I. LIPOIC ACID 577 



vitamin Bis ;^' "• '" however, the latter identity has not been confirmed ex- 

 perimentally.'-'' 



The vitamin was first isolated in pure form by Reed, Clunsalus, et a/.;'^ 

 the term "lipoic acid" Avas also introduced at this point to describe the 

 (fat-soluble) compound as it was obtained from liver after hydrolysis, pos- 

 sibly from ester or amide combination. I'he isolation procedure from yeast''* 

 disclosed the existence of five chromatograj^hically distinct forms of the 

 factor. These have been characterized by (iunsalus et al.^ as: (1) a bound 

 form, (2) a water-soluble, fat solvent-insoluble form, referred to as "water- 

 soluble complex," and three fat-soluble varieties referred to as (3) a weak 

 acid, (4) a strong acid, and (5) a neutral substance. The first form was 

 found very widely distributed; the second was observed principally after 

 enzyme digestion. The three fat-soluble compounds remained after hydroly- 

 sis with acid or alkali. However, Heed c( o/.'^' '^ reported only two forms 

 after hydrolysis. This may possibly have been due to production of some 

 forms during operational treatments, or to conversion of natural conjugates 

 (e.g., esters or amides) to the two principal compounds remaining. The 

 two forms, when bioautographed on paper with water-saturated butanol, 

 exhibited Rf values of 0.90 and 0.00.'^ The Rf 0.60 form was largely con- 

 verted to the Rf 0.90 variety (a more polar substance). A similar intercon- 

 version was reported by the Lederle group in their isolation of protogen;'^ 

 they observed a marked increase in water solubility and designated the 

 process protogen A — > protogen B. 



The potency of pure hpoic acid (the Rp O.GO form, called a-lipoic acid)'^ 

 was found to be 250,000 times that of yeast, based on the stimulation pro- 

 ^^ded in the S. faecalis test for pyruvate oxidation. As a replacing agent for 

 acetate in L. casei, lipoic acid was 15,000,000 times as potent; half-maxi- 

 mum growth was provided by 1.7 X 10"^ y per milliliter of culture medium. 

 This compound thus ranks with biotin, vitamin Bi> , and the folic acid 

 group as an extremely active biocatalyst. 



Studies on the structure of the lipoic acids have been both intensive and 

 numerous. The picture of the interrelationships among the various forms 

 has rapidly emerged, although the important details are not yet all worked 



" E. E. Snell and L. D. Wright, Ann. Rev. Biochem. 19, 307 (1950). 



12 V. H. Cheldolin, Nutrition Revs. 9, 289 (1951). 



•2» Since this manuscript was prepared, a roi)ort lias t)een issued indicating; a relation- 

 ship between vitamin Bi, and orotic acid [L. Manna and S. M. llauKe, ./. Biol. 

 Chcm. 202, 91 (1953)]. 



»» L. J. Reed, B. G. De Busk, I. C. Gunsalus, and C. S. HornherKer, .Jr., Hcivncv 114, 

 93 (1951). 



'^ L. J. Reed, B. G. DeBusk, P. IM. John.ston, and .M. E. Getzendaner, ./. BiiA. ('hem. 

 192, 851 (1951). 



iM.. J. Reed, M. E. Getzendaner, B. G. De Busk, and V. M. Johnston, ./. Biol. 

 C/iem. 192, 859 (1951). 



