This work describes a novel technique for the direct quantification of Acid Green 16 (AG16) adsorbed on a molecularly imprinted polymer (MIP) through the application of diffuse reflectance spectrophotometry (DRS) directly in a solid material. The MIP was synthesized by a bulk method using 1-vinylimidazole as the functional monomer. To conduct DRS analysis, adsorption assays were performed through the application of the MIP in a solution containing different concentrations of AG16 for 120 minutes; subsequently, the MIP was left to dry and a certain quantity of the polymer was analyzed. Under optimized conditions, a linear concentration range of 1.0 μmol L-1 to 10.0 μmol L-1 and limits of detection and quantification of 0.3 μmol L-1 and 1.0 μmol L-1, respectively, were obtained. The repeatability and reproducibility of the method were evaluated and RSD values lower than 4% were obtained. Selectivity studies allowed finding imprinting factor values of 1.9, 2.6, 1.1, and 1.1 for AG16, Direct yellow 50, Acid Blue 1, and Brilliant Green, respectively. The proposed method was applied toward the analysis of river water and textile industry effluents. The advantage and novelty of the technique lie in the fact that the amount of the analyte adsorbed on the selective polymer is directly measured on the solid material for AG16 and not indirectly via the remaining solution as it has always been carried out in previous studies reported in the literature. The findings show that the proposed technique is relatively simple, novel and highly versatile for the quantification of analytes adsorbed on MIPs, as well as for the analysis of the material of interest and quantification of diverse analytes in different matrices.