Selectivity of IMAC columns in trypsin inhibitor purification.
The properties of an adsorbent and the parameters in an adsorption process affect the resolution of chromatographic purifications. This is reflected in the elution profile, which shows the relative affinity of different proteins for a specific adsorbent. In the work presented here, elution profiles for trypsin inhibitor were used to study the effects of the concentration of trypsin inhibitor, ionic strength of the protein solution, slope of the elution gradient, and the regeneration treatment of the chromatography column on the selectivity of the adsorbent Cellufine Chelate-Cu(II)(ida). Cytochrome c was used as a reference protein. Variations in the concentrations of trypsin inhibitor and in the ionic strength of the buffered solution did not have any effects on the elution profile. On the other hand, changes in the slope of the pH gradient used for elution caused shifting of the elution peaks toward lower values of the elution volume, resulting in the best strategy to modify the elution profile of the system. Finally, using a constant slope pH gradient of elution, the variation of the selectivity of the adsorbent for trypsin inhibitor when subjected to cleaning treatments with 0.5 N NaOH was studied. Appropriate cleaning practices used in industry were followed. The adsorbent showed only a slight tendency for resolution loss in the order of 2 x 10(-4) days(-1). The results presented here show a good stability of the adsorbent when compared to other biospecific adsorbents commonly used.
Yeomans-Reina
H
H
Departamento de Ingenieria Quimica y Metalurgia, Departamento de Investigacion en Alimentos, and Departamento de Investigaciones Cientificas y Tecnologicas, Universidad de Sonora, Mexico. Jhyeomans@huatabampo.iq.uson.mx
Ruiz-Manriquez
A
A
Wong
B R
BR
Mansir
A T
AT
D002614:Chelating Agents / Q000737:chemistry; D002846:Chromatography, Affinity / Q000295:instrumentation / Q000379:methods*; D003300:Copper / Q000737:chemistry; D003574:Cytochrome c Group / Q000302:isolation & purification; D006863:Hydrogen-Ion Concentration; D009994:Osmolar Concentration; D014360:Trypsin Inhibitor, Kunitz Soybean / Q000302:isolation & purification*
D002614:Chelating Agents; D003574:Cytochrome c Group; D003300:Copper; D014360:Trypsin Inhibitor, Kunitz Soybean