生物分离工程-双语版 5.pptVIP

Chapter 5 Protein inactivations during chromatography I introduction 1 Advantage of stationary phase combined with HPLC (1) Sensitivity (2) Reproducibility (3) Saving time 2 It is imperative to develop industrial-scale methods of separation and purification 3 Chromatography techniques have tremendous interest 5 Major problems of chromatography (1) denaturation (2) denaturation causes and mechanism of proteins have not been extensively and comprehensively analyzed Denaturation 1 Interfacial phenomena and surface-surface interactions 2 Geometric complementarity and affinity interaction 3 Chromatography retention is largely determined by amino acids on polypeptide surface 4 Interest protein may be denatured and require refolding to regain the native structure 5 Denaturation reasons (1) Competing hydrophobic and coulombic forces on the column are greater than those maintaining proteins structure (2) Mobile phase denaturation (3) Not a single reason to minimize protein denaturation Adsorption isotherm 1 Ion exchange chromatography play a major role in biotechnology industry 2 Langmuir isotherm 3 Freundlich isotherm 4 Adsorption characteristics of proteins affect conformational dynamics on surface II chromatography techniques 1 Popular methods of bioseparation (1) ion exchange (2) gel permeation (3) affinity chromatography (4) hydrophobic interaction chromatography 2 HPLC for bioseparation in rapid development (1) rigid chromatography supports (2) high flow and rapid separation (3) same criteria as conventional liquid chromatography A isoelectric point B net charge at different pH C molecular weight II chromatography techniques 3 Elution (1) continued elution (2) pH gradient (3) ion strength gradient 4 Other application of chromatography techniques (1) antibiotics (2) vitamins (3) steroids (4) peptides A Ion-exch