A single crystal of Bi_1.94Pb_0.06V_0.9Co_0.1O_5.32, in which the Bi and V sites of layer-structured Bi₂VO_5.5 were substituted simultaneously, was prepared for the first time and the effects of the Pb substitution on the conductivities in a(b)- and c-axis directions were investigated. Any improvements of ionic conductivity along c-axis perpendicular to the (Bi₂O₂)²⁺ layer was not observed compared with that in single crystal without Pb substitution. From the results of conductivity and Rietveld refinement of XRD pattern, oxygen vacancies generated by Pb substitution were suggested to locate in the pseudo-perovskite block, not in the (Bi₂O₂)²⁺ layer.