Analytical method considerations regarding carryover for monophosphate prodrugs for in vivo samples by liquid chromatography-tandem mass spectrometry.

Three different components that impact carryover in a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method were evaluated to establish baseline conditions for analyzing in vivo samples for twelve monophosphate prodrug compounds and their corresponding parent compounds. The three components were: wash solvent modifier, column shell material (metal vs. metal free), and tubing composition. These components were tested for their impact on system carryover by using rat plasma extracted samples. It was determined that a wash solution containing hexylamine yielded the lowest average carryover of the solutions tested. In addition, metal free columns and PEEK (poly ether ether ketone) tubing yielded the lowest carryover when compared to metal columns, stainless steel tubing and nickel tubing. These conditions were also tested against the parent molecules for each prodrug in the test set, to ensure that changing the conditions for the prodrugs did not impact the ability to analyze the parent, since there is typically a desire to measure both compounds in study samples. Under all conditions, the carryover of the corresponding parent molecule was not adversely impacted in these studies.