The objectives of this study were to study the removal mechanism of Cr(VI) by natural olive stone (OS) and to present a sequential-batch process for the removal of total chromium (original Cr(VI) and Cr(III) derived from reduction of Cr(VI) during biosorption at acidic conditions). First, experiments were conducted varying pH from 1 to 4, and showed that a combined effect of biosorption and reduction is involved in the Cr(VI) removal. Then, X-ray photoelectron spectroscopy and desorption tests were employed to verify the oxidation state of the chromium bound to OS and to elucidate the removal mechanism of Cr(VI) by this material. The goal of these tests was to confirm that Cr(III) is the species mainly absorbed by OS. Finally, the possibility of total chromium removal by biosorption in a sequential-batch process was analyzed. In the first stage, 96.38% of Cr(VI) is removed by OS and reduced to Cr(III). In the second stage, approximately 31% of the total Cr concentration was removed. However, the Cr(III) released in the first stage is not completely removed, and it could suggest that the Cr(III) could be in a hydrated compound or a complex, which could be more difficult to remove under these conditions.