Preparation and characterization of magnetic nanoparticles for urokinase purification

Bu tezde, manyetik poli(2-hidroksietil metakrilat) [p(HEMA)] nanopartiküller sürfaktansız emülsiyon polimerizasyonu tekniği ile sentezlenmiştir. Manyetik p(HEMA) nanopartiküllere Cibacron Blue F3GA kovalent olarak bağlanmış ve boya molekülleri Cu2+ iyonları ile şelatlaştırılmıştır. Ürokinazın Cu2+ ile şelatlaştırılmış Cibacron Blue bağlı manyetik poli(HEMA) nanopartiküllere adsorpsiyonu gerçekleştirilmiştir. Sentezlenen manyetik nanopartiküllerin 80 nm boyutunda, küresel yapıda ve manyetik özellikte olduğu gözlenmiştir. Manyetik nanopartiküllere bağlanan Cibacron Blue F3GA miktarı 28.125 µmol/g polimer olarak bulunmuştur. Yapıya katılan Cu2+ miktarı 10.229 µmol/g polimer olarak hesaplanmıştır. Sentezlenen Cu2+ ile şelatlaştırılmış Cibacron Blue bağlı manyetik poli(HEMA) nanopartiküllere farklı ortam koşulları (pH, başlangıç ürokinaz derişimi, iyonik şiddet ve sıcaklık) altında ürokinaz adsorpsiyonu gerçekleştirildiğinde maksimum adsorpsiyon kapasitesi 630.43 mg/g polimer olarak bulunmuştur. Langmuir adsorpsiyon izoterminin bu adsorpsiyon işlemine uygulanabilir olduğu gözlenmiştir. Adsorplanan ürokinazın desorpsiyonu 1.0 M NaCl ile gerçekleştirilmiş ve desorpsiyon oranı % 96 olarak bulunmuştur. Beş kez tekrar edilen adsorpsiyon-desorpsiyon işleminden sonra adsorplanan ürokinaz miktarında önemli bir azalma gözlenmemiştir. Bu çalışma, yeni sentezlenen bu destek materyalinin biyolojik kaynaklardan ürokinaz saflaştırılması için kullanılabileceğini vadetmektedir.In this thesis, magnetic poly(2-hydroxyethyl methacrylate) [p(HEMA)] nanoparticles were synthesized by surfactant free emulsion polymerization technique. Cibacron Blue F3GA was covalently attached to the magnetic p(HEMA) nanoparticles and Cu2+ ions were then chelated to dye molecules. Urokinase was adsorbed onto Cu2+-Cibacron Blue F3GA derived magnetic p(HEMA) nanoparticles. Synthesized magnetic nanoparticles was about 80 nm in diameter, spherical and carried magnetic character. Incorporation rate of Cibacron Blue for magnetic nanoparticles was found to be 28.125 µmol/g polymer. The Cu2+ ions loaded was calculated to be 10.229 µmol/g polymer. This Cu2+-Cibacron Blue F3GA derived magnetic p(HEMA) nanoparticles were used for urokinase adsorption under different conditions (i.e. pH, enzyme initial concentration, ionic strength, temperature). Maximum adsorption capacity was found to be 630.43 mg/g polymer for Cu2+-Cibacron Blue F3GA derived magnetic p(HEMA) nanoparticles. It was observed that Langmuir adsorption isotherm was applicable in this adsorption process. The urokinase adsorbed Cu2+-Cibacron Blue F3GA derived magnetic p(HEMA) nanoparticles was desorbed with 1.0 M NaCl and desorption rate was found to be 96 %. It was also analyzed that adsorption capacity did not changed significantly after five adsorption/desorption cycles. This study promised that synthesized support material could be used for urokinase purification from biological samples

Preparation of laccase-based nanobiosensors for determination of catechol, dopamine and (-)-epicatechin

Amaç: Bu çalışma, lakkaz temelli nanobiyosensörlerin katekolün, dopaminin ve (-)-epikateşinin tayininde kullanılabilirliğinin araştırılması amacıyla yapılmıştır. Materyal ve Yöntem: Damlatma tekniği ile Lakkaz/Cys-Ag@Fe3O4/Kitosan/SPE, Lakkaz-Ni/Au/PPy-COOH/NAF/SPE ve Lakkaz-GO/TiS2-NAF/SPE olarak hazırlanan elektrotların sırasıyla katekol, dopamin ve (-)-epikateşin tayini için optimum çalışma koşulları belirlenmiştir. Biyosensörlerin kinetik ve analitik performansları değerlendirilerek musluk suyu ve göl suyunda katekol; ticari insan serumunda ve yapay idrarda dopamin; şarap ve meyve suyunda (-)-epikateşin tayinine yönelik uygulamalar yapılmıştır. Bulgular: Lakkaz/Cys-Ag@Fe3O4/Kitosan/SPE'nin katekol tayini için doğrusal aralığı 0.025–100 µM, LOD değeri 0.060 µM, duyarlığı 60.056 µ\u1d434��mM-cm-2; Lakkaz-Ni/Au/PPy-COOH/NAF/SPE'nin dopamin tayini için doğrusal aralığı 0.01–50 µM, LOD değeri 2.265 nM, duyarlığı 0.902 µ\u1d434��µM-cm-2; Lakkaz-GO/TiS2-NAF/SPE'nin (-)-epitakeşin tayini için doğrusal aralığı 0.1–250 µM, LOD değeri 0.073 µM ve duyarlığı 0.3264 µ\u1d434��µM-cm-2 olarak belirlenmiştir. Sonuç: Lakkaz/Cys-Ag@Fe3O4/Kitosan/SPE, Lakkaz-Ni/Au/PPy-COOH/NAF/SPE ve Lakkaz-GO/TiS2-NAF/SPE biyosensörlerinin katekol, dopamin ve (-)-epikateşinin tayininde başarılı olduğu bulunmuştur.Objective: This study was carried out to investigate the usability of laccase-based nanobiosensors for the determination of catechol, dopamine and (-)-epicatechin. Material and Methods: Laccase/Cys-Ag@Fe3O4/Chitosan/SPE, Laccase-Ni/Au/PPy-COOH/NAF/SPE and Laccase-GO/TiS2-NAF/SPE electrodes were prepared by the drop-casting method. Optimization experiments were carried out for the detection of catechol, dopamine and (-)-epicatechin, respectively, and kinetic and analytical performances of biosensors were investigated. Application assays were carried out for catechol in tap water and lake water; dopamine in commercial human serum and artificial urine and (-)-epicatechin in red wine and fruit juice. Results: Linear range, LOD and sensitivity value of Laccase/Cys-Ag@Fe3O4/Chitosan/SPE for catechol was determined as 0.025–100 µM, 0.06 µM, and 60.056 µ\u1d434�mM-cm-2, respectively. Linear range, LOD and sensitivity of Laccase-Ni/Au/PPy-COOH/NAF/SPE for dopamine was determined as 0.01–50 µM, 2.265 nM, and 0.902 µ\u1d434�µM-cm-2, respectively. Linear range, LOD and sensitivity of Laccase-GO/TiS2-NAF/SPE for (-)-epitachechin was determined as 0.1–250 µM, 0.073 µM, and 0.3264 µ\u1d434�µM-cm-2, respectively. Conclusion: Laccase/Cys-Ag@Fe3O4/Chitosan/SPE, Laccase-Ni/Au/PPy-COOH/NAF/SPE and Laccase-GO/TiS2-NAF/SPE biosensors are effective in the determination of catechol, dopamine and (-)-epicatechin

Lectin attached affinity cryogels for amyloglucosidase adsorption

WOS: 000454929200003Preparation of poly[ethylene glycol dimethacrylate (EGDMA)-glycidyl methacrylate (GMA)] cryogel and its usability for amyloglucosidase adsorption were investigated. Cryogels were prepared by cryocopolymerization technique and then functionalized by lectin concanavalin A (Con A). The synthesized cryogel was characterized by FTIR, SEM and EDX analysis and then used for adsorption of amyloglucosidase in a continuous flow system. The maximal amyloglucosidase adsorption efficiency of Con A attached cryogel was found to be as 30.50mg/g cryogel. Con A modified cryogels were used repeatedly for 30 times without any significant decrease at the amyloglucosidase adsorption capacity. SDS-PAGE and activity studies confirmed that the desorbed amyloglucosidase was active and retained 90% of its initial activity after the adsorption/desorption processes