Organophosphate ester metabolites in human breast milk determined by online solid phase extraction coupled to high pressure liquid chromatography tandem mass spectrometry.

Abstract

The analysis of metabolites of organophosphate esters (OPEs) in human breast milk is essential to evaluate OPE and OPE metabolite exposure of newborns. In the current study, an analytical method which only needs a small amount of breast milk (100μl) was developed and validated for six diester metabolites and three hydroxylated metabolites applying salt-induced liquid-liquid extraction (SI-LLE) and dispersive solid phase extraction (d-SPE) for sample preparation and online solid phase extraction coupled to high pressure chromatography tandem mass spectrometry (online-SPE-HPLC-MS/MS) for quantitative measurement. The final method consisted of an extraction with formic acid (FA)/acetonitrile (1:200, v/v) and a cleanup with C18 d-SPE. The final extracts were trapped on a C18 cartridge with application of a wash step of 2ml 0.1% FA milli-Q/methanol (98:2, v/v). Method detection limits (MDLs) ranging from 21.7ng/l for BBOEHEP to 500ng/l for BCIPP and average recoveries ranging from 58% for 5-OH-EHDPHP to 120% for BCIPP were achieved. Thirty-three breast milk samples from the LINC (Linking EDCs in maternal Nutrition to Child health) cohort collected in three distinct areas in The Netherlands were analyzed using the validated method. BCEP, BCIPP, BCIPHPP, BDCIPP, and 5-OH-EHDPHP were not detected in any of the samples, while BBOEP was the most frequently detected metabolite with a concentration range of <MDL to l.47ng/ml, followed by DPhP and BBOEHEP, detected in ranges of <MDL to 0.09 and <MDL to 0.027ng/ml. The results indicated that OPEs entering the human body are only to a limited extent excreted via breast milk.