Physical and biochemical characteristics of protein adsorption on custom-made hydrophobic matrix in expanded bed chromatography

• Main Authors: Chih-Yu Chen, 陳致宇
• Other Authors: John Chi-Wei Lan
• Format: Others
• Language: zh-TW
• Online Access: http://ndltd.ncl.edu.tw/handle/q352g2

碩士 === 元智大學 === 化學工程與材料科學學系 === 103 === Since 1990s, Expanded Bed Adsorption(EBA) chromatography technique has been developed. Due to following advantages: reduction in operation steps and purification cost improvement in product recovery. Therefore, EBA has great potential to be implemented for biomolecule separation. In this study, the cellulose matrix with stainless steel powder was fabricated as weighting agent by using reverse suspension adsorbent. The physical characteristics included wet density, water content, porosity, pore volume and surface area. The particle size was found in the range between 150 and 300 μm and the wet density is 1.41 g mL-1 as well as water content was calculated 50 % ~ 60%. Subsequently, the adsorbent was activated by coupling 1,1'-Carbonyldiimidazole (CDI) octylamine immobilized. On the matrix optical microscopy, Scanning electron microscope (SEM), Fourier transform infrared spectroscopy (FTIR), contact angle and a gas chromatography (GC) clearly presented the appearance and structure information. The result indicated that the spherical matrix were dispersed uniformly. FTIR data apparently showed absorption peaks (-CH3, -CH2); Contact angle demonstrated after octylamine immobilized on the matrix surface seemed to be in hydrophobicity. According to characteristics homemade adsorbent showed a great opportunity to be expanded bed matrix . The hydrophobic adsorbent was loaded into EBA column to determine fluid properties including height equivalent of a theoretical plate, number of theoretical plates, Bodenstein value and axial dispersion coefficient. Comparison of fluid properties between custom-made hydrophobic matrix and commercial adsorbent was investigated. Isothermal adsorption experiments to explore the relationship of adsorption capacity was also assayed. The study showed that by increasing salt concentration and temperature adsorption capacity could be improved. The BSA maximum adsorption could achieve 96.88 mg mL-1 at 298 K with 2.0M Na2HPO4 buffer, pH 7.5. Moreover, when absorbents were filled into expanded bed to adsorb BSA protein, at sedimentation bed height 1.5 cm and 3.0 cm the BSA adsorption capacity could reach 48 mg mL-1. Regarding system efficiency, due to sedimentation bed height 1.5 cm spend much more time reaching 90 % adsorption saturation than sedimentation bed height 3.0 cm. That is the higher sedimentation bed height could perform well adsorption efficiency.