The Osmotic Pressure of Haemoglobin in the Absence of Salts
Proceedings of the Royal Society A
From the Low Temperature Station, Cambridge). Introduction. In the general theory of haemoglobin solutions the osmotic pressure of the pure protein does not occupy an important place, because it is simply one of many possible combinations of haemoglobin, water, hydrogen ions, and salts. There is, however, the definite advantage in studying pure haemoglobin solutions that, if the protein be in equilibrium with distilled water, there is no need to consider secondary complications in the
... ns in the distribution of salts, due to the presence of the membrane. If a very acid protein were used, it would be necessary to consider the pressure due to hydrogen ions, but the iso-electric point of haemoglobin is so near neutrality that the hydrogen-ion pressure is immeasurably small. The osmotic pressure of purified haemoglobin solutions aroused the interest of early investigators, because it offered a means of determining the molecular weight. The equivalent weight calculated from iron analyses was about 16,700, and the molecular weight must be N times as great, where N is the number of iron atoms in the molecule. According to van't Hoff's formula, the osmotic pressure should be proportional to the concentration, and if results are expressed in the form of millimetres of mercury (reduced to 0° C.) per 1 per cent, of protein, the calculated pressure is 10-21/N. It is usually stated that Hiifner proved that the molecular weight of haemo globin was the same as the equivalent, 16,700. His results are stated below, together with the data obtained by other workers on the osmotic pressure of pure haemoglobin. Hiifner and Gansser, 9-3-12-1 mm. ; Reid, 3-5-4-4 mm. ; Roaf, 5-3 mm.; Wilson, 7-11 mm. The accepted explanation of these variable results, mainly due to Prof. A. Y. Hill, is that haemoglobin forms in pure water a uni-molecular solution, and that the lower pressures obtained by Reid and others were due to an aggregation of the molecules caused by traces of salts. There is, however, an alternative view. Protein preparations almost always contain traces of combined acids or bases, and these diffuse away very slowly when dialysed against distilled water. It is here suggested that these impurities caused irregular osmotic effects, formerly attributed to changes in aggregation.