Differential Autoradiography and Spin‐Column Purification of Sugars on Activated Charcoal
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Methods utilizing activated charcoal for separating radiolabeled sugars from each other and from water are described. The first method alleviates a practical limitation for use of tritiated sugars as a tracer in planta. Tritium exchange with water can now be readily quantified because the method allows a more rapid separation of sugars and water than has been previously described. A spin column containing activated charcoal and powdered cellulose retained (>99%) sucrose during elution (>99%) of 3H2O in water rinses. The retained sucrose could then be eluted (95%) from the column by rinsing with aqueous methanol. The second method partially resolves another limitation in the use of radiolabeled sugars as tracers in planta. We can now ascertain the specified distribution of various radiolabeled sugars within particular organs. Charcoal filter paper was used to adsorb sucrose and hexose sugars for development of differential autoradiographs of sugars blotted from stem cross-sections of sorghum (Sorghum bicolor [L.] Moench). The hexose sugars were selectively removed during rinsing of the charcoal paper in water. Autoradiography was performed before and after rinsing. The autoradiographic image obtained before rinsing reflected the presence on the blot of both radiolabeled sucrose and hexose sugars. In comparison, the image obtained after rinsing no longer reflected the presence of radiolabeled hexose sugars. These methods help retire studies of carbon dynamics in plants by enhancing our ability to detect in planta movement of sugars through particular metabolic pathways and to detect the spatial distribution within organs of particular radiolabeled sugars.
author list (cited authors)
Tarpley, L., & Vietor, D. M.