Phema mikroküre gömülü süper makrogözenekli kriyojel kompozit membranların üretilmesi ve protein adsorpsiyon kapasitelerinin incelenmesi
Yükleniyor...
Dosyalar
Tarih
2011
Yazarlar
Dergi Başlığı
Dergi ISSN
Cilt Başlığı
Yayıncı
Aksaray Üniversitesi Fen Bilimleri Enstitüsü
Erişim Hakkı
info:eu-repo/semantics/openAccess
Özet
Bu çalışmada, lizozimin etkin bir şekilde ayrılması için Afinite kromatografisi yöntemleri arasında, son yıllarda yoğun bir uygulama alanı bulan, İmmobilize metal afinite kromatografisi (İMAK) kullanılmıştır. Bu amaçla, yaklaşık 2 ?m çapında poli(2-hidroksietil metakrilat) (PHEMA) mikroküreler hazırlanmış, bu kürelere iminodiasetik asit (İDA) kovalent olarak bağlanmış, daha sonra da İDA üzerinden Cu2+ takılmıştır. Son olarak hazırlanan bu partiküller ile PHEMA partikül gömülü süpermakrogözenekli kriyojel kompozitler hazırlanmıştır. Bu amaçla, monomer olarak poli(2-hidroksietil metakrilat), çapraz bağlayıcı olarak N,N?-metilen-bis-akrilamid, başlatıcı/aktivatör olarak N,N,N´,N´-tetrametilen diamin (TEMED)/amonyum persülfat (APS) kullanılmış ve radikal kriyo-kopolimerizasyon yöntemiyle 5 mL?lik plastik tüplerde sentezlenmiştir. Hazırlanan Cu2+-takılı PHEMA mikroküreler gömülü süpermakrogözenekli kriyojeller yüzey alan tayini, şişme testleri, taramalı elektron mikroskobu (SEM), FTIR ile karakterize edilmiştir. Sulu çözeltiden lizozim adsorpsiyonuna pH?nın, tamponun derişimin, akış hızının, iyonik şiddetinin, ligand miktarının ve sıcaklığın etkisi incelenmiştir. Maksimum adsorpsiyon kapasitesi pH 9 tris-baz tamponunda, 5 mg/mL başlangıç lizozim derişiminde 107,14 mg/g polimer olarak belirlenmiştir. Ayrıca, Cu2+-takılı PHEMA mikroküreler gömülü süpermakrogözenekli kriyojeller üzerine enzimin defelarca adsorplanıp desorplanabildiği belirlenmiştir.
In this study, Immobilized Metal Affinity Chromatograph (IMAC) finding a wide application area among affinity chromatography methods was used for removal of lysozyme efficiently from aqueous solutions. Poly(2-hydroxyethyl methacrylate) beads with the size of about 2 ?m were prepared, and than iminodiacetic acid (IDA) was covalently attached to these beads. Then, Cu2+ ions were attached to PHEMA beads modified by IDA. And finally, supermacroporous poly (2-hydroxyethyl methacrylate) base monolithic cryogel column embedded with PHEMA-IDA-Cu2+ beads. For this purpose, supermacroporous composite cryogels were prepared by free radical polymerization using N,N,N´,N´-tetramethylene diamine (TEMED) and ammonium persulfate (APS) as initiator/activator pair, N,N´-methylene-bis-acrylamide (MBAAm) as cross-linker in a plastic syringe of 5 mL. Prepared supermacroporous cryogels with embedded PHEMA-IDA-Cu2+ beads were characterized by FTIR, SEM, sweling tests and surface area measurements. The effect of pH, buffer, ionic strength, initial lysozyme concentration, temparature and flow rate on adsorption were investigated. The maximum amount of lysozyme adsorption from aqueous solution was at 107,14 mg/g polymer in Tris-Base buffer at pH 9 with initial lysozyme concentration of 5 mg/mL. It was also observed that enzyme could be repeatedly adsorbed and desorbed on the supermacroporous cryogels with embedded PHEMA-IDA-Cu2+ beads.
In this study, Immobilized Metal Affinity Chromatograph (IMAC) finding a wide application area among affinity chromatography methods was used for removal of lysozyme efficiently from aqueous solutions. Poly(2-hydroxyethyl methacrylate) beads with the size of about 2 ?m were prepared, and than iminodiacetic acid (IDA) was covalently attached to these beads. Then, Cu2+ ions were attached to PHEMA beads modified by IDA. And finally, supermacroporous poly (2-hydroxyethyl methacrylate) base monolithic cryogel column embedded with PHEMA-IDA-Cu2+ beads. For this purpose, supermacroporous composite cryogels were prepared by free radical polymerization using N,N,N´,N´-tetramethylene diamine (TEMED) and ammonium persulfate (APS) as initiator/activator pair, N,N´-methylene-bis-acrylamide (MBAAm) as cross-linker in a plastic syringe of 5 mL. Prepared supermacroporous cryogels with embedded PHEMA-IDA-Cu2+ beads were characterized by FTIR, SEM, sweling tests and surface area measurements. The effect of pH, buffer, ionic strength, initial lysozyme concentration, temparature and flow rate on adsorption were investigated. The maximum amount of lysozyme adsorption from aqueous solution was at 107,14 mg/g polymer in Tris-Base buffer at pH 9 with initial lysozyme concentration of 5 mg/mL. It was also observed that enzyme could be repeatedly adsorbed and desorbed on the supermacroporous cryogels with embedded PHEMA-IDA-Cu2+ beads.
Açıklama
Anahtar Kelimeler
Afinite, Lizozim, Kriyojel, İMAK, Adsorpsiyon, Lysozyme, Cryogel, Affinity, IMAC, Adsorption
