Plasma electrolytic oxidation (PEO) is a relatively new surface modification process that may be used as an alternative to plasma spraying methods to confer bioactivity to Ti alloy implants. The aim of this study was to compare physical, chemical and biological surface characteristics of two coatings applied by PEO processes, containing different calcium phosphate (CaP) and titanium dioxide phases, with a plasma-sprayed hydroxyapatite (HA) coating. Coating characteristics were examined by X-ray diffraction, energy dispersive X-ray spectroscopy, scanning electron microscopy, surface profilometry, and wettability tests. The biological properties were determined using the human osteoblastic cell line MG-63 to assess cell viability, calcium and collagen synthesis. The tests showed that PEO coatings are significantly more hydrophilic (6%) and have 78% lower surface roughness (Ra) than the plasma-sprayed coatings. Cell behavior was demonstrated to be strongly dependent on the phase composition and surface distribution of elements in the PEO coating. MG-63 viability for the TiO2-based PEO coating containing amorphous CaPs was significantly lower than that for the PEO coating containing crystalline HA and the plasma-sprayed coating. However, collagen synthesis on both the CaP and the TiO2 PEO coatings was significantly higher (92% and 71%, respectively) than on the plasma-sprayed coating after 14 days. PEO has been demonstrated to be a promising method for coating of orthopedic implant surfaces.