METABOLIC PRODUCTS 467 



identified by esterification and fractionation of the esters. 

 Some acids, for example spiculisporic acid (see p. 286) 

 from P. spiculisporum, are precipitated on acidification 

 of the metabolism solution with hydrochloric acid. 



4. Non-volatile Neutral Products. — Polysaccharides, 

 if present, will, in general, have been precipitated with the 

 calcium salts of the non-volatile acids on addition of four 

 volumes of alcohol to the metabolism solution. They may 

 be separated from the calcium salts by solution of the 

 precipitate in water, acidification with hydrochloric acid 

 and again precipitating with alcohol, the free acids being 

 soluble in 80 per cent, alcohol. Neutralisation of the 

 solution, freed from the polysaccharides, with calcium 

 carbonate will then result in the precipitation of the 

 calcium salts. 



The residue remaining after the distillation of the 

 volatile acids (see p. 465) is neutralised and the solution 

 evaporated to dryness, preferably under reduced pressure, 

 and the residue repeatedly extracted with hot absolute 

 alcohol. The alcoholic solution is evaporated to dryness 

 and the residue once more extracted with alcohol. 



(a) Mannitol, if present, crystallises out on cooling 

 as long needles, m.p. 168° C. It may be characterised by 

 formation of the tribenzylidene derivative, m.p. 224° C. 



(6) Addition of four or five volumes of ether to the 

 alcoholic extract precipitates glycerol, trimethylene glycol 

 or butylene glycol, which can be separated by distillation 

 in vacuo. Glycerol and trimethylene glycol can be identi- 

 fied also as the l^enzoyl or phenylurethane derivatives, 

 and butylene glycol as the phenylurethane derivative. 



Acetoin may be detected in the metabolism solution 

 by the Voges-Proskauer reaction or by O'Meara's modifica- 

 tion (addition of creatine). 



5. MiscelUuieoas Products. — The addition of fei'ric 

 chloride to a metabolism solution (particularly useful in 



