The Biosynthesis of Carbon-14-Labeled Compounds. III. The Separation and Isolation of Sugars by Ion-exchange Chromatography

One of the maj or difficulties in the preparation of labeled compounds by biosynthetic procedures is the isolation of chemically and radiochemically pure compounds. Often compounds which appear chemically pure are found to contain radioactive contaminants which may make such labeled compounds virtually useless in biological experiments. UDENFRIEND and GIBBS (11) and more recently PUTMAN and HASSID (8) used paper chromatography to effect the separation and isolation of several labeled sugars. The latter workers pointed out that such a procedure enabled them to prepare sugars of very high specific activity since it was unnecessary to add any carriers. Furthermore, they were able to obtain labeled sugars containing extremely smiiall amounts of radioactive contaminants. NOGGLE and BOLOMEY (5) used column chromatography to effect the separation of glucose and fructose while GIBBs et al. (2) prepared labeled sucrose by fermenting out the miionosaccharides. NOGGLE and SCHUMACHER (6) separated the sugars present in plant extracts into a monosaccharide, a disaccharide, and a trisaccharide fraction by column chromatography on charcoal. Certain disadvantages are found in all of these methods, such as limited capacity, inability to obtain radiochemically pure products, or the necessity of adding carrier, so that it seemed desirable to explore other methods that would minimize these difficulties. It has been known for a long time that sugars and certain other polyhydroxy compounds react readily with borate ions to form borate complexes which are negatively charged ions. KHYM and ZILL (4) de1nonstrated that a mixture of these sugar-borate complexes could be resolved by ionexchange chromatography on strong-base anion exchange resins. Subsequently, NOGGLE and ZILL (7) showed that this procedure could be used for the quantitative analysis of the sugars found in plant extracts and wlhen this method was tried as a preparative method for separating labeled sugars prepared biosynthetically, it was found to give results that overcame many of the difficulties encountered in otlher preparative procedures. The experimental results are presented in this paper.