260 PRACTICAL PHYSIOLOGY 



If the product of the action of nitrites be treated with ammonium 

 sulphide, the spectrum passes through a transient oxyhsemoglobin 

 stage, succeeded by reduced haemoglobin, and later becomes nitric 

 oxide haemoglobin. 



5. The Visible Spectrum of Acid-heematin. If some diluted defi- 

 brinated blood be treated with a little glacial acetic acid and gently 

 warmed, it will assume a dark brown colour. If it be diluted 

 sufficiently, and examined spectroscopically, it will present a spec- 

 trum characterised by one band on the red, the wave-length corre- 

 sponding approximately to A 645. The blue end of the spectrum 

 will be very largely absorbed (Spectrum 6 in Chart). 



There is frequently a considerable amount of general absorption in the 

 acid-hsematin prepared as above, and the band referred to may only be made 

 clear by filtering the solution. More satisfactory results are obtained by 

 extracting the colouring matter with ether, or treating with chloroform and 

 excess of acetic acid, as follows : 



(a) Take defibrinated blood, and add about half its volume of glacial acetic 

 acid and about an equal quantity of ether. Shake at once. The ether will 

 extract the colouring matter, and, on examining the same spectroscopically, 

 three distinct bands will be seen one on the red similar to that already 

 described, but apparently shifted nearer the D line (X 640), one on the green 

 (\ 550), another on the green but nearer the blue (X 515). A very indistinct 

 band may be seen on the yellow (X 590) (Spectrum 7 in Chart). 



(6) Take defibrinated blood, warm and add half its volume of glacial acetic 

 acid. Cool and add half the volume of chloroform, and more acetic acid if any 

 precipitate appears. The solution will become clear and give a spectrum 

 similar to that shown in the ethereal extract. 



6. The Visible Spectrum of Alkaline Haematin. Take some diluted 

 defibrinated blood, and add a few drops of strong caustic soda, and 

 warm. The colour will change to a greenish-brown tint, and when 

 the solution is examined spectroscopically, it will show a single 

 band on the orange (wave-length, A 600). A more satisfactory 

 method of preparing the alkaline hsematin is to form a paste of 

 potassium carbonate and defibrinated blood ; dry it over a water- 

 bath and extract with alcohol, when a reddish-brown solution is 

 obtained which shows the distinguishing absorption band clearly 

 (Spectrum 8 in Chart). 



7. The Visible Spectrum of Hsemochromogen (reduced Alkaline 

 Hsematin). If a watery solution of alkaline hsematin be warmed with 

 a few drops of ammonium sulphide, the brownish colour is replaced by 

 a more marked red. If the solution be examined spectroscopically, 

 the one band of alkaline hsematin is found to be replaced by two 

 bands on the green, the wave-lengths of their centres being 

 approximately A 557 and A 525 (Spectrum 9). 



8. The Visible Spectrum of Heematoporphyrin. Take some strong 

 sulphuric acid (10 c.c.) in a test tube and add a few drops of blood, 

 and shake up the mixture. A purple colour will result, due to the 

 decomposition of the haemoglobin and the formation of the iron- 

 free pigment, hsematoporphyrin. This examined spectroscopically 

 will, in the above acid solution, show two bands, the centres being 

 approximately A 605 and A 565 (Spectrum 10 in Chart). 



