Analysis of organophosphate flame retardant diester metabolites in human urine by liquid chromatography electrospray ionisation tandem mass spectrometry

• We have developed a new method for the analysis of dialkyl and diaryl phosphates.
• OASIS WAX, a weak anion exchange sorbent, had the best performance.
• Derivatisation by methylation does not improve the analysis of DAPs by LC–MS/MS.
• Diphenyl phosphate was the main DAP metabolite in urine.
• GC–MS/MS is superior in terms of sensitivity for the chlorinated DAPs.

A new analytical method was developed for the determination of dialkyl and diaryl phosphates (DAPs), which are metabolites of organophosphate triesters (PFRs), in human urine. Target DAPs included dibutyl phosphate (DBP), diphenyl phosphate (DPHP), bis(2-butoxyethyl) phosphate (BBOEP), bis(2-chloroethyl) phosphate (BCEP), bis(1-chloro-2-propyl) phosphate (BCPP), and bis(1,3-dichloro-2-propyl) phosphate (BDCIPP). Sample preparation was based on solid phase extraction using a weak anion exchange sorbent (Oasis WAX). Although several instrumental techniques have been tested, best results were obtained with reversed phase liquid chromatography–negative electrospray ionisation tandem mass spectrometry (LC–ESI-MS/MS) taking the total analysis time into account. Method accuracy at 3 ng/mL in pooled urine ranged between 69 and 119% (recovery), while inter-day imprecision (as relative standard deviation) was <31%. The performance of the LC–MS/MS method was compared to a method based on gas chromatography–electron impact tandem mass spectrometry (GC–MS/MS) and a good correlation (Pearson r = 0.82, p < 0.01) between the results of these two methods was obtained for DPHP. LC–MS/MS analysis was more suitable for DPHP and BBOEP with respective method limits of quantification (mLOQ) of 0.3 and 0.15 ng/mL. In contrast, GC–MS/MS had a better sensitivity for BCEP, BCIPP, and BDCIPP, their respective mLOQs being 0.1, 0.06, 0.02 ng/mL, compared to 1.2, 3.7, and 0.5 ng/mL by LC–MS/MS. A set of urine samples from volunteers was analysed, in which DPHP was the major DAP metabolite. A significant increase of DPHP levels was observed in the group of smokers (geometric mean of 1.55 ng/mL) compared to the non-smokers (geometric mean of 0.88 ng/mL). Metabolic transformation of triphenyl phosphate to DPHP by metabolic enzymes induced in smokers could be an explanation for this observation.