Living cells contain various types of organic cations that may interact with nucleic acids.In order to understand thenucleic acid-binding properties oforganic cations of different sizes, we investigated the ability ofsimple organic cations to inhibit the RNA phosphodiester bond cleavage promoted by Mg2+, Pb2+, and RNA-cleaving serum proteins. Kinetic analysis using chimeric DNA-RNA oligonucleotides showed that the cleavage at ribonucleotide sites was inhibited in the presence of monovalent cations comprising alkyl chains or benzene rings. The comparison of thecleavage rates in the presence ofquaternary ammonium and phosphonium ionsindicated thatthe steric hindrance effect of organic cations on their binding tothe RNA backboneis significant when the cation size is larger than the phosphate-phosphate distance of a single-stranded nucleic acid.The cleavage inhibition was also observed for ribonucleotides located in long loops but not in short loopsof oligonucleotide structures, indicating less efficient binding of bulky cationsto structurally constrained regions.Theseresults revealtheunique nucleic acid-binding properties of bulky cations distinct from those of metal ions.