The aim of this research study, develop electrospun nano-bio membrane (ENBM) from Poly (vinyl) alcohol (PVA), nano-Demineralized Bone Matrix (nano-DBM) and to impart additional strength, incorporation of carbon nanoparticles (CNP) by elctrospinning machine for their potential application in bone tissue regeneration. The effective of ENBM for biocompatibility properties were evaluated using MG 63 osteoblast cell line, which showed 100% biocompatibility and more viable cell present in the electrospun nano-biomembrane. In vitro biomineralization, more apatite formation was observed SEM images. Electrospun nanoscaffold prepared using PVA, nano-DBM and CNP (0.6%) possessed improved mechanical properties viz. 14.58 +/- 0.13 Mpa of tensile strength, 13.87 +/- 0.05% of elongation at break and 36.84 +/- 0.11% water absorption. This research study proved the bone formation of these ENBM in bone tissue regeneration and regenerative medicine.