Electron spin resonance (ESR) measurements were performed on highly oriented pyrolytic graphite (HOPG) samples irradiated with highly charged ions (HCIs). The interaction between a HCI and surfaces results in emission of photons in the range of visible to X-ray, hundreds of secondary electrons, sputtering of secondary ions and modification of surface structure in nanometer scale. In the present experiments, HCIs were produced by electron beam ion source (EBIS) and Ar 8+ and Ar 14+ were used for the irradiation. ESR measurement provides information on unpaired electrons of the sample. We investigated the dependence of defect formation on charge state and fluence of incident HCIs using ESR. The L1 line appeared in HOPG samples irradiated with HCIs at the low temperature region, and the intensity became larger at higher charge state and higher fluence.