Determination of highly polar catecholamine with liquid chromatography–tandem mass spectrometry using weak cation-exchange stationary phase to increase retention time

This paper reports method development and validation work to determine highly polar bases, catecholamine compounds, using weak cation-exchange liquid chromatography of low ionic strength mobile phase with electrospray tandem mass spectrometry. Catecholamine compounds, such as epinephrine and norepinephrine, well-known biomarkers to diagnose hypertension disease, spiked in saline solutions are purified with solid phase extraction (SPE) using alumina powders. The extracts are loaded into a weak cation-exchange liquid chromatographic column via an injection loop and analyzed with electrospray-mass spectrometer. The de-salted extracts contain only small amounts of electrolytes to avoid saturating weak cation-exchange sites in the stationary phase with sodium ions. Using carefully selected mobile-phase solvents with optimized compositions (acetonitrile and water 10:90 v/v) and with dilute acid additives (acetic acid 0.1% v/v), we are able to elute catecholamine at sufficient retention times to avoid co-elution of saline matrix residues while maintaining adequate electrospray ionization efficiency of these compounds. Using epinephrine and norepinephrine standards, these methods are validated at the range of 5 to 500 ng mL− 1. The measurement accuracy and precision of using epinephrine standards are within 12% and 5.3% respectively, whereas the accuracy and precision are within 6.0% and 4.2% respectively using epinephrine standards.The detection limits of epinephrine and norepinephrine are 0.10 ng mL− 1 and 0.45 ng mL− 1 respectively. The recovery percentages of our solid phase extraction methods using alumina powders are higher than 74%. When the validated calibration curves are used to determine epinephrine and norepinephrine in rat blood dialysates, the determination errors of accuracy and precision are both within 4%, while the determination errors are within 3% in rat blood plasma samples.