citation
Ekici, G. (2007). Metakrilik asit aşılanmış kitosan kürelerin karakterizasyonu ve lizozim enziminin saflaştırılmasında kullanılması. Yayınlanmamış yüksek lisans tezi. Uludağ Üniversitesi Fen Bilimleri Enstitüsü.