SIMULTANEOUS DETERMINATION OF CROSS-REACTIVE LEUKOTRIENES IN BIOLOGICAL MATRICES USING ONLINE LIQUID-CHROMATOGRAPHY IMMUNOCHEMICAL DETECTION

Sulfidopeptide leukotrienes, which are important biomarkers for severa l diseases, are commonly measured by microtiter plate immunoassays, Th ese immunoassays, however, cannot distinguish between several structur ally similar leukotrienes and their cross-reactive metabolites and, th erefore, need extensive sample handling and fractionation by means of liquid chromatography (LC), This paper describes the development and a utomation of a continuous-now immunochemical detection (ICD) system an d its subsequent on-line coupling to LC, The online LC-ICD system base d on fluorescence-labeled leukotriene E(4) (LTE(4)) was used to determ ine sulfidopeptide leukotrienes and their cross-reactive metabolites i n a single run, Furthermore, biological matrices, e,g,, urine and huma n cell. extracts, were analyzed, the only sample pretreatment being on -line solid-phase extraction (SPE) on a novel RP-C-4 restricted-access support, The determination limit of LTE(4) in urine was 0.2 ng/mL (80 0 fmol; injection volume, 2000 mu L; signal-to-noise ratio, 10), The s ystem was linear from 0.2 to 1.0 ng/mL LTE(4), Using nonlinear curve-f itting, the range could be expanded to 2.5 ng/mL, It is shown that, be sides quantitation of known analytes, on-line LC-ICD is useful in the discovery of cross-reactive LTE(4) metabolites.