Determination of monobromobimane derivatives of phenylmercapturic and benzylmercapturic acids in urine by high-performance liquid chromatography and fluorimetry
M. Buratti et al., Determination of monobromobimane derivatives of phenylmercapturic and benzylmercapturic acids in urine by high-performance liquid chromatography and fluorimetry, J CHROMAT B, 751(2), 2001, pp. 305-313
A method was developed for the determination in human urine of S-phenylmerc
apturic (PMA) and S-benzylmercapturic (BMA) acids, metabolites respectively
of benzene and toluene. PMA and BMA were determined, after alkaline hydrol
ysis, to give respectively thiophenol and benzylmercaptan, and coupling of
the thiol-containing compounds with monobromobimane (MB), by reversed-phase
HPLC on a diphenyl-silica bonded cartridge (100x4.6 mm I.D., 5 mum particl
e size) with fluorimetric detection. Wavelengths for excitation and emissio
n were 375 and 480 nm, respectively. The recovery of PMA and BMA from spike
d urines was >90% in the 10-500 mug/l range; the quantification limits were
respectively 1 and 0.5 mug/l; day-to-day precision at 42 mug/l was C.V. <7
%. The suitability of the proposed procedure for the biological monitoring
of exposure to low-level airborne concentrations of benzene and toluene, wa
s evaluated by analyzing the urinary excretion of PMA and BMA in subjects e
xposed to different sources of aromatic hydrocarbons, namely occupationally
-unexposed referents (non-smokers, n=15; moderate smokers, n=8; mean number
of cigarettes smoked per-day=17 cig/day) and non-smoker workers occupation
ally exposed to toluene in maintenance operations of rotogravure machines (
non-smokers, n=17). Among referents, non-smokers showed values of PMA rangi
ng from <1 to 4.6 mug/l and BMA from 1.0 to 10.4 mug/l; in smokers, PMA val
ues ranging from 1.2 to 6.7 mug/l and BMA from 9.3 to 39.9 mug/l, were obse
rved. In occupationally exposed non-smoker subjects, BMA median excretion v
alue (23.6 mug/l) was higher than in non-smoker referents (3.5 mug/l) (P<0.
001) and individual BMA values (y, <mu>g/l) were associated and increased w
ith airborne toluene concentration Gs, mg/m(3)) according to the equation y
=6.5+0.65x (r=0.69, P<0.01, n=17). The proposed analytical method appears t
o be a sensitive and specific tool for biological monitoring of low-level e
xposure to benzene and toluene mixtures in occupational and environmental t
oxicology laboratory. (C) 2001 Elsevier Science B.V. All rights reserved.