532 



BIOTIN 



A 0.3% solution of the compound in 0.1 N NaOH showed an optical 

 activity of W^' = +92°.27 



C. CONSTITUTION 



1 Functional Groups 



The easy saponification of biotin methyl ester indicated the loss of one 

 methylene group. Re-esterification of the free acid could be accomplished by 



03 .04 X)5 j06 



ALKALI (MILLI-EQUIVALENTS) 



Fig. 2. Electrometric titration of 17.99 mg. of biotin with NaOH. From D. B. 

 Melville, Vitamins and Hormones 2, 42 (1944). 



the use of diazomethane : the methyl ester produced in this manner was iden- 

 tical with that isolated from esterified liver fractions. Biotin, C10H16O3N2S, 

 in contrast to the methyl ester, did not liberate volatile iodides when treated 

 with HI and consequently does not contain — OCH3, NCH3, or SCH3 

 groups. Electrometric titration of biotin yielded a curve (Fig. 2) resembling 

 that of a monocarboxylic acid.^^ Biotin crystallized from strongly acid 

 solutions in form of the free acid. This fact, together with the shape of the 

 titration curve for biotin, indicates that the basic group of groups present 

 in the molecule were very weak. No inactivation occurred, ^^ and no color 

 was produced^^ when the substance was treated with ninhydrin. Further, 

 no nitrogen was formed when biotin was inactivated by the Van Slyke 



" K. Hofmann, D. B. Melville, and V. du Vigneaud, /. Biol. Chem. 141, 207 (1941). 



