SPA enhanced FPIA-based detection of pesticide residue with ppb/ppt level detection limit.

Pesticide residue in fruits & vegetables is one of the key issues affecting the export of rural products in India. Pesticide exposure or intake causes major nervous system problems in children. The solutions to quantitate them in field are rare and the pesticide residue detection in the parts per billion (ppb) ranges is challenging. Except ELISA, none of the existing methods can detect pesticide residues in ppb range in the field. We employed a new approach of concentrating field samples and used sodium polyacrylate (SPA) as water absorbing material. The SPA beads concentrate the field samples and obtained a sub ppb range detection using an existing FPIA system and could improve overall sensitivity by 10-100 fold. The developed assay can be done in few seconds. We have used three pesticides 2,4-D, atrazine and methyl parathion with 0.1, 0.5 and 3 ppb detection limit respectively. We developed a simple field ready FPIA device and used sodium poly acrylate (SPA) in this biochemical FPIA to enhance sensitivity. Our tests with spiked field samples offers a possibility of using SPA concentration assisted FPIA in field. This study will have far reaching applications of both qualitative & quantitative analysis chemical analytes in field samples.

Hydrophobic interactions in donor-disulphide-acceptor (DSSA) probes looking beyond fluorescence resonance energy transfer theory.

Donor-linker-acceptor (DSSA) is a concept in fluorescence chemistry with acceptor being a fluorescent compound (FRET) or quencher. The DSSA probes used to measure thiol levels in vitro and in vivo. The reduction potential of these dyes are in the range of -0.60 V, much lower than the best thiol reductant reported in literature, the DTT (-0.33 V). DSSA disulphide having an unusually low reduction potential compared to the typical thiol reductants is a puzzle. Secondly, DSSA probes have a cyclized rhodamine ring as acceptor which does not have any spectral overlap with fluorescein, but quenches its absorbance and fluorescence. To understand the structural features of DSSA probes, we have synthesized DSSANa and DSSAOr. The calculated reduction potential of these dyes suggest that DSSA probes have an alternate mechanism from the FRET based quenching, namely hydrophobic interaction or dye to dye quenching. The standard reduction potential change with increasing complexity and steric hindrance of the molecule is small, suggesting that ultra- low Eo' has no contribution from the disulphide linker and is based on structural interactions between fluorescein and cyclized rhodamine. Our results help to understand the DSSA probe quenching mechanism and provide ways to design fluorescent probes.