Table 8 Ion-exchange chromatography (IEX) in proinsulin (PI) and insulin purification
Protein | AEX/CEX | Media | Comments | References |
|---|---|---|---|---|
Folded preproinsulin | AEX | DEAE-Sepharose fast flow/ Source 30 Q | Flow-through mode at pH 8.3, 6.1 mS/cm, in which the preproinsulin was not bound to the gel but washed through the column with the permeate The higher molecular weight impurities were adsorbed | Thurow et al. (2010) |
Folded preproinsulin | CEX | Source 30 S | Elution with linearly increasing NaCl gradient | Thurow et al. (2010) |
Refolded PI | AEX | Q-Sepharose Fast Flow | Equilibration with 20 mM glycine–NaOH, pH 10.0 NaCl elution in 20 mM glycine–NaOH, pH 10.0 | Chen et al. (2016) |
Sulfonated PI | AEX | DEAP-Spheronit | NaCl elution in 7.5 M urea Yield: 95% Purity: 70–95% (order of chromatography affects purity) | Tikhonov et al. (2001) |
Sulfonated PI | AEX | Mono-Q HR | Binding and elution buffers contain 7 M urea NaCl elution | Cowley and Mackin (1997) |
Sulfonated PI | AEX | Q-Sepharose Fast Flow | NaCl elution in 8 M urea | Castellanos-Serra et al. (1996) |
Sulfonated PI | CEX | S-Sepharose | Eluted at a rate of 3 mL/min using a linear gradient of 0.5 M NaCl in 7 M urea/20 mM formic acid buffer (pH 4.0) for 50 min | Redwan et al. (2007) |
Sulfonated PI | CEX | SP Sepharose Fast Flow | NaCl elution Yield: 90% | Petrides et al. (1995) |
Insulin | AEX | DEAE | Purification after citraconylation and trypsin digestion step | Mikiewicz et al. (2017) |
Insulin | AEX | DEAE-Sepharose | Purification after citraconylation and trypsin digestion step Elution with Tris pH 8.6 and 30% isopropanol (conductivity 6 mS) | Zieliński et al. (2019) |
Insulin | AEX | Q | Purification after citraconylation and trypsin digestion step | Mikiewicz et al. (2017) |
Insulin | AEX | Q-Sepharose | Elution with Tris pH 8.6 and 30% isopropanol (conductivity 3 mS) | Zieliński et al. (2019) |
Insulin | AEX | Source 30Q | The load is in 30% ethanol, pH 7.5, < 3 mS/cm and contains Zn-ions in an amount of 2 zinc atoms per six insulin molecules Elution with ammonium acetate, ethanol, triethanolamine at pH 6.4, 6.8 and 7.2 At pH 6.4, the peak is fronting (flat front and steep tail), and a baseline separation is seen between the impurity and the product peak Yield: > 90% | Mollerup and Frederiksen (2016) |
Insulin | CEX | BioSepra CM | Loading diluent and elution solution contain hexylene glycol 2 washes with NaCl Isocratic elution with NaCl Yield: 60–85%; Purity: > 90% | Coleman et al. (2019) |
Insulin | CEX | Capto SP ImpRes | Subsequent purification after enzymatic conversion Binding buffer: Na acetate buffer pH 4, 10% ethanol Elution with 47.5% ethanol and 128 mM NaCl Yield: 102%; Purity: 63–94% | GE application note 29–0018-56 AB |
Insulin | CEX | SP Sepharose Fast Flow | Capture of glargine insulin NaCl elution | Hwang et al. (2016) |
Insulin | NA | NA | NaCl elution Yield: 95% | Petrides et al. (1995) |