324 PRACTICAL PHYSIOLOGY 



(any sulphur which may separate out being removed by nitration) and 

 allowed to stand on ice for two or three days, when a large number of 

 oblique rhombic crystals of creatine will have separated out. (See 

 Fig. 203.) These are collected on a filter (for which purpose a suction 

 pump will be found necessary) and are thoroughly washed with alcohol 

 until no more pigment is removed. The nitrate is preserved for the 

 isolation of the other extractives. 



Xanthine and Hypoxanthine. The creatine -free nitrate is made 

 strongly alkaline with ammonia, and silver nitrate solution added until 

 precipitation is complete. The purine bodies are thus precipitated. 

 The precipitate is collected on a filter paper and thoroughly washed 

 with dilute ammonia. The hypoxanthine and xanthine are sepa- 

 rated by the following method : the precipitate is removed from 

 the filter paper and dissolved in boiling nitric acid (sp. gr. 1-1), a 

 few crystals of urea being added to the solution so as to destroy any 

 nitrous acid which may be present, and which would decompose the 

 purine bodies. When all the precipitate has dissolved the solution is 

 quickly filtered hot, and the filtrate is allowed to stand overnight, when 

 it will be found that a precipitate consisting of fine needle-shaped 

 crystals (Fig. 204) has separated out. This consists of hypoxanthine 

 silver nitrate combined with nitric acid ; to remove the nitric acid wash 

 it with distilled water, transfer it from the filter to a small beaker and 

 boil it with ammonia until the crystals break up and become amor- 

 phous, and then, to remove the silver, pass in H 2 S, filter off the silver 

 sulphide, and evaporate the filtrate slowly to dryness, when a white 

 chalk-like mass of hypoxanthine, will be obtained. In order to obtain 

 the xanthine silver salt the filtrate from hypoxanthine should be 

 treated with ammonia, when a few yellow flakes of the salt will be 

 obtained. To separate the xanthine this precipitate is treated in 

 exactly the same way as for hypoxanthine. 



Test for Hypoxanthine. Place some hypoxanthine in a small 

 evaporating dish with a few drops of pure concentrated nitric acid and 

 evaporate slowly to dryness : a brilliant yellow residue is obtained. 

 Cool, and then add a drop of sodium hydrate solution, when the residue 

 will change to orange. If the residue be dissolved in water and the 

 solution again evaporated to dryness the orange colour persists, thus 

 differing from the murexide stain which, when similarly treated, loses 

 its colour (see p. 259). 



Test for Xanthine. Repeat the same test as for hypoxanthine and 

 note that the sodium hydrate produces in this case a deep red colour, 

 which persists on dissolving in water and evaporating. 



Sarcolactic Acid. The ammoniacal filtrate, from which the alloxuric 

 bodies have been separated, is treated with sulphuretted hydrogen gas 

 so as to remove the silver which it contains : the silver sulphide is 

 filtered off, and the filtrate evaporated till all the ammonia has been 

 expelled. It is then made strongly acid with phosphoric acid, and the 

 lactic acid, which is hereby liberated, is dissolved out by shaking it in a 

 separating funnel with ether. 



After extracting three or four times, the ethereal extracts are com- 

 bined and the ether evaporated away by placing on a waterbath 

 heated to about 60 C., the flame underneath which has been 

 extinguished. An acid syrup remains behind ; this is impure lactic 

 acid. In order to purify it, dilute three times with water, bring the 





