Prelog wanted to work in an academic environment, so he accepted the position of lecturer at the University of Zagreb in 1935. At the Technical Faculty in Zagreb, he lectured on organic chemistry and chemical engineering.
With the help of collaborators and students, Prelog started researching quinine and its related compounds. He was financially supported by the pharmaceutical factory "Kaštel", currently Pliva. He developed a financially successful method of producing Streptazol, one of the first commercial sulfonamides. In 1941, while at Zagreb, Prelog developed the first synthesis of adamantane, a hydrocarbon with an unusual structure that was isolated from Moravian oil fields.