Development of an enzyme-linked immunosorbent assay for hexazinone and its application to water

• Main Authors: Bushway, R.J. (University of Maine, Orono, ME.), Fan, T.S, Katz, L.E, Reed, A.W, Ferguson, B.S, Xu, C.Y, Perkins, L.B, Young, B.E
• Format: Book
• Language: English
• Published: Washington, DC (USA) American Chemical Society 1996
• Subjects: AGUA; EAU; HEXAZINONE
• ISBN: 9780841233799; 0841233799

Polyclonal antibodies to hexazinone were raised in rabbits using a combination of intradermal and subcutaneous injections. An appropriate hexazinone derivative was prepared by refluxing the metabolite 3-(4-hydroxycyclohexyl)- 6-(dimethylamino)-1-methyl-1,3,5-triazine-2, 4-(1H, 3H)-dione and succinic anhydride in pyridine at 100 degrees C for 6 h. The resulting hexazinone hemisuccinate was converted to an active ester by reacting it with N-hydroxysuccinate and 1-ethyl-3-(dimethyl-aminopropyl)carbodiimide.HCl (EDC) in dimethylformamide (DMF) overnight at 20 degrees C. The immunogen was prepared by conjugating the active ester to bovine serum albumin (BSA); the enzyme tracer consisted of the derivative conjugated to horse radish peroxidase (HRP). The immunoassay was performed employing two formats, plates and tubes, and both were precoated with rabbit anti-hexazinone. Thirty-two groundwater samples analyzed for hexazinone using a plate enzyme immunoassay (EIA) correlated well with high-performance liquid chromatography (HPLC) (correlation coefficient of 0.9468). A correlation coefficient of 0.9563 was calculated for 30 water samples run by tube EIA vs. HPLC. Of the eight known hexazinone metabolites, seven were tested for cross-reactivity and five were crossreactive