The agar diffustion method using Escherichia coli was investigated for determination of Vitamin $B_{l2}$ instead of turbidimetric method using Lactobacillus leichannii (USP XXII method). The turbidimetric method is difficult to control the test organism and it has complicated procedure. From this study, it was found that the agar diffusion method on the determination of Vitamin $B_{12}$ in pharamaceutical preparation is simple and convenient as compared with turbidimetric method. Also we found that the coefficient of variation in reproducibility and the standard error in recovery were 2.18% and 1.83%, respectively.