Complementary analysis of mercapturic acids in urine samples

Mercapturic acids background

Mercapturic acids are conjugation products from N-acetyl-L-cysteine that are formed from a wide range of endogenous and exogenous chemicals. The conjugation takes place in the liver and these compounds are subsequently excreted in urine. Many of these marcapturic acids are built from toxicants as a consequence of exposure to mainstream smoke, including:

  • Acrolein (3-HPMA)
  • Acrylamide (AAMA, GAMA)
  • Acrylonitrile (CEMA, HEMA)
  • Benzene (SPMA)
  • 1,3-Butadiene (MHBMA, DHBMA)
  • Crotonaldehyde (HMPMA, CMEMA)
  • N,N-Dimethylformamide (AMCC)
  • Propylene oxide (2-HPMA)
  • Styrene (PHEMA)
  • Toluene (SBMA)
  • Vinyl chloride (HEMA)
  • Methylating / Ethylating agents (MMA, EMA)

Therefore, urinary levels of mercapturic acids can be used as biomarkers of exposure (BoE) to thesse toxicants.

Mercapturic acids analysis by LC-MS/MS

Up to now 4 different methods for the quantification of numerous mercapturic acids in urine samples were available at ABF.

For optimization and acceleration of the determination of mercapturic acids within clinical studies, the 4 methods used at ABF were multiplexed in only 2 different methods. These methods have been succesfully validated according to FDA guidlines and showed excellent results in terms of sensitivity, reproduciubility and robustness.We were able to speed up the analytical workflow,and 6 new analytes were succesfully included in the methods (see table below).

Mercapturic acids covered by the newly developed methods are listed in the following table.

Mercapturic acids (precursors) in the new Methods I and II

Method I

Method II

SPMA (Benzene)

3-HPMA (Acrolein)

SBMA (Toluene)

2-HPMA (Propylene oxide) 1

PHEMA-1 (Styrene)1

AAMA (Acrylamide)

PHEMA-2 (Styrene)1

GAMA (Acrylamide)

CEMA (Acrylonitrile; minor: acrolein)

MHBMA-1 (1,3-Butadiene)

HMPMA (Crotonaldehyde)

MHBMA-2 (1,3-Butadiene)

CMEMA (Crotonaldehyde)

DHBMA (1,3-Butadiene)

EMA (Ethylating agents) 1

AMCC (N,N-Dimethylformamide)1

HEMA (Ethylene oxide, ethylene; minor: acrylonitrile, vinyl chloride)


MMA (Methylating agents) 1


1 Newly included analytes.

The method was published in Analytical & Bioanalytical Chemistry 2015 (DOI: 10.1007/s00216-015-8719-x)

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Max Scherer

May 15th 2015