ISOLATION OF SERINE 53 



acid as possible. The residue is dissolved in water and boiled with 

 yellow lead oxide until a test portion of the cold solution no longer 

 gives a reaction for chlorine. 1 The filtered solution is freed from lead 

 by hydrogen sulphide and the filtrate from lead sulphide is evaporated 

 to a small volume, when aspartic acid crystallises out. The mother 

 liquor contains principally serine, but more aspartic acid and other 

 products are also present. 



Note. Osborne and Liddle [1910, 2] have observed that an intermediate fraction between 

 fractions III. and IV. containing aspartic ester and leucine ester and possibly also phenyl- 

 alanine ester may distil. The separation of leucine and aspartic acid (or glutamic acid) 

 is impossible by fractional crystallisation. The fraction should be treated as described and 

 the leucine separated from the aspartic acid by neutralising with soda and crystallising. 

 Leucine results. On acidifying the filtrate and again crystallising the aspartic acid is 

 obtained. 



(e) Glutamic Acid. 



The filtrate from the barium aspartate is exactly freed from barium 

 by sulphuric acid and the solution is evaporated to dryness in vacuo. 

 The residue is dissolved in water, the solution decolorised, if neces- 

 sary, by boiling with charcoal and the glutamic acid is precipitated 

 as hydrochloride by passing in dry gaseous hydrogen chloride. A 

 further quantity of glutamic acid hydrochloride may be obtained from 

 the mother liquor by concentration and similar treatment. Practically 

 the whole of the glutamic acid present in the protein is thus obtained 

 as hydrochloride. The larger portion is separated directly, before 

 the mixture of amino acids is esterified. 



Glutamic acid is obtained from the hydrochloride by treatment with 

 the calculated quantity of caustic soda to combine with the hydro- 

 chloric acid and by crystallisation from water, in which it is soluble, 

 when pure, with some difficulty. Elementary analysis of the free 

 acid, or of its hydrochloride, determines its identity and its weight 

 gives the amount in the protein. 



(/) Serine. 



It is most difficult to isolate serine and obtain it in a pure state. 

 The solution from which the active aspartic acid has crystallised 

 out is neutralised, if acid, with caustic soda and concentrated. Serine 

 crystallises out in crusts of monoclinic crystals, and is identified by its 

 melting-point of 240 and elementary analysis. 



1 Levene and Van Slyke [1910] point out that, if excess of lead oxide be used, the 

 insoluble lead salt of aspartic acid is formed with consequent loss of this amino acid. 



