A method employing immunoaffinity chromatography for sample clean-up has been developed for a number of triazine herbicides. The approach eliminates: the requirement for solvents such as hexane, dichloromethane, acetone and other solvents commonly used for organic-aqueous partitions or adsorption chromatographic clean-up of sample extracts. The only solvent used was methanol, (the least toxic, most environmentally friendly and least expensive of those mentioned) for the initial sample extraction and clean-up. Acetonitrile was used in the liquid chromatographic mobile phase, although methanol proved to be satisfactory for all triazines except for atrazine which coeluted with simetryn and simazine which coeluted with metribuzin. An immunoaffinity cartridge employing immobilized polyclonal antibodies generated against atrazine but which cross-reacted with several other triazines, was used for the clean-up of extracts of apple, carrot, celery, corn, potato and peas. The triazines atrazine, cyanazine, simazine, propazine, terbutylazine, simetryn and prometone were studied. The method involved extraction of the triazines from the plant material with methanol followed by concentration of an aliquot of the extract with subsequent dilution in aqueous phosphate buffered saline (PBS). The PBS mixture was cleaned up sequentially using a strong anion-exchange solid phase extraction cartridge (SPE-SAX) followed by the immunoaffinity cartridge. Quantitation was carried out using reversed-phase liquid chromatography with UV absorption detection at 220 nm. Atrazine, simazine, propazine and terbutylazine were recovered from extracts consistently above 80% at a spiking level of 25 ng/g each. The average recovery for cyanazine was 71% (n=9), for prometone it was 72% (n=7) and for simetryn it was 65% (n=9). Detection limits were estimated to be 2-10 ng/g depending upon the triazine and the plant tissue analyzed. One immunoaffinity cartridge was used for 49 plant extracts with no loss of activity or effectiveness for extract clean-up. A combination of two immunoaffinity cartridges (one not recognizing the triazines and one recognizing them) was evaluated for the selective clean-up of several of the plant extracts. This approach compared well with the SPE-SAX-immunoaffinity cartridge combination. Spiked (25 ng/g of each triazine) extracts of celery and corn were readily confirmed by GC-mass spectrometry.
