The mutagenicity of ICR-170, a monofunctional nitrogen mustard derivative of acridine (2-methoxy-6-chloro-9-[3-(ethyl-2-chloroethyl) amino propylamino] acridine dihydrochloride), was compared with other specific mutagens, NA, EMS, and HA, to characterize the genetic alterations produced by ICR-170 at the molecular level. A comparison was also made of the mutagenicity of compounds closely related in structure to ICR-170 to determine which parts of ICR-170's molecular structure are important for their different mutagenic activities. Assays for specific revertibility were made on a series of 8 ad-3B mutants of Neurosopora crassa comprised of a tester set of 4 mutants that revert by base-pair substitutions, 2 that revert by base-pair insertion and/or deletion, and 2 that revert only spontaneously. In such tests ICR-170 produces predominantly base-pair insertions or deletions as well as a lower frequency of base-pair substitutions. Tests with the structurally related compounds showed that (1) Atabrine, which has a molecular structure similar to ICR-170 and lacks the nitrogen mustard group, is not mutagenic; (2) that ICR-174 and ICR-172, which have the same aliphatic side chain as ICR-170 but only the left two-thirds or the right two-thirds of the acridine ring, show a low nonspecific mutagenic activity; and (3) that ICR-177, a monofunctional nitrogen mustard with the same aliphatic side chain as ICR-170, and the common bifunctional nitrogen mustard (HN 2 ) induce reversions in all the mutants that revert by base-pair substitutions and in one of the mutants that reverts by base-pair insertion or deletion. The mutagenic activity of the compounds structurally related to ICR-170 has been interpretated as indicating that (1) the nitrogen mustard group is essential for the mutagenicity of ICR-170, (2) that theintact acridine ring is required to produce reversions due to base-pair insertion and/or deletion, and (3) that the lower frequency of reversions produced by ICR-170 in the mutants that revert by base-pair substitution can be attributed to the monofunctional nitrogen mustard group. In summary, both the acridine ring and the nitrogen mustard group of the ICR-170 molecule are essential for its mutagenic activity as well as for its antioplastic activity (as shown previously by Dr. H. J. Creech, Institute for Cancer Research, Philadelphia, Pa., (U.S.A.).