IRON IN BLOOD 199 



chemically pure ferric chloride (FeCls.GH^O). The standard as 

 prepared and used contains an amount of iron equal to that in the 

 hemoglobin of 0.040 c.c. of blood when the hemoglobin content is 

 100 per cent in the Haldane scale; i.e., 13.6 gms. of hemoglobin 

 per 100 c.c. of blood binding 18.5 c.c. of oxygen. The figure 13.6 

 gms. of hemoglobin per 100 c.c. as a basis may be derived from 

 Hiifner's 1 finding that 1 gm. of hemoglobin will combine with 

 1.36 c.c. of oxygen. This has been confirmed by a number of inves- 

 tigators. If 1 gm. of hemoglobin will combine with 1.36 c.c. of 

 oxygen, 18.5 c.c. of oxygen will combine with 13.6 gm. of hemo- 

 globin. Butterfield, 2 as others before him, has shown that the 

 iron content of 1 gm. of hemglobin is constant, lying between 0.33 

 and 0.34 per cent, that is, one three-hundredth of the molecule. 

 One three-hundredth of 13.6 gives 45 mg. of Fe per 100 c.c. of 

 standard blood. This is the standard used in the determination 

 of iron. It may be made up from a concentrated solution of 

 ferric chloride, 4.355 gms. of FeCl3.6H2O in a liter of water, by 

 taking 1 c.c. and making up to 100 c.c. with water, which is the 

 standard solution containing 0.009 mg. of Fe per c.c. 



If the hemoglobin is much higher than the normal, other 

 standards can be used, corresponding respectively to 25, 35, 55, 

 and 65 mg. of Fe per 100 c.c. of blood. In the note on the colori- 

 metric comparison reference will be made to this. 



Ammonium Sulfocyanate. This is prepared as 5 N in order 

 to have a concentrated solution which reduces the degree of dis- 

 sociation of the ferric sulfocyanate into the colorless ions. It 

 should be kept in a dark bottle away from the light. 



Comparison in Colorimeters. Duboscq and Kober colorim- 

 eters proved suitable for this work. A comparison of the standard 

 itself should always be carried out quickly. Readings can be made 

 in daylight, preferably from a north window, or with artificial 

 light filtered through " daylight glass." The average of at least 

 four readings, quickly done before the acetone evaporates, is taken. 

 As in all colorimetric readings, if the reading falls below 12 or 

 above 8 (the standard being at 10) a reading should be made with 

 one of the other standards mentioned above. Indeed, the standard 

 to be used should depend upon the hemoglobin reading; e.g., with 

 50 per cent hemoglobin the standard corresponding to 25 mg. of 



1 Hufner, G., Arch. Physiol., 1894, 130. 



2 Butterfield, E. E., Z. physiol. Chem., 1909, 62, 173. 



