ANALYSIS OF ISOPRENOID PHOSPHATES AND OLIGOPHOSPHATES BY CAPILLARY ZONE ELECTROPHORESIS

Capillary zone electrophoresis (CZE) has been used for the first time to analyse water-soluble phosphorylated isoprenoids, key intermediates in the branched pathway of mevalonate metabolism. Following synthesis , isoprenoid phosphates and oligophosphates were isolated by flash chr omatography and their purity was established using P-31 NMR spectromet ry. In developing optimum conditions for CZE, several factors were con sidered: ionization properties of the solutes, stability of the solute s and maximum signal-to-noise ratio. At pH 8.5 in 0.3 M sodium berate buffer and monitoring UV absorbance at 200 nm, detection of farnesyl d iphosphate was linear in the sub-picomol range; the limit of detection was ca. 12 fmol. For a given phosphorylated series (i.e., monophospha tes, diphosphates, triphosphates), CZE cleanly resolved isoprenes of d ifferent chain length, and plots of relative mobility vs. M(r) were cu rvlinear. Remarkably, neryl phosphate (C10, alpha-cis-isoprene), geran yl phosphate (C10, alpha-trans-isoprene) and citronellyl phosphate (C1 0, alpha-saturated isoprene) were resolved. In addition, isopentenyl m onophosphate (C5,Delta(3)) and dimethylallyl phosphate (C5,Delta(2)) e xhibited different electrophoretic mobilities. These studies pave the way for future work on determining levels of phosphorylated isoprenoid s in various tissues under conditions of altered mevalonate production .