The analytical method for 4-{[(6-methoxy-1,3-benzothiazol-2-yl)imino]methyl}benzene-1,4-diol (MBTIMB) was developed and validated using reverse-phase high-performance liquid chromatography (RP-HPLC). The determination was carried out on a HYPERSIL column with a mobile phase consisting of acetonitrile and water in an 80:20v/v ratio, flowing at a rate of 0.5 ml/min, and UV detection at 254nm. The retention time for 4-{[(6-methoxy-1,3-benzothiazol-2-yl)imino]methyl}benzene-1,4-diol was measured at 8.142 minutes. The method exhibited a linear response for 4-{[(6-methoxy-1,3-benzothiazol-2-yl)imino]methyl}benzene-1,4-diol in the concentration range of 4-24 ppm, with a high correlation coefficient (r-value) of 0.9993. Validation of the developed method included assessments of precision, accuracy, linearity, selectivity, range, and force degradation study. The results indicated that the method was specific, accurate, linear, and precise. The relative standard deviations (RSD) for injector repeatability and inter-assay precision were found to be greater than 2%. The percentage recoveries for 4-{[(6-methoxy-1,3-benzothiazole-2-yl)imino]methyl}benzene-1,4-diol ranged from 98.87% to 100.65%, with an overall mean recovery of 100.02%