A Novel, Automated Dispersive Pipette Extraction Technology Greatly Simplifies Catecholamine Sample Preparation for Downstream LC-MS/MS Analysis.

Abstract

Catecholamines are integral neurotransmitters in the central and peripheral nervous system. Clinically, catecholamine levels are determined to help diagnose disease and measure corresponding therapeutic effectiveness. However, manual extraction of catecholamines and their metabolites may be labor-intensive and user-variable and require a variety of peripheral laboratory devices, especially at low sample concentrations. Here, we propose a novel solid-phase extraction (SPE) method using patented dispersive pipette extraction (DPX) tip technology. The tips are readily integrated into an automated workflow to extract these compounds from urine, which increases analytical throughput while removing human variability and error. Diphenylboronic acid (DPBA) forms a stable, negatively charged complex with catecholamines in the samples, and when aspirated into the DPX tip, the complexed analytes are retained on a styrene divinyl benzene sorbent. Wash buffers remove interfering compounds, after which the complex is eluted from the tip using an acidic aqueous solution and subsequently measured via liquid chromatography with tandem mass spectrometry (LC-MS/MS). The automated DPX method for catecholamine sample preparation from urine has excellent linearity over more than three orders of magnitude with concentrations ranging from 0.5 to 1000 ng/mL, with replicate analyses resulting in coefficients of variation of less than 8%. This high-throughput workflow is appropriate for use in regulated laboratories.