1. 1. Crystalline ribonuclease samples obtained from different sources as well as a sample prepared from calf pancreas in the laboratory were separated by column chromatography. 2. 2. Ribonuclease B of each of the four crystalline ribonucleases liberates guanylic acid from ribonucleic acid (RNA), and it catalyzes synthesis of dinucleoside phosphates from cyclic cytidylic, uridylic, and guanylic acids, but not from cyclic adenylic acid. Ribonuclease B fraction, similar to the ribonuclease A fraction and to the crystalline ribonuclease, hydrolyzes ribonucleic acid, liberating pyrimidine mononucleotides. It also hydrolyzes cyclic guanylic acid to guanosine 3′-phosphate, whereas ribonuclease A does not. 3. 3. The dinucleoside phosphates formed by either ribonuclease A or B are resistant to hydrolysis by ribonuclease B but not by A, those containing cytidine being more resistant to ribonuclease A than those containing uridine. 4. 4. Ribonuclease A hydrolyzes pyrimidine cyclic nucleotides six to ten times faster than does ribonuclease B. 5. 5. It is thus demonstrated that certain of the activities of “crystalline ribonuclease” reside in different protein entities and that some activity toward purine nucleotide esters exists.