Quantification of gastric cancer biomarkers in the urine matrix by high internal phase emulsion polymer-based µsolid-phase extraction followed by HPLC-MS/MS.

This paper reports on the development of an analytical method for the simultaneous determination of 4-methyl phenol, 4-hydroxybenzoic acid and five bile acids, namely cholic acid, glycochenodeoxycholic acid, chenodeoxycholic acid, deoxycholic acid, and hyodeoxycholic acid (logP 1.6-4.9) in urine, exploring porous acrylic polymers as unconventional solid-phase extraction sorbents before HPLC-MS/MS analysis. These materials, obtained by polymerization of high internal phase emulsions with different composition, were used in a miniaturized setting (µSPE), preliminarily in synthetic urine samples enriched with 200 ng mL-1 of each selected compound. It was found that the polyethylene glycol (PEG)-modified polymer combines good analytical performance, in terms of extraction efficiency (76-100% for most analytes), from 1 mL samples and single-fraction elution in pure ethanol. Recovery was assessed in untreated real urine (1 mL, pH ~ 3.5) at three quality control levels (50, 200, 500 ng mL-1) observing inter-day RSD below 20% (n = 3). The polymer exhibited a service life of at least ten sorption/desorption cycles, a further strength point of this sample treatment ascertained by greenness metrics. This streamlined procedure also provided satisfactory sample clean-up, with a negligible matrix-effect, as observed in HPLC-MS/MS. An ad hoc chromatographic method was developed to enable the simultaneous separation and quantification of all targets in a single run. Good linearity was observed in the range 10-600 ng mL-1 (R2 > 0.9919) with suitable sensitivity (a few ng mL-1) to future application for monitoring these endogenous compounds.