Born at Arezzo in Italy, Cesalpino studied anatomy and medicine at the University of Pisa, where he graduated in 1551; in 1555 he became professor of medicine and director of the botanic garden there. In 1592 he became physician to Pope Clement VIII and professor at the Sapienza University.

He is most famous for his plant classification based on fruit and seed characteristics, which is described in his work, De plantis (1583; On Plants). This work also discusses the whole of theoretical botany and had great influence on later botanists.

Cesalpino also wrote a number of anatomical books in which he partly anticipated the theory of the circulation of the blood proposed by William Harvey.