Comprehensive monitoring of specific metabolites of tri-(2-ethylhexyl) trimellitate (TEHTM) in urine by column-switching liquid chromatography-tandem mass spectrometry

Abstract

Tri-(2-ethylhexyl) trimellitate (TOTM or TEHTM) is a substitute for the plasticizer di-(2-ethylhexyl) phthalate (DEHP). Here, a fast and robust HPLC method is presented for the first time enabling the simultaneous quantification of several TEHTM metabolites in urine. These are the three TEHTM monoester isomers 1-mono-(2-ethylhexyl) trimellitate (1-MEHTM), 2-mono-(2-ethylhexyl) trimellitate (2-MEHTM), and 4-mono-(2-ethylhexyl) trimellitate (4-MEHTM) as well as several selected side chain oxidized monoesters of TEHTM, namely, 1-mono-(2-ethyl-5-hydroxyhexyl) trimellitate (5OH-1-MEHTM), 2-mono-(2-ethyl-5-hydroxyhexyl) trimellitate (5OH-2-MEHTM), 1-mono-(2-ethyl-5-oxohexyl) trimellitate (5oxo-1-MEHTM), 2-mono-(2-ethyl-5-oxohexyl) trimellitate (5oxo-2-MEHTM), 1-mono-(2-ethyl-5-carboxypentyl) trimellitate (5cx-1-MEPTM), 2-mono-(2-ethyl-5-carboxypentyl) trimellitate (5cx-2-MEPTM), 2-mono-(2-carboxymethylhexyl) trimellitate (2cx-2-MMHTM), and 1-mono-(2-carboxymethylhexyl) trimellitate (2cx-1-MMHTM). The method is characterized by a short sample preparation, for which the urine samples are enzymatically hydrolyzed and cleaned up by an online column arrangement. Separation of the analytes is enabled using liquid chromatography coupled with tandem mass spectrometry. Thus, in less than 30 min, 11 postulated metabolites of TEHTM can be selectively and sensitively quantified. The method is distinguished by its wide linear working range of up to 1800 μg/L with detection limits ranging from 0.3 μg/L (for 5oxo-1-MEHTM) to 1.5 μg/L (for 1-MEHTM). Precision and repeatability of the method were proven with determined relative standard deviations between 2.5 and 11.3%. The selection of the analytes of this method was based on a pilot study, by which several potential TEHTM metabolites were investigated in human urine of an orally exposed volunteer. Thus, the here presented method is a perfect tool for human biomonitoring of TEHTM exposure. Graphical abstract Analysis of several postulated TEHTM metabolites in human urine using LC-MS/MS.

New SPE-LC-MS/MS method for the simultaneous determination in urine of 22 metabolites of DEHP and alternative plasticizers from PVC medical devices

“…To date, few biomonitoring methods for the quantification of TEHTM metabolites are available. There are two methods for blood [25,38] and one recently published method by Höllerer et al for urine [22]. In the latter, the quantification method was developed for the determination of both specific primary metabolites of TEHTM and selected side chain oxidized monoester metabolites present in human urine.…”

“…Two methods have also been used to identified MEHA in urine [19,20]. More recently, the analysis of DEHTP [20,21] and TOTM [22] metabolites in urine has been described in study reports. Until now, no analytical method has been published for the simultaneous analysis of DEHP, DINP, DEHTP, DEHA, DINCH and TOTM metabolites.…”

Simultaneous and sensitive determination of the main metabolites of the plasticizer DEHP and its substitutes DEHTP, DINCH and TEHTM in human urine by coupling of on-line SPE, UHPLC and tandem mass spectrometry

“…Other analytical methods, that include urinary TEHTM biomarkers usually enable the determination of primary monoester metabolites (1-MEHTM, 2-MEHTM) 40 or even of the diester metabolites only (1,4-DEHTM, 2,4-DEHTM) 55 that were previously shown to be of minor importance as urinary biomarkers of TEHTM. 34,38 In comparison to our previous analytical method for the determination of TEHTM metabolites in human urine, 39 the excellent sensitivity as well as a good precision, and accuracy could be maintained even with the signicantly enlarged analyte spectrum.…”

“…32,33 For TEHTM, the main urinary metabolites in humans were observed to be the primary hydrolysis products 1mono-(2-ethylhexyl) trimellitate (1-MEHTM) and 2-mono-(2-ethylhexyl) trimellitate (2-MEHTM) as well as their respective sidechain oxidized species as secondary metabolites. 34,35 Several validated methods for the determination of plasticizer metabolites are already available, however, these methods usually enable the analysis of urinary metabolites of only one plasticizer (DEHP; 36 DINCH; 30 DEHTP; 37 TEHTM 38,39 ), predominantly using LC-MS/MS as the analytical technique. Up to now, few LC-MS/MS methods enable the determination of multiple plasticizer metabolites.…”

“…40 Recent investigations showed that the secondary TEHTM metabolites are signicant biomarkers of exposure to TEHTM. 38,39 Been et al (2019) 7 presented a LC-MS/ MS method using off-line solid phase extraction (SPE) for the determination of di(2-propylheptyl) phthalate, DINCH, DEHTP and DEHA metabolites in human urine and serum. But, the still relevant plasticizer DEHP and its important alternative TEHTM are not covered by that method.…”