Affinity Chromatography of Hemoglobin on Cross-Linked Amylose

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Abstract
The cross-linked amylose is a semisynthetic polysaccharide realised by cross-linking amylose with epichlorohydrin, generating an insoluble three-dimensional structure based on polyglucose chains linked by α-1, 4-glucosidic bonds. We have evidentiated a selective retention of Hb on Cross-Linked Amylose columns with a capacity charge of 4.5–10 mg Hb/mL gel bed, irrespective to the cross-linking degrees. No Hb retention was observed in identical chromatographic conditions on Sephadex G-25 or Sephadex G-100 (cross-linked dextran based on α-1, 6-glucosidic bonds). Hemoglobin is therefore able to specifically recognize the polyglucose chains based on α-1, 4-glucosidic bonds, suggesting a Hb recognition site for this sequence. This new aspect on the Hb properties can offer a better understanding of Hb-glycoprotein interactions and at the same time, a possibility to purify Hb directly from erythrocytary extracts, by a single step chromatographic procedure on cross-linked amylose columns.