Zeta potential of liposomes is often measured in studies of their properties and/or applications. However, there are not many papers published in which zeta potential was determined during enzymatic reaction of a lipid. Therefore in this paper size, polydispersity index, and zeta potentials of 1,2-dipalmitoylsn-glycero-3-phosphatidylcholine (DPPC) and 1,2-dioleoyl-sn-glycero-3-phosphatidylcholine (DOPC) liposomes were investigated in 1 mM NaCl (pH 6.2) and phosphate buffer (pH 8.1) during 60 min of phospholipase C action at 20 and 37 °C. The hydrocarbon chains saturation differ these two phospholipids what appears in some differences in the zeta potential changes during the hydrolysis reaction run. The polydispersity index of the liposomes indicates that they are relatively stable and monodisperse, except for DPPC in phosphate buffer where up to 30 % of the initial amount forms larger size moieties, possibly aggregates of the liposomes, enzyme and the hydrolysis products. However, in this buffer zeta potential of the liposomes practically does not change during PLC action. Also the changes of zeta potential of DOPC liposomes are minor, although their negative values are much smaller than those of DPPC at both temperatures. These small changes of the potential may be due to compression of the diffuse double layer by present phosphate buffer. However, distinct changes of the zeta potentials in the presence of PLC take place in NaCl solution. The observed changes can be explained by reorientation of the phospholipid polar heads and their different density on DPPC and DOPC surface of the liposomes. Although it appeared that the zeta potential is not a very sensitive parameter for tracking the hydrolysis reaction in phosphate buffer, generally zeta potential enables the characterization of such reactions through determination of electrokinetic properties of liposomes as well as the polydispersity and size distribution of the liposomes do.