Enzymatic Reaction Coupled with Flow-Injection Analysis with Charged Aerosol, Coulometric, or Amperometric Detection for Estimation of Contamination of the Environment by Pesticides
Mikelova, R., Prokop, Z., Stejskal, K., Adam, V., Beklova, M., Trnkova, L., Kulichova, B., Horna, A., Chaloupkova, R., Damborsky, J., Kizek, R.
CHROMATOGRAPHIA 67: S47-S53 (2008)
Because excessive using of pesticides poses a threat to the environment and to human health, development of low-cost and sensitive methods for analysis of pesticides in the environment is needed. Several bacteria can release halide ions from the molecules of halogenated hydrocarbons. This can be used in a device for analysis of halogenated hydrocarbons in the environment by quantification of the halide anions. Here we directed our attention to selecting an instrument for detection of chloride anions. We tested three different detectors, amperometric, and coulometric, both coupled with flow-injection analysis and charged aerosol, coupled with high performance liquid chromatography. Detection limits (3 × S/N) for measurement of chloride anions by use of these detectors was 30 µM (charged aerosol), 100 nM (coulometric), and 1 nM (amperometric). Because of its lowest detection limit for chloride anions and the many technical possibilities of miniaturization, the amperometric detector was used to test of effect of different cations on the chloride signal under the optimized experimental conditions (working electrode potential -365 mV; “Current R” 5 µA; mobile phase 0.2 M phosphate buffer, pH 6; flow rate 0.5 mL min-1). NaCl, SrCl2, NH4Cl, and CsCl were tested as sources of chloride anions. We then used the detector to detect chloride anions catalytically cleaved from 1-chlorohexane by the enzyme haloalkane dehalogenase LinB from the bacterium Sphingobium japonicum UT26. The activity of the enzyme increased with increasing reaction temperature until the maximum was observed at 39°C. The results obtained were in good agreement with data obtained by colorimetric detection.