Enzymatic biosensor for the electrochemical detection of 2,4-dinitrotoluene biodegradation derivatives

In this work, we demonstrate for the first time that 4-methyl-5-nitrocatechol (4M5NC) and 2,4,5-trihydroxytoluene (2,4,5-THT), two compounds obtained from the 2,4-DNT biodegradation are recognized by polyphenol oxidase as substrates. An amperometric biosensor is described for detecting these compounds and for evaluating the efficiency of the 2,4-DNT conversion into 4M5NC in the presence of bacteria able to produce the 2,4-DNT-biotransformation. The biosensor format involves the immobilization of polyphenol oxidase into a composite matrix made of glassy carbon microspheres and mineral oil. The biosensor demonstrated to be highly sensitive for the quantification of 4M5NC and 2,4,5-THT. The analytical parameters for 4M5NC are the following: sensitivity of (7.5 ± 0.1) × 105 nAM−1, linear range between 1.0 × 10−5 and 8.4 × 10−5 M, and detection limit of 4.7 × 10−6 M. The sensitivity for the determination of 2,4,5-THT is (6.2 ± 0.6) × 106 nAM−1, with a linear range between 1.0 × 10−6 and 5.8 × 10−6 M, and a detection limit of 2.0 × 10−7. Under the experimental conditions, it was possible to selectively quantify 4M5NC even in the presence of a large excess of 2,4-DNT. The suitability of the biosensor for detecting the efficiency of 2,4-DNT biotransformation into 4M5NC is demonstrated and compared with HPLC-spectrophotometric detection, with very good correlation. This biosensor holds great promise for decentralized environmental testing of 2,4-DNT.