Sensitive biomonitoring of phthalate metabolites in human urine using packed capillary column switching liquid chromatography coupled to electrospray ionization ion-trap mass spectrometry

Anal Bioanal Chem. 2004 Apr;378(7):1762-8. doi: 10.1007/s00216-003-2488-7. Epub 2004 Jan 31.

Abstract

A rapid and sensitive method for the determination of the phthalate monoesters monoethyl phthalate (MEP), monobutyl phthalate (MBP), monobenzyl phthalate (MBzP) and monoethylhexyl phthalate (MEHP), in human urine, using packed capillary column liquid chromatography coupled to electrospray quadrupole-ion trap mass spectrometry (ESI-QITMS(n)) has been developed. Sample volumes of 200 microL of deconjugated and diluted urine were loaded onto a precolumn of 30 mmx0.32 mm I.D. packed with Hypercarb 5 microm particles, using a sample carrier consisting of acetonitrile/water (15/85, v/v, adjusted to pH 2 using HCl) with a flow rate of 20 microL/min. Backflushed elution onto a 100 mmx0.32 mm I.D. analytical column packed with 5 microm Hypercarb particles was conducted using a tetrahydrofuran/water gradient where both solvents contained 10 mM ammonium acetate, at a flow rate of 4 microL/min. Determination of the monophthalates was achieved within 8 min. Ionization was performed in the negative mode and the analytes were observed as [M-H](-) at m/ z=193.1, 221.1, 255.1 and 277.0 for MEP, MBP, MBzP and MEHP, respectively. Quantification was performed in the multiple reaction monitoring (MRM) mode monitoring the fragments at m/ z=121.1, 177.0, 183.0 and 233.0 for MEP, MBP, MBzP and MEHP, respectively. The method was validated over the concentration range 2.5-125 ng/mL in pretreated urine samples, corresponding to 25-1250 ng/mL untreated urine, yielding correlation coefficients in the range 0.996-0.999. The within-assay ( n=6) and between-assay ( n=6) repeatabilities were in the range 4.0-18% and 4.8-15% RSD, respectively. The mass limits of detection were in the range 32-70 pg, corresponding to concentration limits of detection of 1.6-3.5 ng/mL of untreated urine.

MeSH terms

  • Chromatography, Liquid / methods*
  • Humans
  • Phthalic Acids / urine*
  • Reproducibility of Results
  • Sensitivity and Specificity
  • Spectrometry, Mass, Electrospray Ionization / methods*

Substances

  • Phthalic Acids