High sensitivity HPLC detection of /sup 3/H- and /sup 14/C-radiometabolites of aldosterone using an on-line berthold beta radioactivity detection system

The difficulty of detecting trace quantities of compounds such as hormones, vitamins, pesticides, drugs etc. is common, especially when the compounds lack a chromophore or ability to fluoresce. A variety of studies need to investigate the metabolism, distribution, mode of action, and other properties of compounds using a radioactive tracer as the method of choice. The authors' objective was to separate and quantitate aldosterone and its metabolites which have been extracted from tissues including liver, kidney, plasma, and urine post-injection of a physiological dosage of /sup 3/H- or /sup 14/C-aldosterone. Aldosterone was separated from its polar and reduced metabolites, identified and quantitated using reverse phase HPLC with Dupont Zorbax C/sub 8/ columns eluting with aqueous methanol gradients ranging from 5% to 50%. Jacketed columns were heated (40 to 550C) to enhance resolution of these compounds. The quantities of /sup 3/H- or /sup 14/C-radioactivity encountered in this project necessitated the need for a radioactivity detector which offers sensitivity, reproducibility, together with high efficiency of detection and a low background. For this purpose, the authors have used a Berthold radioactivity Model LB 504 monitor designed for HPLC with automatic data acquisition, data reduction, storage, and printer-plotter output which can be used to integrate each of the separated /sup 3/H- or /sup 14/C-radiometabolites of aldosterone. This technology provided the capability of on-line detection and real-time display of low levels of /sup 3/H- or /sup 14/C-radiometabolites. The Berthold software is ''user friendly'' and provides considerable flexibility to manipulate data and graphics on the screen prior to final printout