Accurate quantification of PGE₂ in the polyposis in rat colon (Pirc) model by surrogate analyte-based UPLC–MS/MS

An accurate and reliable UPLC–MS/MS method is reported for the quantification of endogenous Prostaglandin E2 (PGE₂) in rat colonic mucosa and polyps. This method adopted the “surrogate analyte plus authentic bio-matrix” approach, using two different stable isotopic labeled analogs — PGE₂-d9 as the surrogate analyte and PGE₂-d4 as the internal standard. A quantitative standard curve was constructed with the surrogate analyte in colonic mucosa homogenate, and the method was successfully validated with the authentic bio-matrix. Concentrations of endogenous PGE₂ in both normal and inflammatory tissue homogenates were back-calculated based on the regression equation. Because of no endogenous interference on the surrogate analyte determination, the specificity was particularly good. By using authentic bio-matrix for validation, the matrix effect and exaction recovery are identically same for the quantitative standard curve and actual samples – this notably increased the assay accuracy. The method is easy, fast, robust and reliable for colon PGE₂ determination. This “surrogate analyte” approach was applied to measure the Pirc (an Apc-mutant rat kindred that models human FAP) mucosa and polyps PGE₂, one of the strong biomarkers of colorectal cancer. A similar concept could be applied to endogenous biomarkers in other tissues.