Formation of protoporphyrin from haemoglobin in vitro.

1. The formation of protoporphyrin from red blood cells or purified haemoglobin in aqueous perchloric acid media without the prior isolation of haemin is described. The reaction is carried out in the absence of oxygen and in red light. Even traces of oxygen inhibit the reaction by oxidative destruction of protoporphyrin and by the oxidation of haem to haematin. 2. Perchloric acid releases iron and protoporphyrin from haemoglobin at similar rates, but the amount of protoporphyrin in the filtrate varies with the solubility of protoporphyrin in the concentration of perchloric acid used. The yield of protoporphyrin may reach 50-60%. Less than 5mug. of haemoglobin/ml. can be detected by measuring the fluorescence of the porphyrin released. 3. A porphyrin other than protoporphyrin is obtained in small amounts. Its possible identity is discussed. 4. If sodium sulphite is present as a reducing agent the exclusion of oxygen is not required, but the porphyrin formed is more polar and more soluble in water than protoporphyrin. The presence of oxygen appears to be necessary for the formation of this polar porphyrin.