In the present batch study, eucalyptus leaves (EUL), H2SO4-treated eucalyptus leaves (SEUL), and H3PO4-treated eucalyptus leaves (PEUL) are used as bio-adsorbents for the removal of methylene blue (MB). The bio-adsorption is executed to inspect the results of the variation between different experimental variables such as pH (2–10), adsorbent dose (1–10 g/L), contact time (5–360 min), and temperature (298–318 K) on the bio-adsorption of MB. The Langmuir isotherm (R2 = 0.99) fitted adequately to the bio-adsorption data for the initial MB concentrations of 10–300 mg/L. It is also necessary to mention that the MB bio-adsorption occurred in the order of a monolayer on the EUL, SEUL, and PEUL. The bio-adsorption kinetics have been fitted by the pseudo-second-order model (R2 ≥ 0.99) for various MB concentrations. The maximum bio-adsorption capacity was 194.34 mg/g and was achieved for the H3PO4-treated eucalyptus leaves (PEUL). These results showed that EUL, SEUL, and PEUL may be utilized as a favourable low-cost bio-adsorbent to eliminate MB from aqueous solutions. With safe disposal methods in mind, this investigation has revealed the eco-friendliness of the bio-adsorbents. A prediction of the removal percentage of methylene blue using a genetic algorithm (GA) from the data collected from the experiment has also been tested. The results related to the prediction using the GA-ANN are accurate. © 2019 Canadian Society for Chemical Engineering