Toxisterol __Substance248
True or false, Do Mammals synthesize Vitamins D ?
Do Fish synthesize Vitamin D3 ?

C28H44O
Melting point about 50°.305
Narrow absorption band with maximum at 248 mµ.
Molecular extinction coefficient at 18,300. 45
[ alpha ]22D -16°(in chloroform). 305

In 1927 Morton et al., 213extanding the work of of Pohl, 30showed that when ergosterol is irradiated, its characteristic absorptionbands disappear, and a single new band of great intensity develops at 247or 248 mµ, which in turn fades away to a weak general absorptionin the far ultraviolet region. With inadequate biological evidence, theassumption was easily made that this new band represented Vitamin D. Smakula,229
however, concluded that Substance248 was not the vitamin. Bills et al. 239showed that the appearance of the band at 248 mµ coincided not withthe development, but with the
destruction, of antiricketic potency. They, and also van Wilk andReerink, 306 associated
Substance 248with isoergosterol. Cox and Bills 307contributed further evidence of
relationship to the isoergosterols, but noted a point of difference,namely, that substance
248 does not percipitate with digitonin.

Bills et al. 239 foundthat irradiation products withdrawn for examination at the moment
when the Substance 248was at its maximum concentration still contained some Vitamin D. Furtherirradiation totally eliminated antiricketic activity, while only a smallamount of Substance 248was destroyed. Laquer and Linsert 306attributed a toxic quality to Substance248, and proposed the name Toxisterin ( German forToxisterol ). The productwhich they investigated still contained some Vitamin D, but not enoughto account for the relatively great toxicity. It was evidently a mixturesimilar to the one which
Hoyle 309 obtained by limitedoverirradiation and which he reported to have toxic-calcifying propertiesall out of proportion to the antiricketic potency. Such a mixture, unfortunately,
seems to have been the " Vitamin D " of the I. G. Farbenindustriepatent 310 of 1928
( the Vigantol of toxic repute ). The claims of this patent, andof the corresponding American patent, 311call for the irradiation of ergosterol to be continued just until the absorptionspectrum of what is now recognized as Toxisterolattains its maximum ! The mistake was clearly the result of assuming, inthe absence of proper bioassays, that the spectographically most conspicuousirradiation product was the Vitamin. There is considerable evidence, spectographicand toxicologic, that Toxisterolis formed most readily when alcohol is the solvent in which the ergosterolis irradiated. 238,239, 306,309, 312

The purest described preparation of Toxisterolis that of Linsert ( Farbenindustrie ) patent 305for isolating, from the irradiation products of Vitamin D, a " productwhich is essentially characterized by its increasing action on the bloodlevel of blood calcium ".
According to the example, crystaline VitaminD, made from ergosterol, is irradiated in
heptane in the absence of oxygen until itsabsorption maximum at 265 mµ substantially
disappears. The product is converted into the 3,5-dinitrobenzoate,which is crystallized
free of the suprasterols. Saponificationof the dinitrobenzoate,which is dextrorotary, [ alpha ]22D + 33° in acetone, m. p. 130°,gives the free drug as a whitish powder which levorotary, [alpha ]22D - 16° in chloroform.( A similar anomalous difference in sign of rotation between dinitrobenzoateand free sterol is noted in tachysterol 3 313).

Ergosterol series

Ergosterol2
|
Lumesterol2
|
Protachysterol2
|
Precalciferol2
| | goes both directions
Calciferol2
/ \
Toxisterol ---------- Suprasterols Iand II
( Substance 248 )

The exceptionally low melting point, about 50°, castssome doubt on the purity of the preparation, but the product is statedto be " practically antirachitically inactive ". It is worth noting thatthe patent description of this drug, which was one of a series of bloodcalcium raising substances then under investigation by I. G. Farbenindustrie( following the Vigantol episode ), makes no mention of the term Toxisteroland no reference to the absorption spectrum of the product by which itcould identified as Substance 248. Athough the isolation of Toxisterolhas never been described elsewhere than in this patent and its Europeancounterparts, the German organic chemists appear to have been
very familiar with Toxisterol to the extentof publishing its absorption spectrum showing the extinction coefficient,A. Windaus et al. 45.

• 30 R.Pohl, Nachr.Ges. Wiss. Göttingen, Math physik. Kl. 175 (1927).
• 45 A.Windaus, A. Lüttringhaus, and P. Busse, Nachr. Ges. Wiss.Göttingen, Math physik. Kl., FachgruppeIII, 150 (1932).
• 213 R. A. Morton, I. M. Heilbron,and E. D. Kamm, J. Chem. Soc. 1927, 2000.
• 229 D. Smakula, Nachr. Ges.Wiss. Göttingen, Math physik. Kl. 49 (1928).
• 238 C. E. Bills, E. M. Honeywell,and W. M. Cox, Jr., J. Biol. Chem. 92, 601 (1931).
• 239 C. E. Bills, E. M. Honeywell,and W. M. Cox, Jr., J. Biol. Chem. 96, 307 (1932).
• 305 O. Linsert, U. S. Pat. 2,030,377(1936).
• 306 A. van Wijk, and E. H. Reerink,Nature 122, 648 (1928).
• 307 W. M. Cox Jr., and C. E.Bills, J. Biol. Chem. 88, 709 (1930).
• 308 F. Laquer, and O. Linsert,Klin. Wochschr. 12, 753 (1933).
• 309 J. C. Hoyle, J. Pharmacol.Exptl. Therap. 40, 351 (1930).
• 310 I. G. Farbenindustrie Aktiengesellschaft,British Pat. 296,093 (1928).
• 311 W. Zimmermann and W. Frankenburger,U. S. Pat. 1,896,191 (1933).
• 312 W. E. Dixon, and J. C. Hoyle,Brit Med. J. 2, 832 (1928);
L. J. Harris, and T. Moore, Biochem. J. 22, 1461 (1929);
J. C. Hoyle, and H. Buckland, Bichem. J. 23, 558 (1929);
R. Kern, M. F. Montgomery, and E. U. Still, J. Biol. Chem.93, 365 (1931).
• 313 A. Windaus, M. Deppe, andW. Wunderlich, Ann. 533, 118 (1937).