In this investigation, a novel application of Aliquat 336 (tricaprylmethylammonium chloride)-impregnated chitosan (AICS) for the separation and purification of 99Mo from some fission products, such as 137Cs, 85Sr and 131I, in alkaline solution is presented. Before impregnation, pristine chitosan experienced no adsorption affinity for Mo. However, this situation dramatically changed after the impregnation. The structure of AICS was elucidated by FTIR, SEM and EDX spectra. The influence of contact time, solution pH, weight of AICS, initial Mo concentration and temperature on the adsorption process was studied. Kinetic studies revealed that the rate of adsorption was impressively very fast and only 3 min were sufficient to reach equilibrium. The pH influence showed that Mo could be effectively adsorbed over a wide range of pH 3–11. The equilibrium data fitting to isotherms models followed the order Langmuir > Freundlich > Dubinin-Radushkevich. Based on the Langmuir model, the maximum adsorption capacity was computed at 60.1 mg/L. Thermodynamic studies indicated that the adsorption process is spontaneous and endothermic in nature. Finally, a pure solution of 99Mo with a purity >99 % was obtained from a real sample. The data obtained confirmed that AICS is a promising candidate for separation and purification of 99Mo from alkaline media.