Determination of veterinary drugs in dry milk powder by supercritical fluid extraction - Enzyme-linked immunosorbent assay

The objective of this study was to establish whether supercritical fluid extraction (SFE) coupled with enzyme-linked immunosorbent assay (ELISA) is a viable technique for the determination of sulfamethazine in dry milk powder at concentrations as low as 2.5 ng/g. Extraction and analysis of organic compounds in various matrices by SFE-ELISA has been under evaluation in our laboratory since it results in greater sample throughput, allowing rapid screening of environmental samples. In this study, we demonstrated that sulfamethazine could be extracted quantitatively (average recovery at 5-ng/g spike was 91.7 percent) from spiked dry milk powder at 450 atm and 80 degrees C using supercritical carbon dioxide modified with 10 percent methanol as the extraction fluid. The extracted material was collected in methanol, concentrated to dryness, and redissolved in phosphate buffer for analysis by a competitive ELISA. The precision of the ELISA technique, as established over a period of 17 days, was 15 percent or better for sulfamethazine concentrations ranging from 5 to 15 ng/mL of extract in phosphate buffer.