Jacopo Mazzoni was born in Cesena on November 27, 1548, studied literature in Bologna and philosophy in Padua. According to his contemporaries he had a prodigious memory which enabled him to amass a vast erudition.

He was a member of the Academy of Crusca and the Florentine Academy. Gregory XIII (1572-1585) wanted Mazzoni to come to work for him and tried pressuring him into taking religious vows, but Mazzoni preferred the professor's gown to the churchman's robes. He became a university professor, first at Macerata and then at Pisa, where he taught philosophy from November 1588 to the end of the 1596-97 academic year, and in his first period there made the acquaintance of Galileo, who was then a young mathematics instructor. They started up a cordial friendship, and on May 30, 1597 Galileo wrote Mazzoni a letter in which, besides commenting on the latter's book In universam Platonis et Aristotelis philosophiam praeludia (Venice, 1597), stated his inclination to the Copernican system over the Ptolemaic one.

Jacopo Mazzoni was called to the University La Sapienza in Rome in 1597, but shortly after accepting the chair had to accompany Cardinal Aldobrandini to Ferrara, and from there to proceed on a mission to Venice. On his return he took ill and had himself removed to Cesena, where he died on April 10, 1598.