The main focus of his research in plant secondary metabolites, mainly biosynthesis of lignin and alkaloids (pilocarpine and caffeine). His lab has been working on the understanding of the biosynthesis of lignin and other cell wall components in sugarcane and eucalyptus; mechanisms underlying the tolerance of Eucalyptus globulus to low temperature; the elucidation of the biosynthesis route of the alkaloid pilocarpine in Pilocarpus microphyllus (Jaborandi); and the control of caffeine biosynthesis in coffee. Mazzafera has over 150 scientific papers in international journals. His research on caffeine-free coffee plants is world known since the discovery of natural decaffeinated plants and later the production of mutants practically absent of caffeine. Mazzafera was a member of the Committee of Botany at CNPq, Assistant of the Dean of Unicamp, head of Department and Director of The Institute of Biology. He is also a member of the Agriculture and Veterinary coordination of FAPESP.