ESTIMATION 



and G. E. Clock. "^ The dried tissue was extracted with water and 

 the fractions insoluble in 70 % acetone and soluble in ether were 

 both removed from the extract. Glucose was then removed by- 

 fermentation with yeast and both acidic and basic substances were 

 removed by adsorption on ion exchange materials. The free inositol 

 in the solution was then quantitatively oxidised with periodic acid 

 and the excess estimated iodometrically. Water-soluble combined 

 inositol was estimated after acid hydrolysis of the aqueous extract. 



2VIicrobiological Methods 



With inositol, as with so many other members of the vitamin B 

 complex, biological and chemical methods of estimation have given 

 place to microbiological methods, which are generally less tedious and 

 more accurate than biological methods and more specific than chemical 

 methods. 



The first microbiological assay method was that of D. W. Woolley,^ 

 who used the Hansen No. i strain of Toronto yeast, Saccharomyces 

 cerevisiae. This was grown on a basal medium consisting of glucose, 

 casein hydrolysate, salts, other members of the vitamin B complex 

 and an aqueous extract of malt sprouts to supply " bios II ". The 

 response of the organism to graded doses of the test solution and of a 

 standard inositol solution was measured turbidimetrically. The error 

 was estimated to be not more than 5 or 6 %. A similar procedure was 

 used by R. J. Williams et al.,^ but in this instance the Gebruder-Meyer 

 strain of yeast was employed. V. Jurist and J. R. Foy ^^ also used 

 yeast but in their basal medium the only constituents of uncertain 

 composition were casein hydrolysate and a folic acid concentrate. 

 Atkin et al}^ used 5. carlsbergensis. 



G. W. Beadle ^^ used a mutant of Neurospora crassa, which was 

 claimed to have the advantage over yeast that the basal medium was 

 simpler and the mould did not grow at all in the absence of inositol. 

 Amounts of inositol ranging from 5 to 30 fig. per 20 ml. could be 

 estimated with an error of ± 0-3 fxg. 



Biukholder et al}^ used the yeast, Kloeckera brevis, for the assay 

 of inositol, whilst Emery et al.^^ compared the response obtained with 

 this organism and that given by another yeast, Schizosaccharomyces 

 pombe. 



References to Section 5 



1. D. W. Woolley, /. Biol. Chem., 1940, 136, 113; 1941, 139, 29; 



/. Nutrition, 1941, 21, Suppl., 17. 



2. L. B. Winter, Biochem. J., 1934, 28, 6 ; 1940, 34, 249. 



3. J. Needham, ibid., 1923, 17, 422, 431. 



4. P. Fleury and J. Marque, Compt. rend., 1939, 188, 1686 ; /. Pharm. 



Chim., 1929, [8], 10, 241. 



569 



