560 THE BIOCHEMISTRY OF B VITAMINS 



nervous disorders and loss of weight. A spasticity develops which in the 

 later stages of the deficiency causes the rats to assume a typical kangaroo- 

 like posture, and unless the condition is remedied, death ensues. 74 



The principle in egg white which is responsible for the detrimental effect 

 has the properties characteristic of a protein; that is, it is destroyed by 

 heat or mild hydrolysis with dilute acid and is precipitated by such 

 agents as ammonium sulfate. A naturally occurring substance which 

 exerts a protective action against this egg white injury has been termed 

 vitamin H by Gyorgy. 75 The identity of vitamin H with biotin was 

 finally established 76 > 77, 78 by testing a sample of biotin methyl ester 

 isolated by Kogl. 2 



The protein in raw egg white which renders biotin unavailable to 

 animals was found also to prevent the utilization of biotin by Saccharo- 

 myces cerevisiae 79 and other microorganisms requiring biotin for growth. 

 Thus, a protein constituent of the egg white forms a stable, nondialyzable 

 complex with biotin. The combination between the protein and biotin 

 was found to occur in stoichiometric amounts. With the microbiological 

 test for the egg white factor, the protein which combines with biotin has 

 been isolated by Eakin and associates 79, 80 in crystalline form and named 

 avidin. Purified avidin produced effects in rats similar to those caused 

 by dried egg white. 81 With the assumption that one molecule of biotin 

 combines with one molecule of protein, the molecular weight of avidin 

 has been calculated to be 43,500. The isoelectric point occurs at approxi- 

 mately pH 10. 82 Some loss of activity was inherent in crystallization, since 

 the potency of a crystalline sample of avidin was approximately 4,000 

 units per gram as compared with 7,000 units per gram for amorphous 

 preparations. By definition one unit of avidin combines with 1 y of 

 biotin. 80 



It is interesting to note that avidin passes through the alimentary tract 

 of animals unchanged and can be demonstrated in the feces. 83 Even liver, 

 kidney and proteolytic enzymes are inactive in liberating biotin combined 

 with avidin. 84 However, destruction of avidin by heat treatment liberated 

 material with biotin-like activity. 



The ability of various analogues of biotin to combine with avidin has 

 been determined. For example, it has been reported that sufficient avidin 

 completely inhibits the growth promoting activity of biotin sulfone for 

 Saccharomyces cerevisiae. 47 Since limited amounts of avidin added to a 

 medium containing excess biotin sulfone produce responses in the yeast 

 characteristic of biotin but not of biotin sulfone, it appears that biotin 

 sulfone liberates biotin from the complex. 



A method for determining the avidin-combinability of biotin analogues 

 has been developed by Wright and Skeggs, 85,86 in which the "relative 



