Chemical Reduction of Zoalene to ANOT for Use in Zoalene Residue Studies

A novel chemical synthesis and purification scheme to produce ANOT (3-amino-5-nitro-o-toluamide) was developed to yield an analytical standard grade material for zoalene residue investigations. An original conversion of zoalene (3,5-dinitro-o-toluamide) to ANOT was developed. Several literature references were available for the partial reduction of dinitro aryl compounds, but none detailed the conversion of zoalene to ANOT. A chemical synthesis was chosen over a biological synthesis due to the latter's complex reaction matrix, difficult recovery, and low batch yields. Using general partial reduction of dinitro aryl schemes, it was found that 10% palladium on carbon coupled to a cyclohexene reducing agent provided an optimum ANOT yield. Chromatographic techniques were utilized to isolate ANOT from the product mixture and to purify it to a standard grade material (>99% pure). The chemically synthesized ANOT product was characterized using colorimetric, TLC, FTIR, NMR, and LC/MS techniques. Keywords: ANOT; zoalene; zoaline residue; chemical reduction