Aqueous chlorination of humic acids results in the formation of compounds with direct-acting mutagenic activity in the Ames/Salmonella plate assay for tester strains TA98, TA100, TA1535, TA1537, and TA1538. The addition of a rat liver microsomal fraction (S-9) plus cofactors causes a substantial decrease of activity, the extent of which is tester strain dependent. The mutagenic activity is due predominantly to non-volatile compounds. Production of mutagens is greatly dependent on the chlorination pH, with a pattern of decreasing mutagenic activity with increasing pH. This order of activity could be at least partially explained by the alkali-liability of the compounds. Chlorination of commercial humic acids is proposed as a model for examination of mutagen formation during water chlorination.