06 May 2010
DEVELOPING DETECTION METHOD FOR PESTICIDES

Paraquat is one of most widely used herbicides in the world and highly toxic to humans. Since July 2007 the use of this pesticide in the EU is banned, which does not affect that farmers outside the EU continue to use paraquat and export their products to the EU. Therefore, there it remains being necessary to control that paraquat residues are below the EU maximum residue limits (MRL) .  

Evaluation of the matrix effect on the immunochemical determination of PQ

 The nonspecific interferences caused by the cereals and potato extracts in the PQ microplate-based ELISA has been investigated. The results have shown that this ELISA has an extraordinary tolerance to these matrices and to the presence of methanol. Initial studies performed to know the resistance of the PQ ELISA to organic solvents, already showed that measurements could be made in samples with up to 50% of methanol. Moreover, cereals (barley and wheat) and potato extracts obtained using  a very simple procedure developed CVUA and slightly modified to produce extracts compatible with the immunochemical procedure,  can be directly analyzed with the PQ microplate-based ELISA without further sample treatments. The extraction procedure involves the use of aqueous MeOH-HCl (0.1M) for few minutes at room temperature, centrifugation, filtering of the supernatant and buffering the extract to adjust pH and conductivity before the immunochemical measurements. Using this procedure, the final extracts obtained contain 25% MeOH and can be analyzed directly by the PQ-microplate ELISA. Thus, as it can be observed, calibration curves build using these extracts were almost identical to the standard curve prepared in the assay buffer. Moreover, further dilutions of the matrix with the assay buffer did not affect the calibration of the assay. Only the extracts prepared in barley show a slight matrix effect that can be removed by dilution of the matrix. Since the limit of detection is very good, this does not compromise performance of the assay in compliance with the EC regulations regarding the possibility to measure PQ below the maximum residue limits established for PQ.

Development of an immunosensor for PQ

Two new electrochemical immunosensors configurations for PQ have been developed. The first immunosensor was developed, based on the previous knowledge of the AMRg-CSIC group, using antigen derivatized magnetic particles and HRP (horseradish peroxidise) labelled PQ specific antibodies, that catalyzed the oxidation of the substrate producing an amperometric current. The immunosensor was able to reach a limit of detection of about 0.5 ug L^-1. However, since the detectability was lower than that reached by the PQ microplate ELISA using the same immunoreagents, it was decided to investigate alternative strategies. Thus, the new immunosensor configuration established uses specific antibodies labelled with metal nanoprobes. In this case, the signal recoded is produced by the reduction of the metal in the electrode. The detectability reached using this second immunosensor configuration was very good (around 0.07 ug L^-1); moreover, this second strategy offers the possibility of multiplexation, which may be very useful for further developments for rapid multi-residue analysis 

 

Information supplied by Hans Mol, RIKILT, NL, WP1c Leader and Pilar Marco, CSIC, Sp, WP1c Deputy
Source : CONffIDENCE
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