The oxygen dissociation curve of haemoglobin in dilute solution

Abstract

The spectroscopic determination of the oxygen dissociation curves of haemoglobin has an advantage over the tonometer and gas analysis method, in that much smaller quantities of haemoglobin can be made use of. The spectroscopic method was used to determine the relationship between the foetal and maternal haemoglobins in the sheep during a study of foetal respiration made by Barcroft (1935). The conditions for the comparison of the haemoglobins were a dilute solution of the haemoglobin at p H 9·2 (borate buffer) and at 20° C. These conditions were chosen because of the very accurate determinations of the dissociation curves of dilute haemoglobin of the sheep by Forbes and Roughton (1931) and because these authors recommend p H 9·2 at room temperature as most suitable for a study of the oxygen equilibrium of haemoglobin, all the haemoglobin being in the form of the alkali salt. McCarthy (1933) and Hall (1934) had found previously that the haemoglobins of the foetal and maternal goat were different, the foetal haemoglobin (in the blood and as purified haemoglobin) having a higher affinity for oxygen. The same relationship was found to exist in the sheep haemoglobins in dilute solution at 20° C and p H 9·2. When samples of human foetal and maternal blood (sent by Professor Fleming from the Obstetrical Department of the Royal Free Hospital) were compared in dilute solution it was found that the foetal haemoglobin had a lower affinity for oxygen than the maternal. This was also found by Haurowitz (1935) for dilute solutions of the haemoglobins of mother and new born infant. Haurowitz, however, pointed out that in the corpuscles the affinity for oxygen is less in the infant’s haemoglobin than in that of the mother, but the method used by him did not allow of measurements on suspensions of corpuscles. In the present work the dissociation curves of dilute suspensions of corpuscles have been compared with similar solutions of the haemoglobin. It was found that the relationship of the dissociation curves for human foetal and maternal corpuscles is the same as that found by Barcroft in the goat and in the sheep. It has now been found that by a dilution of human adult haemoglobin the dissociation curve is altered by 200% to a position of higher affinity for oxygen, without any marked change in shape. The haemoglobin of the human foetus, on the other hand, is much less affected by dilution, thus explaining the anomaly of the reversed relationship when solutions of the haemoglobins are used instead of suspensions of corpuscles. It was shown by the work of Bock, Field, and Adair (1924), and by Adair (1925), that a solution of haemoglobin free from stromata and of a similar concentration to blood gives a dissociation curve like whole blood. This makes it clear that in the comparison of dilute haemoglobin solutions with suspensions of corpuscles we are concerned, not simply with a change in the haemoglobin due to haemolysis, but a change due to a dilution of the contents of the corpuscle.