Bismuth borate glasses with composition (100-x)B2O3 – x (Bi2O3) (where x is in mol.% ranging from 25 -50 in steps of 5) have been prepared by using melt quenching technique. Positron annihilation lifetime (PAL), Fourier transform infra-red (FT-IR) and X-ray diffraction (XRD) measurements have been performed to study the amorphous and defect structure as well as the vibrating band structure. PAL results have revealed weak formation of ortho-positronium (o-Ps) at amorphous sites. At low concentration, the pore size at these sites increases from 2.5 nm to 3.5 nm with increase of Bi2O3 content, then show weak decrease to 3nm at higher concentration showing also a reduction in the relative pore fraction from 9.8% to 1% in agreement with XRD spectra, which indicate the presence of some peaks associated with Bi2O3 phase. On the other hand, the variation of the annihilation parameters at defects, suggest strong trapping of the free positron at non-bridging oxygen (NBO) bonds, which have been determined by FT-IR. In addition, IR spectra revealed the presence of some bonds that are assigned to vibrations of Bi-O bonds from [BiO3] pyramidal and [BiO6] octahedral units and B-O bonds from [BO3] and [BO4] units.