Gentle chemical deoxygenation of hemoglobin solutions.
Dithionite is often used to deoxygenate aqueous solutions because it reacts readily with oxygen. However, milder reducing agents, that do not ordinarily react readily with oxygen, may do so in the presence of an appropriate redox catalyst. We show that dithiothreitol reacts rapidly with oxygen in concentrated hemoglobin solutions to produce a mixture of deoxy-, met- and sulf-hemoglobin. The reaction in neutral phosphate buffer is not significantly affected by superoxide dismutase, benzoate or EDTA. However, addition of catalase or horseradish peroxidase decreases the proportions of met- and sulf-hemoglobin produced. We conclude that both hemoglobin and horse radish peroxidase accept dithiothreitol as the reducing substrate in heme catalyzed reactions with their respective oxidizing substrates (dioxygen and hydrogen peroxide). As a result, deoxy-hemoglobin suitable for physical studies can be prepared with a combination of a stoichiometric excess of dithiothreitol and a catalytic amount of horse radish peroxidase.
Herzfeld
J
J
Department of Chemistry, Brandeis University, Waltham, MA 02254-9110.
Seidel
N E
NE
Taylor
M P
MP
Droupadi
P R
PR
Wang
N E
NE
D001565:Benzoates / Q000494:pharmacology; D019817:Benzoic Acid; D001718:Birefringence; D002374:Catalase / Q000494:pharmacology; D004227:Dithionite; D004229:Dithiothreitol; D004492:Edetic Acid / Q000494:pharmacology; D006418:Heme / Q000378:metabolism; D006454:Hemoglobins / Q000302:isolation & purification*; D006735:Horseradish Peroxidase; D006801:Humans; D008706:Methemoglobin / Q000138:chemical synthesis; D010084:Oxidation-Reduction; D013435:Sulfhemoglobin / Q000138:chemical synthesis; D013482:Superoxide Dismutase / Q000494:pharmacology
D001565:Benzoates; D006454:Hemoglobins; D004227:Dithionite; D006418:Heme; D019817:Benzoic Acid; C100003:deoxyhemoglobin; D008706:Methemoglobin; D013435:Sulfhemoglobin; D004492:Edetic Acid; D006735:Horseradish Peroxidase; D002374:Catalase; D013482:Superoxide Dismutase; D004229:Dithiothreitol