An LC-MS/MS Method for a Comprehensive Determination of Metabolites of BTEX Anaerobic Degradation in Bacterial Cultures and Groundwater

Simon Godin,Anthony Ranchou-Peyruse,Joanna Szpunar,Guilhem Caumette,Pawel Kubica,Isabelle Le Hecho,Delphine Patriarche,Ryszard Lobinski

doi:10.3390/w12071869

Abstract

BTEX (benzene, toluene, ethylbenzene, and the different xylene isomers), known for carcinogenic and neurotoxic effects, are common environmental contaminants. The first step for the development of the bioremediation technologies is the detection of intense microbial degradation in contaminated waters in the quest for the most active bacterial strains. This requires the multispecies analysis for BTEX metabolites which are considered as markers of microbial degradation. A direct (50 µL injection) HPLC–electrospray MS/MS analytical method was developed for the simultaneous analysis of 11 BTEX metabolites (o-, m-, p-toluic, salicylic, benzoate, benzyl, and phenyl succinic acids, 2-(1-phenylethyl)-, 2-(2-methylbenzyl), and 2-(3-methylbenzyl)-, 2-(4-methyl benzyl)-succinic acids) in bacterial cultures and ground waters down to 0.1 ng/mL. The optimization of the chromatographic conditions allowed for the resolution of position isomers of toluic and methylbenzyl-succinic acids. The stability of the analytes during sample storage tested in different conditions showed the instability of some of them when stored at room temperature. The feasibility of the method was demonstrated by the detection of all the investigated metabolites in a water sample of a deep aquifer hosting natural gas storage. A model laboratory study emphasized the importance of 2-(2-methylbenzyl)-succinic acid as a marker of anaerobic microbial degradation.

Highlights

Aromatic hydrocarbons, such as benzene, toluene, ethylbenzene, and xylenes, are recognized as environmental pollutants co-occurring at hazardous waste sites
In the presence of bacteria, BTEX undergo natural degradation, which is accompanied by the production of different metabolites
The detection and quantification of the metabolites that are characteristic of in situ anaerobicbenzene transformation is a definitive way to demonstrate the occurrence of the self-bioremediation of a BTEX-impacted site; it is of paramount importance—along with the stable isotope fractionation analysis—for model laboratory studies aiming at elucidating the metabolic pathways [4]

Summary

Introduction

Aromatic hydrocarbons, such as benzene, toluene, ethylbenzene, and xylenes (aka BTEX), are recognized as environmental pollutants co-occurring at hazardous waste sites. Their chemical inertness, relatively high solubility in water, and toxicity make them common targets of the environmental monitoring [1,2]. Water 2020, 12, 1869 is an issue of wide interest Green technologies, such as using bacteria metabolizing BTEXs, are promising. The detection and quantification of the metabolites that are characteristic of in situ anaerobic (alkyl)benzene transformation is a definitive way to demonstrate the occurrence of the self-bioremediation of a BTEX-impacted site; it is of paramount importance—along with the stable isotope fractionation analysis—for model laboratory studies aiming at elucidating the metabolic pathways [4]. The understanding of the natural attenuation processes is a key to the development of efficient bioremediation and environmental engineering technologies [5]

Objectives

Methods

Results