Analysis of benzylsuccinates in groundwater by liquid chromatography/tandem mass spectrometry and its use for monitoring in situ BTEX biodegradation

Benzylsuccinic acid (BS) and methylbenzylsuccinic acid (MeBS) isomers have been proposed as distinctive indicators of anaerobic toluene and xylene metabolism in fuel-contaminated aquifers; however, labor-intensive analytical procedures have limited their analysis at field sites. In this article, a rapid electrospray LC/MS/MS (liquid chromatography/mass spectrometry/mass spectrometry) method for benzylsuccinates is described that involves selected reaction monitoring, internal standard quantification with [ring-H-2(5)]BS, small sample size (<1 mL), and no extraction/concentration steps. The highly selective LC/MS/MS method was shown to be sensitive (detection limits ca. 0.3 mug/L), accurate, and precise. The method was used to characterize geographic and temporal distributions of BS and MeBS isomers in an anaerobic, hydrocarbon-contaminated aquifer. BS was never detected, and MeBS isomers were detected in the three wells with the highest concentrations of BTEX (benzene, toluene, ethylbenzene, and xylenes); MeBS concentrations ranged from <0.3 to 205 mug/L. A strong linear correlation (r(2) = 0.94; n = 12) was found between concentrations of total MeBS isomers and their parent compounds, xylenes, which suggests that anaerobic xylene metabolism at this site was probably first-order rather than zero-order. The novel LC/MS/MS method for BS and MeBS isomers is a promising technique for rapid and reliable monitoring of in situ bioremediation of gasoline-contaminated groundwater.