Use of a silylation procedure and C-13-NMR spectroscopy to characterize bound and sequestered residues of cyprodinil in soil

Soil-bound residues of the fungicide cyprodinil (4-cyclopropyl-6-methyl-2-phenylaminopyrimidine), which was labeled with C-13 and C-14 either in the phenyl ring or the pyrimidyl ring, were analyzed by a silylation procedure and C-13-NMR spectroscopy. After a 6-month incubation of soil with 3 or 500 ppm cyprodinil, bound residues amounted to about 50% and 18% of the initial radioactivity, respectively. The isolated humic acid fraction and the NaOH-extracted soil (the humin fraction) were suspended in chloroform and silylated by overnight shaking with trimethylchlorosilane. Analysis of the silylated extracts by C-13-NMR revealed that the formation of soil-bound residues in the 500 ppm samples involved: (1) sequestration of the unaltered or slightly altered fungicide in the humin fraction and (2) cleavage of the cyprodinil molecule between the aromatic rings followed by covalent binding of the separated moieties to humic acid. The sequestered fungicide (phenyl label) generated triplet NMR signals at 121.0, 125.0, 128.7, and 135.8 ppm, which closely coincided with the resonances shown by the silylated standard of cyprodinil. TLC analysis indicated that, in the 3-ppm samples, cleavage products were also subject to sequestration in humin. As determined by size-exclusion chromatography, the molecular size of the humic acid fraction with the incorporated cleavage products was in the range of 2 x 10(3) Da.