IMMUNOCHEMICAL EVIDENCE SUPPORTING 2-PENTYLPYRROLE FORMATION ON PROTEINS EXPOSED TO 4-HYDROXY-2-NONENAL

Previous model studies suggested the formation of lysine-based 2-penty lpyrroles as novel late adduction products formed upon exposure of pro teins to the lipid peroxidation product 4-hydroxy-2-nonenal (HNE). Two 8-pentylpyrrole immunogens were prepared, one by treating keyhole lim pet hemocyanin (KLH) directly with 4-oxononanal and the other by prefo rmation of 6-(2-pentylpyrrol-1-yl)hexanoic acid from 6-aminocaproic ac id and 4-oxononanal, followed by carbodiimide coupling to KLH. Pyrrole content-and lysine modification in KLH were assayed independently. Fo llowing immunization of rabbits, antibody titer increased and plateaue d over a 4 month period. The structural specificity of the IgG fractio ns of the antisera was evaluated through comprehensive competitive ELI SA studies. These antibodies were used to verify the time-dependent ap pearance of the 2-pentylpyrrole epitope in protein exposed to HNE. Pot ential advantages of antibodies recognizing ''advanced lipid peroxidat ion end products'' over those recognizing ''early'' HNE adduction prod ucts are discussed.