In the study of the metabolic disposition of ivermectin in cattle, sheep, and rats, a group of nonpolar metabolites was detected in the fat tissues of these animals. Upon saponification or esterase treatment, these nonpolar metabolites gave rise to polar products that were similar to the ivermectin metabolites present in the liver. A hypothesis was thus proposed that the polar ivermectin metabolites produced in the liver were esterified and stored in the fat as nonpolar entities, which can be reconverted back to the polar metabolites chemically or enzymatically. To substantiate this hypothesis, the regeneration of polar metabolites from the nonpolar metabolites was studied and hydrolysis products were characterized to establish the basic structure of the alcohol portion of the metabolites. Furthermore, chromatographic comparisons were made between radiolabeled in vivo metabolites and synthetic fatty acid ester samples. These results established the general structural class of the ivermectin nonpolar metabolites and also confirmed the unusual metabolic pathway of ivermectin disposition in fat tissue.