A polyprenyltransferase catalysing the formation of Z-double bonds was partially purified from mulberry leaves, Morus bombysis. The enzyme catalysed a consecutive condensation of isopentenyl diphosphate with geranyl- geranyl diphosphate as an allylic primer to produce a series of ficaprenol-type Z, E-mixed polyprenyl diphosphates with carbon chain length ranging from C 40 to C 60. Not only all-E-geranylgeranyl diphosphate but also E, E-farnesyl- and geranyl diphosphates were accepted as substrates. Addition of Triton X-100 stimulated the enzymatic activity. The stereochemistry of hydrogen elimination from the 2-position of isopentenyl diphosphate during the Z-prenyl chain formation was examined directly by experiments using this synthase and ( S)-[1- 14C, 2- 3H]isopentenyl diphosphate and it was demonstrated that the 2-pro- S hydrogen was lost. Feeding experiments of stereospecifically 3H-labelled mevalonic acid with intact mulberry leaves, however, showed that mevalonic acid was incorporated into polyprenols with elimination of the 4-pro- S hydrogen of the acid.