Purification and characterization of human hemoglobin: effect of the hemolysis conditions

International Journal of Biological Macromolecules
C T AndradeE G Azero

Abstract

Human hemoglobin was isolated and purified by anion exchange chromatography. To isolate hemoglobin, outdated red blood cells (RBC) were transformed into carbonylhemoglobin, by reaction with carbon monoxide, and submitted to washing/centrifugation procedures, to eliminate other plasma proteins. Albumin was quantified in each supernatant, by the bromcresol green method. Hemolysis was performed in three different hypotonic media (water, 0.01 M NaCl and 5 mM Tris/HCl buffer at pH 7.4), at 8 degrees C for 24 h. Sonication for 5 min was also used to lyse RBC. After isolation of hemoglobin, additional purification was carried out by anion exchange chromatography on AG MP-1, Q-SFF and both exchangers. Hemoglobin concentration of hemolysates and of purified solutions were determined by the hemiglobincyanide method. Residual phospholipids were extracted from the four different hemolysates, as well as from the purified hemoglobin solutions, and were analyzed by high performance liquid chromatography. Native and SDS-polyacrylamide gel electrophoresis experiments were performed on purified hemoglobin samples to verify the presence of proteins other than hemoglobin. According to the results, the hemolysis conditions have influence on the pur...Continue Reading

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Citations

Jan 19, 2012·Applied Biochemistry and Biotechnology·Radoslava StojanovićBranko Bugarski
Apr 10, 2012·Journal of Hazardous Materials·Jiayin LiuZhengqiang Li
Feb 19, 2009·Biotechnology Progress·Andre F PalmerDavid R Harris
Sep 23, 2016·Frontiers in Physiology·Gene A Morrill, Adele B Kostellow
Sep 25, 2016·Artificial Cells, Nanomedicine, and Biotechnology·Kadir Erol, Kazım Köse
Dec 18, 2013·International Journal of Biological Macromolecules·Yuping WeiXia Xu
Aug 19, 2007·Artificial Cells, Blood Substitutes, and Immobilization Biotechnology·Maria Celiana P Lima, Cristina T Andrade
Jul 10, 2012·The Journal of Immunology : Official Journal of the American Association of Immunologists·Tian LinH Shaw Warren
Apr 1, 2021·The Journal of Physical Chemistry. B·Jakub DybasPiotr J Mak

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