Stroma-free hemoglobin from bovine blood.
Isolation and purification of bovine hemoglobin (HbBv) was carried out after reaction of whole blood with carbon monoxide. Washing/centrifugation steps were used to eliminate leukocytes, platelets, and plasma proteins. Hypotonic media and ultrasound radiation were used to lyse red blood cells. Lyse by ultrasound was shown to lead to solutions at the highest concentrations in HbBv, and the least concentrations in major phospholipids contaminants. Additional purification procedures were performed to remove membrane proteins and phospholipids. In the first case, proteins were denatured by thermal treatment, and filtered. To eliminate phospholipids, liquid chromatography was used with strong anion exchangers. Purity of HbBv was evaluated by normal phase high performance liquid chromatography (HPLC), electrophoresis, and size-exclusion HPLC.
Lima
Maria Celiana P
MC
Instituto de Macromoléculas Professora Eloisa Mano, Universidade Federal do Rio de Janeiro, Centro de Tecnologia, Rio de Janeiro, Brazil.
Andrade
Cristina T
CT
D000818:Animals; D001769:Blood; D001798:Blood Proteins; D002248:Carbon Monoxide; D002417:Cattle; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D002852:Chromatography, Ion Exchange / Q000379:methods*; D006454:Hemoglobins / Q000302:isolation & purification*; D010743:Phospholipids
D001798:Blood Proteins; D006454:Hemoglobins; D010743:Phospholipids; D002248:Carbon Monoxide