A Test to Differentiate Ergosterol From Cholesterol.

When sulphuric acid or a mixture of selenious and sulphuric acids is used as a reagent for cholesterol dissolved in chloroform, the two layers, that is the chloroform and the acid layer, assume different colorations. With sulphuric acid alone the chloroform layer is cherry red and the acid layer shows green fluorescence. With the selenious-sulphuric acid mixture, the chloroform layer assumes a deep beautiful purple, while the lower layer is red brown but does not display green fluorescence.

When, however, ergosterol is used with either reagent, the acid layer is colored but the chloroform layer remains free from color. With concentrated sulphuric acid and an equal volume of chloroform solution, the acid layer ranges in color from a brownish red in a 1%, to cherry red in a 0.5%, to light cherry red in a 0.1%, to orange in a 0.04%, and to yellowish orange in a 0.025% solution of ergosterol. In all these concentrations the typical green fluorescence is also observed in the acid layer.

With concentrated sulphuric acid containing 125 mg. of sodium selenite per 25 cc. of acid, the chloroform layer ranges in color from dark red brown to light red brown in the above concentrations of ergosterol while no green fluorescence is noted. The chloroform layer remains uncolored.

From the above results the conclusion may be drawn that cholesterol on treatment with acid forms a decomposition product or products giving rise to colored solutions in chloroform and in sulphuric acid or in both. Ergosterol, on the other hand, seems to give a decomposition product or products dissolving with color formation only in sulphuric acid.

Solutions of cholesterol in chloroform remain colorless, while solutions of ergosterol rapidly assume a deep yellow-brown tinge.