Abstract: Benzene is an important industrial solvent and a ubiquitous environmental pollutant. Urinary
S-phenyl mercapturic acid (SPMA) and trans, trans - muconic acid (t,t-MA) are specific and sensitive biomarkers
for the determination of low levels of benzene exposure. This paper describes a specific and sensitive LC-MS/
MS method for simultaneous determination of t,t-MA and SPMA from human urine samples using dual columns
of two different internal diameters to reduce the matrix effect on ionization. This method was applied for the
measurements of t,t-MA and SPMA from the random urine samples of exposed subjects of the footwear industry
and comparable control subjects. The recoveries (Mean ± SD) of spiked standards of t,t-MA and SPMA in urine
were 95.4 ± 12.3% (n = 6) ranging from 79.4 - 114% and 69.7 ± 9.5% (n=6) ranging from 60.1 - 89% respectively.
The Limit of Detection (LOD) was found to be 1 ng/ml and 0.03 ng/ml and the Limit of Quantitation (LOQ) was
found to be 5 ng/ml and 0.1 ng/ml for t,t-MA and SPMA at the S/N ratio of 3 and 10 respectively. The observed
values of t, t-MA and SPMA in the exposed subjects of footwear industry were below the values of biological
exposure indices (BEI) as described by ACGIH for benzene exposure.