Abstract: In order to reduce the use of fossil fuels and to solve environmental problems, the utilization of the renewable energy source has great significance. As an energy carrier, hydrogen is considered to be one of the most promising substitutes of fossil fuel in the future. The two-stage process involving biomass pyrolysis and bio-oil catalytic steam reforming is an important and economical technology for hydrogen production. Ni catalysts have been used extensively for bio-oil steam reforming process because of their high catalytic activity, along with low prices. However, the limitation of nickel catalysts is the carbon formation on the catalyst surface. Hence, it is theoretical and practical significance to prepare an cheap catalyst with high hydrogen yield and low char yield. In this paper, the catalyst carriers were natural montmorillonite (Mmt) and sepiolite (Spt). The Ni/Mmt-Spt catalyst was prepared by alkali-treated montmorillonite-sepiolite (Mmt-Spt) as the support and nickel as the active ingredient. The impact of Ni/Mmt-Spt for hydrogen production from bio-oil steam reforming was inspected. The specific surface area analysis (BET) results showed that the Ni/Mmt-Spt catalyst had a large specific surface area and the scanning electron microscope (SEM) results showed that Ni/Mmt-Spt catalyst retained the fibrous structure after the reaction. The NH₃-temperature programmed desorption (NH₃-TPD) results showed that the acidic center of the alkali-treated Ni/Mmt-Spt catalyst reduced. The X-ray diffraction (XRD) results showed the particle size of NiO on Ni/Mmt-Spt catalyst was small. The presence of Mg and Ca in the carrier can promote the water gas shift reaction and can help to eliminate carbon deposition to reduce the char yield. The reaction conditions for the steam reforming of bio-oil with the Ni/Mmt-Spt catalyst were optimized as the follows: temperature of 900 ℃, water/carbon mass ratio of 6 and weight hourly space velocity (WHSV) of 2 h⁻¹. The hydrogen yield of 81.5%, potential hydrogen yield of 90.4% and char yield of 3.8% can be obtained at the optimum conditions. The hydrogen yield of Ni/Mmt-Spt catalyst was 7.1% higher than that of Ni/Al₂O₃ catalyst. While the char yield of Ni/Mmt-Spt catalyst was 1.8% lower than that of Ni/Al₂O₃ catalyst.