The general objective of this research project is the elucidation of the specificity and mechanism of action of the cyclase enzymes involved in the biosynthesis of the diterpenes kaurene, beyerene, sandaracopimaradiene, trachylobane, and casbene and of the oxidase enzymes which convert kaurene to kauren-19-oic acid. Thus, the stereochemistry of the cyclizations at the prochiral positions is being determined by use of substrates bearing deuterium, tritium or carbon-13 tracers. The complete stereochemistry of the intramolecular cyclopropanation reaction by which the phytoalexin casbene is produced by a soluble enzyme extract from Ricinus communis is being established by a combination of labelling experiments, degradation, and spectroscopy. The stereochemical course of the two CH3 yields CH2 elimination steps in the biosynthesis of kaurene, the precursor to the giberellin family of plant growth regulating substances, is to be clarified by use of mevalonic acid bearing a chiral methyl group. The substrate specificity of the various diterpene cyclases is being explored by experiments with analogues of geranylgeranyl and copalyl pyrophosphates. The stereochemistry and isotope effects associated with the oxidation of the axial C-4 methyl group of kaurene through the sequence CH3 yields CH2OH yields CHO yields CO2H is being studied by labelling methods and spectroscopic assays. The results will afford insight into the mechanism of action of the oxidases involved in the biosynthesis of terpenes and sterols.