PRACTICAL EXERCISES 67 



addition of ammonium sulphide, since carbonic oxide haemoglobin 

 is a more stable compound than oxyhaemoglobin. 



(d) Methmceoglobin. Put some blood into a test-tube, add a few 

 drops of a solution of ferricyanide of potassium, and heat gently. On 

 diluting a well-marked band will be seen in the red. On addition 

 of ammonium sulphide this band disappears ; the oxyhaemoglobin 

 bands are seen for a moment, and then give place to the band of 

 reduced haemoglobin (Fig. 7). 



(e) Acid Hcematin. To a little diluted blood add strong acetic acid 

 and heat gently. The colour becomes brownish. The spectrum 

 shows a band in the red between C and D, not far from the position 

 of the band of methaemoglobin. The addition of a drop or two of 

 ammonium sulphide causes no change in the spectrum, and this is a 

 means of distinguishing acid-haematin from methaemoglobin. If 

 more ammonium sulphide be added, haematin will be precipitated 

 when the acid solution has been rendered neutral, and a further 

 addition of ammonium sulphide or sodium hydroxide will cause the 

 haematin to be again dissolved, a solution of alkaline haematin being 

 formed. This in its turn maybe reduced by an excess of ammonium sul- 

 phide, and the spectrum of haemochromogen may be obtained (Fig. 7) . 



Since the watery solution of acid 

 haematin obtained as above is usually 

 somewhat turbid, a solution in acid ^ 



ether is sometimes employed for ^A ^ 



spectroscopic examination. Add to ^ f^ 

 a little undiluted defibrinated blood 

 about half its volume of glacial x<^ ^ 



acetic acid, and then not less than r ^\ 



an equal volume of ether. Mix well, w| 



pour off the ethereal extract and 



examine it with the spectroscope, 



diluting, if necessary, with ether 



and glacial acetic acid. It shows a FlG I6 .1 CRYS TALS OF 



strong band in the red somewhat (FREY) 



farther from the D line than the 



methaemoglobin band. On dilution, three additional fainter bands 



may be seen. 



(/) Alkaline Hamatin. To diluted blood add strong acetic acid 

 and warm gently for a few minutes. Then, when the spectroscopic 

 examination of a sample shows that acid-haematin has been formed, 

 neutralize with sodium hydroxide. A brownish precipitate of haematin 

 is thrown down, which dissolves in an excess of sodium hydroxide, 

 giving a solution of alkaline haematin (or alkali-haematin). 



Or add sodium hydroxide to blood directly, and warm for a couple of 

 minutes after the colour has changed decidedly to brownish-black. 

 The spectrum of alkaline haematin is a broad but ill-defined band 

 just overlapping the D line, and situated chiefly to the red side of it 

 (Fig. 7). The solution should be shaken up with air before being 

 examined, as some of the alkali-haematin is changed into haemo- 

 chromogen by reducing substances formed by the action of the alkali 

 on the blood. 



(g) Hcemochromogen. To a solution of alkaline haematin add a 

 drop or two of ammonium sulphide. The band near D disappears, 

 and two bands make their appearance in the green (Fig. 7) . 



(h) Hcematoporphyrin. Put some strong sulphuric acid in a test- 

 tube. Add a few drops of blood, agitate the test-tube till the blood 



52 



