Fertilization of the sea urchin egg results in a transient decline in the amount of phosphatidylinositol (PI) to a level equal to about 50% of that present in the unfertilized egg. This response begins as early as 15 seconds after insemination. The level of PI reaches a minimum at 30 seconds post-insemination, and returns to the original value between 2 and 5 min later. Pulse labelling studies with 32PO4 and [3H]-inositol showed that the incorporation of these two isotopes into 1-(3-sn-phosphatidyl)-L-myo-inositol 4,5-biphosphate [PtdIns(4,5)P2] increased as much as 50% within one minute after insemination. This suggests that at least part of the reduction in PI levels represents the phosphorylation of PI to form PtdIns(4,5)P2. We also found that the production of [3H]-labelled 1D-myoinositol 1,4,5 triphosphate [Ins(1,4,5)P3] present in the trichloroacetic acid (TCA) soluble fraction of eggs increased over five-fold during the first 10 min post insemination. The temporal correlation between the early burst of PtdIns(4,5)P2 and Ins(1,4,5)P3 formation and the transient increase in intracellular free calcium known to occur in the fertilized egg suggest that the production of PtdIns(4,5)P2 and ultimately Ins(1,4,5)P3 may be associated with calcium mobilization within the egg.