Three distinct N-malonyltransferases were purified from peanut seedlings, accepting either anthranilic acid, D-tryptophan, or 3,4-dichloroaniline, respectively, as a substrate. Partially purified malonyl-CoA:D-tryptophan malonyltransferase also catalyzed the formation of the corresponding malonic acid conjugate when 1-aminocyclopropane-1-carboxylic acid was employed as a substrate. These N-malonyltransferases were clearly distinguished from several O-malonyltransferase activities also present in the same seedlings. N-Malonic acid conjugates have been previously isolated from peanut either as a natural constituent or after feeding with xenobiotics. By analogy to the results reported with cultured parsley cells, multiple malonyltransferases in peanut may have a role in vacuolar transport. Crude extracts of young peanut seedlings were incapable of hydrolyzing the respective N-malonic acid conjugates. However, dialyzed extracts of older plants released malonic acid from malonyl-1-aminocyclopropane-1-carboxylic acid but not from malonyl-3,4-dichloroaniline, suggesting that some N-malonic acid conjugates may be metabolized in plants which are approaching senescence.