The isolation and identification of 2,2'-dithiolisobutyric acid from asparagus.

A non-protein sulfhydryl substance occurs in edible asparagus. Fresh press-juice gives a test for both sulfhydryl and disulfide groups, so that an equilibrium apparently exists in the plant between the sulfhydryl and the disulfide (oxidized) form. However, when press-juice is exposed to atmospheric oxygen, the sulfhydryl form goes over completely to the oxidized form. Attempts to identify this substance as cystine or oxidized glutathione either by isolation or by polarographic analysis have been unsuccessful.’ It has now been found that the disulfide compound is acidic, thus permitting its isolation and purification by the extraction of concentrated acidified asparagus juice with butanol, removal of the substance from the butanol with sodium bicarbonate solution, subsequent acidification of the bicarbonate solution, and reextraction of the disulfide compound therefrom with butanol. A repetition of this distribution with several solvents gave a material which appeared to be pure. However, it was not found possible to crystallize the disulfide compound or its derivat,ives such as the methyl ester, since in the format,ion of the disulfide material from the sulfhydryl compound a relatively large polymer appeared to form. Even ethereal solutions of the disulfide compound became viscous on standing. However, upon reduction wit’h sodium in liquid ammonia, 2,2’-dithiolisobutyric acid (I) was isolated in pure crystalline form. The proof of the structure of this material was obtained from the accompanying series of reactions. Hydrogenolysis of 2,2’-dithiolisobutyric acid (I) with Raney’s nickel (1) gave isobutyric acid (III) which was identified as the p-phenylphenacyl ester (2). Refluxing (I) with formaldehyde (3) gave a product whose analysis was in agreement with that of the previously undescribed 1,3-dithiane-5-carboxylic acid (II). The reaction of the sodium salt of the reduced product in liquid ammonia with methyliodide (4) gave dimethyl-2,2’-dithiolisobutyric acid (IV). This was shown by