Recently, microalgae have tended to be used as a biological treatment for wastewater decontamination. The present study aimed to investigate the Cr(VI) removal using the freshwater microalgae ‘Craticula subminuscula’ and their biobased adsorbant, isolated from a Moroccan river in the High Atlas Mountain. The optimum operational conditions for maximum Cr(VI) biosorption by the biobased adsorbent form (95.32%) were determined at (pH = 1.09, adsorbent dose = 10.91 mg L−1, and treatment duration = 129.47 min) using response surface methodology (RSM). Under those optimal conditions, the biosorption process of Cr(VI) by C. subminuscula is endothermic, spontaneous and follows Langmuir and a pseudo-second-order model with a constant rate; the theoretical and experimental biosorption capacity of 0.0004 g/mg/min was 289.01 mg g−1 and 277.57 mg g−1, respectively. Fourier Transform Infrared Spectroscopy (FTIR) analyses of the biomass and scanning electron microscopy (SEM) revealed that the principal mechanism to remove Cr(VI) by C. subminuscula was the affinity of Cr(VI) by the cell walls of microalgae. Thus, the positive results of desorption cycles promise increased potential utilization of these algae in continuous systems within industrial processes. The findings contribute valuable insights into the effectiveness of C. subminuscula as a biobased remediation agent for Cr(VI) in wastewater treatment.