H^MOCHROMOGEN 25 



tions. Indeed they cannot claim to have stood the test of repetition. 

 Quite lately, however, claims have been put forward both by Anson 

 and Mirsky and by Hill and Holden (6) to have prepared haemoglobin 

 and by completely different methods. The evidence is entirely spectro- 

 scopic. It is not claimed by Anson and Mirsky that the crystalline 

 oxyhaemoglobin was obtained, nor is that claim made by Hill and 

 Holden. What I ask of both is that they should produce the material 

 (and preferably in the crystalline form) in sufficient quantity for me 

 to determine the dissociation curve ! The point on which Hill and 

 Holden lay stress is that the globin used must be of the particular type 

 known as " undenatured " — I must admit that to me this word was 

 once a little frightening. The globin is prepared from oxyhaemoglobin 

 and the precautions necessary to prevent its denaturation are fully set 

 out in Hill and Holden's paper. When added to hsematin (made by 

 dissolving hsemin in sodium carbonate) methsemoglobin results. The 

 remaining processes are familiar: methsemoglobin treated with a 

 reducing agent gives reduced haemoglobin which when oxidised yields 

 oxyhaemoglobin. Not only have Hill and Holden made ordinary 

 oxyhaemoglobin in this way but, using the same undenatured protein, 

 they have attached it to the analogues of haematin made not from 

 protoporphyrin but from haemato- and mesoporphyrins, and so 

 obtained the haemoglobins, the spectra of which were described in the 

 last chapter. No small performance I think. 



REFERENCES 



(1) Bertin-Sans and Moitessier. C. R. Ac. Sci. Paris, cxvi. 591. 1893. 



(2) DiLLiNG. Atlas of Hcemochromogens. Stuttgart, 1910. Atlas der Krystallformen 



u. Ahsorptionhdnder der Hdmochromogene. 



(3) Anson, Barcroft, Mirsky and Oinuma. Proc. Roy. Soc. B. xovn. 61. 1925. 

 Anson and Mirsky. Journ. Physiol, lx. 50. 1925. 



(4) Keilin. Proc. Roy. Soc. B. c. 129. 1926. 



(5) V. Klavbren. Zeit. Physiol. Chem. xxxiii. 293. 1901. 



(6) Hill and Holden. Biochem. Journ. xx. 1326. 1926. 



