Differentiation between ribose-3-phosphate and ribose-5-phosphate by means of the orcinol-pentose reaction.

During a study of the energy-rich phosphorus compounds of higher plants, it became necessary to differentiate between ribose-3-phosphate and ribosed-phosphate. The only method available for distinguishing between these esters at low levels (quantities of 1 mg. or less) is that of Levene and Stiller (1). This method depends on the difference in the rate at which these compounds release their phosphorus in weak acid at 100’ over the course of several hours, the phosphorus from the 3-ester splitting off much more readily than that of the 5-ester. This method may be applied not only to the free ribose phosphates, but also to related compounds which possess them. Schlenk (2) used such a procedure to determine Dhe position of the phosphorus on t,he ribose of diphosphopyridine nucleotide. LePage and Umbreit (3) measured the rate of phosphorus release from the ribose phosphate derived from the adenosine triphosphate of Thiobacillus thioozidans, and concluded, partly on the basis of such data, that this organism possesses an adenosine-3-triphosphate. The adenylic acids possessing the 3or 5-ribosephosphate may also be differentiated by several other methods. Klimek and Parnas (4) devised a method based on the formation of a blue soluble complex by the 5-adenylic acid (myoadenylic acid) in alkaline solution in the presence of copper sulfate. Under the same conditions, only an insoluble precipitate is formed by the 3-adenylate (yeast adenylic acid), which after centrifugation leaves a clear colorless supernatant solution. This procedure has recently been more completely standardized by Berlin and Westerberg (5). The most recent method for differentiation between the two types of adenylic acid is that of Kalckar (G), utilizing a purified deaminase first described by Schmidt (7) which removes the amino group from myoadenylic acid, but not from yeast adenylic acid. The reaction is measured spectrophotometrically. The enzyme is without effect on either adenosine dior triphosphate. The present paper describes a new method for differentiating between ribose-3-phosphate and ribose-5-phosphate. It is based on a difference in