Abstract: In a Tris-HCl buffer medium of pH 5, an ion-association complex was formed by the reaction of bromocresol green and spectinomycin (SPM), having its positive and negative absorption maxima at the wavelengths of 580 nm and 449 nm respectively, with apparent molar absorptivities of (ε580) 3.41×104L·mol-1·cm-1 and (ε449) 2.86×104L·mol-1·cm-1. Beer′s law was obeyed in the ranges of 0.6-9.2 mg·L-1 (measured at 580 nm) and 0.6-11.1 mg·L-1 (measured at 449 nm). When dual wavelengths spectrophotometry was applied and absorbance was measured by the double absorption peak method, value of apparent molar absorptivity was increased to 6.26×104L·mol-1·cm-1. The proposed method was applied to the analysis of a sample of injection of SPM, giving values of SPM content in consistency with the labelled value.